Antimicrobial Therapy in Veterinary Medicine [6 ed.] 1119654599, 9781119654599

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Chapter 15 Chloramphenicol, Thiamphenicol, and Florfenicol
References

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Antimicrobial Therapy in Veterinary Medicine

Sixth Edition

Edited by Patricia M. Dowling, DVM, MSc, DACVIM, DACVCP Western College of Veterinary Medicine University of Saskatchewan

John F. Prescott, MA, VetMB, PhD, FCAHS Ontario Veterinary College University of Guelph

Keith E. Baptiste, BVMS, MSc, PhD, MRCVS, DACVIM, DECEIM Danish Medicines Agency (Lægemiddelstyrelsen)

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Antimicrobial Therapy in Veterinary Medicine

Published by John Wiley & Sons, Inc., Hoboken, New Jersey. Published simultaneously in Canada. No part of this publication may be reproduced, stored in a retrieval system, or transmitted in any form or by any means, electronic, mechanical, photocopying, recording, scanning, or otherwise, except as permitted under Section 107 or 108 of the 1976 United States Copyright Act, without either the prior written permission of the Publisher, or authorization through payment of the appropriate per-­copy fee to the Copyright Clearance Center, Inc., 222 Rosewood Drive, Danvers, MA 01923, (978) 750-­8400, fax (978) 750-­4470, or on the web at www.copyright. com. Requests to the Publisher for permission should be addressed to the Permissions Department, John Wiley & Sons, Inc., 111 River Street, Hoboken, NJ 07030, (201) 748-­6011, fax (201) 748-­6008, or online at http://www.wiley. com/go/permission. Trademarks: Wiley and the Wiley logo are trademarks or registered trademarks of John Wiley & Sons, Inc. and/ or its affiliates in the United States and other countries and may not be used without written permission. All other trademarks are the property of their respective owners. John Wiley & Sons, Inc. is not associated with any product or vendor mentioned in this book. Limit of Liability/Disclaimer of Warranty: While the publisher and author have used their best efforts in preparing this book, they make no representations or warranties with respect to the accuracy or completeness of the contents of this book and specifically disclaim any implied warranties of merchantability or fitness for a particular purpose. No warranty may be created or extended by sales representatives or written sales materials. The advice and strategies contained herein may not be suitable for your situation. You should consult with a professional where appropriate. Further, readers should be aware that websites listed in this work may have changed or disappeared between when this work was written and when it is read. Neither the publisher nor authors shall be liable for any loss of profit or any other commercial damages, including but not limited to special, incidental, consequential, or other damages. For general information on our other products and services or for technical support, please contact our Customer Care Department within the United States at (800) 762–­2974, outside the United States at (317) 572–­3993 or fax (317) 572–­4002. Wiley also publishes its books in a variety of electronic formats. Some content that appears in print may not be available in electronic formats. For more information about Wiley products, visit our web site at www.wiley.com. Library of Congress Cataloging-­in-­Publication Data Applied for: Hardback ISBN: 9781119654599 Cover Design: Wiley Cover Images: World Map: © teekid/Getty Images; Farm Animals: © Vectorig/Getty Images; Livestock: © mollyw/ Shutterstock; Fish: © RobinOlimb/Getty Images; Reptiles: © RobinOlimb/Getty Images; Insects: © Kristtaps/Getty Images; Zoo Animals: © JulieWeiss/Getty Images; Medicine pills: © Alexander Raths/Shutterstock Set in 9.5/12.5pt STIXTwoText by Straive, Pondicherry, India

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Copyright © 2025 by John Wiley & Sons, Inc. All rights reserved, including rights for text and data mining and training of artificial intelligence technologies or similar technologies.

Contents Contributors  ix Preface  xiii Acknowledgments  xv Important Notice  xvii List of Abbreviations  xix Section I  General Principles of Antimicrobial Therapy  1 1 Antimicrobial Drug Action and Interaction: An Introduction  3 John F. Prescott 2 Antimicrobial Susceptibility Testing Methods and Interpretation of Results  13 Joseph E. Rubin and Peter Damborg 3 Antimicrobial Resistance and Its Epidemiology  29 Marisa Haenni and Patrick Boerlin 4 Pharmacokinetics of Antimicrobials  51 Patricia M. Dowling 5 Pharmacodynamics of Antimicrobials  81 Andrew P. Woodward and Ted Whittem 6 Principles of Antimicrobial Drug Selection and Use  109 J. Scott Weese and Patricia M. Dowling Section II  Classes of Antimicrobial Agents  119 7 Beta-­lactam Antibiotics: Penam Penicillins  121 Laura Y. Hardefeldt and John F. Prescott 8 Beta-­lactam Antibiotics: Cephalosporins  143 Laura Y. Hardefeldt and John F. Prescott

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Contents

9 Other Beta-­lactam Antibiotics: Beta-­lactamase Inhibitors, Carbapenems, and Monobactams  169 John F. Prescott and Laura Y. Hardefeldt 10 Peptide Antibiotics: Polymyxins, Glycopeptides, Bacitracin, and Fosfomycin  187 Patricia M. Dowling 11 Lincosamides, Pleuromutilins, and Streptogramins  203 Grazieli Maboni and Leticia Trevisan Gressler 12 Macrolides, Azalides, and Ketolides  223 John F. Prescott and Keith E. Baptiste 13 Aminoglycosides and Aminocyclitols  249 Patricia M. Dowling 14 Tetracyclines  273 Ronan J. J. Chapuis and Joe S. Smith 15 Chloramphenicol, Thiamphenicol, and Florfenicol  291 Patricia M. Dowling and Hélène Lardé 16 Sulfonamides, Diaminopyrimidines, and Their Combinations  305 Jennifer M. Reinhart and John F. Prescott 17 Fluoroquinolones  325 Patricia M. Dowling 18 Miscellaneous Antimicrobials: Ionophores, Nitrofurans, Nitroimidazoles, Rifamycins, and Others  345 Patricia M. Dowling and Keith E. Baptiste 19 Antifungal Chemotherapy  371 Keith E. Baptiste Section III  Antimicrobial Stewardship  401 20 General Concepts in Antimicrobial Stewardship  403 Laura Y. Hardefeldt, J. Scott Weese, and Stephen W. Page 21 Global Aspects of One Health Antimicrobial Stewardship  425 Jeffrey T. LeJeune, Mark E. Caudell, Suzanne N. Eckford, and Stephen W. Page 22 Antimicrobial Stewardship in Companion Animals  445 Peter Damborg, J. Scott Weese, and John F. Prescott

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23 Antimicrobial Stewardship in Food-producing Animals  459 David C. Speksnijder, Stephen W. Page, Jaap A. Wagenaar, and John F. Prescott 24 Antimicrobial Prophylaxis, Metaphylaxis, and the Treatment of Immunocompromised Patients  487 Diego E. Gomez and J. Scott Weese 25 Regulation of Antimicrobial Use in Animals  513 Alan Chicoine and Keith E. Baptiste 26 Antimicrobial Drug Residues in Foods of Animal Origin  527 Joe S. Smith and Patricia M. Dowling

Section IV  Antimicrobial Therapy in Selected Animal Species  545 27 Antimicrobial Drug Use in Horses  547 Keith E. Baptiste 28 Antimicrobial Therapy in Dogs and Cats  577 Jane E. Sykes and Peter Damborg 29 Antimicrobial Therapy in Beef Cattle  611 Michael D. Apley, Brian V. Lubbers, and Nora D. Schrag 30 Antimicrobial Therapy in Dairy Cattle  635 Sarah Wagner and Sarah Depenbrock 31 Antimicrobial Therapy in Sheep and Goats  655 Joe S. Smith and Amanda J. Kreuder 32 Antimicrobial Therapy in New World Camelids  671 Joe S. Smith 33 Antimicrobial Therapy in Swine  685 Ken Steen Pedersen 34 Antimicrobial Therapy in Poultry  697 Jenny A. Nicholds, David French, Daniel Parker, and Peter O’Kane 35 Antimicrobial Therapy in Companion Birds  721 Marike Visser 36 Antimicrobial Therapy in Rabbits, Rodents, and Ferrets  735 Colette L. Wheler and Patricia M. Dowling

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Contents

Contents

37 Antimicrobial Therapy in Reptiles  767 J. Scott Weese 38 Antimicrobial Therapy in Zoo and Wildlife Species  791 Ellen Wiedner and Robert P. Hunter 39 Antimicrobial Therapy in Aquaculture  803 Patrick Whittaker, Timothy S. Kniffen, and Simon Otto 40 Antimicrobial Therapy in Honey Bees  819 Elemir Simko, Sarah Wood, and Ivanna V. Kozii Index  831

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C ­ ontributors­ Chapter numbers are in parentheses. Michael D. Apley (29) Frick Professor of Clinical Sciences Department of Clinical Sciences Kansas State University College of Veterinary Medicine Manhattan, Kansas, USA

Alan Chicoine (25) Assistant Professor Veterinary Biomedical Sciences Western College of Veterinary Medicine University of Saskatchewan Saskatoon, Saskatchewan, Canada

Keith E. Baptiste (12, 18, 19, 25, 27) Specialist Veterinary Consultant Department of Veterinary Medicine Danish Medicines Agency (Lægemiddelstyrelsen) Copenhagen South, Denmark

Peter Damborg (2, 22, 28) Associate Professor Department of Veterinary and Animal Sciences Faculty of Health and Medical Sciences University of Copenhagen Copenhagen, Denmark

Patrick Boerlin (3) Associate Professor Department of Pathobiology Ontario Veterinary College University of Guelph Guelph, Ontario, Canada Mark E. Caudell (21) Food and Agricultural Organization of the United Nations Gigiri, Nairobi, Kenya Ronan J.J. Chapuis (14) Assistant Professor Biomedical Sciences Ross University School of Veterinary Medicine Basseterre, St Kitts, West Indies

Sarah Depenbrock (30) Assistant Professor Medicine & Epidemiology School of Veterinary Medicine Davis, California, USA Patricia M. Dowling (4, 6, 10, 13, 15, 17, 18, 26, 36) Professor Veterinary Biomedical Sciences Western College of Veterinary Medicine University of Saskatchewan Saskatoon, Saskatchewan, Canada Suzanne N. Eckford (21) Head, International Office Veterinary Medicines Directorate Addlestone, Surrey, United Kingdom

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ix

­Contributors

David French (34) Clinical Associate Professor Department of Population Health Poultry Diagnostic and Research Center Athens, Georgia, USA

Ivanna V. Kozii (40) Department of Veterinary Pathology Western College of Veterinary Medicine University of Saskatchewan Saskatoon, Saskatchewan, Canada

Diego E. Gomez (24) Assistant Professor Department of Clinical Studies Ontario Veterinary College University of Guelph Guelph, Ontario, Canada

Amanda J. Kreuder (31) Assistant Professor Veterinary Microbiology and Preventive Medicine College of Veterinary Medicine Iowa State University Ames, Iowa, USA

Leticia Trevisan Gressler (11) Adjunct Professor Laboratory of Veterinary Microbiology and Immunology Instituto Federal Farroupilha Frederico Westphalen, RS, Brazil Marisa Haenni (3) Deputy Head, Antibioresistance et Virulence Bactériennes Unit Agence Nationale de Sécurité Sanitaire de l’Alimentation, de l’Environnement et du Travail Maisons-­Alfort, France Laura Y. Hardefeldt (7, 8, 9, 20) National Centre for Antimicrobial Stewardship Faculty of Veterinary and Agricultural Sciences Asia-­Pacific Centre for Animal Health University of Melbourne Melbourne, Victoria, Australia

Hélène Lardé (15) Assistant Professor Veterinary Biomedical Sciences Faculté de médecine Vétérinaire Université de Montréal Rimouski, Quebec, Canada Jeffrey T. LeJeune (21) Food Safety and Quality Officer Food and Agriculture Organization of the United Nations Rome, Italy Brian V. Lubbers (29) Associate Professor Food Animal Therapeutics College of Veterinary Medicine Kansas State University Manhattan, Kansas, USA

Robert P. Hunter (38) Veterinary Pharmacologist One Medicine Consulting Olathe, Kansas, USA

Grazieli Maboni (11) Assistant Professor Department of Infectious Diseases College of Veterinary Medicine University of Georgia Athens, Georgia

Timothy S. Kniffen (39) Technical Services Veterinarian Global Aquaculture Marketing Merck Animal Health De Soto, Kansas, USA

Jenny A. Nicholds (34) Clinical Associate Professor, Avian Medicine Department of Population Health Poultry Diagnostic and Research Center Athens, Georgia, USA

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Peter O’Kane (34) Senior Associate Veterinarian Slate Hall Veterinary Services Metheringham, Lincolnshire, United Kingdom Simon Otto (39) Associate Professor School of Public Health University of Alberta Edmonton, Alberta, Canada Stephen W. Page (20, 21, 23) Director Advanced Veterinary Therapeutics Newtown, New South Wales, Australia Daniel Parker (34) Director Emeritus Slate Hall Veterinary Services Metheringham, Lincolnshire, United Kingdom Ken Steen Pedersen (33) Professor of Herd Diagnostics and Antibiotic Use in Pigs Faculty of Health and Medical Sciences University of Copenhagen Frederiksberg C, Denmark John F. Prescott (1, 7, 8, 9, 12, 16, 22, 23) University Professor Emeritus Department of Pathobiology Ontario Veterinary College University of Guelph Guelph, Ontario, Canada Jennifer M. Reinhart (16) Assistant Professor Small Animal Internal Medicine Department of Veterinary Clinical Medicine College of Veterinary Medicine University of Illinois Urbana-­Champaign, Illinois, USA

Joseph E. Rubin (2) Professor Department of Veterinary Microbiology Western College of Veterinary Medicine University of Saskatchewan Saskatoon, Saskatchewan, Canada Nora D. Schrag (29) Veterinary Consultant Livestock Veterinary Resources, LLC Olsburg, Kansas, USA Elemir Simko (40) Professor Department of Veterinary Pathology Western College of Veterinary Medicine University of Saskatchewan Saskatoon, Saskatchewan, Canada Joe S. Smith (14, 26, 31, 32) Assistant Professor Large Animal Clinical Sciences College of Veterinary Medicine University of Tennessee Knoxville, Tennessee, USA David C. Speksnijder (23) Assistant Professor Faculty of Veterinary Medicine, Department Biomolecular Health Sciences, Division of Infectious Diseases and Immunology Utrecht University Utrecht, The Netherlands Jane E. Sykes (28) Professor Department of Medicine & Epidemiology University of California, Davis Davis, California, USA Marike Visser (35) Veterinary Clinical Research Manager Zoetis Inc. Kalamazoo, Michigan, USA

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­Contributors

­Contributors

Jaap A. Wagenaar (23) Department of Infectious Diseases and Immunology Faculty of Veterinary Medicine University of Utrecht Utrecht, The Netherlands Sarah Wagner (30) Professor of Pharmacology Texas Tech School of Veterinary Medicine Amarillo, Texas, USA J. Scott Weese (6, 20, 22, 24, 37) Professor Department of Pathobiology Ontario Veterinary College University of Guelph Guelph, Ontario, Canada Colette L. Wheler (36) Clinical Research Veterinarian Vaccine and Infectious Disease Organization University of Saskatchewan Saskatoon, Saskatchewan, Canada Patrick Whittaker (39) Head Veterinarian Grieg Seafood BC Ltd. Campbell River, British Columbia, Canada

Ted Whittem (5) Professor and Dean College of Public Health, Medical and Veterinary Sciences Townsville, North Queensland, Australia Ellen Wiedner (38) Hyrax Consulting, LLC. Durango, Colorado, USA Sarah Wood (40) WCVM Research Chair in Pollinator Health/ Associate Professor Western College of Veterinary Medicine University of Saskatchewan Saskatoon, Saskatchewan, Canada Andrew P. Woodward (5) Applied Biostatistics Researcher Faculty of Health University of Canberra Canberra, Australian Capital Territory, Australia

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xii

Preface The first edition of Antimicrobial Therapy in Veterinary Medicine was published in 1988 and was followed by four subsequent editions as the knowledge and practice of evidence-­ based animal treatment evolved. The increasing impact of antimicrobial resistance on human and animal health prompted the need for a sixth edition with an increased global focus on all the earlier topics covered in this book but with the addition of three chapters focusing specifically on antimicrobial stewardship concepts in different types or spheres of veterinary practice at both national and international level. To achieve this global perspective, we’ve assembled a group of ­editors and chapter authors with strong ­international experience. This enhances the traditional goal of this ­textbook  –­ that practicing veterinarians, veterinary specialists, drug regulators, educators, and students find it to be an indispensable reference on the general principles and clinical applications of all types of antimicrobials used in veterinary medicine. The book is divided into four sections. The first provides general principles of antimicrobial therapy including susceptibility testing, antimicrobial resistance, and antimicrobial pharmacokinetics and pharmacodynamics. The second section describes each class of antimicrobial agents, revised to include the most up-­to-­date information on the drugs

used in veterinary medicine. The third section deals with the increasingly critical topic of ­antimicrobial stewardship. As part of this ­section, the regulation of antimicrobials in ­animals and antimicrobial drug residues in foods of animal origin is reviewed from a global perspective. The final section addresses the specific principles of antimicrobial therapy in major veterinary species. A chapter on antimicrobial therapy in bees has been added to this edition to reflect the increase in veterinary involvement in the health of bees. Before beginning this new edition, we were greatly saddened by the untimely death of Dr  Steeve Giguère, the editor in chief of the fifth edition. It is our goal to maintain the high standard of scholarship set by Steeve so we have added Dr Keith E. Baptiste to the editorial team. We also mourn the death in 2016 of Dr  J. Desmond Baggot, the distinguished ­veterinary pharmacologist and one of the founders of this book. We are grateful to all the contributors for the care and effort they have put into their chapters. We thank the staff of Wiley Blackwell Publishing, particularly Merryl Le Roux, for their help, patience, and support of this book. We encourage ­readers to send us comments or suggestions for improvements so that future editions can be improved. Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste

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xiii

Acknowledgments Patricia M. Dowling: I thank my husband, Brian Zwaan, for his support of my work on this text. And I thank the contributors for their work through the global pandemic to support antimicrobial stewardship in veterinary medicine.

John F. Prescott: I thank my wife Cathy Prescott for her unwavering support during the production of the six editions since the ­inception of this book.

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xv

I­ mportant Notice The indications and dosages of all drugs in this book are the recommendations of the authors and do not always agree with those given on package inserts prepared by pharmaceutical manufacturers in different countries. The medications described do not necessarily have the specific approval of national regulatory authorities, for use in the diseases and dosages

recommended. In addition, while every effort has been made to check the contents of this book, errors may have been missed. The ­package insert for each drug product should therefore be consulted for use, route of administration, dosage, and (for food animals) ­withdrawal period, as approved by the reader’s national regulatory authorities.

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xvii

L ­ ist of Abbreviations ADI EDI EFSA EU FAO FDA JECFA LOD LOQ MBC MDR MIC MPC MRL NOAEL USA VDD WHO

acceptable daily intake estimated daily intake European Food Safety Authority European Union Food and Agriculture Organization of the United Nations Food and Drug Administration Joint FAO/WHO Expert Committee on Food Additives limit of detection limit of quantification minimum bactericidal concentration multidrug resistant minimum inhibitory concentration mutant prevention concentration maximum residue limit no-­observed-­adverse-­effect level United States of America Veterinary Drugs Directorate World Health Organization

For dosages: PO IM IV SC SID BID TID QID q 6 h, q 8 h, q 12 h, etc.

per os, oral administration intramuscular administration intravenous administration subcutaneous administration single daily administration twice-­daily administration (every 12 hours) 3 times daily administration (every 8 hours) 4 times daily administration (every 6 hours) Every 6, 8, 12 hours, etc.

For example, a dosage of “10 mg/kg TID IM” means 10 milligrams of the drug per kilogram of body weight, administered every 8 hours intramuscularly.

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xix

Section I

General Principles of Antimicrobial Therapy

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1 Antimicrobial Drug Action and Interaction: An Introduction John F. Prescott

Antimicrobial drugs exploit differences in structure or biochemical function between host and parasite. Modern chemotherapy is traced to Paul Ehrlich, a pupil of Robert Koch, who devoted his career to discovering agents that possessed selective toxicity so that they might act as so-­called “magic bullets” in the fight against infectious diseases. The remarkable efficacy of modern antimicrobial drugs still  retains the sense of the miraculous. Sulfonamides, the first clinically successful broad-­spectrum antibacterial agents, were produced in Germany in 1935. However, it was the discovery of the antimicrobial penicillin, a fungal metabolite, by Fleming in 1929 and its subsequent development by Chain and Florey during World War II that led to the “antibiotic revolution.” Within a few years of the introduction of penicillin, many other antimicrobials were described. This was followed by the development of semisynthetic and synthetic antimicrobial agents which has resulted in an increasingly powerful and effective array of compounds used to treat infectious diseases. The term antibiotic has been defined as a low molecular weight substance produced by a microorganism that at low concentrations inhibits or kills other microorganisms. In contrast, the word antimicrobial has a broader definition than antibiotic and includes any

substance of natural, semisynthetic, or synthetic origin that kills or inhibits the growth of a microorganism but causes little or no damage to the host. Antimicrobial agent and antibiotic are commonly used synonymously. The term antimicrobial is preferentially used in this book as the more encompassing term. The marked structural and biochemical differences between prokaryotic and eukaryotic cells give antimicrobial agents greater opportunities for selective toxicity against bacteria than against other microorganisms such as fungi, which are nucleated like mammalian cells, or viruses, which require their host’s genetic material for replication. Nevertheless, in recent years increasingly effective antifungal and antiviral drugs have been introduced into clinical practice. Important milestones in the development of antibacterial drugs are shown in Figure  1.1. Because of the enormous costs of development, the therapeutic use of these agents in veterinary medicine has usually followed their use  in  human medicine. However, some ­antimicrobials have been developed specifically for animal health and production (e.g., tylosin, tiamulin, tilmicosin, ceftiofur, tulathromycin, gamithromycin, tildipirosin), although all these are related to drug classes used in human medicine. A few classes not used because of toxicity for humans, such as the orthosomycins, have been relegated to oral use

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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Antimicrobial Drug Action and Interaction: An Introduction Human infectious diseases

Antibacterial agents 8 1930

Penicillin discovered

2 4 6 Serious infections respond to sulfonamide

8

Florey demonstrates penicillin's effectiveness

'40

First sulfonamide released

2 4

Streptomycin, first aminoglycoside

6

Chloramphenicol

8

Chlortetracycline, first tetracycline

'50 2 Penicillin-resistant infections become clinically significant

Erythromycin, first macrolide

4 6

Vancomycin

8 '60 2 4 Gentamicin-resistant Pseudomonas and methicillin-resistant staphylococcal infections become clinically significant Beginning in early 1970s, increasing trend of nosocomial infections due to opportunistic pathogens Ampicillin-resistant infections become frequent

6

2 4 6 '80 2 4 6

Expansion of methicillin-resistant staphylococcal infections

Vancomycin-resistant enterococci Multidrug-resistant Mycobacterium tuberculosis Penicillin-resistant Streptococcus pneumoniae Spread of extended-spectrum beta-lactamases among Gram-negatives Multidrug-resistant Pseudomonas, Acinetobacter baumanii, and S. pneumoniae

Gentamicin, antipseudomonal penicillin Ampicillin Cephalothin, first cephalosporin

8 '70

8 AIDS-related bacterial infections

Methicillin, penicillinase-resistant penicillin

8 '90 2

Amikacin, aminoglycoside for gentamicin-resistant strains Carbenicillin, first antipseudomonal beta-lactam Cefoxitin, expanded-spectrum cephalosporin Cefaclor, oral cephalosporin with improved activity Cefotaxime, antipseudomonal cephalosporin Clavulanic-acid-amoxicillin, broad beta-lactamase inhibitor Imipenem-cilastatin Norfloxacin, newer quinolone for urinary tract infections Aztreonam, first monobactam Newer fluoroquinolone for systemic use Improved macrolides

4 6 8 2000 2 4 6 8 10

Oral extended-spectrum cephalosporins Effective antiviral drugs for HIV Quinupristin-dalfopristin Linezolid, first approved oxazolidinone Broader-spectrum fluoroquinolones Telithromycin, first ketolide Tigecycline, first glycylcycline Retapamulin, first pleuromutilin (topical) Doripenem Telavancin, semi-synthetic derivative of vancomycin Ceftaroline

Figure 1.1  Milestones in human infectious disease and their relationship to development of antimicrobial drugs, 1930–2010, illustrating the relationship between the introduction of an antibacterial drug and the emergence of resistance. Source: Modified and reproduced with permission from Kammer (1982).

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in animals for treatment of enteric ­infections. Figure 1.1 highlights the relationship between antimicrobial use and the ­development of resistance in many target microorganisms.

­ pectrum of Activity S of Antimicrobial Drugs Antimicrobial drugs may be classified in a variety of ways, based on four basic features.

Class of Microorganism Antiviral and antifungal drugs generally are active only against viruses and fungi, respectively. However, some imidazole antifungal agents have activity against staphylococci and nocardioform bacteria. Antibacterial agents can be described as narrow spectrum if they inhibit only Gram-­positive and Gram-­negative bacteria or broad spectrum if they also inhibit a wider range of bacteria such as chlamydia, mycoplasma, and rickettsia (Table 1.1).

Antibacterial Activity Within the class description of antibacterial drug activity, antimicrobial drugs can further also be described as narrow spectrum if they  inhibit only either Gram-­positive or Gram-­negative bacteria and as broad-­spectrum drugs if they inhibit both Gram-­positive and Gram-­negative bacteria. This distinction is often not absolute since, although some agents may be primarily active against Gram-­positive bacteria, they may also inhibit some Gram negatives (Table  1.2). It seems likely that some antimicrobial drugs developed in the future may be narrow spectrum and targeted to particular pathogens, avoiding the considerable “bystander” effect of broad-­spectrum antimicrobials on the nonpathogenic microflora.

Bacteriostatic or Bactericidal Activity The minimum inhibitory concentration (MIC) is the lowest concentration of an antimicrobial agent required to prevent the growth of the  pathogen. In contrast, the minimum

Table 1.1  Spectrum of activity of common antibacterial drugs. Class of Microorganism Drug

Bacteria

Fungi

Mycoplasma

Rickettsia

Chlamydia

Protozoa

Aminoglycosides

+

–­

+

–­

–­

–­

Beta-­lactams

+

–­

–­

–­

–­

–­

Chloramphenicol

+

–­

+

+

+

–­

Fluoroquinolones

+

–­

Glycylcyclines

+

+

+

+

–­

+

+

+

+/–­

Lincosamides

+

–­

+

–­

–­

+/–­

Macrolides

+

–­

+

–­

+

+/–­

Oxazolidinones

+

–­

+

–­

–­

–­

Pleuromutilins

+

–­

+

–­

+

–­

Tetracyclines

+

–­

+

+

+

+/–­

Streptogramins

+

–­

+

–­

+

+/–­

Sulfonamides

+

–­

+

–­

+

+

Trimethoprim

+

–­

–­

–­

–­

+

+/–, activity against some protozoa.

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­Spectrum of Activity of Antimicrobial Drug  5

Antimicrobial Drug Action and Interaction: An Introduction

Table 1.2  Antibacterial activity of selected antibiotics. Aerobic Bacteria

Anaerobic Bacteria

Spectrum

Gram +

Gram –

Gram +

Gram –

Examples

Very broad

+

+

+

+

Carbapenems; chloramphenicol; third-­ generation fluoroquinolones; glycylcyclines

Intermediately broad

+

+

+

(+)

Third-­and fourth-­generation cephalosporins

+

(+)

+

(+)

Second-­generation cephalosporins

(+)

(+)

(+)

(+)

Tetracyclines

+

+/–­

+

(+)

Ampicillin; amoxicillin; first-­generation cephalosporins

+

–­

+

(+)

Penicillin; lincosamides; glycopeptides; streptogramins; oxazolidinones

+

+/–­

+

(+)

Macrolides

+/–­

+

–­

–­

Monobactams; aminoglycosides

(+)

+

–­

–­

Second-­generation fluoroquinolones

(+)

(+)

–­

–­

Trimethoprim-­sulfa

–­

–­

+

+

Nitroimidazoles

+

–­

(+)

(+)

Rifamycin

Narrow

+, excellent activity; (+), moderate activity; +/−, limited activity; −, no or negligible activity.

bactericidal concentration (MBC) is the lowest concentration of an antimicrobial agent required to kill the pathogen. Antimicrobials are usually regarded as bactericidal if the MBC is no more than four times the MIC. This distinction is rarely important for treatment of clinical conditions. Some drugs are routinely bactericidal (e.g., beta-­lactams, aminoglycosides) whereas others are usually bacteriostatic (e.g., chloramphenicol, tetracyclines), but this distinction depends on both the drug concentration at the site of infection and the microorganism involved. For example, benzyl penicillin is bactericidal at usual therapeutic concentrations but bacteriostatic at lower concentrations.

Time-­or Concentration-­dependent Activity Antimicrobial agents are often classified as exerting either time-­dependent or concentration-­dependent activity, depending

on their pharmacodynamic properties. These properties of a drug address the relationship between drug concentration and antimicrobial activity (Chapter  5). Drug pharmacokinetic features, such as serum concentrations over time and area under the serum concentration-­time curve (AUC), when integrated with MIC values, can predict the probability of bacterial eradication and ­clinical success. These pharmacokinetic and pharmacodynamic relationships are also important in preventing the selection and spread of resistant strains. The most significant factor determining the ­efficacy of ­beta-­lactams, some macrolides, tetracyclines, trimethoprim-­sulfonamide combinations, and chloramphenicol is the length of time that serum concentrations exceed the MIC of a given pathogen. Increasing the concentration of the drug several-­fold above the MIC does not significantly increase the rate of microbial killing. Rather, it is the length of  time that bacteria are exposed to

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6

concentrations of these drugs above the MIC that dictates their rate of killing. Optimal dosing of such antimicrobial agents involves frequent administration. Other antimicrobial agents such as the aminoglycosides, fluoroquinolones, and metronidazole exert concentration-­dependent killing characteristics. Their rate of killing increases as the drug concentration increases above the MIC for the pathogen and it is not necessary or even beneficial to maintain drug levels above the MIC between doses. Thus, optimal dosing of aminoglycosides and fluoroquinolones involves administration of high doses at long dosing intervals. Some drugs exert characteristics of both time-­ and concentration-­dependent activity. The best predictor of efficacy for these drugs is the 24-­hour area under the serum concentration-­time curve (AUC)/MIC ratio. Glycopeptides, rifampin, and, in some situations, fluoroquinolones fall within this category (Chapter 5).

­ echanisms of Action M of Antimicrobial Drugs Antibacterial Drugs Figure  1.2 summarizes the diverse sites of action of commonly used antibacterial drugs. Their mechanisms of action fall into four categories: inhibition of cell wall synthesis, damage to cell membrane function, inhibition of nucleic acid synthesis or function, and inhibition of protein synthesis. Antibacterial drugs that affect cell wall synthesis (beta-­lactam antimicrobials, bacitracin, glycopeptides) or inhibit protein synthesis (aminoglycosides, chloramphenicol, lincosamides, glycylcyclines, macrolides, oxazolidinones, streptogramins, pleuromutilins, tetracyclines) are more numerous than those that affect cell membrane function (polymy­ xins) or nucleic acid function (fluoroquinolones, nitroimidazoles, nitrofurans, rifampin). Agents that affect intermediate metabolism

(sulfonamides, trimethoprim) have greater selective toxicity than those that affect nucleic acid synthesis.

Developing New Antibacterial Drugs Infection caused by antimicrobial-­resistant bacteria has been an increasing and rapidly developing problem and has reached a crisis in medicine. The speed with which some bacteria develop resistance considerably outpaces the slow development of new antimicrobial drugs. Since 1980, the number of antimicrobial agents approved for use in people has fallen steadily. What has been approved are variations of existing drugs; no new classes of antimicrobials have been discovered since the 1980s. Several factors contribute to driving large pharmaceutical companies out of the antimicrobial drug market. These include expensive regulatory requirements, the challenges of drug discovery and the high cost of drug development coupled with the low rate of return on investment compared with drugs for the treatment of chronic “life-­style” conditions. This has left limited treatment options for infections caused by methicillin-­ resistant staphylococci and vancomycin-­resistant enterococci. The picture is even bleaker for infections caused by some Gram-­negative bacteria such as Pseudomonas aeruginosa, Acinetobacter baumanii, extended-­spectrum beta-­lactamase (ESBL)-­resistant E. coli, Klebsiella spp., and Enterobacter spp., which are occasionally resistant to all safe antimicrobial agents. Judicious use of the antimicrobials currently available and better infection control practices, discussed in Chapters 20–24, will prolong the effectiveness of the drugs that are currently available. However, even if we improve these practices, resistant bacteria will continue to emerge and to spread, and new drugs will be needed. While improvements in some existing classes of antimicrobial drugs continue to be laboriously made, numerous technological advances and improved understanding of bacterial ­pathogens hold considerable promise for the

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­Mechanisms of Action of Antimicrobial Drug  7

Antimicrobial Drug Action and Interaction: An Introduction Nitroimidazoles, nitrofurans Cell wall Sulfonamides, trimethoprim

Beta-lactam antibiotics, glycopeptides, bacitracin

Cell membrane

Purine synthesis

DNA

Polyenes

Fluoroquinolones Novobiocin

Ribosome

Rifampin

Messenger RNA

New protein

30S

Transfer RNA 50S Amino acids

Tetracyclines, aminoglycosides

Oxazolidinones Lincosamides, macrolides, streptogramins

Chloramphenicol

Figure 1.2  Sites of action of commonly used antibacterial drugs that affect virtually all the important processes in a bacterial cell. Source: Modified and reproduced with permission after Aharonowitz and Cohen (1981).

development of novel antimicrobial drugs. However, such development is challenging and extremely expensive. Novel targets for antimicrobial drugs that have been identified include those involved in essential amino acid biosynthesis, in cell wall lipid biosynthesis, in metal chelator biosynthesis, in quorum sensing, in efflux pumps, and in regulation of gene

expression, among others. The investigation of novel antimicrobial sources has undergone a revival, and many novel antimicrobials have been identified, including numerous peptides. Development of antimicrobial drugs targeting specific pathogens is more straightforward than developing broad-­spectrum compounds and, combined with increasing sensitivity of specific

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8

agent diagnosis, is likely to be an important part of the future of antimicrobial therapy in human medicine. Despite a degree of optimism about the future development of new antimicrobials, the costs are considerable and this and other reasons are likely to preclude veterinary application. Bringing a novel antimicrobial into human clinical use takes an estimated 10–15 years and costs an estimated US$1 billion, with the constant threat of development of resistance among important pathogens, which appear increasingly adept at spreading resistance. Many multinational companies have abandoned the search for new antimicrobials. If candidate drugs found in preclinical development are identified, they are moved into three phases of human clinical trials, the last of which can consume 80% of the total research and development costs, which include the high costs of regulatory approval, all of which have to be recovered. Historically, only about 60% of drugs entering Phase 3 clinical trials are approved. Such expensive antimicrobials will therefore tend to have restricted use as “last resort” drugs, further limiting the return on investment. A record of bankruptcy of companies that have brought novel drugs to market does not inspire private investment. Public–private philanthropic initiatives such as CARB-­X (Combating Antibiotic­resistant Bacteria Biopharmaceutical Accelerator; www.carb-­x.org) have been developed to help support drug discovery. In 2022, there were 62  antibacterial compounds of various types under clinical development for human medicine, with three antibacterials introduced since 2020 (Butler et al., 2023). As a result of funding initiatives, an encouraging increasing number of compounds are entering early Phase 1 evaluation studies. However, the clinical pipelines and recently introduced drugs are insufficient to address the emergence and spread of antimicrobial resistance. Most are direct-­acting small molecules including peptides, but others include bacteriophage-­related or antivirulence products. The development of new antimicrobial drugs is inevitably focused on human rather

than veterinary medicine, but research continues into new antimicrobial drugs targeted to topic use in specific veterinary infections (Greco et al., 2019; Bellavita et al., 2020).

Antifungal Drugs Most currently used systemic antifungal drugs damage cell membrane function by binding ergosterols that are unique to the fungal cell membrane (polyenes, azoles) (Chapter  19). The increase in the number of HIV-­infected individuals and of people undergoing organ or bone marrow transplants has resulted in increased numbers of immunosuppressed individuals in many societies. The susceptibility of these people to fungal infections has renewed interest in the discovery and development of new antifungal agents. The focus of antifungal drug development has shifted to  cell  wall structures unique to fungi (1,3-­beta-­D -­glucan synthase inhibitors, chitin synthase inhibitors, mannoprotein binders).

­ ntimicrobial Drug Combinations: A Synergism, Antagonism, and Indifference In general, the use of combinations should be avoided because the toxicity of the antimicrobials will be at least additive and may be synergistic, because the ready availability of broad-­spectrum bactericidal drugs has made use of combinations largely unnecessary, and because they may be more likely to lead to bacterial superinfection. There are, however, well-­ established circumstances, discussed in Chapter 6, in which combinations of drugs are more effective and often less toxic than drugs administered alone. Knowledge of the different mechanisms of action of antimicrobials provides some ability to predict their interaction when they are used in combination. It was clear from the early days of their use that combinations of

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­Antimicrobial Drug Combinations: Synergism, Antagonism, and Indifferenc  9

Antimicrobial Drug Action and Interaction: An Introduction

antimicrobials might give antagonistic rather than additive or synergistic effects. Concerns regarding combinations include the difficulty in defining synergism and antagonism, particularly their method of determination in vitro; the difficulty of predicting the effect of a combination against a particular organism; and the uncertainty of the clinical relevance of in vitro findings. The clinical use of antimicrobial drug combinations is described in Chapter  6. Antimicrobial combinations are used most frequently to provide broad-­ spectrum empiric coverage in the treatment of patients who are critically ill. With the availability of broad-­spectrum antimicrobial drugs, combinations of different drugs are less commonly used than in the early days of antimicrobial therapy, except for specific purposes. An antimicrobial combination is additive or indifferent if the combined effects of the drugs equal the sum of their independent activities measured separately; synergistic if the combined effects are significantly greater than the independent effects; and antagonistic if the combined effects are significantly less than their independent effects. Synergism and antagonism are not absolute characteristics. Such interactions are often hard to predict, vary with bacterial species and strains, and may occur only over a narrow range of concentrations or ratios of drug components. Because antimicrobial drugs may interact with each other in many ways, it is apparent that no single in  vitro method will detect all such interactions. Although the techniques to quantify and detect interactions are relatively crude, the observed interactions occur clinically. The two methods commonly used, the checkerboard and the killing curve, measure two different effects (growth inhibition and killing, respectively) and have sometimes shown poor clinical and laboratory correlation. In the absence of simple methods for detecting synergism or antagonism, the following general guidelines may be used.

Synergism of Antimicrobial Combinations Antimicrobial combinations are frequently ­synergistic if they involve: (1) sequential inhibition of successive steps in metabolism (e.g., trimethoprim-­sulfonamide); (2) sequential inhibition of cell wall synthesis (e.g., mecillinam-­ ampicillin); (3) facilitation of drug entry of one antibiotic by another (e.g., beta-­lactam-­ aminoglycoside); (4) inhibition of inactivating enzymes (e.g., amoxicillin-­clavulanic acid); and (5) prevention of emergence of resistant populations (e.g., combination therapy in the treatment of tuberculosis).

Antagonism of Antimicrobial Combinations To some extent, the definition of antagonism as it relates to antimicrobial combinations reflects a laboratory artifact. However, there have been only a few well-­documented clinical situations where antagonism is clinically important. Antagonism may occur if antimicrobial combinations involve: (1) inhibition of bactericidal activity such as combination therapy with tetracycline and penicillin in which the bacteriostatic tetracycline prevents the critically required bactericidal activity of the penicillin; (2) competition for drug-­binding sites such as macrolide-­chloramphenicol combinations (of uncertain clinical significance); (3) inhibition of cell permeability mechanisms  such as chloramphenicol-­aminoglycoside combinations (of uncertain clinical significance); and (4) induction of beta-­lactamases by beta-­lactam drugs such as imipenem and cefoxitin combined with older beta-­lactam drugs which are beta-­lactamase unstable. The impressive complexity of the interactions of antimicrobials, the fact that such effects may vary depending on the bacterial species, and the uncertainty of the applicability of in  vitro findings to clinical settings make predicting the effects of some combinations hazardous. For example, the same

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10

combination may cause both antagonism and synergism in different strains of the same bacterial species. Laboratory determinations are required but may give conflicting results

depending on the test used. Knowledge of the mechanism of action is probably the best approach to predicting the outcome of the interaction in the absence of other guidelines.

­References and Bibliography Aharonowitz Y, Cohen G. 1981. The microbiological production of pharmaceuticals. Sci Am 245:141. Bellavita R, et al. 2020. Novel antimicrobial peptide from temporin L in the treatment of Staphylococcus pseudintermedius and Malassezia pachydermatis in polymicrobial inter-­Kingdom infection. Antibioitcs 9:530. Butler MS, et al. Antibiotics in the clinical pipeline as of December 2022. J Antibiotics 76:431. Costa BO, et al. 2019. Development of peptides that inhibit aminoglycoside-­modifying enzymes and β-­lactamases for control of resistant bacteria. Curr Protein Pept Sci 21:1011. Dheman N, et al. 2020. An analysis of antibacterial drug development trends in the United States, 1980–2019. Clin Infect Dis 73(11):e4444–e4450.

Douafer H, et al. 2019. Antibiotic adjuvants: make antibiotics great again. J Med Chem 62:8665. Greco I, et al. 2020. Structure-­activity study, characterization, and mechanism of action of an antimicrobial peptoid D2 and its D-­and L-­peptide analogues. Molecules 24:1121. Kammer RB. 1982. Milestones in antimicrobial therapy. In: Morin RB, Gorman M, (eds). Chemistry and Biology of Beta-­Lactam Antibiotics. Vol 3. Orlando, FL: Academic Press. Matravadi PK, et al. 2019. The quest for novel antimicrobial compounds: emerging trends in research, development and technologies. Antibiotics 8:8. O’Neill AM, et al. 2021. Antimicrobials from a feline commensal bacterium inhibit skin infection by drug-­resistant Staphylococcus pseudintermedius. eLife 10:e66793.

11

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  ­References and Bibliograph

2 Antimicrobial Susceptibility Testing Methods and Interpretation of Results Joseph E. Rubin and Peter Damborg

The veterinary diagnostic microbiology laboratory plays a key role in the practice of  evidence-­based antimicrobial therapy by providing culture and susceptibility information to practitioners. Before the introduction of antimicrobials, we were largely powerless to treat invasive infections. The antimicrobial age began with the familiar story of the discovery of penicillin in 1928 by Alexander Fleming. By the early 1940s, that Penicillium notatum extract was success­fully used against infections caused by organisms ranging from Staphylococcus aureus to Neisseria gonorrhoeae (Aronson, 1992; Bryskier, 2005). Unfortunately, the evolutionary power of bacteria resulted in the rapid emergence of antimicrobial resistance. Susceptibility testing  is now vital for effective therapeutic decision making. Although veterinary laboratories utilize many of the same basic microbiological techniques as human diagnostic labs, they face some unique challenges. These challenges include the difficulty in cultivation of fastidious veterinary-­specific organisms, availability of species customized antimicrobial panels for susceptibility testing, and lack of validated veterinary breakpoints. In the clinical setting, the goal of antimicrobial susceptibility testing (AST) is to help clinicians choose the optimal antimicrobial therapy. The decision to undertake culture

and susceptibility testing depends on the site of infection, state of the patient, prior ­history  of infections and antimicrobial use, co-­morbidities and underlying disease, and the predictability of the susceptibility patterns of the most likely pathogen(s). For example, susceptibility testing is generally not indicated in horses with “strangles,” as Streptococcus equi is uniformly susceptible to the drug of choice,  namely penicillin (Erol et  al.,  2012). Conversely, culture and susceptibility testing should always be done in cases of recurrent urinary tract infections in dogs. Early methods used to assess the suscepti­ bility of organisms to antimicrobials were developed by individual labs and lacked standardization; the first effort to standardize susceptibility testing was published in 1971 (Ericsson et  al.,  1971). National standards organizations responsible for guidelines for conducting and interpreting antimicrobial susceptibility tests were subsequently created. In the United States, the Clinical and Laboratory Standards Institute (CLSI) formed in the late 1960s as the National Committee for Clinical Laboratory Standards (NCCLS) and was tasked with developing a standard for disk diffusion  antimicrobial susceptibility testing (Barry, 2007). While standardization of methods yields more comparable data between labs,

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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13

Antimicrobial Susceptibility Testing Methods and Interpretation of Results

heterogeneity in interpretive criteria persists. In 1997, the European Committee on Antimicrobial Susceptibility Testing (EUCAST) was formed to harmonize both testing methods and interpretive criteria throughout Europe. In North America, the CLSI methodologies are used for both human and veterinary diagnostics. For the most common nonfastidious veterinary bacterial pathogens (e.g. Enterobacterales and staphylococci), susceptibility testing is straightforward using a common method. For testing fastidious and anaerobic organisms, specific standards have been developed but are not covered in this chapter.

A ­ ntimicrobial Susceptibility Testing Methods The available standardized antimicrobial susceptibility tests yield quantitative data (minimum inhibitory concentration [MIC] or inhibition zone diameter) that can be interpreted in different ways as described later in this chapter. Testing methods can be divided into two distinct categories: diffusion and dilution based.

Diffusion-­based Methods Two types of diffusion tests are available. The size of the inhibitory zone is a function of the rate of drug diffusion, thickness of the media, concentration of drug in the disk, and the susceptibility of the organism, making method standardization necessary for interpretive criteria to be applied (Figures 2.1, 2.2). Disk diffusion testing is done using 4  mm thick Mueller–Hinton agar plates and antimicrobial impregnated filter paper disks (CLSI, 2018a,b). Room-­temperature plates are inoculated with a lawn of bacteria drawn from a McFarland 0.5 (approximately 108 CFU/ ml) suspension using a sterile swab. Plates are allowed to dry for up to 15 minutes before the disk is applied and are then incubated for 16–20 hours at 35°C (+/–­ 2°C) at room

(A)

(B)

(C)

(D)

(E)

(F)

Figure 2.1  Disk diffusion: The results of the disk diffusion test can be influenced by he depth of the medium (A and B, increase in zone of inhibition; C, decrease in zone of inhibition) or the quality of the inoculum (D, false increase in zone of inhibition; E, false decrease in zone of inhibition; F, mixed culture, false decrease in zone of inhibition).

atmosphere. After incubation, the diameter of the inhibitory zone is measured (Figure 2.2A). Owing to differences in antimicrobial diffusion rate, amount of drug included in disks, and pharmacodynamic interactions, the size of the inhibitory zone corresponding to resistance breakpoints is unique to each drug/organism combination. The relative clinical appropriateness of different antimicrobials can therefore not be determined by simply comparing inhibitory zone diameters. Gradient tests (e.g., Etest®) are conducted in the same way as disk tests. These strips contain a gradient of antimicrobial from low to high concentrations corresponding to printed MIC values on the strip. Following incubation, the apex of the teardrop zone of inhibition indicates the MIC of the organism (Figure 2.2B).

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14

(A)

(C)

(B)

2 µg/ml

Disk diffusion (Bauer-Kirby procedure)

(D)

Method 

Antimicrobial gradient method Etest®

4 µg/ml

8 µg/ml

2 µg/ml, 4 µg/ml, 8 µg/ml Agar dilution

(E)

Broth macrodilution

Broth microdilution

Figure 2.2  Antimicrobial susceptibility testing methods.

Diffusion-­based tests are technically simple to perform and versatile, allowing customization of test panels to bacterial and patient species and type of infection.

Dilution-­based Methods Dilutional susceptibility testing can be done using either broth or agar media and yields MIC data. By convention, MICs are measured on a doubling dilution series (. . . 0.12 μg/ml, 0.25 μg/ml, 0.5 μg/ml, 1 μg/ml, 2 μg/ml . . .). An antimicrobial free control medium must always be included as a positive control for bacterial growth. The lowest concentration without bacterial growth defines the MIC, except for the sulfonamides and trimethoprim, where an 80% reduction in growth compared to the control constitutes inhibition.

For agar media dilution, Mueller–Hinton agar plates are prepared incorporating doubling dilutions of antimicrobial. Antimicrobial stock solutions at 10 times the test concentration are prepared using the solvents and diluents recommended by the CLSI (CLSI, 2018b). To prepare media, antimicrobial stock solution is added in a 1:9 ratio to molten Mueller– Hinton agar no hotter than 50 °C, and poured into sterile petri dishes. Separate plates are prepared for each antimicrobial concentration test. Plates must not be stored for more than seven days prior to use and for some drugs (e.g., imipenem), they must be prepared fresh on the day of use. Room-­temperature plates are inoculated with approximately ~104 CFU using either a multi-­spot replicator or manually by pipette. To prevent discrete samples from mixing, plates are left on the bench top

15

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­Antimicrobial Susceptibility Testing

Antimicrobial Susceptibility Testing Methods and Interpretation of Results

for up to 30 minutes for the bacterial spots to be absorbed prior to incubation. Plates are incubated in room air at 35 °C for 16–20 hours and examined for growth (Figure  2.2C). This technique is very labor intensive, hence its use is mainly limited to research. For broth dilution, Mueller–Hinton broths containing doubling dilutions of antimicrobial are prepared. As in agar dilution, antimicrobial stock solutions at 10 times the final concentration are prepared and added to test medium in a 1:9 ratio. Each antimicrobial concentration is dispensed into separate vials and inoculated with bacteria to yield a final concentration of 5 × 105 CFU/ml. A McFarland 0.5 inoculum is typically made in either sterile water or saline and then aliquoted into the Mueller–Hinton broth to yield the final concentration. Growth is indicated by turbidity or a cell pellet, and the MIC is defined by the lowest concentration where growth is not seen. Commercially prepared microdilution plates (Figure 2.2D) allow a large number of bacterial isolates to be tested efficiently without the need to prepare antimicrobial dilutions or large volumes of media in house. In some systems, microdilution plates can be inoculated and read automatically, thereby reducing hands-­on time. Some automated systems (e.g., Vitek® 2) work by regularly comparing the growth of test and antimicrobial-­free control cultures to provide a growth index from which susceptibility is predicted. In this way, results can be produced faster than the 16–20 hours typically required for incubation of MIC assays. Unfortunately, these commercially prepared plates are more expensive compared to the supplies required for disk diffusion, and the flexibility to change antimicrobial panels is limited (Figure 2.2E).

I­ nterpretation of Susceptibility Testing Results Antimicrobial susceptibility testing results can be interpreted in different ways.

The epidemiological cut-­off (ECOFF) for an antimicrobial agent is defined as the highest MIC for organisms without phenotypically detectable acquired resistance.1 Isolates with MICs above the ECOFF have therefore acquired resistance mechanisms that differentiate them from wild-­type organisms of the same species. ECOFFs are established by evaluating the MIC distributions of large isolate collections. While ECOFFs are invaluable research tools, e.g., for surveillance of emerging resistance, they do not take into account achievable drug concentrations in target tissue and should not be used as a first tool to guide therapy of patients. In fact, as shown in Figure 2.3, ECOFFs may differ from clinical breakpoints (CBPs), which are used for categorical interpretation of antimicrobial susceptibility test results for clinical use: susceptible = high probability of success following treatment, intermediate = treatment possible when the drug concentrates at the target site or when the dosage can be increased, and resistant = low probability of success following treatment. In 2024, a new category called susceptible dose dependent (SDD) was added for some drug–organism combinations by the breakpoint-­setting association CLSI. SDD implies that susceptibility of an isolate depends on the dosage regimen that is used in the patient. One reason for adding the SDD category was that to the clinician, “intermediate” too often means “resistant” because they do not appreciate the full definition of “intermediate.” Nevertheless, the SDD definition can appear confusing, as it partially overlaps with the intermediate category. For further elaboration, readers are encouraged to consult material provided by the CLSI (CLSI, 2024). CBPs are established by taking into account ECOFFs as well as pharmacokinetic data and, 1  How to: ECOFFs – the why, the how, and the don’ts of EUCAST epidemiological cutoff values. www. eucast.org/eucast_news/news_singleview/?tx_ ttnews%5Btt_news%5D=482&cHash=33c06c275 0a843056a5ec64da3321a12

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16

Gentamicin MIC Distribution for Escherichia coli 35000 30000

ECOFF

25000 20000

Clinical Breakpoint

15000 10000 5000 0

128

64

32

16

8

4

Resistant Acquired Resistance 2

1

0.5

0.25

0.12

0.06

Susceptible Wildtype

Figure 2.3  In this MIC distribution graph, the observed gentamicin MICs for 80 001 E. coli isolates are displayed. MICs are listed on the x-­axis in μg/ml and the number of isolates inhibited at each concentration is listed on the y-­axis. The EUCAST epidemiological cut-­off (ECOFF) differentiating wild-­type organisms from those with an acquired resistance mechanism is compared with the CLSI clinical resistance breakpoint (CLSI, 2024). In this example, E. coli may possess an acquired gentamicin resistance mechanism without crossing the threshold into clinical resistance.

when available, clinical outcome data following treatment. CBPs are specific to animal ­species, dosing regimen (dose, route of ­administration, and frequency), disease, and target pathogen. Internationally recognized veterinary-­specific CBPs are currently only published by the CLSI (CLSI, 2024), although the veterinary subcommittee of EUCAST (VetCAST) also has veterinary CBPs in the pipeline. Unfortunately, a general lack of validated veterinary-­specific CBPs is a serious limitation for veterinarians trying to practice evidence-­ based medicine. For example, there are no validated CBPs for any pathogens causing enteric disease, for important (but difficult to culture) pathogens like Mycoplasma, or for pathogens in certain animal species including fish and exotic/wildlife species (e.g., rodents, lagomorphs, reptiles, amphibians, and birds). Current veterinary-­specific CBPs from the CLSI are listed in Table 2.1, but as breakpoints are continuously being developed, readers are

advised to always seek the most updated ­guideline documents. Extrapolation of nonapproved breakpoints should be done with extreme caution; in some instances CBPs from other infection sites, bacterial species, animal species or humans can be adapted. One should consider consulting with a clinical microbio­ logist or clinical pharmacologist when ­deviating from animal-­, infection-­, and dosage-­ specific CBPs. In practice, the application of antimicrobial susceptibility test results for clinical use is often reduced to susceptible = good treatment choice and resistant = bad treatment choice, rather than a thorough analysis of the susceptibility profile. Interpretive reading is a more biological approach that incorporates knowledge of intrinsic drug resistance, exceptional resistance phenotypes, indicator drugs, and consideration of antimicrobial selection pressure (Livermore et al., 2001). Interpretive reading is used to detect specific resistance phenotypes such as methicillin resistance or

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­Interpretation of Susceptibility Testing Result  17

Animal species   Drug

Dog

Cat

Amikacin

Ent, PA, Staph spp., Streptococcus spp.

Amoxicillin-­ clavulanate

Ent, Staph spp. (SST), EC, Enterococcus spp., K. pneumoniae, Proteus mirabilis/vulgaris (ur)

Ent, PM, Staph spp., Strep spp. (SST), EC, Enterococcus spp., K. pneumoniae, PM, P. mirabilis/vulgaris, Staph spp., Strep spp. (ur)

Ampicillin

Ent, Enterococcus spp. SP, Strep spp. (SST), EC, Enterococcus spp., Proteus mirabilis (ur)

Ent, PM, Staph spp., Strep spp. (SST), EC, Enterococcus spp., PM, P. mirabilis, Staph spp., Strep spp. (ur)

Cefazolin

Ent (SST), EC, K. pneumoniae, P. mirabilis (ur), SA, SP (resp, SST, ur), beta-­hem Strep spp. (gen, resp, SST, ur), PM (resp, SST)

Ent (metritis), HS/ MH/PM (resp)

EC, P. mirabilis (ur), SP, beta-­hem Strep spp. (SST)

Cefpodoxime

EC, P. mirabilis (ur, wound/ abscess), SA, SP, S. canis, PM (wound/abscess)

Ceftazidime

Ent, PA (SST)

SA (resp, SST), S. equi subsp. equi and zooepidemicus (resp), Ent Ent, beta-­hem Strep spp. (gen, resp)

EC, Staph spp., S. agalactiae, S. dysgalactiae, S. uberis (mastitis)

Cefovecin

Horse

Pig

Ent, PA, SA, S. equi subsp. equi and zooepidemicus

Cefoperazone

0005781542.INDD 18

Cattle

EC (ur), PM (SST)

APP, BB, PM, S. suis (resp)

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Table 2.1  Drugs with veterinary-­specific CLSI resistance breakpoints according to the VET01S-­Ed7 document (CLSI, 2024). Abbreviations used for bacteria are shown in the footnote.

11-08-2024 13:20:10

EC, SA, S. agalactiae, S. dysgalactiae, S. uberis (mastitis), HS/MH/PM (resp)

Cephalexin

EC, K. pneumoniae, P. mirabilis (ur), EC, SA, SP, beta-­hem Strep spp. (SST)

Cephalothin

SA, SP, beta-­hem Strep spp. (SST)

Chloramphenicol

Ent, Enterococcus spp., Staph spp.

Clindamycin

Staph spp., beta-­hem Strep spp. (SST)

Danofloxacin

APP, PM, S. Choleraesuis, S. suis (resp)

MH, PM (resp)

Difloxacin

Ent, Staph spp., beta-­hem Strep spp. (SST, ur)

Doxycycline

Staph spp. (SST), Ent

Enrofloxacin

Ent, PA, Staph spp., beta-­hem Strep spp. (resp, SST, ur)

Ent, Staph spp., S. equi subsp. equi and zooepidemicus Ent, PA, Staph spp., Strep spp. (SST)

Florfenicol

HS/MH/PM (resp)

Ent, PA, Staph spp., S. equi (resp, SST)

HS/MH/PM (resp)

Gamithromycin Gentamicin

S. equi subsp. zooepidemicus (resp)

APP, BB, PM, S. Choleraesuis, S. suis (resp)

HS/MH/PM (resp) Ent, PA

Actinobacillus spp., Ent, PA

Kanamycin-­cephalexin

EC, Staph spp., S. dysgalactiae, S. uberis (mastitis)

Levofloxacin

Ent, PA (SST)

Marbofloxacin

Ent, PA, Staph spp., beta-­hem Strep spp. (SST, ur)

Ent, Staph spp., Strep spp. (SST)

APP, PM, S. suis, (resp)

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Ceftiofur

(Continued)

0005781542.INDD 19

11-08-2024 13:20:10

Animal species   Drug

Dog

Minocycline

Staph spp. (SST)

Orbifloxacin

Ent, Staph spp., beta-­hem Strep spp. (SST, ur)

Cat

Cattle

Ent, Staph spp., Strep spp. (SST) HS/MH/PM (resp)

Penicillin-­novobiocin

SA, S. agalactiae, S. dysgalactiae, S. uberis (mastitis)

Pirlimycin

SA, S. agalactiae, S. dysgalactiae, S. uberis (mastitis) Ent, PA, Staph spp. (SST, ur)

Pradofloxacin

EC, SP (skin, ur)

Spectinomycin Tetracycline

EC, PM, SA, S. canis, S. felis, SP (resp, skin)

HS/PM (resp)

Staph spp., Strep spp. (resp, soft tissue)

PM, S. suis (resp)

APP (resp)

HS/MH/PM (resp) Staph spp. (SST)

HS/MH/PM (resp)

APP, PM, S. suis (resp)

HS/MH/PM (resp)

APP, BB, PM (resp)

Tiamulin Tildipirosin

Pig

Ent, Staph spp., Strep spp. (resp, SST)

Penicillin G

Piperacillin-­ tazobactam

Horse

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Table 2.1  (Continued)

APP (resp)

Tilmicosin

MH (resp)

APP, PM (resp)

Tulathromycin

HS/MH/PM (resp)

APP, BB, PM (resp)

APP, Actinobacillus pleuropneumoniae; BB, Bordetella bronchiseptica; EC, Escherichia coli; Ent, Enterobacterales; gen, genital tract; HS, Histophilus somni; MH, Mannheimia haemolytica; PA, Pseudomonas aeruginosa; PM, Pasteurella multocida; resp, respiratory tract; SA, Staphylococcus aureus; SP, Staphylococcus pseudintermedius; SST, skin and soft tissue; ur, lower urinary tract. Note: a single avian breakpoint for enrofloxacin for E. coli has not been included in this table.

0005781542.INDD 20

11-08-2024 13:20:10

the production of extended-­spectrum beta-­ lactamases (ESBLs). Some of these tests are organism specific, and use across species or genera may not yield reliable results. For example, the CLSI recommends that either cefoxitin or oxacillin resistance may be used as an indicator of mecA-­mediated methicillin resistance in S. aureus, while only oxacillin resistance reliably predicts mecA in S. pseudintermedius (CLSI, 2023, 2024; Papich,  2010). In Enterobacterales, the susceptibility profile to a combination of third-­generation cephalosporins with and without clavulanic acid, cefoxitin, and cloxacillin is useful for phenotypically differentiating ESBLs from AmpC type beta-­ lactamases (CLSI, 2023, 2024). Intrinsic resistance is an innate structural or functional characteristic of a bacterial species causing all or the vast majority of bacteria from that species to resist the action of an antimicrobial agent; this differs from acquired resistance, which may be present or not, and which differentiates an organism from wild type (Leclercq et al., 2013). It is unnecessary to test the susceptibility of an organism to a drug to which it is intrinsically resistant; reporting “resistant” may spuriously lead a prescriber to believe that the isolate is multidrug resistant, and to inappropriately select a second-­ or third-­line therapy. Knowledge of intrinsic resistance is invaluable to the practitioner for guiding empirical treatment (e.g., what drugs should I immediately rule out for therapy?), and to the laboratory as an indicator of quality control (e.g., susceptibility of an organism which is intrinsically resistant may indicate misidentification). Intrinsic resistance may be a consequence of the absence of drug targets, impermeability of cell membrane or endogenous efflux pumps, lack of transport mechanisms or endogenous production of degradative enzymes. For example, resistance by absence occurs in the case of Mycoplasma, which lacks a peptidoglycan-­ containing cell wall and is therefore intrinsically beta-­lactam resistant. The Gram-­negative cell membrane prevents many antimicrobials

(vancomycin and the macrolides, for example) from entering the cell (Cox et al., 2013) and is one of the best described mechanisms of intrinsic resistance. Pseudomonas aeruginosa is widely recognized for its extensive intrinsic resistance, partly due to its outer membrane, estimated to be only 8% as permeable as E. coli (Cox et al., 2013). For drugs which are actively transported across the cell wall, the absence of transport mechanisms can confer resistance; anaerobic organisms are intrinsically aminoglycoside resistant due to oxygen-­dependent uptake of these drugs (Bryan et al., 1981). Finally, beta-­lactamase production is frequently encountered, particularly among Gram negatives. Of particular note are the “SPICE organisms” (Serratia, Providencia, indole positive Proteae [Proteus vulgaris and  Morganella spp.], Citrobacter, and Enterobacter). These bacteria endogenously express AmpC type beta-­lactamases and are consequently intrinsically resistant to first-­ generation cephalosporins and ampicillin/ amoxicillin, including with beta-­lactamase inhibitors. A detailed description of intrinsic resistance phenotypes is published by EUCAST and available at www.eucast.org. Some commonly encountered veterinary pathogens with intrinsic resistance to antimicrobials are included in Table 2.2. An appreciation of exceptional (unexpected) resistance phenotypes allows unusual isolates or test results to be identified and investigated further. Vancomycin-­resistant staphylococci, penicillin-­resistant group A streptococci, and metronidazole-­resistant anaerobes are all exceptional phenotypes that should be confirmed before starting antimicrobial therapy. While such results can be due to the emergence of resistance, it is more likely that these results reflect errors in reporting, testing, isolate identification, or testing mixed cultures (Livermore et al., 2001). The CLSI M100 document and the EUCAST expert rules describe exceptional phenotypes (CLSI, 2023; Leclercq et al., 2013).

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­Interpretation of Susceptibility Testing Result  21

Antimicrobial Susceptibility Testing Methods and Interpretation of Results

Klebsiella pneumoniae

R

R

Proteus vulgaris

R

R

Serratia marcescens

R

R

Yersinia pseudotuberculosis

R

SPICE

R

R

R

Aeromonas hydrophila

R

R

R

R R

R R

Glycopeptides

Nitrofurantoin

R

Polymyxins

R

R

Sulfonamides

R

Chloramphenicol

R

Tetracycline

R

Aminoglycosides

Lincosamides

All cephalosporins

1GC, 2GC

Amox + clavulanate

R

Macrolides

All Enterobacterales

Ampicillin

Penicillin

Table 2.2  Intrinsic resistance phenotypes of importance to veterinary medicine.

R R R

R

R R

R

R

R

R R

R

R

R

R

R

R

R

R

R

R

Acinetobacter baumannii

R

R

R

R

R

R

R

R

R

Pseudomonas aeruginosa

R

R

R

R

R

R

R

R

R

Stenotrophomonas maltophila

R

R

R

R

R

R

R

R

R

Staphylococcus spp.

R

Streptococcus spp. Enterococcus faecalis

R* R

R

Enterococcus faecium

R

R

Enterococcus gallinarum

R

R

Listeria monocytogenes

R

R

R

R*

R

R

R*

R

R

R

R*

R

R

R

R

For each organism or group of organisms, intrinsic resistance to a drug or drug class is indicated by an “R.” These data were extracted and modified from EUCAST expert rules on intrinsic resistance and VET01S ED5:2020 (CLSI, 2020). Enterobacterales – a bacterial order including E. coli, Salmonella, and Klebsiella; SPICE – subgroup of organisms in the order Enterobacterales composed of Serratia, Providencia, indole-­positive Proteae (Proteus vulgaris and Morganella spp.), Citrobacter, and Enterobacter; 1GC – first-­generation cephalosporins (e.g., cefazolin, cephalothin or cefalexin); 2GC (e.g., cefuroxime). R* indicates low-­level resistance.

Bacterial resistance mechanisms often predictably confer resistance to multiple antimicrobials such that resistance to one may indicate resistance to others. By testing indicator drugs, susceptibility test results can be extrapolated to a broader panel of antimicrobials than could practically be tested (Table 2.3). Diagnostic laboratories face a dilemma of wanting to report simple antibiograms (“S, I, R”) that can be easily followed by clinicians, while also wanting to provide useful details pertaining to interpretive reading. One solution could be to keep a simple antibiogram layout while adding footnotes or supplementary

text explaining, for example, intrinsic resistance, the observation of methicillin resistance, or if a drug may be used as an indicator for other drugs. MICs should generally be excluded from antibiograms, as they are prone to misinterpretation by clinicians. As an example, the highly potent fluoroquinolones tend to have the lowest MICs, but other drugs with higher MICs in the susceptible category may be equally or even more effective for a particular organism. A final consideration for antibiograms is to report drugs selectively. For example, ­excluding critically important drugs like carbapenems could be a way to minimize use of drugs within

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22

Table 2.3  Indicator drugsa. Prediction Based on Indicator Organism

Indicator Drug

Agents Affected

R

Staphylococcus spp.

Oxacillin or cefoxitin

All beta-­lactams

X

Gentamicin

All aminoglycosides

X

Beta-­hemolytic streptococci (S. agalactiae, dysgalactiae, equi, canis)

Clindamycin

Lincomycin

X

Tetracycline

Chlortetracycline, oxytetracycline, minocycline, doxycycline

X

Benzylpenicillin

Aminopenicillins and cephalosporins

X

Clindamycin

Lincomycin

Streptococcus spp. (except S. pneumoniae)

Tetracycline

Doxycycline, minocycline

Streptococcus and Enterococcus spp.

Gentamicin

All aminoglycosides except streptomycin

X

Enterococcus spp.

Penicillin or ampicillin

Penicillin, ampicillin, amoxicillin, amoxicillin clavulanate

X

Tetracycline

Doxycycline, minocycline

All aminoglycosides

Gentamicin

X

Amoxicillin-­clavulanate

Ampicillin and amoxicillin

X

Cefazolin

Oral cephalosporins

X

Doxycycline

Chlortetracycline, oxytetracycline, minocycline

Enterobacterales, Pseudomonas aeruginosa

Tobramycin or gentamicin

Amikacin

X

Bordetella spp., Pasteurella multocida, Actinobacillus pleuropneumoniae

Ampicillin

Aminopenicillins

X

Mannheimia haemolytica

Tetracycline

Tetracycline class

X

All bacteria

Erythromycin

Azithromycin, clarithromycin

Enterobacterales

S

X

X X

X

X

X

X X

a

 Indicator drugs can be used to infer susceptibility or resistance of an organism to other agents. If used to infer susceptibility, susceptibility to the indicator drug may be extrapolated to other agents. Similarly, indicator drugs may be used to identify cross-­resistance. Critically, these criteria may only be valid for extrapolating either susceptibility or resistance; for example, penicillin-­susceptible Streptococcus canis would be predicted to be susceptible to ampicillin while penicillin-­resistant S. canis may or may not be resistant to ampicillin.

this class, while retaining the results for ­surveillance purposes. Selective reporting may also be used to customize antibiograms by reporting only drugs of relevance to the

infection. This requires detailed information to be provided by the clinician, and good knowledge in the laboratory concerning clinically relevant and available drugs.

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­Interpretation of Susceptibility Testing Result  23

Antimicrobial Susceptibility Testing Methods and Interpretation of Results

Overall, most of these “reporting issues” may be solved by good and frequent communication between clinicians and microbiologists in diagnostic laboratories.

A ­ djunctive Susceptibility Testing Methods Apart from the traditional dilution-­ and diffusion-­based methods for antimicrobial susceptibility testing, several adjunctive

methods exist. Some of these are described in the following text and examples are also ­provided in Table  2.4. Selective media have been designed to quickly identify particular antimicrobial-­resistant organisms from ­clinical samples. A detailed description of screening media for extended-­spectrum beta-­ lactamases in Enterobacterales, methicillin resistance in staphylococci, and high-­level aminoglycoside and vancomycin resistance in enterococci is published by the CLSI (CLSI, 2023, 2024).

Table 2.4  Examples of other susceptibility testing methods.

Principle for detection of AMR

Approximate time from pure culture until postanalysis interpretationa

Method type

Method

Phenotypic

Selective agar plate

Agar plate containing antibiotics and other ingredients that suppress or kill most microorganisms other than the target resistant organism

NA (used directly on samples before pure culture is obtained)

Simple, rapid, and cheap but confirmation of antimicrobial susceptibility usually needs verification by other methods

Latex agglutination

Latex beads with antibodies specific to a certain target antigen like a protein conferring resistance. Antigen-­ antibody binding causes visible agglutination

90% protein bound? No

Yes Does the drug have a narrow therapeutic index?

No

Yes Would a transient increase in free drug concentration be clinically relevant?

Low

What is the hepatic extraction ratio?

No

No clinically significant interaction

High No

Is the route of administration IV? Yes Potential for a clinically significant interaction. Look for a clinical study to confirm.

Figure 4.7  Algorithm for determining clinically relevant protein-­binding interactions. Source: Adapted from Rolan (1994). 100

Drug Concentration (μg/ml)

IV ORAL 10

1

Area = ½ base(height’ + height”)

0

0

5

10

15 Time (hr)

20

25

30

Figure 4.8  Plasma drug concentration profiles intravenous versus oral administration of ampicillin in a dog. Calculation of the area under the curve (AUC) for each curve is done by the trapezoidal rule. It is illustrated here for the oral curve, where the trapezoidal area between data points is calculated and summed to give the total AUC.

The method of corresponding areas is used to determine F: F

AUC PO AUC IV

Dose IV Dose PO

where AUC is the total area under the plasma concentration-­time curve relating to the route of drug administration (IV versus PO, IM, or SC). The systemic availability of orally administered antimicrobial drugs is often incomplete

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­Drug Distributio  61

Pharmacokinetics of Antimicrobials

(0.7, medium if E = 0.3 and low if E 0.35, medium if about 0.15, and low if t½ 1 hr), little to no drug accumulation occurs. There is little lag time to reach the targeted plasma concentrations but there is marked fluctuation between Cmax and Cmin.

­ esigning Dosage Regimens D for Clinical Patients The success of antimicrobial drug therapy is highly dependent on the dosage regimen design which is based on pharmacokinetic data as described. A drug dosage regimen is composed of a dose and a dosing frequency. Some drugs are given as single doses, so a

specific plasma concentration will be targeted. When multiple doses of a drug are given, the dosage frequency and drug accumulation must be considered. Not all antimicrobials need rigid individualization of the dosage regimen. In the case of antimicrobials with a broad safety range, such as the penicillins and cephalosporins, the dosage is not titrated precisely but rather is based

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72

on the label directions or clinical judgment to maintain an effective plasma concentration above the bacterial pathogen’s minimum inhibitory concentration (MIC). For drugs with a narrow therapeutic margin, such as the aminoglycosides which are frequently used in an extra-­label manner, the individualization of the dosage regimen is very important. The objective of the dosage regimen for these drugs is to produce a safe plasma drug concentration that does not exceed the minimum toxic concentration or fall below a critical minimum concentration below which the drug is not effective.

Single-­dose Regimen For drugs given as single doses, only the target drug concentration and the Vd of the drug in that patient are needed with consideration of the effects of physiology or pathophysiology on the Vd. This is also how a drug dosage regimen is calculated for concentration-­dependent antimicrobials with a prolonged postantibiotic effect whose efficacy is based upon an ideal Cmax:MIC and we assume once-­daily dosing (e.g., aminoglycosides). Dose

Vd C

where C equals the target plasma concentration, which for the aminoglycosides is >10 times the MIC.

Continuous Intravenous Infusion Continuous intravenous infusion offers the most precise control of drug levels in the body and is essential for precise control of drugs with a narrow safety margin or very rapid elimination. It is not feasible for most drugs and most clinical situations in veterinary medicine. But when the desired response needs a constant drug concentration, the antimicrobial may be infused at a constant rate following the initial intravenous dose:

R

C Vd K or R since Vd K Cl

C Cl ,

where R (rate) is essentially the rate of drug loss from the body (mg/min or mg/hr). Therefore to maintain the established concentration of drug in the body, it is necessary to infuse the antimicrobial at the rate equal to its loss (clearance).

Multiple-­dose Regimens For concentration-­dependent antimicrobials whose efficacy is determined by an ideal AUC0–­24 hr:MIC, the following equation can be used to calculate a dose per day: Dose

AUC 0

24 hr

: MIC MIC Cl

F 24 hr

where F is bioavailability. This approach is often used to calculate doses for fluoroquinolones, which have a concentration-­dependent effect on Gram-­negative aerobic bacteria, and where an AUC:MIC value of 100 or 125 is used. For time-­dependent antimicrobials whose efficacy is determined by an ideal time that the drug concentration remains above the bacterial MIC, the following equation is used to target the percentage of time: %time

MIC

ln(dose /[VD MIC]) (t½ / ln2) (100/ DI )

where ln is the natural logarithm and DI is the dosage interval in hours. This approach is often used to calculate the doses for penicillins, cephalosporins, macrolides, tetracycline, phenicols, and potentiated sulfonamides.

Loading Dose Followed by Maintenance Dose Regimen Although infrequently done with antimicrobials, drugs that have long half-­lives such as sulfonamides will have a lag time to reach

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­Designing Dosage Regimens for Clinical Patient  73

Pharmacokinetics of Antimicrobials

therapeutic drug concentrations. Therefore they may be given by a large loading dose followed by maintenance doses. Dose loading

Dose maintenance 1 e

Kt

Sustained-­release and Prolonged-­action Dosage Formulations If maintenance dosage regimens are not practical for a patient, then sustained-­release or prolonged-­action preparations of antimicrobials that have short half-­lives provide a longer duration of drug action and eliminate the need for dosing at short intervals, for administering unduly large doses of the usual drug formulation, or for resorting to an intravenous formulation. Prolonged-­action parenteral antimicrobial drugs are commonly used where repeated handling of animals is difficult or undesirable. The potential for toxicity of sustained-­release formulation is greater, as release from the dosage form and absorption are unpredictable in most clinical situations. Prolonged concentrations of subtherapeutic antimicrobials may also contribute to antimicrobial resistance.

Duration of Therapy The success of antimicrobial therapy depends upon the administration of a drug to which the causative pathogenic microorganisms are susceptible at the concentrations attained at the site of the infection, but also on the duration of treatment. While both the microbiological and pharmacokinetic properties of the antimicrobial agent selected are taken into account in the dosage regimen, the duration of treatment is largely empirical (discussed further in Chapter 6). It is imperative that antimicrobial therapy be maintained for an adequate duration, which should be based upon monitoring the response both by clinical assessment of the animal

patient (resolution of fever, leukocytosis and other signs of acute inflammation) and in some cases also bacterial culture of properly collected specimens. Definitive diagnosis at an early stage of infection and the application of specific therapy, based on knowledge of the causative pathogenic microorganism and its susceptibility, will decrease the overall duration of treatment and minimize residual sequelae. An extended course of treatment is generally required in immunocompromised animals. There are certain infections that, due to the relative inaccessibility of the causative pathogenic microorganisms to antimicrobial agents, invariably require a prolonged duration (3–5 weeks, rather than 6–10 days) of therapy. They include prostatitis, osteomyelitis and skin infections in dogs, and Rhodococcus equi pneumonia in foals.

Adjusting Dosage Regimens for Clinical Patients Age Considerations

The age of the animal may have a profound effect on drug disposition. The definition of “geriatric” varies between species, and in dogs it varies between breeds. Body composition and regional blood flow change in geriatrics. Cardiac output decreases, so regional and organ blood flow also decreases. These changes will impact drug absorption, distribution, and elimination. Blood flow is preferably redistributed to the brain and heart, so there is an increased risk of drug toxicity in these organs. Gastrointestinal motility and absorptive capacity are reduced. Hepatocyte number and function decrease along with hepatic and splanchnic blood flow. As renal blood flow decreases, glomerular filtration rate and active secretory capacity of the nephron decrease, resulting in lowered renal clearance of drugs. Lean body mass decreases while fatty tissues increase. Increased body fat results in a decrease in total body water and cell mass. The plasma concentrations of water-­soluble (low

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74

volume of distribution) drugs tend to increase, while the plasma concentrations of lipid-­ soluble (high volume of distribution) drugs tend to decrease. Serum albumin decreases while gamma-­globulins increase, so that total plasma protein concentrations remain the same. The neonatal period also varies with species and age, but all the determinants of drug disposition are altered as the animal matures. This period is approximately 1–2  weeks in foals; about eight weeks in calves, kids, lambs, and piglets; and 10–12  weeks in puppies and kittens. However, the most profound adaptive changes in physiological variables occur during the first 24 hours after birth in all species. Blood flow to the heart and brain is greater and faster, making the pediatric patient more susceptible to drug-­induced cardiotoxicity and neurotoxicity. Gastrointestinal absorption tends to be decreased due to decreased gastric emptying and decreased intestinal peristalsis. Since the rumen takes 4–8  weeks to develop and become functional, the bioavailability (rate and extent of absorption) of drugs administered orally to preruminant calves resembles that in monogastric species rather than in cattle. Absorption from intramuscular and subcutaneous sites changes as muscle mass and blood flow change. Neonates have less fat and greater total body water (primarily extracelluar fluid) than adults. Therefore, low Vd drugs will distribute into a large volume, making it necessary to increase the dose to avoid therapeutic failure. Because of low body fat stores, ­lipid-­soluble drugs will have higher plasma concentrations. Drug elimination by both hepatic metabolism and renal excretion is limited in pediatric animals. Antimicrobials that undergo extensive ­first-­pass metabolism by hepatic microsomal oxidative reactions would be expected to have higher systemic availability in neonatal animals. This applies to trimethoprim, which has far higher systemic availability in newborn kids than in older kids and adult goats. Although there are species differences in the

degree to which some drug metabolic ­pathways are deficient in neonatal animals, a relative lack of development of hepatic smooth-­ surfaced endoplasmic reticulum and its associated drug metabolizing enzyme systems (mediate oxidative reactions and glucuronide conjugation) appears to be a characteristic of the neonatal period in all mammalian species. Because of the low activity of most metabolic pathways, the half-­lives of drugs that undergo extensive hepatic metabolism are prolonged in neonatal animals, particularly during the first 24 hours after birth. The maturation of the various metabolic pathways could be related to hormonal ­influence on postnatal enzyme induction. In most species (ruminant animals, pigs, dogs, and presumably cats), the hepatic microsomal-­ associated metabolic pathways develop rapidly during the first 3–4  weeks after birth, and at 8–12  weeks of age have activity approaching that of adult animals. The foal appears to be an exception in at least the rate of development of glucuronide synthesis, which develops very rapidly during the first week after birth. In addition to lower hepatic microsomal oxidative activity, the ruminal microflora have not developed in neonatal ruminant species and this can impact antimicrobial disposition. The renal excretion mechanisms (glomerular filtration and active, carrier-­mediated tubular secretion) are incompletely developed at birth in all mammalian species. During the neonatal period, renal excretion mechanisms mature independently at rates that are species related. GFR, based on inulin clearance, attains adult values at two days in calves; 2–4 days in lambs, kids, and piglets; and may take at least 14 days in puppies. Proximal tubular secretion, based on clearance of para-­aminohippurate, matures within two weeks after birth in ruminant species and pigs but may take up to six weeks in puppies. Indirect evidence, provided by pharmacokinetic studies of some antimicrobial agents, suggests that renal function develops rapidly in foals at a rate similar to that in ruminant species. This implies that the neonatal

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­Designing Dosage Regimens for Clinical Patient  75

Pharmacokinetics of Antimicrobials

foal, like the calf, has relatively mature renal function compared with neonates of most other species. Even though renal function is immature in neonatal, particularly newborn, animals, it has the capacity adequate to meet physiological requirements. However, when lipid-­soluble drugs are administered to neonatal animals, the combined effect of slow hepatic microsomal associated metabolic reactions (oxidation and glucuronide conjugation) and relatively inefficient renal excretion mechanisms considerably decreases the rate of elimination of the parent drugs and their polar metabolites. Urinary pH is acidic in neonates of all species, so favors renal tubular reabsorption and extends the elimination half-­life of drugs that are weak organic acids and of sufficient lipid solubility to be reabsorbed by passive diffusion (e.g., most sulfonamides). Antimicrobial treatment regimens in neonates are best determined by clinical trials in such patients. If such trials are not available, general recommendations are to increase the dose of low Vd drugs according to the increased ECF of the neonate and to increase the dosage interval for any drugs where accumulation could result in toxicity. Considerations in Pregnant Animals

Physiological adaptations that occur during pregnancy can influence the oral bioavailability and disposition of drugs due to an increase in gastric pH, an increase in the circulating blood (plasma) volume and in renal blood flow, an alteration in body fluid compartments, and hormonal-­induced change in hepatic microsomal enzyme activity. A major concern in the use of drugs during pregnancy is the potential for adverse effects on the fetus, since all drugs administered to the mother cross the placental barrier, although at different rates, and the fetus is ill equipped to eliminate drugs. To what extent enzymes located in the placental membranes (e.g., microsomal drug-­ metabolizing system, which mediates various oxidative reactions, and cholinesterase)

contribute to the conversion of drugs to inactive, more active or potentially toxic metabolites has not been established in domestic animal species. Placental drug transfer by passive diffusion is similar to passage across any epithelial barrier, and in many respects resembles passage from the systemic circulation into milk of ­lactating animals. Drug diffusion across the placenta from mother to fetus is favored by lipid solubility, a large concentration gradient of unbound drug between the maternal and fetal circulations, and the presence of drug in the nonionized form in the maternal circulation. Blood flow to the placenta limits the rate of delivery of drug to the fetal circulation. Conversely, molecules that are ionized (penicillins, cephalosporins), hydrophilic (aminoglycosides), and present in low free drug concentrations (doxycycline, macrolides, lincosamides) have restricted access to the fetus. Differences in the extent of plasma protein binding by the mother and fetus (in which it  is lower) affect the total plasma drug concentrations in the maternal and fetal circulations. Regardless of the physicochemical properties of a drug, the duration of maternal therapy with the drug influences the concentrations that will be attained in the fetus. Some drugs known to diffuse well in other body fluids such as synovial and abdominal fluid do not necessarily reach therapeutic concentrations in fetal fluids. For example, concentrations of ceftiofur and related metabolites in placenta, fetal ­fluids, and fetal tissues are well below therapeutic concentrations after IM administration of ceftiofur to pregnant mares (Macpherson et al., 2013). In contrast, penicillin G and gentamicin undergo effective placental transfer in pregnant mares (Murchie et al., 2006). Because toxic effects could be produced in the fetus, caution should be exercised with use in pregnant animals of a wide variety of antimicrobial agents, while some others (fluoroquinolones, tetracyclines, griseofulvin) are contraindicated.

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Considerations with Renal Impairment

Polar drugs and drug metabolites are largely confined to extracellular fluid and undergo elimination by renal excretion. This is because of their limited capacity to passively diffuse through lipid membranes. Even though lipid-­soluble drugs are mainly eliminated by hepatic metabolism, a fraction of the systemically available dose is usually eliminated by renal excretion. Therefore, it is not surprising that renal disease profoundly impacts the disposition of most drugs administered to animals with renal insufficiency or failure. With reduced renal clearance, the parent drug and/or its metabolites may accumulate in the patient and cause toxicity. Loss of proteins and electrolytes in urine and the alterations in acid– base balance associated with renal failure affect the pharma­cokinetics and pharmacodynamics of drugs. Enhanced drug activity or toxicity can occur due to synergy with uremic complications. Altogether, these effects make it difficult to determine safe and effective drug dosages for veterinary patients in renal insufficiency or failure. The renal excretion of drugs and drug metabolites involves glomerular filtration and, for some drugs and most metabolites, carrier-­mediated proximal tubular secretion. Extensive binding to plasma proteins limits the availability of drug molecules for glomerular filtration but might not hinder their secretion by proximal renal tubules because of rapid dissociation of the drug–protein complex. The primary pathophysiological sequela relevant to antimicrobial dosing in renal dysfunction is a decreased GFR, which results in a decreased clearance of drugs eliminated by the kidney. Because of the large renal functional reserve, 75% of GFR must generally be lost before signs of clinical disease are readily evident. The goal of dosage adjustment is to provide a drug concentration-­time profile in the renal failure patient that is as similar as possible to  a normal patient. The best approach to

modifying drug therapy in renal failure patients is to carry out therapeutic drug monitoring and adjust the dosage for each individual patient. This is possible with some drugs such as gentamicin and amikacin but is impractical and cost-­prohibitive for most drugs used in veterinary practice. The best approach for most drugs is to estimate a corrected dose from available renal function tests and then to monitor the patient closely for evidence of efficacy or toxicity. For drugs that are eliminated primarily by renal mechanisms, creatinine clearance correlates well with drug clearance. Creatinine (Cr) is an endogenous product of creatinine phosphate metabolism in muscle. It is removed by glomerular filtration and serum concentrations are relatively constant in healthy people and animals. The elimination half-­life of a drug that is eliminated in urine remains stable until creatinine clearance is reduced to 30–40% of normal, which is why drug dosage regimens are typically not adjusted until two-­thirds of renal function has been lost. In human patients, creatinine clearance is quantified by determining urinary creatinine excretion over a 24-­hour period. The measured creatinine clearance is then used in formulas to make drug dosage adjustments. Unlike in human medicine, values for creatinine clearance are not usually available for veterinary patients. When creatinine clearance is not available, a single value of the patient’s serum creatinine can be substituted into the formulas. However, the relationship between serum creatinine is not linear once serum creatinine is above 4  mg/dl (275 μmol/l), so the adjustment formulas are even less accurate for predicting an ideal dose adjustment. These formulas do not account for changes in the volume of distribution and nonrenal clearance mechanisms of the drug that may be caused by the renal dysfunction. Therefore, these dosage adjustments must be regarded as preliminary estimations to be followed by adjustments based on observed clinical response.

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­Designing Dosage Regimens for Clinical Patient  77

Pharmacokinetics of Antimicrobials

Dose Reduction Method

With this method, the normal dosage regimen is adjusted by reducing the drug dose and maintaining the drug dosing interval. Adjusted dose normal dose patient’s Cr clearance / normal Cr clearance or Adjusted dose normal dose normal Cr / patient’s Cr

Interval Extension Method

With this method, the normal dosage regimen is adjusted by reducing the drug dose and maintaining the drug dosing interval. Adjusted interval normal interval patient’s Cr clearance / 1/ normal Cr clearance or Adjusted interval normal interval normal serum Cr / 1/ patient’s serum Cr Both methods attempt to keep the average plasma drug concentrations constant. The interval extension method produces Cmax and Cmin values similar to those seen in healthy patients. However, it does produce substantial periods of time where drug concentrations may be subtherapeutic. This is the preferred method with aminoglycosides, which have a long postantibiotic effect and where a low trough concentration is desirable to reduce the risk of nephrotoxicity. Depending on the relationship of the elimination half-­life to the dosage interval, significant drug accumulation may occur with the dose reduction method, but at steady state there are no periods of time where concentrations are subtherapeutic. This is the preferred method for the beta-­lactam antibiotics, where maintaining

the plasma concentration above the pathogen’s MIC correlates with efficacy and the drugs are relatively nontoxic even if accumulation occurs. To decide which method to use, the practitioner should determine if drug efficacy and toxicity are related to peak, trough or average plasma concentrations and then select the method which balances efficacy against potential toxicity. The interval extension method is more client convenient, since the normal recommended dose is simply administered less frequently. And if the drugs are available only in fixed dosage forms (e.g., capsules, unbreakable tablets), it is easier to adjust the dosage interval. Since the elimination half-­life is prolonged in patients with renal disease and it always takes five half-­lives to reach 99% of steady-­state concentrations, there is a delay in reaching steady state in renal failure patients compared to animals with normal renal function. Therefore, a loading dose may be needed to rapidly achieve therapeutic drug concentrations. If the dose reduction method is used, this is achieved by giving the usual dose initially, followed by the reduced dose at the next time. If the interval extension method is used, this is accomplished by giving a double dose initially. For renal failure patients in general: ●●

●●

●●

●●

avoid using any drugs at all unless there are definite therapeutic indications. If you must use a drug, try to select one that is hepatically metabolized and excreted in bile rather than eliminated by the kidneys (e.g., doxycycline) if therapeutic drug monitoring is available, tailor the drug dosage regimen to the specific patient if therapeutic drug monitoring is unavailable, determine if there are clinically proven, adjusted dosage regimens for specific drugs. The package insert on human pharmaceuticals often gives guidelines for adjusting dosages if the drug has not been sufficiently studied to have dosage adjustment recommendations, determine if there is sufficient

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78

●●

information about its kinetics to estimate the proper drug dose in renal failure carefully monitor treated patients for signs of efficacy and toxicity.

Considerations with Hepatic Impairment

Antimicrobial agents that are mainly eliminated by hepatic metabolism include some fluoroquinolones (enrofloxacin, difloxacin, marbofloxacin), trimethoprim, sulfonamides, minocycline, chloramphenicol and its derivatives, clindamycin, metronidazole, rifampin, and azole antifungal drugs with the notable exception of fluconazole. Macrolides and lincosamides, nafcillin, cefoperazone, and ceftriaxone are eliminated by biliary excretion but do not involve hepatic metabolism, while marbofloxacin is partly eliminated by excretion in bile as well as in urine. Even though tetracyclines (except minocycline and doxycycline) are excreted in bile, they are reabsorbed from the intestine and returned to the liver (enterohepatic recirculation) for reentry into the systemic circulation. Moderate or severe liver damage reduces the capacity of the liver to eliminate antipyrine (a marker substance for microsomal oxidative activity) and indocyanine green (marker substance for biliary secretion that may be influenced by liver blood flow). However, it is very difficult to quantify the degree of hepatic dysfunction and its influence on the clearance of lipid-­soluble drugs, making it hard to determine specific dosage adjustments. In general, the dosage interval for a drug that is mainly eliminated by hepatic metabolism should be increased in the presence of impaired liver

function if a renally eliminated drug is not an acceptable choice. Likewise, the dosage interval of an antimicrobial agent that is extensively metabolized by the liver should be increased when it is used concomitantly with a drug that inhibits microsomal oxidative reactions (such as ketoconazole, omeprazole, or cimetidine). Rifampin is known to induce hepatic microsomal oxidative activity and may interfere with the bioavailability of other antimicrobials administered concurrently.

Summary of Adjusting Dosage Regimens

Adjusting the dosage regimen is frequently required since antimicrobials are usually administered to diseased animals, while the dosage regimens often have been worked out in relatively normal animals. It is especially indicated when drug elimination or the drug’s volume of distribution is significantly altered due to disease in the animal, but specific guidelines have not been developed for most veterinary patients and even the most common pathological conditions. In general: ●●

●●

if the volume of distribution changes  – change the drug dose if the clearance or elimination half-­life of the drug changes – change the dosing interval.

For drug dosage regimens determined by therapeutic drug monitoring (e.g., aminoglycosides), modifications can be made on a percentage basis: New dose

old dose target concentration / measured concentration

R ­ eferences Chiesa OA, et al. 2006. Use of tissue-­fluid correlations to estimate gentamicin residues in kidney tissue of Holstein steers. J Vet Pharmacol Ther 29:99.

Macpherson M, et al. 2013. Disposition of desfuroylceftiofur acetamide in serum, placental tissue, fetal fluids, and fetal tissues after administration of ceftiofur crystalline

79

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  ­Reference

Pharmacokinetics of Antimicrobials

free acid (CCFA) to pony mares with placentitis. J Vet Pharmacol Ther 36:59. Murchie TA, et al. 2006. Continuous monitoring of penicillin G and gentamicin in allantoic fluid of pregnant pony mares by in vivo microdialysis. Equine Vet J 38:520. Nielsen P, Gyrd-­Hansen N. 1996. Bioavailability of oxytetracycline, tetracycline and chlortetracycline after oral administration to fed and fasted pigs. J Vet Pharmacol Ther 19:305. Rolan PE. 1994. Plasma protein binding displacement interactions – why are they still regarded as clinically important? Br J Clin Pharmacol 37:125. Schmidt S, et al. 2010. Significance of protein binding in pharmacokinetics and pharmacodynamics. J Pharmaceut Sci 99:1107.

Toutain PL, Bousquet-­Melou A. 2002. Free drug fraction vs free drug concentration: a matter of frequent confusion. J Vet Pharmacol Ther 25–460. Toutain PL, Bousquet-­Melou A. 2004a. Bioavailability and its assessment. J Vet Pharmacol Ther 27:455. Toutain PL, Bousquet-­Melou A. 2004b. Plasma clearance. J Vet Pharmacol Ther 27:415. Toutain PL, Bousquet-­Melou A. 2004c. Plasma terminal half-­life. J Vet Pharmacol Ther 27:427. Toutain PL, Bousquet-­Melou A. 2004d. Volumes of distribution. J Vet Pharmacol Ther 27:441. Zeitlinger MA, et al. 2011. Protein binding: do we ever learn? Antimicrob Agents Chemother 55:3067.

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80

5 Pharmacodynamics of Antimicrobials Andrew P. Woodward and Ted Whittem

Classically, in teaching, pharmacodynamics has been defined as “what the drug does to the body,” in contrast to pharmacokinetics which is “what the body does to the drug.” In modern scientific practice, these concepts overlap under the broader banner of quantitative phar­ macology and are integrated in antimicrobial dosage design. The antimicrobials, along with the antiparasitic and antineoplastic agents, are unique among the therapeutics in that their pharmacodynamics do not express only those effects on the biochemistry or physiology of the patient but also that of a foreign invader. Therefore, considering the action of the drug on only the “body” is not quite correct, as the pharmacodynamics are complex actions of the drug on the interaction of the body and patho­ gen. Nonetheless, similar principles may be applied. An important implication for veterinary clinical practice is that the pharmacodynamics ensuring the efficacy of antimicrobials are largely pathogen specific, rather than host spe­ cific, so similar principles are applicable for diverse patients. In this chapter, we will explore the foll­ owing  key concepts in antimicrobial pharmacodynamics. ●●

The mathematical foundations of pharma­ codynamics, and how popular measures of

●●

●●

antimicrobial effect are related to the funda­ mental theory. Popular clinical models of pharmacody­ namics, and how dosage regimen recom­ mendations are derived from basic principles. Scientific techniques that have been applied to study pharmacodynamics and the impli­ cations they have for clinical reasoning.

­ easurements and Models M in Pharmacodynamics A Clinical Scenario The treatment of disease in wildlife species is an emerging area of importance for veterinary therapeutics, with diseases such as chytridio­ mycosis, sarcoptic mange, chlamydiosis, and others important in Australia, for example (Smith et  al.,  2009). A challenging aspect of this undertaking is that clinical evidence in the form of randomized trials may not be availa­ ble, and patient and pathogen diversity mean that direct extrapolation from clinical experi­ ence may be unreliable. Fortunately, as explored in this chapter, we can apply a ­bottom-­up style of reasoning to utilize our knowledge of antimicrobial drug action, even when we only have partial information.

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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81

Pharmacodynamics of Antimicrobials

Consider the following example: A colleague working in a zoological institution contacts you for help with a disease outbreak in a cap­ tive population of lizards, which are managed in a captive breeding program. This lizard spe­ cies is critically endangered; no individuals are known to exist outside this managed popula­ tion. The disease outbreak is potentially devas­ tating for the future of this species; several individuals have died and at least a dozen more appear to have consistent clinical signs. Postmortem examination is consistent with bacterial pneumonia and secondary sepsis, so you send a series of samples to a reference microbiology lab for investigation. The following afternoon, you receive a phone call from the microbiologist, who seems quite excited. Apparently, the laboratory team has isolated heavy pure growth of fast-­growing Gram-­negative bacteria, genus Pseudomonas, from all the postmortem samples. Previously, this organism has only been associated with disease in invertebrates. The laboratory team is keen to pursue a deep investigation to publish its findings about a novel pathogen; however, you have a more pressing problem. What action should you take to protect this popula­ tion from disease? Isolation and biosecurity may help to prevent spread but will be of no benefit to those already infected. Vaccination might protect the whole population but inevi­ tably requires substantial time and resources. Antimicrobial therapy seems necessary. But which drug? What dosage regimen? You’re just starting your literature search when you receive an email from the laboratory: “We just finished running some broth microdilution tests on that organism. I believe enrofloxacin is widely used in reptiles? Looks like the enro­ floxacin minimum inhibitory concentration (MIC) is 1 μg/ml. The guideline doesn’t include any breakpoints for this organism but maybe you have some other information about the appropriate dose for this drug?” This type of situation presents a substantial therapeutic challenge. There is a compelling need for antimicrobial therapy but past clinical

experience provides little direct guidance. Evidence-­based medicine principles suggest that controlled trials are key support for thera­ peutic decisions but no relevant studies are available. How can more accessible pharmaco­ logical information be applied to make reason­ able, bottom-­up decisions about antimicrobial therapy? ●●

●●

●●

What does the MIC, among other laboratory measures, tell us about antimicrobial drug action? Can the MIC, measured in vitro, provide use­ ful predictions about drug activity in vivo? What is required to translate our knowledge of antimicrobial drug effect to clinical action?

Similarly, after the identification of chytridi­ omycosis, a critically important fungal disease of amphibians, veterinarians attempted to treat the last known members of an Australian species, and wrote about their experience (Banks and McCracken,  2002). Though these examples seem extreme, every patient is ulti­ mately unique and so every clinical decision requires consideration of a network of sup­ porting evidence. These same questions are useful to consider in all our clinical applica­ tions of antimicrobials.

The Minimum Inhibitory Concentration: A Fundamental Quantity In the clinical scenario, our colleagues from the microbiology laboratory reported the MIC to describe the effect of enrofloxacin. The MIC is the simplest quantitative summary of the effect of an antimicrobial drug in common use. As we will explore in this chapter, the MIC summarizes a functional relationship with just one value but the ease of generation, standard­ ization, and interpretation has made the MIC the most widely reported and utilized measure of antimicrobial effect. To determine the MIC, the microorganism of interest, typically bacteria, are inoculated into a suitable growth medium (Chapter  2). The experiment is usually conducted on

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82

a  micro-­volume cell culture plate, such as a ­96-­well plate. The growth medium contains antimicrobial drug in a dilution series, typi­ cally a two-­fold dilution. In clinical laborato­ ries, for antimicrobial agents that are well characterized, the actual drug concentrations are standardized. In the development or research setting, the appropriate concentration range to test may be an open question, which may warrant use of a different dilution series (such as a five-­fold or ten-­fold series). The final assay therefore includes a growth assessment for each of a series of drug concentrations. The assessment is usually by a simple visual exami­ nation of each well, and is binary: either growth is observed (negative response) or it is not (positive response). An important characteristic of the MIC is that the observations actually represent a range

rather than a point value. Presuming that there is a “true” underlying MIC value, MICtrue, that the assay is attempting to estimate, the observed MIC, MICobs, represents an interval in which MICtrue is estimated to occur. Because the assay response at the MIC was negative, the true MIC must be lower than that; con­ versely, because the response at the concentra­ tion below the MIC was positive, the true MIC must be higher than that. The observed MIC is therefore interval censored; the true minimum concentration could be the observed MIC but it must be smaller than the next concentration above the observed MIC. An MICobs value of 2  μg/ml, for example, represents any value greater than 1, and any value less than or equal to 2, and could be expressed as (1,2] μg/ml (Figure  5.1). This is an important considera­ tion in the epidemiology of MICs for resistance

0.20

Probability Density

0.15

0.10

0.05

32

16

8

4

2

1

0.5

0.25

0.125

0.0625

0.03125

0.015625

0.0078125

0.00

Concentration (μg/mL)

Figure 5.1  Histogram capturing an observed MIC distribution for a sample of a hypothetical bacterial population. The underlying MICtrue was simulated from the normal distribution, on log2 scale, with mean 0.5 and standard deviation 2. The solid line captures the observed probability density of the MICtrue, while the histogram represents the MICobs under the usual case of interval censoring. Histogram break labels are centered, reflecting the MICobs. Adapted from van de Kassteele, 2012.

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­Measurements and Models in Pharmacodynamic  83

Pharmacodynamics of Antimicrobials

monitoring, and experimental determination of MIC for new antimicrobials (van de Kassteele et al., 2012). The MIC has substantial importance, as both the most common statistical summary of anti­ microbial potency and as a parameter in phar­ macodynamic models that express our understanding of drug effect in vivo. Potency in pharmacodynamics refers generically to the dose or drug concentration at which drug effect occurs. A widely discussed concept is that drug potency does not indicate efficacy, because it reflects only the dose at which effect occurs, not the size of the effect. A classic example of this principle is the use of the low-­potency opioids morphine and methadone in the dog and cat; analgesic efficacy is substantially better than the higher-­potency buprenorphine or butorph­ anol but a larger dose is required. The MIC can be considered as a hybrid measure, which reflects both a statement about the effect size and a statement about potency, and is closely related to other pharmacodynamic measures.

Quantitative Descriptions of Drug Action Similarly to pharmacokinetics, pharmacody­ namics has strong underlying mathematical theory, and much of our understanding and the way we communicate about pharmacody­ namics are linked to that theory. Also like pharmacokinetics, the theory can be applied quite directly to clinical questions. Here is a simple conceptual model of drug effect: Equation 1:

E X

f X

where X is the concentration of drug at its site of action, E is the magnitude of the drug effect, and f (X) is the functional relationship between concentration and effect. This expression is read as a statement that the drug effect at some drug concentration depends upon the concentration. Intuitively, when X is zero, E is also zero (though in the real biology, other processes

might also influence the observed effect). Pharmacodynamics can be simply defined as the study of f(X) – the quantitative relationship between effect and drug concentration. In con­ trast, pharmacokinetics can be considered as the study of factors that control X. In living sys­ tems, drug action is inevitably occurring along­ side drug disposition, meaning that X is not constant or controlled but is subject to diverse biological processes. By contrast, in the labora­ tory, pharmacodynamics is studied in isolated in  vitro systems in which drug concentration can be closely controlled. Our understanding of f(X) as it occurs in vivo is closely linked to our understanding of the time course of X, and popular clinical models for antimicrobial effect actually pool information regarding X and f(X). The receptor–ligand theory of drug action (Rang,  2006) poses that all pharmacological effects occur because of a direct chemical interaction between the drug and some bio­ chemical process of a living system. Most drug action occurs via reversible chemical interac­ tion between the drug molecule and one or more target molecules, which are usually pro­ teins. These interactions typically involve hydrogen bonds, in which a weak electrostatic relationship forms between a hydrogen atom and an electronegative oxygen, nitrogen or fluorine atom. The biochemical target of drug action may be a membrane-­bound receptor, a cytoplasmic second messenger, a membrane-­ spanning channel, or an enzyme, among other possibilities. Depending on the receptor type and the specific drug, the function of the recep­ tor is suppressed, triggered or modified when drug binds to it. Formally, these interactions may be described as equilibrium processes: Equation 2:



D

R

Kd

DR

The interaction of free drug D with receptor R results in the formation of a drug–receptor complex DR, and the complex DR dissociates to return free drug D and receptor R; the con­ centration [DR] at equilibrium (the state at which there is no net change occurring over

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time) is dependent on the concentrations of drug [D]  and receptor [R], and on the drug’s binding affinity. The binding affinity is deter­ mined by how energetically favorable the product is relative to the reactants. Binding affinity is expressed by an equilibrium disasso­ ciation constant Kd. Note that the bidirectional Equation 2 can be equivalently expressed as two linked unidirectional reactions, each with their own rate, and Kd is the ratio of those rates. Because the reaction is bidirectional, Equation 2 implies that after any individual drug–receptor interaction has occurred, both the drug and target molecules are recovered unchanged. The action of the majority of anti­ microbials is of this reversible type, with some important exceptions. Drug action occurs via formation of the complex DR – either agonism, in which drug binding stimulates the activity of R, or antagonism, in which drug binding inhibits the activity of R. Probably the most fundamental task in phar­ macodynamics is to determine the shape of the relationship f(X); in other words, to learn the relationship between drug concentration and drug effect from experimental data. This is a sta­ tistical task; we obtain an understanding of the system by examining observations of its behav­ ior. Similar to learning from experimental data in pharmacokinetics, a simple mathematical model is a useful way to ground our analysis in theory. Some important clues are given by Equation 2. For an equilibrium reaction, in a low-­concentration state for the drug, the con­ centration of the drug–receptor complex at equilibrium is also relatively low. If the effect of a drug is causally related to the concentration of the drug–receptor complex, the observed drug effect would be correspondingly low. At a high drug concentration, the concentration of the drug–receptor complex is also relatively high, because the equilibrium reaction energetically favors formation of the product (balance is shifted to the right), and a correspondingly high drug action would be observed. An interesting implication of Equation 2 is that the shape of the relationship between drug

concentration and effect is not linear. In the transition from a very low concentration state to a very high concentration state, drug effect will also increase but the amount of increase will not be constant. This is difficult to appreci­ ate directly from Equation 2. A reformulation of Equation 2 called the Langmuir equation, a classic model from physical chemistry, can be specified in terms of drug concentration [D] and dissociation Kd: Equation 3:

D Kd

D

where the proportion (between 0 and 1) of receptor binding θ at equilibrium is a func­ tion of both the drug concentration [D] and the disassociation rate constant Kd. This model presumes that the receptor concentration is constant, which is mostly reasonable in pharmacology. Figure 5.2 is a classic visualization of the general form of this relationship between a drug concen­ tration [D] and the proportion of the recep­ tor occupied, θ. The most important feature of Figure 5.2 is that for drugs with a single site of action, the transition from minimal drug effect to maximal drug effect occurs within a relatively narrow concentra­ tion range. In practice, this underlying equilibrium reaction is difficult to work with, and is only indirectly related to our process of interest, the relationship between concentration and antimicrobial effect. Fortunately, a sigmoidal dose–response relationship can be conveni­ ently defined using its own equation. The classic sigmoidal dose–response curve can be expressed as (note the similarity to Equation 3): Equation 4:

E X E MAX

X X EC50

An interesting feature is that the drug effect is expressed as a proportion of its maximum. Rearranging the equation by multiplication of

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­Measurements and Models in Pharmacodynamic  85

Pharmacodynamics of Antimicrobials

1.00

Proportion of Receptor Occupied

Kd = 1 0.75

Kd = 10

0.50

0.25

Kd = 100

0 1e-01

1e+01 1e+03 Drug Concentration (Arbitrary unit)

Figure 5.2  A simple drug–receptor interaction model, Langmuir’s isotherm as in Equation 3, describing the proportion of receptor occupied θ as a function of drug concentration [D] (here in arbitrary units). The equilibrium dissociation constant Kd is set to one of three values (1, 10, or 100), which appear ordered from left to right. Note the logarithmic scale of the x-­axis.

both sides by EMAX allows this parameter to be determined: Equation 5:

E X

X E MAX X

EC50

This expression contains two parameters; a parameter is a quantity in our system that takes a constant but potentially unknown, value. EMAX defines the maximum possible effect, which is the effect E when the concentration X is infi­ nitely large. This expression proposes that the effect E is zero where drug concentration is zero. In many cases, this is not reasonable. For exam­ ple, the antifungal drug terbinafine (Chapter 19), a popular therapy for dermatophytosis, inhibits the enzyme squalene epoxidase which converts the metabolite squalene into precursors for ergosterol, a key cell membrane component. Squalene is present in the normal fungal cell but is toxic at high concentrations, so inhibiting this enzyme results in fungal cell damage. We could define the drug effect of terbinafine simply as

the squalene concentration, which could be measured in an in vitro experiment. As terbin­ afine concentration increases, the squalene con­ centration will also increase. However, some squalene is present even if the terbinafine con­ centration is zero. In this case, Equation 6 would be more suitable: Equation 6: E X

E MIN

X E MAX X

E MIN

EC50

Now we have parameters representing both the maximum effect (EMAX) and the minimum effect (EMIN). The value of E actually approaches these limits asymptotically (they are not reached at any real concentration), which is practically reasonable, as with real measurements we cannot distinguish zero from arbitrarily small values. The remaining parameter EC50 is typically of most interest. This can be interpreted as the

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86

EMAX = 150

150

Response (Arbitrary unit)

EMAX = 125

100

EMAX = 100

50

0 1e-01

1e+01 1e+03 Drug Concentration (Arbitrary unit)

Figure 5.3  A typical sigmoid dose–response model, as in Equation 6, describing the system response E as a function of drug concentration X (both here in arbitrary units). The maximum possible effect EMAX is set to one of three values (150, 125, or 100), which appear ordered from top to bottom. EMIN is set to zero and EC50 to 10. Note the logarithmic scale of the x-­axis.

concentration at which the response is half of the maximum response. Strictly speaking, it is the concentration at which the effect is equi­ distant between the EMIN and EMAX, as speci­ fied in Equation 6. In some systems, depending on how response is defined, the EMIN may not be zero, in which case the effect at EC50 will not represent “half” but in practice the data are often scaled to set the minimum effect at zero. In that case the parameters then represent relative effects. This reflects the emphasis on the estimation of EC50 as a single summary of drug effect but it does not contain any information about the magnitude of the effect. Practical studies in drug development often focus on this parameter when reporting pharmacodynamics of novel drugs or systems. A good way to understand the implications of these equations is to visualize them. Modifying the EMAX has the effect we would expect, which is that the y-­axis position of the

upper asymptote changes directly with the value of EMAX (Figure 5.3). Similarly, modifying the EMIN simply changes the y-­axis position of the lower asymptote (Figure 5.4). An important observation is that these adjustments to the EMAX and EMIN have no influence on the horizontal position of the rela­ tionship, that is, the drug concentration at which drug effects occur. The x-­axis position of the relationship is exclusively modified by the EC50 (Figure 5.5). Increasing the EC50 shifts the x-­axis posi­ tion of the relationship to the right, meaning that the same drug effect occurs at higher con­ centration. Decreasing the EC50 shifts the x-­axis position to the left, representing effects occurring at lower concentration. The EC50 is therefore often utilized as an expression of drug potency. Qualitatively, potency is simply the inverse of EC50, i.e., high-­potency drugs

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­Measurements and Models in Pharmacodynamic  87

Pharmacodynamics of Antimicrobials

Response (Arbitrary unit)

150

100

50

EMIN = 50 EMIN = 25

EMIN = 0 0 1e-01

1e+01 1e+03 Drug Concentration (Arbitrary unit)

Figure 5.4  A typical sigmoid dose–response model, as in Equation 6, describing the system response E as a function of drug concentration X (both here in arbitrary units). The minimum possible effect EMIN is set to one of three values (50, 25, or 0), which appear ordered from top to bottom. EMAX is set to 150 and EC50 to 10. Note the logarithmic scale of the x-­axis.

100

EC50 = 1

Response (Arbitrary unit)

75

EC50 = 10

50

EC50 = 100

25

0 1e-01

1e+01 1e+03 Drug Concentration (Arbitrary unit)

Figure 5.5  A typical sigmoid dose–response model, as in Equation 6, describing the system response E as a function of drug concentration X (both here in arbitrary units). The concentration at which effect is half-­maximal, EC50, is set to one of three values (1, 10, or 100), which appear ordered from left to right. EMAX is set to 100 and EMIN to 10. Note the logarithmic scale of the x-­axis.

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88

have a low EC50, meaning that their effect occurs at low concentration; conversely, low-­ potency drugs have high EC50, i.e., their effect occurs at high concentrations. This highlights the common misuse of potency as a synonym for efficacy, which is in fact is related to the magnitude of the maximum drug effect (EMAX), not to the amount of drug required to generate that effect. High-­potency agents may have low clinical efficacy if their maximum effect is weak, and low-­potency agents may have high clinical efficacy if their maximum effect is large. Therefore, in pharmacody­ namic studies we should be careful to assess the entire estimated relationship, rather than focus on individual parameters. In clinical practice, potency is one of the most important factors that dictates drug dose. Although the EC50 is often considered a default reporting standard for drug potency, which is convenient as it is a primary parame­ ter in popular models like Equation 6, some­ times other potency measures are used. For example, if the concentration causing a mostly complete effect is of more interest, the EC90 or EC95 might be selected instead (note that the EC100 is always infinite, by definition). Often the highest drug concentration achievable in vivo is constrained by a solubility limit, tox­ icity, or pharmaceutical considerations; if a near-­maximal effect is desired, this informa­ tion is poorly communicated by EC50. Equation 6 implies that the relationship between a drug effect and concentration can be completely expressed by the magnitude of the effect, and the midpoint between the asymp­ totic minimum and maximum effects. This omits an important process, which is the slope of the relationship between effect and concen­ tration. We can imagine a case in which the EC50, EMIN, and EMAX for two drugs under the same conditions are identical but the steepness of the change in effect with concentration is dif­ ferent between them. In equilibrium chemistry, a modification known as the Hill–Langmuir model (Hill, 1913) utilizes an exponent on X to modify the shape of the basic sigmoid function.

This expression, initially applied to the study of hemoglobin oxygen binding, is foundational to quantitative pharmacology. This model has the form: Equation 7: E X

E MIN

Xn

E MAX X

n

E MIN

EC50 n

The simpler model in Equation 6 can be thought of as a special case of this model, where the exponent n is equal to 1. The value of this exponent must be positive. We can visu­ alize how this value affects the shape of the concentration–effect relationship (Figure 5.6).

Applications of Pharmacodynamic Models The model expressed in Equation 7, among other similar forms, is a common pharmacody­ namic (dose–response) model in practical use, similarly to the compartmental model broadly applied in pharmacokinetics. Compared to Figure 5.4, note that in Figure 5.5 the horizon­ tal position of the function is not exclusively contributed by the EC50 except at X equal to the EC50, which highlights the potential impor­ tance of secondary parameters such as EC90. In Hill (1913), the exponent n is proposed to have a physical interpretation in terms of noninde­ pendent binding; that is, once some binding has already occurred, additional binding events are more or less likely than the first. The original use for Equation 7 was the description of oxygen-­binding kinetics of hemoglobin, in which multiple oxygen mole­ cules are bound to each hemoglobin molecule and n reflects the number of binding sites. The physical interpretability of these parameters is controversial (Weiss, 1997). In typical pharma­ codynamic studies, the parameters, especially n, should not be taken as physically meaning­ ful; they capture useful information about the behavior of the system but their value is not directly linked to any real quantity.

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­Measurements and Models in Pharmacodynamic  89

Pharmacodynamics of Antimicrobials

100 n=2

n=1

n = 0.5

Response (Arbitrary unit)

75

50

25

0 1e-01

1e+01 1e+03 Drug Concentration (Arbitrary unit)

Figure 5.6  A typical sigmoid dose–response model including Hill coefficient, as in Equation 7, describing the system response E as a function of drug concentration X (both here in arbitrary units). The coefficient n which controls the steepness of the function is set to one of three values (0.5, 1, or 2) in order of increasing steepness. EMAX is set to 100, EMIN to 0, and EC50 to 10. Note the logarithmic scale of the x-­axis.

This issue highlights an important scientific principle, which has substantial implications for our understanding of pharmacology generally but especially pharmacokinetics and pharmaco­ dynamics. Our sigmoid concentration–response model is a strongly empirical model. This model is a useful tool to formalize our understanding of experimental data but it has no clear relation­ ship to real physical characteristics of the actual system being studied. Introducing the exponent parameter n increases the flexibility of our model, so that it is able to align more closely with our observations, but this does not mean that the value of the parameter n corresponds to any particular physical process or event. The popular pharmacokinetic parameters, such as clearance and volumes of distribution, have sim­ ilar limitations. Though it is very clear that parameters of common statistical models, such as linear models, have no physical interpretabil­ ity at all, pharmacokinetic and pharmacody­ namic parameters are partially mechanistic, so

the limits of their interpretation need to be care­ fully appreciated. The same underlying model is often applied more generally, with indirect measures of con­ centration and response substituted. For exam­ ple, a classic concept is a “dose–response” model, in which the predictor variable X is replaced with an administered dose, which is more direct. A further abstraction is to replace the drug effect with a population statistic rep­ resenting the proportion of subjects that have positively responded. For example, in a dose-­ finding study observing a large number of treated patients with a range of administered doses, the response may be defined as the cumulative proportion of subjects who have a successful outcome, which may be a useful approach to evaluate the benefit of antimicro­ bial therapy at different doses in clinical usage. The final model expressed in Equation 7, and various reparameterizations, can be applied to data from diverse experimental designs. Like

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90

the compartmental models applied in pharma­ cokinetics, this model is statistically nonlinear, so requires iterative techniques for statistical analysis; multiple sets of parameter values are systematically searched to find those that pro­ vide the best fit to the data, given the proposed model structure. Similarly to statistical practice in pharmacokinetics (Chapter  4), two-­stage methods in which subject-­level data are indi­ vidually assessed have predominated in the past and remain common, and are widely sup­ ported in many software environments. More cohesive and statistically rigorous estimation is provided by population methods, which use multilevel models to conduct analysis of all data together (Bon et al., 2018).

The Minimum Inhibitory Concentration is a Hybrid Parameter The various pharmacodynamic parameters that we have explored are often described in drug development studies in the preclinical setting. The common quantitative parameters are more high level and easier to communicate in the con­ text of clinical or epidemiology studies. We have already met one of the most popular, the MIC. As we have explored, the MIC is easy to conceptual­ ize, based on the experimental design from which it is determined. But is it a good summary of drug action? Does it have meaning in terms of our more fundamental model? If we’re willing to make some simplifying assumptions, we can express the MIC as fairly directly related to the sigmoidal model. It has a particularly close relationship with the classic measure of drug potency, the EC50 (Equation 3). Previously we expressed drug effect E as a real value, either directly or as a proportion of the maximum possible response. However, the experimental method of MIC determination pro­ poses that the drug effect is binary; at any drug concentration, either growth occurred or it did not. But the actual effects of antimicrobials, as implied by the sigmoid models, are not binary. We can consider MIC as a dichotomised version of a truly smooth underlying relationship

between concentration and effect. Instead of observing the effect directly, a threshold is applied, and the observed result for any concen­ tration is whether or not the response threshold was exceeded. The MIC therefore represents combined information regarding the antimicrobial effect. It is a potency measure, similar to the EC50, but also describes the size of the effect, which is an antimicrobial effect of sufficient magnitude that “no” pathogen replication occurs. Generally, in pharmacodynamics “potency”, as indicated by the EC50or similar parameter, represents noth­ ing about the degree of effect. In contrast, the MIC implies a large effect, at least in vitro. We can visualize the MIC, with reference to the underlying “true” relationship, as Figure 5.7. In contrast to the discrete MIC determined in the microdilution assay, the disk diffusion (Kirby–Bauer) method generates a smooth (continuous) measurement, which is propor­ tional to antimicrobial drug concentration but does not indicate drug MIC concentration directly. In the disk diffusion method, an agar plate containing appropriate growth medium is inoculated with a suspended culture of the pathogen, to achieve uniform deposition across the plate surface area. Antimicrobial-­ impregnated polymer disks are then placed on the agar surface. During the incubation period, antimicrobial drug diffuses from the disk into the agar medium. As the diffusion rate through the agar is slow, a concentration gradient develops during the incubation period, with drug concentration declining with increasing distance from the disk. Microorganism growth occurs across the agar plate surface, except in those regions where antimicrobial concentra­ tion is too high, corresponding to an estimate of the MIC. It is difficult to determine drug concentration at some position on the plate but the zone diameter is not usually reported directly but instead a judgment is made regard­ ing its clinical interpretation using a break­ point. which will be discussed later. The disk diffusion assay essentially repre­ sents a rearrangement of the microdilution

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­Measurements and Models in Pharmacodynamic  91

Pharmacodynamics of Antimicrobials

100 Sufficient Effect (No Growth) Insufficient Effect (Growth)

Growth Inhibition (%)

75

50

25 Growth (below MIC)

No Growth (at or above MIC)

0 0.0156 0.0312 0.0625 0.125 0.25 0.5 Drug Concentration (μg/mL)

1

2

4

Figure 5.7  The relationship between the MICtrue, MICobs, and a proposed underlying sigmoid concentration response model for a hypothetical antimicrobial drug. In the practical determination of MIC, only a discrete set of concentrations are observed, corresponding to the x-­axis labels. In this example, observed growth occurs unless the degree of growth inhibition exceeds 90% (dashed line). For this set of observed concentrations, the nominated MICobs is 0.5 μg/ml, as the smallest observed drug concentration at which the effect was sufficient to preclude visible growth (dotted line). At the next smaller concentration 0.25 μg/ml, the degree of growth inhibition was not sufficient. The latent MICtrue is between 0.5 μg/ml and 0.25 μg/ml.

assay, in which the unknown continuous con­ centration gradient replaces the known dis­ crete concentrations. In the laboratory, these may be run simultaneously, so that the zone diameter can be transformed to an estimate of the MIC. This is a more complete use of the information than the classic interpretation of discretising the zone diameter as “resistant” or “susceptible.” The continuous zone diameter is a surrogate variable for the MIC, and therefore antimicrobial potency.

The Minimum Inhibitory Concentration in Context We have explored the most common general quantitative description of drug action, and

the use of simple mathematical models as a theoretical foundation for concentration– effect relationships. These models contain a high degree of abstraction, and present quanti­ ties that might be difficult to estimate in ­routine scientific practice. However, the phar­ macodynamic measures that are frequently used in clinical and research practice are closely derived from the models we have explored, and considering them in this frame­ work illuminates their implications and assumptions. An interesting limitation of the MIC con­ cept, which is common to all the simple in vitro assays for antimicrobial potency, is that the  drug effect represented by the MIC is highly constrained. The “effect” expressed in

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Figure  5.7 has a very specific meaning: the degree of increase in density of a microorgan­ ism in culture, under specific environmental conditions (medium, temperature, oxygena­ tion, etc.), that occurred after a specified period of growing time, with a controlled starting cul­ ture density. Substantial effort is invested by the clinical laboratory community to achieve uniformity in these conditions, so that MIC values obtained on different occasions, by dif­ ferent analysts, and at different laboratories share common meaning. These reflect internal validity; that is, the reliability of the measure­ ment. The external validity of MIC, meaning the degree to which MIC information is clini­ cally meaningful, is a more complex issue, because this depends on how closely the study design reflects the events occurring in the patient. Small changes in experimental conditions may result in large changes in estimated MIC, and because those conditions have little ­relationship with the pathophysiology of infections, it is difficult to demonstrate how reasonable they are. Numerous effects may contribute to poor alignment of in vitro MIC to the magnitude of antimicrobial effects in vivo (Martinez et al., 2013). For example, the assay is usually conducted in a homogenous liquid culture, so the microorganism is exposed directly to the dissolved antimicrobial but in  vivo the microorganisms may be seques­ tered within the three-­dimensional geometry of the infected tissue, adding additional diffu­ sion distance and barriers. Some bacteria secrete biofilms, modifying their local envi­ ronment and acting as a diffusion barrier; however, some biofilms enhance the activity of specific antimicrobials. In static culture, the drug concentration is constant and metabolic wastes accumulate over time, whereas in vivo, circulation and tissue fluid turnover imply that drug concentration is subject to continu­ ous change, and nutrients and metabolic waste products undergo continuous turnover. In most infections, the effect of exogenous antimicrobials occurs together with an

inflammatory and immune response, which contribute endogenous antimicrobial effects with an important role in the outcome of infections but are not represented by the MIC. A key characteristic of the MIC is that it rep­ resents all antimicrobial effects equivalently as inhibition of growth. The specific effects of antimicrobials are in fact diverse although some do in fact inhibit metabolism, preventing cell activity and division, others inflict fatal cell damage over a shorter time. The MIC is a sur­ rogate in  vitro measure for relative efficacy, which is helpful for clinical planning because different antimicrobial–microorganism inter­ actions have a common interpretation. However, the MIC is not informative about the actual mechanism of effects. We have explored how the MIC is related to more fundamental, theoretical models of drug effect. The MIC combines a statement about the magnitude of effect, representing appar­ ently complete inhibition of microbial growth, with a statement about potency, the drug con­ centration at which the effect occurs. Let’s reconsider our starting example in these terms. The laboratory has reported that our novel bacterial pathogen infecting the lizards has an enrofloxacin MIC of 1 μg/ml. This demon­ strates that in continuous exposure in an in  vitro medium, the lowest concentration of enrofloxacin that will completely inhibit visi­ ble growth of our pathogen is somewhere between 0.5 μg/ml and 1 μg/ml. As described in Chapter 17, the fluoroquinolones, including enrofloxacin, are clinically most relevant for Gram-­negative infections. After reviewing the enrofloxacin MIC as reported from other Gram-­negative pathogens in reptiles, you find that most Pseudomonas isolates are classified as susceptible (Tang et al., 2020). ●●

It seems like our evaluation has just led to further questions: Pseudomonas spp., a com­ mon Gram-­negative pathogen, is often found to be “susceptible”; can we apply that inter­ pretation to our new pathogen? Why is this finding reported, and not the MIC?

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­Measurements and Models in Pharmacodynamic  93

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●●

“Susceptible” seems to imply that treatment would be effective. How can a prediction about clinical effectiveness be made from only in vitro data? If this patient species has never been stud­ ied, are our predictions trustworthy?

We will now move forward to consider these new questions, which we will classify as clinical pharmacodynamics.

­Clinical Pharmacodynamics In many clinical situations, including our liz­ ard example, we are forced to consider antimi­ crobial therapy from a bottom-­up perspective, based on theoretical knowledge, as we have no direct experience with our patient or pathogen. Despite obtaining some quantitative data about drug effect, it seems that we need to apply more information to evaluate our anti­ microbial drug selection. A key insight is to recognise that selecting an appropriate antimicrobial drug is not enough. The dose, frequency of administration, route of administration, and duration of therapy must all be specified. The objective of quantitative pharmacology, including pharmacodynamics, is to convert our understanding of drug–patient and drug–pathogen interactions into clinical decision making that maximizes the efficacy and safety of therapy. We have explored models that quantitatively describe antimicrobial effect; our next objective is to utilize those mod­ els to support rational clinical decision making with quantitative evidence. A useful conceptualization is that the disease system has three parts: a host, a pathogen, and the antimicrobial drug. The outcome of the treatment of infectious disease results from interactions between these systems. The inter­ action between host and drug is known as pharmacokinetics, as we have explored in Chapter 4, which characterizes the time course of drug concentrations as functions of parame­ ters which are estimated from experimental

studies. The interactions between pathogen and drug are known as pharmacodynamics, and as we have described so far in this chapter, this represents the relationship between drug concentration and effect, including the drug susceptibility of the pathogen. The final com­ ponent is the host–pathogen interaction; though typically considered outside the domain of pharmacology, this includes critical factors such as the pathophysiology, host immune response, and the clinical effects of disease. In practice, the outcome of infection treat­ ment depends on the simultaneous, combined effects of all these interactions. This is an over­ all scientific model by which the outcome of treatment for an infectious disease process can be understood, described, and predicted. This framework can be applied quantitatively, by defining formal mathematical and statistical models for the components, or qualitatively, as a tool to support clinical reasoning. These methods have been widely utilized in veteri­ nary applications (Toutain et al., 2021). After searching the scientific literature, you have located some pharmacokinetic reports that describe the pharmacokinetics of enro­ floxacin in various lizards and snakes; nothing about the species of interest but a report in a species of similar body size (Agius et al., 2020), a good example of the style of information typically available. Our objective is to deter­ mine what strategy should be applied in clini­ cal practice in order to achieve success with this drug. What are the pharmacodynamic principles that we need to consider? A conceptually simple approach to this prob­ lem is to conduct a randomized controlled trial, or a series of trials, and observe the clinical out­ come in an entirely empirical fashion. Where there is descriptive toxicology information avail­ able, which can be used to propose a maximal safe dosage or exposure, it is easy to select a small range of candidate dose regimens and apply them to either clinical patients or suitable experimental animals. The mode of reasoning being applied is simply that a treatment that was clinically effective in the past can reasonably be

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predicted to be effective in the future; no particu­ lar knowledge is required about the mechanism of its effectiveness. This is referred to as top-­ down reasoning. This remains the predominant path of clinical development for many veteri­ nary antimicrobial drugs; for example, intramam­ mary drug development has emphasized clinical trials almost exclusively, as suitable pharmacoki­ netic modeling has only recently become availa­ ble (Woodward et al., 2020). The testing of candidate dosing strategies for antimicrobial therapies using randomized con­ trolled trials is appealing; no knowledge about the mechanism of action is required, the study design is quite simple, communicating the findings is easy, and little specialized knowl­ edge is required to understand the results. However, the practical drawbacks of this approach are substantial. As the dose and dos­ ing frequency are approximately continuous variables, there is a practically infinite candi­ date set of dose regimens that could be tested, so the chance that the optimum strategy, or even close to an optimum strategy, will be selected is low. It is possible that efficacy might be reported as poor simply because a good strat­ egy was not selected for study. Further, the design provides little evidence regarding the expected outcome if conditions change, because there is no theoretical framework to ground predictions. Other problems with using randomized con­ trolled trials are statistical; as the design does not propose any knowledge about the relation­ ship between the dose strategy and outcome, the amount of information obtained from each subject is low, though these data are expensive to collect. Finally, practical challenges for ade­ quate use of randomized controlled trials abound: the number of subjects required is generally large because of the low information obtained from each subject, it is ethically chal­ lenging to apply experimental therapies to large numbers of subjects, and external valid­ ity may be poor as no information is available about the impact of changing conditions. Though a simple and direct strategy is to

extrapolate or adopt the dose regimen from another application, especially from human medicine, this takes little advantage of any species-­specific knowledge that is available, especially regarding pharmacokinetics. A clear issue in our scenario above, which is common in the discipline, is that there is sim­ ply no time or opportunity to conduct such a study anyway. These limitations may be resolved by imple­ menting a more bottom-­up approach: utiliza­ tion of a theoretical model to justify what relationships are expected, and feed in back­ ground information to our design. We will now explore the basis of these models.

Time-­Dependent Antimicrobial Activity Earlier, we explored how the MIC is a hybrid measure related to the potency of a particular antimicrobial drug for a particular pathogen of interest. Major advantages of the MIC are the ease of measurement for the majority of clini­ cally relevant bacteria, and the ease of stand­ ardization. As the MIC is closely associated with the antimicrobial potency, the concentra­ tion of drug above which relevant antimicro­ bial effects occur, it is natural to utilize pharmacodynamic information contained by the MIC in dose regimen evaluation. For simplicity, we will propose that the drug concentration to which the pathogen of inter­ est is exposed is equal to the free drug concen­ tration in plasma (CP). Remembering our sigmoidal model for the magnitude of drug effect (ignoring for now the coefficient n): Equation 8:  E X

E MIN

X

E MAX

E MIN

X EC50

In this case, we will substitute the plasma concentration CP: Equation 9:  E CP

E MIN

CP

E MAX CP

E MIN

EC50

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­Clinical Pharmacodynamic  95

Pharmacodynamics of Antimicrobials

By replacing the hypothetical drug concentra­ tion X with the plasma concentration CP, we have taken the first step towards an in vivo pharmaco­ dynamic model. Previously, we discussed the typical usage of the EC50 as a measure of drug potency and noted that the EC50 contains no information about the magnitude of the effect, only the concentration around which effect occurs. In contrast, the MIC reflects information about both the potency and the effect magnitude, being the lowest concentration with a high degree of inhibitory effect. With the MIC for the pathogen isolate of interest available, we can pro­ pose the concentration of antimicrobial drug CP that is likely to be required for efficacy. As we have explored, the MIC is closely related to Equation 8. This provides our starting point for assessing the in  vivo concentration–effect rela­ tionship. Assuming that the in vivo and in vitro relationships are similar, we can infer that a meaningful antimicrobial effect might occur in vivo and set the dose regimen accordingly. Continuing our case study above, let’s sup­ pose that you have made contact with a

former colleague who has recently completed a pharmacokinetic study in a related species. They have conducted a single-­dose, intrave­ nous bolus administration of a novel cephalo­ sporin drug, and in reviewing your case notes you were reminded of the Gram-­negative anti­ microbial activity of the later-­generation cephalosporins (Chapter  8). In this pharma­ cokinetic experiment, several different doses (0.5, 1, 1.5, and 3 mg/kg) were administered to a cohort of healthy lizards, and the average plasma concentration–time relationship was visualized for each dose (Figure 5.8). This system appears to be compatible with a one-­compartment first-­order pharmacokinetic model. You were able to request that the micro­ biology lab obtain an estimate of the MIC for our pathogen of interest for this cephalosporin. In this case, the MIC was approximately 1 μg/ml. Simple visual assessment of the concentration– time relationships shows that some of them mostly exceed the MIC but others do not. To apply this drug as a clinical therapy, which of these

Plasma Concentration (μg/mL)

4

3

2

1

0 0

18 6 12 Time Since Drug Administration (hr)

24

Figure 5.8  Predicted average plasma concentrations of a hypothetical cephalosporin drug after intravenous bolus administration in dogs, after doses of (ascending from bottom) 0.5, 1, 1.5 or 3 mg/kg body weight. The dashed line denotes the estimated MIC90 of a bacterial pathogen of interest.

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96

doses should be preferred? Is it possible to suggest how appropriate the dose is based simply on the time course of drug concentrations? The MIC is a basic summary of antimicrobial effect, so comparing the observed concentra­ tions to the MIC seems like a sensible start. However, complications become apparent immediately. The MIC captures information about effect but effect is defined in a very spe­ cific way: no visible growth after a defined period of constant exposure to a single drug concentration. In a living patient, drug concen­ tration is rarely constant, as the plasma concen­ tration of drug changes constantly throughout the interdosing interval. Though the MIC assay is conducted using a fixed time duration, actual infections have a different, characteristically longer, time course, so the effect indicated by the MIC does not map directly to the effect occurring in  vivo. These observations suggest that a formal strategy is needed to interpret the relationship between the time-­course of plasma concentration and the MIC. A reasonable interpretation is that the MIC represents a threshold concentration, above which pathogen growth sufficient to generate visible density cannot occur, dichotomizing the concentration–response relationship. Where the drug concentration in plasma exceeds the MIC, a high degree of antimicro­ bial effect is occurring. Where the drug con­ centration does not exceed the MIC, the degree of antimicrobial effect would not be sufficient in continuous exposure to completely prevent visible growth. This suggests that a sensible strategy for antimicrobial administration is to maintain plasma concentration greater than the relevant MIC, for as much of the interdos­ ing interval as possible, therefore matching the inhibitory effect observed in vitro. This strategy arises directly as an extrapolation of the concept of the in  vitro determination of MIC to the in vivo antimicrobial effect. In phar­ macokinetics, we imagine that the animal is a system of fluid-­filled containers; here, we have applied the same reasoning, by imagining the patient is a container of bacterial growth

medium, as in the MIC experiment! If the mag­ nitude of effect at the MIC is presumed to be large and clinically significant, this simple model may be applied to suggest the reasonableness of some proposed dose, or frequency of dosing. Relatively superior strategies would maintain drug concentration exceeding the MIC for a higher proportion of, or the entire, interdosing interval. Drug concentrations lower than the MIC could allow some nonnegligible rate of pathogen replication, if the in vitro conditions are indicative. This concept, the duration of time that drug concentration in the interdosing inter­ val exceeds the MIC, is referred to as the time-­ above-­MIC (T>MIC), and is a widely used model for antimicrobial effect. For some drug–pathogen combinations, the utility of this strategy is sup­ ported by direct clinical evidence, especially in humans (Muller et al., 2013; Tannous et al., 2020). The T>MIC is often referred to as a pharmaco­ dynamic, or pharmacokinetic-­pharmacodynamic (PK/PD), index, which reflects the fact that it is not actually a system parameter (an unknown constant in our equations) but instead emerges as a function of different factors. Depending on drug-­specific pharmacokinetic factors, a larger dose, more frequent administration, constant infusion, or delayed-­release preparations would each increase the T>MIC. Though more fre­ quent dosing or steady-­state administrations effi­ ciently increase the T>MIC, an increased dose also increases T>MIC. A clinically useful rule of thumb, assuming constant bioavailability and plasma clearance, is that doubling the dose increases the T>MIC by one terminal elimina­ tion half-­life. Though the T>MIC model is clearly applica­ ble for comparing the merit of two proposed dose regimens, it doesn’t allow an absolute judgment of any given dose regimen. Though a dose regimen that results in plasma concentra­ tions entirely below the MIC is likely subopti­ mal, for how long should the MIC be exceeded for the antimicrobial effect to be clinically suf­ ficient? A typical recommendation for the cephalosporins is for T>MIC of at least 60% of the inter-­dosing interval, and at least 50% for

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­Clinical Pharmacodynamic  97

Pharmacodynamics of Antimicrobials

the penicillins. These are somewhat context-­ dependent and depend on host and pathogen factors, and are drawn from a range of different lines of evidence including animal and labora­ tory models, so have been unsurprisingly con­ troversial. Recent studies have suggested that targeting T>MIC of the entire dosing interval is most reasonable but obtaining relevant out­ come data from patients is challenging (Barreto et  al.,  2021). An important consideration is that this evidence is drawn mostly from humans, and veterinary-­specific evidence is sparse. Those recommendations cannot be derived mechanistically, so have been deter­ mined based on expert assessment of a combi­ nation of animal and in  vitro models and observed outcomes. It is widely thought that the T>MIC model is a reasonable predictor of clinical effect of the beta-­lactam antibiotics. This was proposed very early in their development history, and remains the predominant model in clinical use in human and veterinary patients. The reason­ ableness of this model is supported by a diver­ sity of evidence, ranging from computational modelling of in vitro data, experimental animal models of severe infections, and evaluation of the clinical outcome in human patients. The archetypal beta-­lactam antibiotic is penicillin G, which remains widely used in large animal medicine and is available in multiple prepara­ tions. In a classic study by Love and colleagues (Love et  al.,  1983), the terminal elimination half-­life and time above MIC for several prepa­ rations were compared. It was clear that some formulations demonstrated flip-­flop kinetics (Chapter 4). As the sodium or potassium salt, when administered by intramuscular injection the absorption was rapid, with terminal elimi­ nation half-­life of about 1.5 hr, so plasma con­ centration declined rapidly below MIC. The typical formulation of penicillin G used in the horse, the procaine salt, had terminal elimina­ tion half-­life of about 20 hr, because compared to the sodium or penicillin salt the disassocia­ tion to the ionized form is slow and this delays absorption. Even greater slowing of absorption

is provided by the benzathine salt but even with larger doses administered, observed peni­ cillin concentrations were erratic, and lower than other preparations. In all cases, the merit of any given drug and dose regimen are context-­dependent, and patient and pathogen factors have an important influence on the appropriateness of any drug and dose regimen selection. Though the antimi­ crobial effect of the beta-­lactams including pen­ icillin G is predicted by the T>MIC, which would imply that slower-­absorption prepara­ tions would be preferred as they extend drug exposure over a longer time range, the achieved plasma concentration remains critical. Counter-­ intuitively, very slowly-­absorbed preparations risk having a short effective duration of action, if they are absorbed so slowly that drug concen­ tration is not sufficiently high to reach MIC. On a similar note, the description of antimicrobials, especially beta-­lactams, as “long-­acting” based on their pharmacokinetics alone is problematic, as the duration of antimicrobial action under the T>MIC model depends on the MIC; as the MIC increases, the duration of action, other fac­ tors being equal, declines, and the duration of action is zero if the MIC is never reached. The T>MIC paradigm provides evidential support for the selection of the dose and inter-­ dosing interval. However, it does not provide any support for selecting the duration of ther­ apy. Consider the conceptual relationship between the T>MIC model and the experi­ mental conditions under which the MIC is generated; MIC experiments are conducted over fixed time duration, which has no in vivo relevance. An interesting implication regards the use of delayed-­release or slow-­elimination preparations of the beta-­lactam antibiotics, in which the prolonged target action blurs the distinction between the inter-­dosing interval and the duration of therapy. For example, the antibiotic cefovecin has very slow elimination in the dog and cat due to its high degree of pro­ tein binding (Chapter  4), so the T>MIC con­ cept can be applied to suggest the duration of effective action for a single dose, More broadly,

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98

selection of the duration of antimicrobial ther­ apy has limited evidential basis. Clinical trial systems typically have poor external validity for infections seen in clinical practice, so the duration of therapy is predominantly chosen based on clinical judgment or expert recom­ mendation, with resulting wide variability (Barzelai and Whittem, 2017).

Concentration-­Dependent Antimicrobial Activity The T>MIC was the first quantitative model proposed to describe antimicrobial action. Several studies, via the development of then-­ novel animal models, demonstrated that anti­ microbials were in fact diverse in their quantitative mechanism of action, and other aspects of the concentration–time relationship were more predictive of efficacy. In clinical practice in the horse, gentamicin and other aminoglycosides have been widely applied, primarily for the treatment of Gram-­ negative infections. In all mammals, including the horse, the aminoglycosides are clinically characterized by substantial toxicity. Though aminoglycosides were marketed and available before the emergence of pharmacodynamic methods for dose determination, their high toxicity suggested them as good candidates for quantitative optimization, in an effort to maxi­ mize antimicrobial effect while minimizing toxicity. Current dosing recommendations for aminoglycoside therapy in the horse are based primarily on quantitative optimization. In the simple model we proposed for T>MIC as a description of the antimicrobial effect, a key implication is that once the necessary threshold value is reached, increasing the concentration further would not provide any additional bene­ fit. This has been referred to as time-­dependent activity. In the context of the simple sigmoid model, where the inhibitory effect is a simple constant function of concentration, it is reason­ able to expect that a suitable strategy would be simply to maintain a constant high drug

concentration. However, if drug exposure causes a long-­lasting dysfunction, then sus­ tained high drug concentration may be unnec­ essary, and periodic peaks may be optimal. Taking advantage of this suggests a strategy of targeting the highest possible drug concentra­ tion to ensure maximum effect but relying on the persistent dysfunction to continue the inhibitory effect during the interdosing interval. It is generally accepted that this postantibi­ otic effect (PAE) is a key contributor to the concentration-­dependent activity of the ami­ noglycosides, which is described by the maxi­ mum plasma concentration to MIC ratio (CMAX:MIC). In contrast to the T>MIC strat­ egy, in the CMAX:MIC strategy, only the high­ est achieved concentration is considered and the time dimension is formally ignored. This concept is referred to as concentration-­ dependent activity. The CMAX:MIC strategy demands the highest safely achievable drug concentration at the site of exposure, and to reduce toxicity, this peak concentration should be maintained for only a short time. This can be achieved by using rapidly absorbed prepara­ tions, or intravenous bolus administration where feasible. As the initial drug concentra­ tion after an IV bolus is a function of only the central volume of distribution (VC), and not the clearance, it is simple to suggest the merit of any particular dose, if an estimate of the VC is available. Greater antimicrobial effect is achieved simply by using a larger dose. Of course, in a real clinical case a dosing fre­ quency is also required; once-­daily is typical but this is a matter of clinical judgment and is not part of the CMAX:MIC model. The ongoing use of the CMAX:MIC model in the clinical setting is somewhat controversial, with practical recommendations for the ami­ noglycosides now emphasizing the related but distinct AUC:MIC model (Bland et al., 2018), which will be reviewed in the next section. General consensus has been reached that for the CMAX:MIC model, sufficient antibacterial effect requires that the CMAX:MIC exceeds 10–12 for the aminoglycosides. This model has

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Concentration-­Dependent Antimicrobial Activity  99

Pharmacodynamics of Antimicrobials

been widely applied for dosage optimization for gentamicin and amikacin, especially in adult horses and foals (Bucki et al., 2004). Among the antifungal agents, a substantial rapid-­killing effect is most associated with the polyene class, most importantly with ­amphotericin-­B, which is the only member of this class that has been used systemically. Both the aminoglycosides and amphotericin-­B are strong candidates for quantitative optimi­ zation, as excessive duration of exposure is associated with substantial toxicity. However, conservatism in the dose regimen is not gener­ ally warranted, as the use of these agents in advanced systemic infections, especially sep­ sis, implies that failure to achieve a sufficient antimicrobial effect is likely to have a highly adverse outcome. The conflict between maxi­ mizing drug concentration to ensure efficacy while simultaneously minimizing toxicity has motivated widespread application of pharma­ cokinetics of these antimicrobials to guide critical dose decisions, especially in special populations. Similarly to the time-­dependent effects model, the concentration-­dependent effects model offers no direct guidance regarding the duration of therapy. Further, as the evalua­ tion of the size of the antimicrobial effect is drawn only from a single point concentration value, it is not informative regarding the fre­ quency of drug administration either. This model therefore only supports selection of the dose. In practice, usage of these agents has emphasized large intermittent doses that generate high peak plasma concentration, with interdosing intervals sufficiently long that most of the administered dose is elimi­ nated before the next dose, to minimize potential for accumulation of drug and resulting toxicity. It is proposed that an important contribu­ tor to the concentration dependence, and the appropriate duration of the dosing interval, is the duration of the postantibiotic effect. In our pharmacodynamic models, antimicrobial effects are related directly to the current drug

concentration, which in  vivo is typically the plasma concentration. PAE is simply persis­ tence of drug effect after drug concentration has reduced. The antimicrobial effect at any time is related not only to the drug concen­ tration at that time but also to the recent his­ tory of drug concentration. The PAE has been observed extensively, for many antimicrobi­ als, using various laboratory methods to monitor bacterial physiology (Stubbings, 2006). There is relatively limited evidence of the mechanism of the PAE. The most compelling current explanation is that the PAE reflects the time taken for antimicro­ bial drug to leave the target cell or to be dis­ sociated from the site of action; though drug concentrations in the organism’s environ­ ment have declined, a disproportionate amount of drug remains closely associated with the pathogen (Srimani et  al.,  2017). In current clinical practice, the dosing interval and total duration of therapy are a largely matter of judgment regarding progress of the individual case. While this judgment may be supported by epidemiological evidence, it is generally not supported by quantitative optimization.

Exposure-­Dependent Activity The time-­dependent and concentration-­ dependent models essentially represent two extremes: one case in which the clinical bene­ fit is predicted solely by a function of time, and the opposite case in which only the maxi­ mum drug concentration is important. Of course, in the in vivo case, antimicrobial drug exposure, the time-­course of X in our Equation 1, comprises both time and concen­ tration dimensions. The area under the curve (AUC) offers a description of the combination of concentration and time; a larger AUC may result from more drug being present, slower drug elimination, or both. This can be appreci­ ated by visualizing the AUC as a polygon formed from the time (x-­axis) and concentra­ tion (y-­axis).

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100

As the AUC is a function of both the drug concentration and time, it is strongly corre­ lated with both the maximum plasma concen­ tration and the time over which the MIC is exceeded. Strategies intended to increase the maximum plasma concentration by, for exam­ ple, increasing the dose also proportionally increase the AUC. An emphasis on increasing the T>MIC also potentially increases the AUC, depending on what is measured. For example, if the AUC is calculated as the total area over 24 hours (AUC24), then decreasing the inter­ dosing interval (say, by dosing eight-­hourly instead of 12-­hourly) will proportionally increase the AUC24. However, for single-­dosing events, the AUC is influenced only by the dose (assuming constant clearance); regarding preparations of penicillin G for example, as we explored earlier, which differ only in their absorption rate, the AUC is not different between preparations despite the substantially different concentration–time profile. Most famously, efficacy of the fluoroqui­ nolone antimicrobials, including the arche­ typal drug ciprofloxacin and the veterinary analogue enrofloxacin, is well described by an AUC:MIC model. The high degree of associa­ tion between AUC and CMAX for single doses makes these difficult to distinguish practically. The utility of the AUC:MIC ratio was thor­ oughly characterized by Drusano and col­ leagues (1993), among others, and has been widely adopted for the fluoroquinolones, including in veterinary applications. Currently for Gram-­negative aerobic pathogens, typical exposure targets for fluoroquinolones, includ­ ing enrofloxacin, are an AUC:MIC ratio of at least 100 hr (Madaras-­Kelly et  al.,  1996). Note that the AUC:MIC has a time unit, con­ ventionally hours; this may be unintuitive but can be shown using dimensional analysis. Interestingly, lower AUC:MIC values are tar­ geted for staphylococci. This does not reflect any relative resistance but highlights that the  exposure target is a feature of the drug–­ pathogen combination and is empirical, rather than mechanistic.

Some other important antimicrobials, including the tetracyclines and vancomycin, were formerly proposed to be time dependent but later evidence has demonstrated that an exposure-­dependent model is preferable (Vandecasteele et al., 2013), and AUC:MIC are now used for optimization of these agents in practice. The utility of the AUC:MIC model extends beyond the antibacterial drugs, and has been applied extensively for dose optimiza­ tion for treatment of systemic fungal infec­ tions, especially with the triazole class (Lepak and Andes, 2015). The AUC:MIC can be calculated in different ways, which may yield very different results, and the optimum strategy has proved controversial. A challenging aspect of the interpretation of AUC is that it expresses a functional relationship rather than a real quantity, so it is difficult to interpret mechanistically. Considering that the AUC is independent of the rate of absorption, it is possible to propose, at least theoretically, situa­ tions in which a relatively large AUC:MIC ratio could be obtained without drug concentration ever actually reaching MIC because of very slow absorption. It is expected that limited antimicro­ bial effect would occur in this case. To overcome this difficulty, one method utilizes what has been termed the area under the inhibitory curve (AUIC), which considers only that area for which the concentration exceeds the MIC. Nevertheless, the currently favored methodology is simply the AUC24:MIC. The usage of terminology for these models in the literature is inconsistent and often confusing; a very effective breakdown is found in Corvaisier et al. (1998). An additional complexity is the selection of the total or free (protein-­ unbound) plasma concentration. PK:PD indices have been nominated for both, and while the free concentration is considered more representative of the concentration at the site of effect, either may be reasonable, depending on the application and the available data. Similar to the approaches proposed to pre­ dict the antimicrobial effect, PK/PD modeling has been applied to predict exposure targets that may minimize the development of

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Concentration-­Dependent Antimicrobial Activity  101

Pharmacodynamics of Antimicrobials

resistance, especially for the fluoroquinolones. Instead of optimizing towards the AUC:MIC, the AUC:MPC is applied instead; the mutant prevention concentration (MPC) represents the projected MIC of a hypothetical pathogen isolate which is derived from the wild-­type iso­ late by simple mutation (Liang et al., 2011).

­Clinical Breakpoints An important component of antimicrobial stewardship is culture and susceptibility test­ ing. Although the combination of clinical observations and microbiology knowledge can sometimes provide good predictions about the identity of a pathogen, and suggest appropriate therapy, the diversity of antimicrobial suscep­ tibility of many organisms means that objec­ tive data are often necessary. We have earlier reviewed the principles of the MIC from the perspective of antimicrobial drug action. Alternately, we could consider the drug factors to be fixed, and propose the MIC as a pathogen factor, summarizing the degree to which the isolate is drug resistant. For a given drug and dosing strategy, the chance of success is expected to be lower for pathogens with higher MIC. However, this is a relative judgment. Can a meaningful clinical interpretation be applied to an MIC in absolute terms? As observed in our lizard population, severe systemic disease may be associated with Pseudomonas infections and demand antimi­ crobial therapy. Pseudomonas, especially P. aeruginosa, is well known for diversity of both intrinsic and acquired drug resistance (Pang et al., 2019), and empirical therapy may be ineffective for this reason. The fluoroqui­ nolones are an important therapeutic option for Pseudomonas infections. For a Pseudomonas isolate with an enrofloxacin MIC of 4 μg/ml, do we expect that enrofloxacin therapy will be similarly effective as for an isolate with MIC of 0.25 μg/ml? Under the AUC:MIC model for the antimicrobial effect, presuming that the elimi­ nation of this drug is dose-­linear, an equivalent

effect would require a dose 16 times larger. Does this warrant selecting a different drug? What information would be required to make this judgment? Interpreting the MIC directly by applying it to a pharmacokinetic-­pharmacodynamic model is a possible strategy to evaluate the appropriate­ ness of a chosen dose regimen. This can take substantial effort, and may be challenging for the clinician, especially under time constraints. A degree of mathematics or computing experi­ ence is also required, and the interpretation of pharmacokinetic data and application to a spe­ cific case can be technically challenging. In some cases, the relevant data may not be readily available. The concept of clinical breakpoints proposes that the primary task for the clinician is rational drug selection, and that the dose regi­ men is relatively fixed. Earlier, we explored the optimization of dosage regimen, under the proposition that the MIC of the target patho­ gen and the clinical outcome resulting from drug exposure are fixed. The concept of the clinical breakpoint is a rearrangement, in which the dose regimen is fixed and the prob­ able clinical outcome is predicted, based on the MIC (Turnidge and Paterson,  2007). Those MIC values which correspond to a good pre­ dicted outcome may then be classed as “sus­ ceptible” and those corresponding to a poor predicted outcome as “resistant.” The critical value that distinguishes these is the breakpoint. Often in clinical practice, a measured MIC is not made available to the clinician after sub­ mission of a sample for culture and susceptibil­ ity testing. Instead, the result for a given drug for the submitted isolate is often simply returned as “resistant” (R) or “susceptible” (S), or sometimes as “intermediate” (I), depending on the test setup. These may be obtained by classification of the MIC as measured by a microdilution-­type test, or by scaling the break­ point to the inhibition zone diameter obtained from a disk diffusion test; this requires valida­ tion to match the inhibition zone diameter to the MIC but the disk diffusion approach is

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102

inexpensive and accessible (though improved technologies have made direct observation of MIC more efficient). It is not possible to obtain the MIC from the susceptibility result expressed in this form, only to state if it is lower or higher than the breakpoint. It is intended that culture and susceptibility testing results provide highly interpreted infor­ mation with direct clinical relevance, so that a result of “R” suggests a poor probability of a good clinical outcome, and a result of “S” sug­ gests a reasonable probability of a good clinical outcome. As we have explored, the likely clini­ cal outcome is a result of numerous interacting factors; the MIC, an indicator of the potency of antimicrobial effect, is only one part of the pic­ ture. With this in mind, substantial informa­ tion must be added to the MIC to provide an interpretation with clinical utility. There must be knowledge of the pharmacokinetics suffi­ cient to predict antimicrobial drug exposure, especially at the proposed dose regimen. Then a prediction of the treatment outcome is required, given the predicted drug exposure. The specification of a clinical breakpoint for some drug, species, and pathogen combination therefore requires that pharmacokinetic and pharmacodynamic data of direct relevance are available. Without pharmacokinetic data applicable to the target patient, the achievable drug exposure in vivo is not very predictable, so there is no clear way to assess if a pharmacody­ namic target will be reached or not. As the tar­ get effect of an antimicrobial is pathogen rather than host specific, it is reasonable to expect that the pharmacodynamic component is fairly constant but host and disease variabil­ ity may contribute to lack of robustness of the breakpoint. As we have explored, our PK:PD models are highly empirical, and they are reliant on actual outcome data to provide their quantitative tar­ gets; the less specific data available to support the breakpoint, the poorer its external validity. For example, directly taking the breakpoint from another patient species, especially from humans, essentially proposes that there is no

interspecies variability in pharmacokinetics or pharmacodynamics, which is a strong assump­ tion. Because the selection of a breakpoint relies on multiple lines of evidence, and in many if not most cases the information is not complete, the clinical breakpoints applied for veterinary clinical practice are a result of expert assessment. This process as implemented in practice has been recently described (Toutain et al., 2017). Though culture and susceptibility testing produces useful patient-­specific infor­ mation, in our opinion, the unthinking use of breakpoint-­interpreted susceptibility results to make clinical drug choices is likely to be mis­ leading in many individual cases. It is just one among many factors for the clinician to consider. It is sufficiently important to repeat: the interpretation of clinical breakpoints has simi­ lar limitations and assumptions to the inter­ pretation of dosage regimen predictions reliant on PK:PD principles. Of specific note is that the concept of the breakpoint demands that a dose regimen be specified. As only a single breakpoint is generally used per drug, species, and pathogen combination, no information is expressed regarding the potential merit of a dose regimen, other than the one assumed by the breakpoint. Further, as the breakpoints only define thresholds, it is not possible to determine where within the breakpoint range any given observation lies. Doubling the dose, for example, may overcome a two-­fold MIC barrier. However, without the actual observed MIC, the degree of drug resistance cannot be known, so it is difficult to predict if a strategy such as increasing the dose would be sufficient.

­ cientific Methods S in Pharmacodynamics We have explored various quantitative models that link pharmacokinetic and in vitro data to generate predictions of in  vivo effect, and the application of those models to the clinical

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­Scientific Methods in Pharmacodynamic  103

Pharmacodynamics of Antimicrobials

setting. Determination of these models requires careful experimental design, and the capability of these study designs has important implica­ tions for the validity and applicability of their theoretical predictions. In this final section, we will explore some of the methods that have been applied to obtain evidence underpinning the clinical pharmacodynamic models. With the objective of understanding the extent of antimicrobial effect in  vivo, given knowledge of the pharmacokinetics and the MIC, the simplest overall approach is to repli­ cate the clinical syndrome of interest in an experimental animal model. In principle, an animal model is a close representative of the clinical patient but has a number of important scientific advantages, beyond simply mitiga­ tion of the ethical impact of implementing such a trial in clinical patients. However, as this design involves inducing an infection, in a sub­ stantial proportion of subjects the antimicro­ bial treatment will fail by design. Consequently, a critical disadvantage of these study designs is their high ethical and welfare penalty. Animal models with induced central infec­ tions, sepsis, skin structure infections, and pneumonia are frequently used to characterize antimicrobial effects, as these have few con­ founding factors. A major benefit of these infection models is that the target tissue for treatment is close to systemic circulation, so it is reasonable to expect that free drug concen­ trations at the site of infection are proportional to those in plasma. As these infections are clini­ cally severe, recovery in the absence of effective antimicrobial treatment is unlikely. Therefore, where treatment success versus failure is the measured outcome, these severe models ensure that treatment effect signal is clearly distin­ guishable from background noise; the higher the probability of recovery in untreated con­ trols, the more subjects are required to identify an effective treatment intervention. A major advantage of the animal model approach is its strong external validity. Because the MIC and pharmacokinetics are closely known, and the outcome is directly observed,

the complete design can eliminate the influence of effects that are difficult to observe or quantify, such as the pathophysiology of disease. If we can observe the pharmacokinetics and pharma­ codynamics as pieces of our three-­component model, and we can also observe the sum of all the components in the form of the treatment outcome, then knowledge of the patient–­ pathogen interaction, which is the most difficult component to observe, is not needed. Although the in vivo design brings substan­ tial advantages as a realistic model of a clinical patient, the observed outcome combines the antimicrobial effect and the host immune response. Often the experimental design is extended to eliminate host immune responses by using large doses of a myelosuppressive drug such as cyclophosphamide, or by using innately immunodeficient animals such as the nude mouse. A key example of this animal model approach to PK/PD is the classic study of Drusano et al. (1993) which evaluated success of lomefloxacin therapy in rats, with experimentally induced Pseudomonas spp. sepsis and cyclophosphamide-­ induced neutropenia. In this large experiment, rats were treated with the same total daily dose but divided into a once, twice, or four times daily regimen, so that the AUC24s were compa­ rable but the CMAX and T>MIC were inversely ranked, and untreated controls were included. The best outcome was observed in those ani­ mals receiving the regimen maximizing CMAX:MIC. Interestingly, later experiments reported in this study using the same design did not distinguish CMAX:MIC from AUC:MIC as predictive of outcome. These studies supported the reasonableness of the AUC:MIC as predic­ tive of the effect of the fluoroquinolones but the observations emphasize the strong correlations among these indices in vivo, a clear limitation of the animal modeling approach. A critical disadvantage of a fully in  vivo approach is that the pharmacokinetic compo­ nent is strongly limited by the actual pharma­ cokinetics occurring in the subject animals. As the major candidate predictive indices, the

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CMAX, AUC, and T>MIC are strongly corre­ lated with one another in vivo, distinguishing the strength of association of each factor with the magnitude of antimicrobial effect is diffi­ cult. Purely in vitro approaches based on “time-­ kill” experiments attempt to define a complete time-­course of effect, rather than summarizing the effect simply with the MIC. By carefully controlling drug concentration and obtaining measurement of bacterial viability across the time of exposure, a more complete under­ standing can be obtained with reference to, for example, a sigmoid model (Wen et al., 2016). Further developments have emphasized cul­ ture systems that allow controllable noncon­ stant drug concentration in the culture medium (Vidaillac et al., 2009). Flow-­through diffusion systems comprise a high surface area growth chamber, which exchanges with circulating replacement medium across a diffusion mem­ brane. Drug concentration in the growth cham­ ber changes with time due to drug diffusion to and from the circulating medium, mimicking the exchange of drug from plasma in vivo. As the system volume and flow rate are controlla­ ble, and the culture may be easily sampled, these systems provide greater flexibility than animal models, in which the pharmacokinetics are relatively fixed and sampling may be tech­ nically limited. These experiments allow fine control over both the time-­course of drug expo­ sure and the pathogen growth conditions. In clinical practice, the PK/PD indices CMAX:MIC, AUC:MIC, and T>MIC are pre­ dominant, forming the mainstay of clinical understanding of the quantitative behavior of the antimicrobials, in both human and veteri­ nary applications. The use of these broad mod­ els is so pervasive that it is easy to forget that they are highly empirical and make numerous simplifying assumptions, and are more heuris­ tics summarizing relevant information than actual characteristics of antimicrobials. Some important cases highlight the weaknesses of these models. We have already noted that the CMAX:MIC does not inform us about the interdosing

interval, because it operates only on a single estimate of drug concentration, and is there­ fore only useful to select the dose. Generally, the dosing interval for the aminoglycosides is set as 24 hours but this has no specific justifi­ cation from the CMAX:MIC model, so other forms of evidence are required. As the rela­ tionship between the AUC and MIC can be calculated in several ways, and there is no mechanistic basis with which to select a pre­ ferred method, use of AUC:MIC-­based strate­ gies requires agreement on how the AUC will be determined and what systems it will be applied to. For example, a specific AUC:MIC corresponds to different drug concentration– time profile under steady state rather than single-­dose conditions but the AUC:MIC in its typical form does not accommodate this; the meaning of the AUC:MIC depends on how its time unit is interpreted (Toutain et al., 2007). Vancomycin was previously proposed to operate under a time-­dependent mechanism but the suitability of a T>MIC or AUC:MIC model is apparently dependent on the density of the starting inoculum, with the AUC:MIC model currently predominant (Vandecasteele et  al.,  2013). As these classic models don’t mechanistically consider the pathogen density, they lose external validity under conditions different from their experimental design. These and numerous other situations high­ light the limitations of the classic indices and emphasize that these models are descriptive of drug action, rather than being representative of an inherent characteristic of antimicro­ bial agents. A response to these and similar issues is to step back to a model closer to the actual drug effect. Using an appropriate in  vitro system, a sigmoid-­type dose–response model may be defined to describe antimicrobial effect based on drug concentration. With a simultaneous pharmacokinetic model, PK/PD optimization may be conducted without any need to summa­ rize the concentration–time relationship. A mathematical model, as part of a computa­ tional simulation, can predict both drug

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concentration and drug effect simultaneously, and be validated using experimental data. An early implementation of a real-­time model of drug effects was reported by Zhi and colleagues (1988), with various extensions and adapta­ tions reported since following this structure. Novel techniques have taken advantage of modern computational resources to implement more explicit models, with less abstraction and empiricism than the common PK/PD indices. The fundamental feature is a mathematical description of the density of the pathogen pop­ ulation, and the change of the density of this

population over time. Recent developments highlight that the classic PK/PD indices can be considered as special cases of these more gen­ eral models (Nielsen et al., 2011). Direct veterinary applications of these mod­ els have recently emerged (Pelligand et al., 2019). Though these offer a more com­ plete and flexible understanding of antimicro­ bial effect, they are more demanding both in the complexity of the data required to develop them, and in the mathematical and statistical sophistication of their implementation, which are both limiting factors for broad usage.

­References Agius JE, et al. 2020. Pharmacokinetic profile of enrofloxacin and its metabolite ciprofloxacin in Asian house geckos (Hemidactylus frenatus) after single-­dose oral administration of enrofloxacin. Vet Anim Sc. 9:100116. Banks CB, McCracken HE. 2002. Captive management and pathology of sharp-­snouted torrent frogs, Taudactylus acutirostris, at Melbourne and Taronga Zoos. In: Frogs in the Community. Brisbane, Australia: Queensland Frog Society. Barreto EF, et al. 2021. Setting the beta-­lactam therapeutic range for critically ill patients: is there a floor or even a ceiling? Crit Care Explor 3:e0446. Barzelai ID, Whittem T. 2017. Survey of systemic antimicrobial prescribing for dogs by Victorian veterinarians. Aust Vet J 95 :375. Bland C, et al. 2018. Reappraisal of contemporary pharmacokinetic and pharmacodynamic principles for informing aminoglycoside dosing. Pharmacotherapy 38:1229. Bon C, et al. 2018. Mathematical modeling and simulation in animal health. Part III: Using nonlinear mixed-­effects to characterize and quantify variability in drug pharmacokinetics. J Vet Pharmacol Ther 41:171. Bucki EP, et al. 2004. Pharmacokinetics of once-­daily amikacin in healthy foals and

therapeutic drug monitoring in hospitalized equine neonates. J Vet Intern Med 18:728. Corvaisier S, et al. 1998. Comparisons between antimicrobial pharmacodynamic indices and bacterial killing as described by using the Zhi model. Antimicrob Agents Chemother 42:1731. Drusano GL, et al. 1993. Pharmacodynamics of a fluoroquinolone antimicrobial agent in a neutropenic rat model of Pseudomonas sepsis. Antimicrob Agents Chemother 37:483. Hill AV. 1913. The combinations of haemoglobin with oxygen and carbon dioxide. Biochem J 7:471. Lepak AJ, Andes DR. 2015. Antifungal pharmacokinetics and pharmacodynamics. Cold Spring Harb Perspect Med 5:a019653. Liang B, et al. 2011. Mutant prevention concentration-­based pharmacokinetic/ pharmacodynamic indices as dosing targets for suppressing the enrichment of levofloxacin-­resistant subpopulations of Staphylococcus aureus. Antimicrob Agents Chemother 55:2409. Love DN, et al. 1983. Serum concentrations of penicillin in the horse after administration of a variety of penicillin preparations. Equine Vet J 15:43. Madaras-­Kelly KJ, et al. 1996. Twenty-­four-­hour area under the concentration-­time curve/MIC ratio as a generic predictor of fluoroquinolone

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antimicrobial effect by using three strains of Pseudomonas aeruginosa and an in vitro pharmacodynamic model. Antimicrob Agents Chemother 40:627. Martinez MN, et al. 2013. The pharmacodynamics of antimicrobial agents. In: Giguere S, Prescott J, Dowling P (eds). Antimicrobial Therapy in Veterinary Medicine. Hoboken: John Wiley, pp. 79–103. Muller AE, et al. 2013. Optimal exposures of ceftazidime predict the probability of microbiological and clinical outcome in the treatment of nosocomial pneumonia. J Antimicrob Chemother 68:900. Nielsen EI, et al. 2011. Pharmacokinetic/ pharmacodynamic (PK/PD) indices of antibiotics predicted by a semimechanistic PKPD model: a step toward model-­based dose optimization. Antimicrob Agents Chemother 55:4619. Pang Z, et al. 2019. Antibiotic resistance in Pseudomonas aeruginosa: mechanisms and alternative therapeutic strategies. Biotechnol Adv 37:177. Pelligand L, et al. 2019. Semi-­mechanistic modeling of florfenicol time-­kill curves and in silico dose fractionation for calf respiratory pathogens. Front Microbiol 10:1237. Rang HP. 2006. The receptor concept: pharmacology’s big idea. Br J Pharmacol 147:S9. Smith KF, et al. 2009. The role of infectious diseases in biological conservation. Anim Conserv 12:1–12. Srimani JK, et al. 2017. Drug detoxification dynamics explain the postantibiotic effect. Mol Syst Biol 13:948. Stubbings W. 2006. Mechanisms of the post-­ antibiotic effects induced by rifampicin and gentamicin in Escherichia coli. J Antimicrob Chemother 58:444. Tang PK, et al. 2020. Antimicrobial susceptibility patterns for aerobic bacteria isolated from reptilian samples submitted to a veterinary diagnostic laboratory: 129 cases (2005–2016). J Am Vet Med Assoc 257:305. Tannous E, et al. 2020. Time above the MIC of piperacillin-­tazobactam as a predictor of

outcome in Pseudomonas aeruginosa bacteremia. Antimicrob Agents Chemother 64:e02571. Toutain PL, et al. 2007. AUC/MIC: a PK/PD index for antibiotics with a time dimension or simply a dimensionless scoring factor? J Antimicrob Chemother 60:1185. Toutain PL, et al. 2017. En route towards European clinical breakpoints for veterinary antimicrobial susceptibility testing: a position paper explaining the VetCAST approach. Front Microbiol 8:2344. Toutain PL, et al. 2021. The pharmacokinetic/ pharmacodynamic paradigm for antimicrobial drugs in veterinary medicine: recent advances and critical appraisal. J Vet Pharmacol Ther 44:172. Turnidge J, Paterson DL. 2007. Setting and revising antibacterial susceptibility breakpoints. Clin Microbiol Rev 20:391. Vandecasteele SJ, et al. 2013. The pharmacokinetics and pharmacodynamics of vancomycin in clinical practice: evidence and uncertainties. J Antimicrob Chemother 68:743. van de Kassteele J, et al. 2012. New statistical technique for analyzing MIC-­based susceptibility data. Antimicrob Agents Chemother 56:1557. Vidaillac C, et al. 2009. In vitro activity of ceftaroline against methicillin-­resistant Staphylococcus aureus and heterogeneous vancomycin-­intermediate S. aureus in a hollow fiber model. Antimicrob Agents Chemother 53:4712. Weiss JN. 1997. The Hill equation revisited: uses and misuses. FASEB J 11:835. Wen X, et al. 2016. Limitations of MIC as sole metric of pharmacodynamic response across the range of antimicrobial susceptibilities within a single bacterial species. Sci Rep 6:37907. Woodward AP, et al. 2020. Population physiologically based modeling of pirlimycin milk concentrations in dairy cows. J Dairy Sci 103:10639. Zhi J, et al. 1988. Microbial pharmacodynamics of piperacillin in neutropenic mice of systematic infection due to Pseudomonas aeruginosa. J Pharmacokinet Biopharm 16:355.

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  ­Reference

6 Principles of Antimicrobial Drug Selection and Use J. Scott Weese and Patricia M. Dowling

In recent years there have been important changes in antimicrobial therapy based on availability of new antimicrobials, a greater database of species-­ specific pharmacokinetic (PK) and pharmacodynamic (PD) information available for antimicrobials used in veterinary medicine and refinement of practices based on PK/PD information, principles of antimicrobial therapy, and antimicrobial stewardship efforts. These have allowed more accurate drug dosing, aiming to maximize antimicrobial efficacy while minimizing adverse effects. Concerns over drug residues in food animals and the continued development of bacterial resistance to antimicrobials have heightened the awareness of rational use of antimicrobials. In many countries, antimicrobial stewardship efforts have resulted in increasing veterinary oversight of antimicrobial use in animals and veterinarians are expected to document an evidence-­based need before using or prescribing antimicrobials. For good antimicrobial stewardship, numerous considerations need to be taken in to account prior to antimicrobial use (Figure 6.1).

­ oes the Diagnosis Warrant D Antimicrobial Therapy? A substantial amount of antimicrobial use is likely directed at nonbacterial infections or infections where specific treatment is unnecessary

(e.g., self-­limiting infections). Antimicrobials are often used in the absence of a definitive diagnosis, and that is often justifiable. However, a reasonable diagnosis and justification for treatment must be established before administering or prescribing antimicrobial therapy. Using antimicrobials to treat minor infections or purely viral or inflammatory diseases is irrational and ­expensive, can be hazardous to the patient and promotes antimicrobial resistance. Clients/producers may expect antimicrobials for trivial infections or  “just in case” an infection may develop. Veterinary practitioners must resist such pressure to use or prescribe unnecessary antimicrobials. Clinicians may similarly use a “just in case” approach in situations where they know infection is unlikely to be present or at high risk of developing, or where prophylaxis is not likely to be indicated. However, use remains common as a form of defensive medicine, whereby the clinician is prescribing antimicrobials more for themselves than their patient.

­ hat Organism(s) is/are Likely W to be Involved? It is not always necessary to culture samples from all patients with infectious diseases in  order to identify the organism involved. Even when specimens can be submitted for

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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109

Principles of Antimicrobial Drug Selection and Use

Are antimicrobials warranted?

• Reasonable diagnosis • Resist defensive medicine

What’s the pathogen?

• Organism known or suspected • Historical susceptibility or susceptibility test results • Clinical experience with type of infection

Does the drug get the pathogen? (Pharmacodynamics)

• MIC/MBC/MPC • Time-or concentrationdependent activity • Post-antibiotic effect • Post-antibiotic leukocyte enhancement

Does the drug get to the pathogen? (Pharmacokinetics)

• Volume of distribution, clearance, elimination half-life • Physicochemical properties of the drug (lipid solubility, ionization, protein binding) • Location of infection (intracellular, extracellular, sequestered site) • Route of administration (IV, IM, SC, PO, topical)

Treatment considerations

• Dosage regimen • Adjustments due to physiology or pathophysiology • Immune status of patient • Rationale for combination treatments • Cost of treatment including value of animal

Risks associated with treatment

• Promotion of antimicrobial resistance • Toxic effects • Human health risks • Drug interactions • Impairment of host defenses • Considerations for food animals

Figure 6.1  Considerations for antimicrobial use to promote good antimicrobial stewardship.

culture, treatment is usually started prior to results being available, necessitating an empirical decision. Often, the practitioner can use clinical experience from similar cases to identify the likely bacterium and typically effective treatments. The signs of some infectious diseases are so obvious and the susceptibility of the causative agent is predictable (e.g., “strangles” in horses from Streptococcus equi subsp. equi) that the need for microbiological identification is minimal from the standpoint of making an antimicrobial selection. However, for those infectious diseases of unknown cause or for those attributable to organisms with unpredictable antimicrobial susceptibility, there is no substitute for isolation and identification of the causative pathogen, if proper specimens can be collected. For these organisms, initial empirical therapy while waiting for culture results may include an antimicrobial with a broad spectrum of activity that is

expected to be effective against the most common pathogens. However, broad-­spectrum drugs are usually more toxic and more expensive, so antimicrobial deescalation to more selective therapy should be done once culture results are obtained. The severity of disease, likelihood of resistance, and likely outcome if the initial antimicrobial selection is incorrect needs to be considered. In life-­threatening infections, broader spectrum treatment that covers all reasonable causes is indicated because of the potential serious (or fatal) outcome if effective treatment is not started.

­ hat is the Antimicrobial W Susceptibility of the Organism(s)? While clinical experience may aid the clinician in suspecting a given pathogen, it is optimal to obtain specimens for culture and susceptibility

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testing in order to select the most appropriate drug and dosage regimen. Preferably, samples for bacteriological culture should be collected before administering an antimicrobial drug. Cytology of an appropriately collected sample may provide insight as to the etiological agent and aid in interpretation of culture results. Susceptibility testing can provide values for minimal inhibitory concentration (MIC), minimal bactericidal concentration (MBC), and/or mutant prevention concentration (MPC) (Chapter 2). Other important pharmacodynamic (Chapter  5) considerations include whether the antimicrobial shows time-­ or concentration-­dependent activity and other microbial effects including postantibiotic effect (PAE) and postantibiotic leukocyte enhancement.

­ ill the Antimicrobial Reach W the Site of Infection? Will It Be Active in the Infection Environment? The pharmacokinetics of the antimicrobial (absorption, distribution, metabolism, excretion) will determine if the it will reach the site of infection in effective concentrations for the necessary amount of time (Chapter 4). An antimicrobial’s physiochemical properties (e.g., lipid solubility, ionization, degree of protein binding) and the route of administration both impact an antimicrobial’s pharmacokinetic profile. Treatment of sequestered infections such as prostatitis, mastitis or meningitis requires antimicrobials that readily cross biological barriers. Antimicrobials characterized by low values for volume of distribution due to their physiochemical properties are unlikely to reach therapeutic concentrations in such sites. For some antimicrobials, the local infection environment reduces their efficacy. Sulfonamides are ineffective in purulent debris, since paraamino benzoic acid (PABA) released from decaying neutrophils serves as a PABA source for bacteria and reduces the competitive effect

of the sulfonamide. Aminoglycosides are ineffective in an abscess due to the acidic, anaerobic environment along with the presence of nucleic acid material from decaying cells which inactivates the aminoglycosides.

­ hat Dosage Regimen will W Maintain the Appropriate Antimicrobial Concentration for the Proper Duration of Time? Dosage regimens are calculated from PK/PD integration (Chapter  5) and may need to be adjusted for physiological variation (e.g., neonates, geriatrics) or pathophysiology (e.g., hepatic and/or renal insufficiency). In some cases, combinations of antimicrobials may be synergistic or additive in their antimicrobial activity. Use of multiple antimicrobial drugs should be limited to: ●●

●●

●●

●●

known synergism against specific organisms (e.g., beta-­lactams plus aminoglycosides in the treatment of enterococcal endocarditis) (Goldstein et al., 2003) prevention of the rapid development of bacterial resistance (e.g., trimethoprim combined with a sulfonamide) (Worthington and Melander, 2013) extending the antimicrobial spectrum of initial empirical therapy of life-­threatening conditions (e.g., beta-­lactams plus aminoglycosides in the treatment of septic foals) (Magdesian, 2017) treating mixed bacterial infections (e.g., a penicillin or cephalosporin plus enrofloxacin in the treatment of aspiration pneumonia in dogs) (Sherman and Karagiannis, 2017).

Known antagonistic combinations of antimicrobials should be avoided, such as penicillin with tetracycline (penicillin acts on actively dividing cell walls while tetracyclines are bacteriostatic in action) (Gal, 1965). In addition to increased costs, there is usually also an increased risk of adverse effects from the administration of multiple antimicrobials (e.g., antimicrobial-­associated diarrhea)

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What Dosage Regimen will Maintain the Appropriate Antimicrobial Concentration for the Proper Duration of Time?  111

Principles of Antimicrobial Drug Selection and Use

(Wilson et al., 1996). For any antimicrobial dosage regimen, patient acceptance, client compliance with dosing, and the cost of the treatment relative to the value of the animal must be considered. The availability of evidence-­based appropriate dosage data must also be considered. Quality of evidence can range from randomized controlled trials for the species diseases and animal species, to pharmacokinetic studies of a small number of healthy animals to extrapolation from other species. Interspecies differences can be substantial so species-­specific clinical efficacy or pharmacokinetic study data should be used whenever available, and limitations in available dosing data should be considered when determining whether to use an antimicrobial. This book attempts to incorporate the latest recommendations on dosages and to point out any current uncertainties about such dosages. Dosing recommendations can also change over time based on new information about the use of that drug in the individual animal species or broader understanding of use of the drug (e.g., the change to once-­daily administration of aminoglycosides from older q8h recommendations). Clinicians should ensure that up-­to-­date treatment recommendations are used. Duration of therapy is a key aspect of the treatment regimen and is typically accompanied by the least (or lowest quality) evidence. Inadequate durations reduce the likelihood of clinical resolution while excessive durations add cost, inconvenience for the person administering treatments, adverse drug reaction risks, and enhanced antimicrobial resistance selection pressure. As opposed to human medicine, where large trials have compared durations for a range of clinical conditions, data are very limited in veterinary medicine. This likely results in the use of excessively long durations either largely for historical reasons or because of a precautionary or defensive approach, particularly as avoiding treatment failure is a more obvious and motivating concern than most of the potential negative effects of treatment. There is increasing movement to using shorter treatment durations in both human and

veterinary medicine. Some veterinary treatment guidelines have reduced the recommended durations, albeit based more on expert opinion and extrapolation from human medicine than species-­specific data (Hillier et al., 2014; Lappin et  al.,  2017; Weese et  al.,  2019; Schuller et al., 2015). Long treatment durations may be required in some situations, and clinical efficacy should not be sacrificed in an attempt to shorten treatment duration. However, it is likely that shorter durations can be used compared to historical regimens, particularly in companion animals and horses. There is greater use of shorter durations in livestock species, often necessitated by logistical and cost issues, which can also provide some support for the efficacy of short durations in other species (e.g., short duration of treatment of bovine pneumonia provides some additional confidence in shortening previously long durations of treatment of pneumonia in dogs). Clinical assessment and diagnostic testing can be used to assess patient response and need for treatment. However, the goals of treatment must be considered. Treatment is typically aiming to control the infection, not necessarily to completely eradicate bacteria from the site or resolve all evidence of disease (e.g., radiographic changes may persist after elimination of the active infection). Given the absence of evidence-­ based data and the variation between infections in different animals, determining a standard optimal treatment duration remains a challenge, but clinicians should consider newer recommendations, patient response, and goals of treatment when determining how long to treat.

­ hat Risks are Associated W with Antimicrobial Treatment? Promotion of Antimicrobial Resistance Development of resistance is a common collateral effect of antimicrobial drug use. Any antimicrobial use can lead to increased risk of resistance. However, the risk is not the same

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for all treatment approaches and for all antimicrobials and the emergence of resistance can be minimized with appropriate dosing regimens. As explained in Chapter 2, current measurement of antimicrobial drug activity against bacterial pathogens relies on measurement of the MIC and, by comparison with established breakpoints, bacteria are classified as susceptible, intermediately susceptible, or resistant. At drug concentrations higher than the MIC, susceptible bacteria should in principle be ­inhibited, whereas a very small proportion of mutants harboring resistance mechanisms may not be inhibited. Nevertheless, these resistant variants will be inhibited at higher drug concentrations (i.e., the MIC of the resistant mutants). From a clinical standpoint, this highlights the need for appropriate dosing regimens, which are dependent on adequate dosing information and accurate patient weight. However, with suboptimal dosing based on use of inadequate doses or improper estimation of weight, or in situations where drug concentrations at the site of infection are low because of disease factors or physiological (e.g., blood– brain, blood–prostate) barriers, resistance emergence is more likely.

Toxic Effects An adverse drug reaction (ADR) is defined as any response to a drug that is unintended or noxious and occurs at doses used for prophylaxis or therapy. Pharmacovigilance or postmarket surveillance includes the collection and analysis of ADRs. The number of ADRs occurring in veterinary medicine is largely unknown due to a lack of recognition and inadequate reporting. Even ADRs of moderately high incidence can easily be missed in licensing studies because of the small sample sizes, and extra-­label use of drugs or doses compounds the data gaps. The risks of ADRs from antimicrobials are often underappreciated. A serious ADR may  complicate treatment of the original ­problem and even be fatal. Failure to

communicate the risks of ADRs to clients is a common cause of litigation. The selective toxicity of antimicrobials is variable. Some drugs, such as beta-­lactams, are generally considered to be extremely safe with a high therapeutic index, whereas others, such as the aminoglycosides, are potentially toxic at therapeutic doses. Antimicrobial drugs can directly damage the function of many organs or tissues, particularly the kidneys (e.g., aminoglycosides, amphotericin B), nervous system (e.g., aminoglycosides, polymyxins), liver (e.g., rifampin), heart (e.g., monensin, tilmicosin), hematopoietic ­system (e.g., sulfonamides, chloramphenicol), retina (e.g., enrofloxacin), and joint cartilage (e.g., fluoroquinolones). Penicillin is particularly associated with anaphylactic reactions. However, any drug can be associated with adverse effects such as allergic reactions and immune-­mediated diseases. The toxicity of antimicrobials with a narrow margin of safety can be minimized by using the lowest effective doses and the shortest duration of treatment, by substituting equally effective but less toxic agents, or by using a combination of antimicrobial agents that work synergistically against the pathogen without increased toxicity to the host (e.g., sulfonamide combined with trimethoprim). Toxicity can also be minimized by identifying high-­risk situations (e.g., aminoglycosides in hypotensive patients, enrofloxacin in cats, fluoroquinolones in growing animals) and avoiding use unless absolutely necessary.

Human Health Risks Veterinarians, technicians, and clients are all exposed to veterinary medicines, some of which can cause ADRs. Veterinary practitioners must warn their personnel of the dangers associated with some of the medications that they work with every day, and the precautions that should be taken to prevent or minimize exposure. Similarly, clients should be warned of these dangers if they are treating their animals at home and dispensed drugs should

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What Risks are Associated with Antimicrobial Treatment?  113

Principles of Antimicrobial Drug Selection and Use

carry proper warning labels regarding risks and safe drug handling. The greatest risk of human exposure to veterinary antimicrobials is sensitization and subsequent hypersensitivity reactions. Exposure to penicillin during treatment of animals has been known to cause sensitization in people with allergies to penicillin, resulting in severe anaphylactic reactions on repeat exposure (Woodward,  2005). Contact dermatitis and urticaria have also occurred after exposure to chlorhexidine, penicillin, and furazolidone. Chloramphenicol causes dose-­dependent bone marrow suppression in people and animals; however, a subset of people will develop idiosyncratic, irreversible aplastic anemia secondary to exposure to very small amounts of the drug (e.g., ophthalmic ointment) (Yunis, 1989). Accidental injection of the macrolide antibiotic tilmicosin has caused severe cardiac abnormalities (chest pains, ECG abnormalities, intraventricular conduction delays, death) in people (Veenhuizen et al., 2006).

Drug Interactions (Table 6.1) Antimicrobials may be physicochemically incompatible with other agents in  vitro (Tomczak et  al.,  2021). For example, tetracyclines are incompatible with any solution containing calcium or magnesium. Although it is appropriate and common practice to use a combination of a cephalosporin and an aminoglycoside in vivo, many cephalosporins are not compatible with aminoglycosides in suspension. Thus, it is not good practice to mix antimicrobial agents in the same syringe or tubing. The lack of an obvious interaction, for example, precipitate, does not mean a chemical inactivation has not occurred. Antimicrobials may also have PK and PD interactions with other drugs. Rifampin reduces the oral bioavailability of clarithromycin in foals by 90% (Peters et al., 2011). Enrofloxacin reduces the metabolism of theophylline in dogs (Intorre et  al.,  1995). Chloramphenicol reduces the hepatic metabolism of phenobarbital and

Table 6.1  Examples of adverse in vivo effects of drug interactions between antibiotics and other agents. Antimicrobial Drug

Interacting Drug

Adverse Effect

Aminoglycoside

Cephaloridine, cephalothin, polymyxins, furosemide

Nephrotoxicity

Polymyxins, curare-­like drugs, anesthetics

Neuromuscular blockade

Amphotericin B

Aminoglycosides

Nephrotoxicity

Azoles (except fluconazole)

Acid suppressant

Decreased absorption

Chloramphenicol

Dicoumarol, barbiturates

Prolonged anesthesia, anticoagulation

Griseofulvin

Dicoumarol, barbiturates

Reduced anticoagulant effect

Lincomycin

Kaolin-­pectate

Decreased lincomycin absorption

Monensin

Tiamulin

Neurotoxicity

Polymyxins

Aminoglycosides

Nephrotoxicity, neuromuscular blockade

Rifampin

Macrolides, many others

Decreased plasma concentrations

Sulfonamides

Oral anticoagulants

Prolonged anticoagulant effect

Tetracyclines

Barbiturates, oral iron, calcium, magnesium

Anesthetic potentiation Decreased tetracycline absorption

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prolongs anesthesia. Tilmicosin sensitizes the cardiac myocytes to epinephrine and increases the risk of mortality.

route, site, and volume of antimicrobial to be injected and proper injection techniques should be used to minimize tissue damage.

Impairment of Host Defenses

­Adjunctive Treatment

Antimicrobial drugs may enhance or suppress host defenses (Chapter 5). These effects may be associated with alterations in cytokine production or the production of other inflammatory mediators. The ability of some antimicrobials (e.g., fluoroquinolones, macrolides) to penetrate and to concentrate within phagocytic cells, while not guaranteeing efficacy, is an important consideration in the treatment of intracellular bacterial infections. For example, phagocyte alteration of pathogen metabolism or structure may render the pathogen more susceptible to the effect of the antimicrobial agent and drug concentrations too low to have a bactericidal effect may render the microbe more susceptible to leukocyte action, an event associated with postantibiotic leukocyte enhancement.

Considerations for Food Animals When antimicrobials are used in food-­producing animals, there are numerous significant public health considerations. Whenever possible, antimicrobials should be used according to their label directions and label withdrawal times (for meat, milk, eggs) but there are many instances where label directions are not appropriate for a specific situation and extra-­label drug use may be necessary (Chapter  25). The regulations surrounding extra-­label drug use vary greatly between countries so veterinarians must know the regulations for their practice area. Veterinarians are responsible for the prevention of violative drug residues when prescribing antimicrobials to be used in an extra-­label manner. Injectable antimicrobials may also be associated with detrimental effects on carcass quality around the site of injection (Van Donkersgoed et  al.,  1999). Label directions should be carefully followed regarding the

Adjunctive treatments to antimicrobial therapy are essential in successfully controlling infection and promoting healing. They include source control, debriding necrotic tissues, removing purulent exudate, removing foreign bodies (including surgical implants), correcting acid–base and fluid imbalances and providing rest and nursing, when appropriate. Predisposing causes (e.g., hyperadrenocorticism, atopy) should be identified and treated whenever possible to minimize the requirement for antimicrobial therapy for opportunistic infections.

­Failure of Antimicrobial Therapy Failure of antimicrobial treatment can have many causes and the veterinarian should determine the reasons for a suboptimal response (Schlossberg,  2006). The following scenarios should be considered in the event of treatment failures.

Infectious Diseases Not Responsive to Antimicrobials Some infections will fail antimicrobial therapy because they are not specifically treatable. These include viral infections, especially upper respiratory infections and viral meningitis, and toxin-­ induced illness such as staphylococcal toxic shock syndrome. Some infections may appear to respond to antimicrobials such as doxycycline because of antiinflammatory rather than antibacterial activity of the drug (Henehan et al., 2017).

Noninfectious Diseases that Mimic Infectious Diseases Many noninfectious diseases may mimic infection and therefore fail to respond to antimicrobial

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­Failure of Antimicrobial Therap  115

Principles of Antimicrobial Drug Selection and Use

therapy. Systemic inflammatory response syndrome, some neoplasias, vasculitis, drug hypersensitivity, malignant hyperthermia, surgical trauma, and adrenal insufficiency can all produce fever and generalized clinical signs in affected patients. Many pulmonary conditions may cause fever, including pulmonary embolism, aspiration, and atelectasis. Cellulitis is mimicked by contact dermatitis, drug hypersensitivity, and insect stings.

Treatable Infection That Fails to Respond Clinically Wrong Antimicrobial

For many reasons, the wrong antimicrobial may have been selected. With empirical therapy, the suspected pathogen and its susceptibility may not be the actual infecting organism. In a polymicrobial infection, one or more pathogens may not be susceptible to the chosen antimicrobial. Fastidious organisms, such as anaerobes, may have failed to grow on culture and their presence must be inferred from cytology or by the clinical situation where they would be expected (e.g., infectious pododermatitis in cattle). The wrong antimicrobial may have been chosen if the microbiology laboratory makes errors in identifying the organism and/or its susceptibility pattern. Some antimicrobials may appear susceptible in vitro but are ineffective in  vivo (Chapter  2). Antimicrobial resistance may emerge during therapy or genotypic resistance may be unmasked (e.g., inducible resistance to clindamycin by staphylococci). Correct Antimicrobial But Incomplete Response

An incomplete response to antimicrobial therapy may signal the need to search for a surgical component to the disease. The presence of a foreign body or iatrogenic materials (e.g., suture material, pins, screws, intravenous and urinary catheters) can cause persistent foci of infection. Most infections characterized by abscessation require surgical drainage. Obstruction requiring surgical relief can be seen with cholecystitis, cholangitis,

and pyelonephritis. Surgical debridement of necrotic tissues is critical to treatment of many soft tissue infections. Inadequate drug penetration of the infection site (e.g., abscess, prostate, CNS) or inactivation of the drug at the site (e.g., aminoglycosides and purulent debris) may also play a role. Inadequate Dosing Regimen and Poor Owner Compliance

Dosing errors can occur and these can account for poor or no response to treatment. Similarly, poor owner compliance can be associated with lack of understanding of the treatment regimen, difficulty administering the antimicrobial, poor communication or lack of effort. Reviewing the recommended regimen and assessing likely owner compliance are important parts of evaluating situations where response to treatment is poor. Immunosuppression or Failure to Address Underlying Causes

Even if the correct antimicrobial or antimicrobial concentration is selected, treatment may fail if the patient is immunocompromised. This can be iatrogenic from corticosteroid or other immunosuppressive drug administration or due to disease states including neoplasia. Infections that occur secondary to an inciting cause (e.g., allergic skin disease) may be difficult to eliminate completely if the inciting cause is not addressed. Similarly, lack of source control or persistent infection of a nidus (e.g., surgical implant) can prevent an effective response. Biofilm

Biofilm is a complex carbohydrate-­dominated matrix that is produced by some types of bacteria. Biofilm-­associated infections can be difficult to eliminate with antimicrobials since bacteria within biofilms can be highly resistant to treatment, even if they are susceptible in their planktonic (‘normal’, free-­living) form. Biofilm provides physical protection by inhibiting the penetration of antimicrobials.

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Additionally, bacteria within biofilms typically have altered, usually downregulated metabolism and growth which render them less susceptible to antimicrobials, particularly those that exert their antibacterial effects during bacterial growth. The role of biofilm in infections is poorly understood but it is of particular concern with surgical implant-­associated infections (Singh et  al.,  2013; Morrison et  al.,  2016; Walker et al., 2016). In those situations, removal of the infected implant is often the best, or only, way to eliminate the infection. Local approaches (e.g., antimicrobial-­impregnated materials) may be necessary to provide high local antimicrobial

levels to have a chance of eliminating biofilm-­ associated infections where removal of the nidus is not possible. Untreatable Infection

Antimicrobial therapy may simply fail if it is instituted too late in the disease process to be effective or when disease is so severe that the patient succumbs to the infection before antimicrobials have a chance to be effective. Veterinarians are the best stewards of antimicrobial use in animals as they have the ability to examine and diagnose infectious disease and triage treatment based on the probability of a successful treatment outcome.

­References Gal K. 1965. Combined antibiotic therapy. Can Med Assoc J 93:844. Goldstein EJC, et al. 2003. Combination antibiotic therapy for infective endocarditis. Clin Infect Dis 36:615. Henehan M, et al. 2017. Doxycycline as an anti-­inflammatory agent: updates in dermatology. J Eur Acad Dermatol Venereol 31:1800. Hillier A, et al. 2014. Guidelines for the diagnosis and antimicrobial therapy of canine superficial bacterial folliculitis (Antimicrobial Guidelines Working Group of the International Society for Companion Animal Infectious Diseases). Vet Dermatol 25:163. Intorre L, et al. 1995. Enrofloxacin-­theophylline interaction: influence of enrofloxacin on theophylline steady-­state pharmacokinetics in the beagle dog. J Vet Pharmacol Ther 18:352. Lappin MR, et al. 2017. Antimicrobial use Guidelines for Treatment of Respiratory Tract Disease in Dogs and Cats: Antimicrobial Guidelines Working Group of the International Society for Companion Animal Infectious Diseases. J Vet Intern Med 31:279. Magdesian KG. 2017. Antimicrobial pharmacology for the neonatal foal. Vet Clin North Am Equine Pract 33:47.

Morrison S, et al. 2016. Adherence of methicillin-­resistant Staphylococcus pseudintermedius to suture materials commonly used in small animal surgery. Am J Vet Res 77:194. Peters J, et al. 2011. Oral absorption of clarithromycin is nearly abolished by chronic comedication of rifampicin in foals. Drug Metab Dispos 39:1643. Schlossberg D. 2006. Clinical approach to antibiotic failure. Med Clin North Am 90:1265. Schuller S, et al. 2015. European consensus statement on leptospirosis in dogs and cats. J Small Anim Pract 56:159. Sherman R, Karagiannis M. 2017. Aspiration pneumonia in the dog: a review. Top Companion Anim Med 32:1. Singh A, et al. 2013. Characterization of the biofilm forming ability of Staphylococcus pseudintermedius from dogs. BMC Vet Res 9 :93. Tomczak S, et al. 2021. Stability and compatibility aspects of drugs: the case of selected cephalosporins. Antibiotics 10:549. Van Donkersgoed J, et al. 1999. The effect of vaccines and antimicrobials on the formation of injection site lesions in subprimals of

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  ­Reference

Principles of Antimicrobial Drug Selection and Use

experimentally injected beef calves. Can Vet J 40:245. Veenhuizen MF, et al. 2006. Analysis of reports of human exposure to Micotil 300 (tilmicosin injection). J Am Vet Med Assoc 229:1737. Walker M, et al. 2016. Evaluation of the impact of methicillin-­resistant Staphylococcus pseudintermedius biofilm formation on antimicrobial susceptibility. Vet Surg 45:968. Weese JS, et al. 2019. International Society for Companion Animal Infectious Diseases (ISCAID) guidelines for the diagnosis and management of bacterial urinary tract infections in dogs and cats. Vet J 247:8.

Wilson DA, et al. 1996. Case control and historical cohort study of diarrhea associated with administration of trimethoprim-­ potentiated sulphonamides to horses and ponies. J Vet Intern Med 10:258. Woodward KN. 2005. Veterinary pharmacovigilance. Part 4. Adverse reactions in humans to veterinary medicinal products. J Vet Pharmacol Therapeut 28:185. Worthington RJ, Melander C. 2013. Combination approaches to combat multidrug-­resistant bacteria. Trends Biotechnol 31:177. Yunis AA. 1989. Chloramphenicol toxicity: 25 years of research. Am J Med 87:4N.

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Section II

Classes of Antimicrobial Agents

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7 Beta-­lactam Antibiotics: Penam Penicillins Laura Y. Hardefeldt and John F. Prescott

B ­ eta-­lactam Antibiotics Alexander Fleming’s observation in 1928 that colonies of staphylococci were lyzed on a plate contaminated with a Penicillium mold was the discovery that led to the development of antibiotics. In 1940, Chain and Florey and their associates were the first to produce enough penicillin from cultures of Penicillium notatum. Almost a decade later, penicillin G became widely available for clinical use. In its clinical application, this antibiotic was found to have limitations which included relative instability in gastric acid, susceptibility to inactivation by beta-­ lactamase (penicillinases), and relative inactivity against clinically important Gram-­negative bacteria. This inactivity against Gram-­negative rods was subsequently found to result from inability to penetrate the Gram-­negative cell wall, lack of available binding sites (penicillin binding proteins) or enzymatic inactivation. Intensive research led to the isolation of the active moiety, 6-­aminopenicillanic acid, within the penicillin molecule. This moiety, which consists of a thiazolidine ring (1) attached to a beta-­lactam ring (2) that carries a secondary amino group (R-­NH-­), is essential for antibacterial activity (Figure  7.1). Isolation of the active moiety has resulted in the design and development of semisynthetic penicillins that

overcome some of the limitations associated with penicillin G. The development of the cephalosporin family, which shares the beta-­lactam ring with penicillins (Figure 7.2), led to a remarkable array of molecules with varying ability to penetrate ­different Gram-­negative bacterial species and to  resist several beta-­lactamase enzymes (Chapter  8). Other naturally occurring beta-­ lactam antibiotics lacking the bicyclic ring of the classic beta-­lactam penicillins and cephalosporins have subsequently been described. Many have potent antibacterial activity and are highly inhibitory to beta-­lactamase enzymes. Some, such as the carbapenems, oxacephems, penems, and monobactams, have potent antibacterial activity whereas others, such as the oxapenam clavulanic acid, have no intrinsic antibacterial activity but possess potent beta-­ lactamase inhibitory activity (Chapter 9). These latter drugs are combined with older beta-­ lactams to increase their range of antibacterial activity. Beta-­lactam antibiotics are in widespread use because of their selectivity, versatility, and low toxicity.

Chemistry The penicillins, cephalosporins, carbapenems, monobactams, and penems are referred to as  beta-­lactam antibiotics. Rupture of the

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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Beta-­lactam Antibiotics: Penam Penicillins H C

R O

B

H C

S A

N

C

beta-­lactam ring for specific purposes, such as increasing resistance to beta-­lactamases of clinically important families or species of bacteria; enhancing activity against selected pathogens; or ensuring favorable pharmacokinetic properties. Thus, some semisynthetic beta-­ lactam drugs have been designed for specific purposes.

CH3

C

CH3 COOH

C H

β-lactam ring Thiazolidine ring

Figure 7.1  Structural formula of penicillin.

Mechanism of Action

beta-­lactam ring, which is brought about enzymatically by bacterial beta-­lactamases, results in loss of antibacterial activity. Hypersensitivity reactions appear to be associated with the active moieties of the beta-­lactam drugs, and although the risk is small, caution should be exercised when administering cephalosporins to penicillin-­sensitive animals because these drugs are of similar structure. In drug development, substitutions can be made on the

Beta-­lactam antibiotics prevent the bacterial cell wall from forming by interfering with the final stage of peptidoglycan synthesis. They inhibit the activity of the transpeptidase and other peptidoglycan-­active enzymes called penicillin-­binding proteins (PBPs) (transpeptidases, carboxypeptidases), which catalyze cross-­linkage of the glycopeptide polymer units that form the cell wall. The PBPs are further classified according to their molecular mass

N O

H β-lactam O

O R C HN

S

R C HN

S

N

N

R2

O

O

CO2H

CO2H Penicillin

Cephalosporin

OH2

H H3C

R

X

R3

R2 - HN

N N

O CO2H Carbapenem

R1

O Monobactam

Figure 7.2  Core structures of naturally occurring beta-­lactams.

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into high molecular weight PBPs and low molecular weight PBPs. In the presence of beta-­lactam drugs, the PBPs are inactivated when covalently bound to the beta-­lactam ring. The drugs exert a bactericidal action but cause lysis only of growing cells, that is, cells which are undergoing active cell wall synthesis. Also, the change in the bacterial cell wall attracts phagocytic activity by immune cells. In Gram-­ positive bacteria the beta-­lactams not only prevent final peptidoglycan cross-­linking, but also stimulate lipoteichoic acid release, causing a suicide response by degradation of peptidoglycan by autolysins. Variation in the activity of different beta-­ lactams results, in part, from differences in affinity of the molecules for the PBPs. The difference in susceptibility between Gram-­positive and Gram-­negative bacteria depends on differences in receptor sites (PBPs), on the relative amount of peptidoglycan present (Gram-­positive bacteria possess far more), on the ability of the drugs to penetrate the outer cell membrane of Gram-­ negative bacteria, and on resistance to the different types of beta-­lactamase enzymes produced by the bacteria. These differences are summarized in Figures 7.3 and 7.4. Beta-­lactam antibiotics are bactericidal drugs with slower kill rates than those exhibited by aminoglycosides or fluoroquinolones. Killing activity starts after a lag period. Penicillins tend to be slightly more active in a slightly acidic environment (pH 5.5–6.5), perhaps because of enhanced membrane penetration. Against Gram-­positive bacteria, all beta-­lactams exhibit an in vitro postantibiotic effect. This does not carry over for the streptococci in  vivo but does for strains of staphylococci that are susceptible to penicillins. The beta-­lactams do not exhibit a postantibiotic effect against Gram-­negative bacteria, with the possible exception of ­carbapenems against Pseudomonas. Optimal antibacterial efficacy is time and not ­concentration dependent (Chapter  5) and therefore requires that serum concentrations exceed the MIC of the pathogen for

(A)

Gram-positive bacteria

Penicillin PBP

(B)

Gram-positive bacteria

Penicillin PBP Penicillinase

(C)

Gram-positive bacteria

Cephalosporin PBP Penicillinase

Figure 7.3  Summary of action and resistance to beta-­lactam drugs: Gram-­positive bacteria. (A) Susceptible bacterium. (B) Exogenous beta-­ lactamase-­producing bacterium, e.g., Staphylococcus aureus. (C) Penicillinase-­producing bacterium susceptible to cephalosporin. PBP, penicillin-­binding proteins. Source: After R D. Walker, with permission.

essentially the entire dosing interval for bactericidal effects. These drugs are best administered frequently or by continuous infusion.

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­Beta-­lactam Antibiotic 

Beta-­lactam Antibiotics: Penam Penicillins

(A)

Gram-negative bacteria β-lactam β-lactamase PBP

(B)

Gram-negative bacteria β-lactam β-lactamase PBP

(C)

Gram-negative bacteria β-lactam β-lactamase PBP

Figure 7.4  Summary of action and resistance to beta-­lactam drugs: Gram-­negative bacteria. (A) Bacterium constitutively resistant to penetration by beta-­lactam. (B) Penetration by beta-­lactam but destruction by periplasmic beta-­lactamase. (C) Susceptible Gram-­negative bacterium. PBP, penicillin-­binding proteins. Source: After R D. Walker, with permission.

Resistance to Beta-­lactam Antibiotics Changes to PBPs can confer resistance. A change in the number of PBPs impacts the amount of drug that can bind to that target. This can occur by an increase in PBPs that have a decrease in drug-­binding ability or decrease in PBPs with normal drug binding. A change in structure (e.g., PBP2a in S. aureus by acquisition of the mecA gene) may decrease the ability of the drug to bind, or totally inhibit drug binding. The level of resistance is determined by how many and to what extent targets are modified. In Gram-­positive bacteria, especially S. aureus, resistance to penicillin G is mainly through the production of beta-­lactamase enzymes that break the beta-­lactam ring of most penicillins. Staphylococcus spp. secretes beta-­lactamase enzymes extracellularly as inducible exoenzymes that are plasmid mediated (Figure 7.3). Inherent resistance to penicillin G of many Gram-­negative bacteria results from low permeability of the Gram-­negative cell wall, lack of PBPs, and a wide variety of beta-­lactamase enzymes (Figure  7.4). Most Gram-­negative bacteria inherently express low levels of species-­specific, chromosomally mediated beta-­lactamase enzymes within the periplasmic space, which sometimes contribute to resistance. These enzymes hydrolyze susceptible cephalosporins more rapidly than penicillin G, but they poorly hydrolyze ampicillin, carbenicillin, and beta-­lactamase-­resistant penicillins. Production of plasmid-­m ediated beta-­ lactamases is widespread among common Gram-­ negative bacterial pathogens. The enzymes are constitutively expressed, present in the periplasmic space, and cause high-­level resistance. The majority are penicillinases rather than cephalosporinases (Figure 7.4). The most widespread are those classified based on their hydrolytic activity as TEM-­type beta-­lactamases, which readily

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hydrolyze penicillin G and ampicillin rather than methicillin, cloxacillin, or carbenicillin. The less widespread OXA-­type beta-­lactamases hydrolyze penicillinase-­stable penicillins (oxacillin, cloxacillin, and related drugs). More details on beta-­lactamases are given in Chapter  9. Beta-­ lactamases probably evolved from PBPs as a protective mechanism for soil organisms exposed to beta-­lactams in nature. Because of the spread of transferable resistance, beta-­lactamase production by pathogens is now widespread and extensive. A major advance has been the discovery of broad-­spectrum beta-­lactamase-­inhibitory drugs (e.g., clavulanic acid, sulbactam, tazobactam). These drugs have weak antibacterial activity but show extraordinary synergism when administered with penicillin G, ampicillin (or amoxicillin), and ticarcillin, because of the irreversible binding of the beta-­lactamase enzymes of resistant bacteria. Other beta-­lactamase inhibitors, such as cefotaxime and carbapenems, have potent antibacterial activity in their own right (Chapter 9).

P ­ enam Penicillins General Considerations The acidic radical (R), attached to the amino group of 6-­aminopenicillanic acid (Figure  7.1) determines the susceptibility of the resulting penicillin to hydrolytic degradation or enzymatic inactivation by bacterial beta-­lactamase, and the antibacterial activity of the molecule. Both these factors influence the clinical effectiveness of penicillins, which is also determined by the concentration attained at the site of infection. The nature of the acidic radical has little influence on the rate of elimination of penicillins but determines the extent of plasma albumin binding and, to a lesser degree, membrane-­penetrating ability. The

6-­aminopenicillanic acid moiety and structure of the acid radicals of some penicillins are shown in Figure 7.5. Penam penicillins are readily distinguished on the basis of antimicrobial activity into six groups (“generations”), which largely correspond to their time of introduction into clinical use (Table  7.1): (1) benzyl penicillin and its long-­acting parenteral forms; (2) orally absorbed penicillins similar to benzyl penicillin; (3) staphylococcal penicillinase-­resistant isoxazolyl penicillins; (4) extended-­ or broad-­spectrum penicillins; (5) antipseudomonal penicillins; (6) beta-­lactamase resistant penicillins. Since the 1940s, there has been progressive development of penicillins for clinical use, resulting in derivatives with similar activity to benzyl penicillin, but which can be administered orally and/or are resistant to S. aureus beta-­lactamase (penicillinase). Subsequently, orally administered penicillins were developed with a broader spectrum of activity, which involved greater Gram-­negative antibacterial activity, and penicillins active against P. aeruginosa. Despite considerable effort at identifying beta-­lactamase resistant penam penicillins, with the exception of temocillin, extended-­ spectrum penicillins are susceptible to beta-­ lactamase-­producing Gram-­negative bacteria. For this reason, the use of penicillins against common Gram-­negative bacteria is limited in favor of more recently introduced cephalosporin beta-­lactams (Chapter 8) or combination with beta-­lactamase inhibitors (Chapter 9).

Mechanism of Action The targets of all beta-­lactam drugs are the PBPs found on the outside of the cytoplasmic membrane, which are involved in synthesizing and remodeling the cell wall. Susceptibility of a bacterium to a penicillin depends on a combination of affinity for the PBP, ability to

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­Penam Penicillin 

Beta-­lactam Antibiotics: Penam Penicillins

(A) Site of amidase action

S

H R

CH

N

C

CH A

B C

N

O

CH

CH3 CH3 COOH

Site of penicillinase action (break in β-lactam ring)

6-Aminopenicillanic acid

(B)

Side chain

Name

CH2 CO

Benzyl penicillin, penicillin G

O

CH2

CO

C

C

CO

N

C O

CH

Phenoxymethyl penicillin, penicillin V

Oxacillin CH3

CO

Carbenicillin

COO–Na+ OCH3 CO

Methicillin

OCH3 O CH

C

(6-APA)

Ampicillin

(6-APA)

Amoxicillin

NH2 O HO

CH

C

NH2

Figure 7.5  Structural formulae of some penicillins. (A) Basic structure of penicillin G. (B) Structures that can be substituted at the R to produce a new penicillin.

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Table 7.1  Classification of the six groups of penam penicillins (6-­aminopenicillanic acid derivatives). Group

Important Derivatives

Antimicrobial Advantage

1. Benzyl penicillins

Procaine (long-­acting form)

Gram-­positive bacteria

2. Orally absorbed benzyl penicillins

Phenoxymethyl penicillin

Gram-­positive bacteria

3. Antistaphylococcal isoxazolyl penicillins

Cloxacillin, dicloxacillin, oxacillin, methicillin, nafcillin

Activity against penicillinase-­ producing (but not methicillin-­resistant) S. aureus and S. pseudintermedius

4. Extended-­(broad-­) spectrum penicillins

Aminobenzylpenicillins (ampicillin, hetacillin, pivampicillin, amoxicillin); amidopenicillins (mecillinam)

Broader spectrum than benzyl penicillins, but beta-­lactamase sensitive

5. Antipseudomonal penicillins

Ureidopenicillins (azlocillin, mezlocillin, piperacillin); carboxypenicillins (carbenicillin, ticarcillin)

P. aeruginosa activity, reduced Gram-­positive activity

6. Beta-­lactamase-­ resistant penicillins

Temocillin

Beta-­lactamase resistance (but not methicillin resistance)

penetrate the cell wall, and ability to resist beta-­lactamase enzymes (Figure  7.3, 7.4). There are usually 4–7  PBPs present in the bacterial cell wall that are the targets for penicillins. The bactericidal effect in Gram-­ negative bacteria results from osmotically induced lysis of cells weakened by loss of their peptidoglycan layer. Gram-­positive bacteria have considerably greater quantities of peptidoglycan in their cell wall than Gram-­ negative bacteria and an effect of beta-­ lactams is to prevent the final peptidoglycan cross-­linking, which gives peptidoglycan its strength. Also, beta-­lactams cause the release of lipoteichoic acid, leading to a suicide response by degradation of peptidoglycan by autolysins (endogenous endopeptidase, carboxypeptidase PBPs). For some Gram-­positive cocci, exposure to beta-­lactam antibiotics, above an optimal killing concentration, results in a reduction of killing, which can be considerable (the “Eagle” or paradoxical effect). Its basis appears to be interference of growth by penicillin binding to PBPs other than the major target PBP. Since beta-­ lactams are effective only against growing, actively cell wall-­synthesizing bacteria, failure to

grow results in failure to be killed. The Eagle effect is an important concept, since there may be a tendency to overdose with beta-­lactam antibiotics, because they are generally nontoxic.

Antimicrobial Activity Benzyl penicillin, penethamate hydriodide, and orally administered benzyl penicillins (phenoxymethyl penicillin) have outstanding activity against many susceptible Gram-­positive bacteria, notably beta-­hemolytic streptococci, nonresistant staphylococci, Actinomyces spp., Arcanobacterium spp., Bacillus spp., Clostridium spp., Corynebacterium spp., and Erysipelothrix rhusiopathiae. Susceptible Gram-­negative species include some Bacteroides spp., some Fusobacterium spp., and a variety of Gram-­ negative aerobic bacteria such as Haemophilus spp., and many Pasteurella spp. Enterobacterales, Bacteroides fragilis, most Campylobacter spp., Nocardia spp. and Pseudomonas spp. are resistant. Penicillinase-­resistant, antistaphylococcal isoxazolyl penicillins (cloxacillin, dicloxacillin, methicillin, nafcillin, oxacillin) have activity similar to but slightly less than that of benzyl

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­Penam Penicillin 

Beta-­lactam Antibiotics: Penam Penicillins

penicillin, with the exception that they are active against penicillinase-­producing S. aureus. Extended-­spectrum penicillins (aminobenzylpenicillins such as ampicillin and its esters, and amoxicillin) retain the activity of benzyl penicillin against Gram-­positive bacteria but have increased activity against Gram-­negative bacteria including E. coli, Proteus spp., and Salmonella spp. They are, however, ineffective against P. aeruginosa and are inactivated by beta-­ lactamases. Mecillinam, another member of the extended penicillin group, differs from aminobenzylpenicillins in its lower activity against Gram-­positive bacteria but considerably greater activity against Gram-­negative bacteria, including a broad spectrum of the Enterobacterales, although it is still inactivated by many beta-­ lactamases. Penicillins (carboxypenicillins, ureidopenicillins) active against P. aeruginosa (carbenicillin, azlocillin, mezlocillin, piperacillin) are effective against both Gram-­positive and Gram-­negative bacteria, including P. aeruginosa. It is important to note that acquired antimicrobial resistance is emerging globally but varies greatly between countries. For example, rates of penicillin resistance to staphylococci vary in different countries and even within different species in the same country (e.g., horses and dogs). Veterinarians should consult local antibiograms (where available) to assist with empirical prescribing decisions. Beta-­haemolytic streptococci species remain highly susceptible to penicillin universally. Although resistance has been described in S. equi subsp. equi isolated from the upper respiratory tract in horses in the United Kingdom (Fonseca et  al.,  2020), these isolates did not undergo repeated susceptibility testing or further analysis to confirm this finding, which should be performed in all cases where penicillin resistance in beta-­haemolytic streptococci species is detected.

Resistance to Penam Penicillins Most resistance results from production of a beta-­lactamase enzyme, although modification of PBPs with reduced drug affinity or reduced

bacterial permeability are additional and sometimes concurrent mechanisms of intrinsic or acquired resistance to penam penicillins. Efflux mechanisms and modification of porins in Gram-­negative bacteria that prevent entry of penicillins are also recognized. Beta-­lactamases are discussed in Chapter 9. Resistance because of exogenously produced beta-­lactamase is now widespread in S. aureus, particularly in clinical isolates, because of mobile genetic elements. Among Gram-­negative bacteria, plasmids encoding beta-­lactamases have also become widespread and are the cause of extensive acquired resistance. Modification of PBPs is recognized to be increasingly important as another mechanism of resistance to penam penicillins, particularly among Gram-­positive organisms. For example, a mutation in PBP5 in Enterococcus faecalis causes loss of affinity and beta-­lactam resistance. A third mechanism is to prevent the beta-­lactam antibiotic from reaching the target by altering the permeability of the outer membrane or increasing efflux pump activity. This is one of the main causes of resistance in Pseudomonas aeruginosa and other pathogenic Gram-­negative bacteria. The most important type of penam penicillin resistance in human medicine is methicillin (oxacillin) resistance in S. aureus (MRSA), which is widespread in humans in some countries, notably Japan and the United States. Also, methicillin resistance is well established in animal populations, notably in dogs, horses, and swine, and appears to reflect the incidence of infection in humans from whom these strains were acquired (Price et al., 2012; Chen and Wu,  2020). Methicillin resistance in S. aureus causing bovine mastitis varies by region, with the highest prevalence in Asia (6.5%) and the lowest in Europe (1.2%), but prevalence appears to be increasing (Zaatout and Hezil, 2022). The reason(s) for the emergence of MRSA in animals since 2000, and of livestock-­associated (LA) MRSA infections, are still unclear but represent host adaptation of particular clonal

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types to livestock, with antimicrobial resistance developing through selection by antimicrobial use. LA-­MRSA has been spreading rapidly among pig herds in concurrence with the common use of high-­dose zinc oxide veterinary medicinal products, used to prevent postweaning diarrhea in piglets. The gene coding for zinc and cadmium resistance is located within the same mobile genetic element as a certain type of Mec, which is a precursor for the methicillin resistance gene mecA, conferring resistance to broad-­spectrum β-­lactams. In addition, animal MRSA strains are often hospital associated and can contaminate veterinary hospital environments, including hospital personnel, to a remarkable extent. Human subclinical and even clinical infections have been acquired from animal sources. MRSA are regarded as resistant to all beta-­lactam antimicrobials and are commonly, but not always, resistant to other antimicrobials. Methicillin-­ resistant S. pseudintermedius (MRSP) is also increasingly isolated from dogs and cats and, like MRSA, is regarded as resistant to all beta-­ lactam antibiotics. Emergence of new and diverse lineages is well documented (Phophi et  al.,  2023). They commonly also have other multidrug resistances. Methicillin resistance is more frequent in Staphylococcus spp. coagulase-­negative Coagulase-­negative Staphylococcus spp. have rarely been associated with clinical disease in the past but may be increasing in significance in companion animal practice where Staphylococcus schleiferi appears to be a significant pathogen in some cases of canine otitis externa (Lee et al., 2019).

Pharmacokinetic Properties The penicillins are organic acids that are generally available as the sodium or potassium salt of the free acid. In dry, crystalline form, penicillins are stable but lose their activity rapidly when dissolved. Apart from the isoxazolyl penicillins (cloxacillin, dicloxacillin, oxacillin) and penicillin V, acid hydrolysis in the stomach

limits the systemic availability of most penicillins from oral preparations. The penicillins (pKa 2.7) are predominantly ionized in plasma, have relatively small apparent volumes of distribution (0.2–0.3 l/kg), and have short half-­lives (0.5–1.2 hours) in all species of domestic animals. After absorption, they are widely distributed in the extracellular fluids of the body, but cross biological membranes poorly since they are ionized and poorly lipid soluble. Concentration in milk, for example, is about one-­fifth that of serum. Entry across biological membranes or through the blood–brain or blood–cerebrospinal fluid barrier is enhanced by inflammation, so that inhibitory drug concentrations may be attained at these sites that are normally inaccessible to penicillin. Penethamate hydriodide, a prodrug of benzyl pencillin, is an exception in that, unlike penicillin salts, it is a weak base that exists in a nonionized state in plasma. After intramuscular injection, it is rapidly absorbed. Due to the pH gradient present between milk and plasma combined with its highly liposoluble properties and its basic state, undissociated penethamate easily crosses the blood–milk barrier. The drug partially dissociates into penicillin G and the remaining undissociated penethamate effectively penetrates the gland parenchyma and into the mammary cells where it is rapidly ionized to penicillin G, thereby limiting its return to interstitial fluid and thus getting trapped intracellularly and intramammary in increasing concentrations. Field studies that investigated the efficacy of systemic penethamate hydriodide treatment in cows with subclinical and clinical mastitis have demonstrated high bacteriological cure rates for staphylococci and streptococci (St Rose et  al.,  2003; Serieys et al., 2005). Penicillins are eliminated almost entirely by the kidneys, which results in very high levels in the urine. Nafcillin is an exception, in that it is excreted mainly in bile. Renal excretion mechanisms include glomerular filtration and tubular secretion. The latter is subject to

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­Penam Penicillin 

Beta-­lactam Antibiotics: Penam Penicillins

competitive inhibition by other organic acids, such as probenecid. Impaired renal function delays excretion of the penicillins, but the wide margin of safety of this class of drug offsets the absolute need to adjust dosage.

Drug Interactions Penicillins are usually synergistic with the aminoglycosides against many bacteria, which are susceptible to each drug alone, because disruption of the bacterial cell walls enhances penetration of the aminoglycoside. Such synergism may even occur with penicillinase-­producing S. aureus. Penicillins are synergistic against these organisms (except MRSA) with drugs that bind beta-­lactamase enzymes, such as cloxacillin, clavulanic acid, sulbactam, tazobactam, and some cephalosporins. Aminobenzyl penicillins and ureidopenicillins are increasingly combined with beta-­lactamase inhibitors (Chapter 9).

Toxicity and Adverse Effects Penicillins and beta-­lactam antibiotics generally are remarkably free of toxic effects even at doses grossly more than those recommended. The major adverse effects are acute anaphylaxis and collapse; hypersensitivity reactions (urticaria, fever, angioneurotic edema) are more common. All penicillins are cross-­sensitizing and cross-­ reacting, but cross-­reactions occur in only about 5–8% of human patients treated with cephalosporins. Anaphylactic reactions are less common after oral than parenteral administration. Penicillins must not be used in animals known to be sensitive. Other reported adverse reactions include immune-­mediated hemolytic anemia and thrombocytopenia. These generally resolve with discontinuation of penicillin.

Dosage Considerations Beta-­lactams produce killing and lysis of susceptible bacteria at concentrations above MIC. Postantibiotic effects are observed only for

staphylococci in  vivo, and for carbapenems against Pseudomonas spp., so that dosage requires that drug concentrations exceed MIC for most of the dosage interval (“time-­dependent antibiotics”) for optimal bactericidal effects. However, bacteriostatic levels may be all that is required in nonneutropenic patients in which case levels need to exceed MIC for approximately 20%, 25–30%, and 25-­40% of the dosing interval for carbapenems, penicillins, and cephalosporins respectively, based on animal studies (Abdul-­ Aziz et al., 2015). Excessive drug concentrations may be counterproductive because of the ‘Eagle’ effect described earlier, in which sometimes dramatic reduction of killing occurs in the presence of high, supra-­MIC concentrations.

Clinical Usage Penicillins (Table 7.1) are important antibacterial drugs in the treatment of infections in animals. The often exquisite susceptibility of Gram-­positive bacteria, such as the beta-­ hemolytic streptococci, means that benzyl penicillin is the drug of choice for these infections, because of its high potency and low toxicity. Antistaphylococcal penicillins are in widespread use in the prevention and treatment of staphylococcal infections in cows. The extended-­spectrum penicillins, particularly aminobenzyl penicillins, have lost much of their potency against Gram-­negative bacteria over the decades, but have been revitalized by their combination with beta-­lactamase inhibitors (Chapter  9). The antipseudomonal penicillins remain important for this activity but are less efficacious than antipseudomonal cephalosporins.

­ roup 1: Penicillin G and LongG ­acting Parenteral Forms Sodium benzyl penicillin G (also known as penicillin G) is available as the benzyl, the procaine benzyl, and now rarely as the tribenzyl ethylenediamine (benzathine) forms. Frequent

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dosing of benzyl penicillin is required due to its rapid excretion, so that long-­acting delayed absorption (benzathine) forms have been developed, with procaine penicillin being the most extensively used because dosing frequency is usually q12 hours (equine) and q24 hours (bovine). The principle behind the use of procaine and benzathine penicillin is that both forms delay absorption from the injection site. Thus, while the elimination half-­life is the same, the absorption half-­life is much longer, thus reducing the need for frequent dosing. Delayed absorption also means a lower peak concentration which is of particular concern for benzathine penicillin since it fails to meet therapeutic concentrations for many common pathogens. Penethamate is a prodrug of benzyl penicillin which dissociates to penicillin G in the mammary gland, resulting in prolonged therapeutic concentrations.

Antimicrobial Activity The activity of penicillin G (benzyl penicillin) was originally defined in units. Crystalline sodium penicillin G contains approximately 1600 units/mg (1 unit = 0.6 μg; 1 million units of penicillin = 600 mg or 0.6 g). Most semisynthetic penicillins are prescribed by weight (mg/kg) rather than units. Antimicrobial susceptibility categories given below are guidelines only; bacterial susceptibility in veterinary medicine may vary by region, species, and over time (Chapter 2). ●●

Good susceptibility (MIC 4 μg/ml) is shown by Enterobacterales (other than some Proteus spp.), Bacteroides fragilis, Bordetella spp., most Campylobacter spp., and Nocardia spp.

Antimicrobial Resistance Despite extensive use of penicillin in veterinary medicine for many years, most Gram-­positive bacteria remain susceptible to the drug. Staphylococcus aureus and S. pseudintermedius are notable exceptions. The beta-­lactamase enzymes of S. aureus are mainly active against penicillin G, ampicillin, and carbenicillin but  hydrolyze penicillinase-­stable penicillins (methicillin, cloxacillin) and cephalosporins poorly. Methicillin-­resistant S. aureus (MRSA) and methicillin-­resistant S. pseudintermedius (MRSP) have spread within animals, and have become increasingly problematic, particularly since they are both resistant to all beta-­lactams and may also be multidrug resistant. Resistance in usually susceptible Gram-­negative bacteria such as Haemophilus and Pasteurella is the result of R plasmid-­mediated production of beta-­lactamases.

Pharmacokinetic Properties These were discussed earlier under general properties of penam penicillins. Acid hydrolysis within the stomach limits the systemic availability of benzyl penicillin administered orally.

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Drug Interactions Penicillin G is synergistic with the aminoglycosides against many Gram-­positive bacteria, except those showing high-­level aminoglycoside resistance. Such synergism may be seen even with penicillinase-­producing S. aureus. Penicillin is synergistic against these organisms with drugs that bind beta-­lactamase enzymes (Chapter  8). Penicillin G has been combined with streptomycin for use in animals but there is little clinical evidence supporting the clinical value of the combination. For this reason, and because streptomycin is associated with tissue residues, the combination is no longer available in most countries. In addition, there are significant differences in pharmacokinetic properties between different combined preparations.

Toxicities and Adverse Effects The parent penicillin G and its numerous derivatives are relatively safe drugs; toxic effects were described under General Considerations. Many of the acute toxicities reported in animals are the result of the toxic effects of the potassium or procaine with which penicillin is combined in the dosage form. To avoid cardiac arrest, care should be taken with the rate at which potassium penicillin G is injected IV in small sized animals; administration of the sodium salt is safer. Procaine penicillin G should never be given IV. Procaine reactions may occur when high doses are given IM. Particularly in horses, this may cause nervous excitement (incoordination, ataxia, excitability) and can result in death but usually from misadventure rather than drug effects. There is no specific treatment. Procaine is usually restricted in racing horses and long withholding periods must be observed to avoid procaine-­ positive drug test results. Procaine penicillin G should be stored in the refrigerator and not used past expiry dates; repeated use of the same IM injection site should be avoided, especially in horses as this may

predispose to inadvertent IV administration. Severe, immune-­mediated hemolytic anemia with icterus has been reported in horses. Dysbiosis is common in pocket pets including mice, rabbits, chinchillas, guinea pigs, and hamsters.

Administration and Dosage Recommended dosages are shown in Table 7.2. Because of the relative lack of toxicity of penicillins, their dosage can be tailored, to some extent, to the susceptibility of the infecting bacteria more than with any other class of antibiotic. The effectiveness of penicillin therapy is related to the time that tissue concentration exceeds the MIC of the pathogen. Because of the short half-­lives of penicillins, preparations that provide rapid absorption must be administered at short intervals (q6 hours). Low systemic availability from oral forms must be compensated for by increasing the size of the dose, except where treating GI infections (e.g., in pigs and poultry). Penicillin G is available as a potassium or sodium salt that can be administered parenterally as freshly prepared solutions. Procaine penicillin G is a special form developed to prolong absorption from the IM injection site. A single dose of 25 000 units/kg provides effective serum concentrations against susceptible bacteria for at least 12 hours (e.g., horses) and for up to 24 hours (e.g., cattle) in all species of domestic animals. For moderately susceptible bacteria, high doses of procaine penicillin given once daily may be useful; an example is administration of 45 000 units/kg in the once-­daily treatment of bovine Mannheimia haemolytica pneumonia but more clinical data are needed on the efficacy of such high dosing, since the Eagle effect may reduce the efficacy of the drug. Benzathine penicillin is a long-­acting, slow-­ release formulation of penicillin G administered every 72 hours. The benzathine penicillin never reaches therapeutic concentrations yet is slowly eliminated and frequently results in violative residues in food animals. It should not be used in lactating dairy cattle. Label doses can result in residues 30–60 times the maximum

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132

Table 7.2  Usual dosages of penam penicillins in animals. Note that these uses and dosages do not apply to all species; check species-­specific chapters. Drug

Route

Dose (IU or mg/kg)

Interval (h)

Comment

Penicillin G, sodium aqueous

IM, IV

15–20 000 IU

6–8

Procaine penicillin G

IM

25 000 IU

24

Every 12 hours for serious infections

Benzathine penicillin

IM

40 000 IU

72

Highly susceptible bacteria only; best avoided

Penicillin V

Oral

10

6–8

Erratic absorption; amoxicillin preferred

Cloxacillin, dicloxacillin, methicillin, oxacillin

Oral

15–25

6–8

Monogastrates only

Ampicillin sodium

IM, IV

10–20

6–8

Ampicillin (hetacillin)

Oral

10–20

8

Monogastrates only

Amoxicillin

Oral

10–20

8–12

Monogastrates only

Amoxicillin

IM (SC)

10

12

Amoxicillin, long-­acting

IM

15

48

Very susceptible bacteria only

Amoxicillin trihydrate

IM

10–20

12

Pivampicillin

Oral

25

12

Monogastrates only

Carbenicillin, indanyl sodium

Oral

33

6–8

Urinary tract only

Carbenicillin

IM, IV

33

6–8

Ticarcillin

IV (IM, SC)

25–40

8

Often used with clavulanic acid

Piperacillin

IV (IM)

50

8

May be used with tazobactam

IM, intramuscular; IV, intravenous; SC, subcutaneous.

residue limits at the injection site even when label withdrawal times are followed.

Clinical Applications The general clinical applications of penicillin G are shown in Table 7.3. Penicillin G is the drug of choice in treating infections caused by Gram-­positive bacteria such as streptococci, corynebacteria, Erysipelothrix, clostridia, and perhaps Listeria, and some Gram-­ negative bacteria such as H. somni, Pasteurella, and many anaerobes. The advantages of penicillin G are its potent and bactericidal activity against susceptible bacteria and its wide margin

of safety; dosage can be tailored to the susceptibility of the pathogen by selecting the form of drug to be administered. Disadvantages are activity only against actively growing bacteria, its need for injection, widespread resistance in S. aureus and Gram-­negative bacteria, and the drug’s failure to cross biological membranes well, except in acute inflammation. Cattle, Sheep, and Goats

Penicillin G is the most commonly used antibiotic for food animals. It was initially licensed at an inappropriately low dosage in many countries. The label has been updated in Canada and New Zealand but remains low in most other

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­Group 1: Penicillin G and Long-­acting Parenteral Form  133

Beta-­lactam Antibiotics: Penam Penicillins

Table 7.3  Applications of penicillin G in clinical infections in animals. Species

Primary Applications

Secondary Applications

Cattle, sheep, goats

Anthrax, clostridial and corynebacterial infections, T. pyogenes, streptococcal mastitis, hemorrhagic septicemia, listeriosis

Actinobacillosis, anaerobic infections, possibly infectious keratoconjunctivitis, leptospirosis

Swine

Streptococcal, clostridial infections, erysipelas, T. pyogenes, A. suis

Glasser disease, pasteurellosis, anaerobic infections

Horses

Streptococcal and clostridial infections

Actinobacillosis, anaerobic infections

Dogs, cats

Streptococcal and clostridial infections

Cat bite abscess, anaerobic infections, leptospirosis

jurisdictions. Parenterally administered penicillin G is the drug of choice for the treatment of disease caused by susceptible bacteria including anthrax, clostridial infections, Corynebacterium renale infection, H. somni infection, pneumonic pasteurellosis caused by susceptible Mannheimia and Pasteurella, ­septicemic pasteurellosis (hemorrhagic septicemia), and infections caused by nonspore-­ forming anaerobes such as Fusobacterium necrophorum and Porphyromonas asaccharolytica. Penicillin G’s poor activity against slowly multiplying bacteria and relative inability to penetrate biological membranes and biofilms may explain its often disappointing effect in treating T. pyogenes, actinomycosis, or chronic S. aureus mastitis. Listeriosis has been successfully treated with a daily dose of 44 000 units/kg of penicillin G administered for 7–14  days, but ampicillin is preferred and oxytetracycline also has good efficacy. Penicillin G is effective against acute leptospirosis, although again, ampicillin is probably preferable. Procaine penicillin G (300 000–600 000 units in 1–2 ml) administered subconjunctivally has been used extensively in the treatment of Moraxella bovis keratoconjunctivitis since this maintains therapeutic concentrations for up to 36 hours. Pneumonic pasteurellosis has been treated successfully with daily intramuscular or subcutaneous injections of 45 000 units/kg of

procaine penicillin. However, resistance among M. haemolytica is increasing and ­further increases in dose are not justified. Serious, acute mastitis caused by streptococci or susceptible S. aureus can be treated by IM procaine penicillin 20–25 000  IU/kg, q12 or q24 hours. Penicillin is more commonly administered intramammarily, often combined with novobiocin, and has given excellent results in the treatment of streptococcal infections during lactation but only modest results against S. aureus. Intramammary treatment of susceptible Gram-­positive cocci with procaine penicillin G and neomycin showed no advantage over procaine penicillin G alone (Taponen et al., 2003). Penicillin G in fixed combination with streptomycin has been used successfully against severe Dermatophilus infection but this combination is no longer available in many countries. Penethamate hydriodide penetrates well into the mammary gland, and dissociates to penicillin G, with activity against Gram-­positive bacteria causing clinical and subclinical mastitis. Swine

Penicillin is the parenteral drug of choice in preventing and treating erysipelas, and streptococcal, clostridial, and corynebacterial infections. For acute erysipelas and streptococcal infections, procaine penicillin G is preferred and benzathine penicillin should be avoided as

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134

therapeutic plasma concentrations are not likely to be achieved. Streptococcus suis meningitis may be treated successfully with daily injections of procaine penicillin given early in the disease. Horses

Procaine penicillin G is used in the treatment of infections caused by S. equi subsp. zooepidemicus and S. equi subsp. equi (“strangles”) such as wounds and respiratory tract infections. The human IV formulations of potassium or sodium penicillin G are used in combination with an aminoglycoside (e.g., amikacin or gentamicin) in the treatment of severe infections such as neonatal sepsis or bacterial pleuropneumonia and for surgical prophylaxis prior to colic surgery. They are also used for intrauterine infusions for metritis and for sinus lavage in cases of sinusitis. Penicillin, while not an antidote, is the preferred antibiotic in tetanus. Injection of procaine penicillin G in the neck or biceps gave higher serum concentrations than injection in the gluteal muscle or SC (Firth et al., 1986). Penicillin should not be administered orally to horses because of its poor absorption and the digestive disturbances it may cause. Dogs and Cats

Due to the availability of amoxicillin and ampicillin, procaine penicillin G is rarely used in small animals.

­ roup 2: Orally G Absorbed Penicillins Phenoxymethyl penicillin (penicillin V) resists stomach acid hydrolysis and is therefore administered orally. It has a spectrum of activity similar to penicillin G and is therefore used for the same purposes in monogastrates. Penicillin is used by oral administration in the prevention and treatment of necrotic enteritis, ulcerative enteritis, and intestinal spirochetosis and in treating erysipelas in turkeys.

­ roup 3: Antistaphylococcal G Isoxazolyl Penicillins: Cloxacillin, Dicloxacillin, Flucloxacillin, Methicillin, Nafcillin, and Oxacillin The antistaphylococcal penicillins are resistant to staphylococcal penicillinase and are used mainly in the treatment or prevention of bovine staphylococcal mastitis. The isoxazolyl penicillins (cloxacillin, oxacillin) are acid stable and may be given orally to monogastric animals, for example in the treatment of staphylococcal skin infections in dogs. Penicillinase production in S. aureus may be detected by using nitrocefin-­impregnated paper disks. All are resistant to S. aureus and S. pseudintermedius penicillinase, although activity against other penicillin-­susceptible bacteria is less than that of penicillin G. Activity of the different drugs is similar in vivo. As described earlier, carriage of, and infections from, methicillin-­resistant S. aureus (MRSA) are reported increasingly, including in the community, but particularly in dogs and in horses that are or have been in veterinary hospitals, as well as in farm livestock (LA-­MRSA), notably horses (LA-­MRSA in Europe), swine and veal calves (Price et  al.,  2012; Cuny et al., 2015). Resistance to methicillin in bovine S. aureus isolates is unusual, especially in isolates from intramammary infections, although MRSA are increasingly isolated from veal calves in certain countries and association with mastitis appears to be increasing (Zaatout and Hezil, 2022). Figures purporting to show extensive resistance in bovine isolates probably reflect inappropriate test conditions or drug inactivity, as methicillin deteriorates readily in storage. As noted earlier, methicillin-­resistant S. pseudintermedius (MRSP) is also increasingly isolated from dogs and cats and, like MRSA, is resistant to all beta-­lactam antibiotics. Canine isolates of MRSA and MRSP are commonly multidrug resistant (Morris et  al.,  2017) but such isolates are not more virulent than susceptible strains. With the emergence of MRSA in animals since about 2000, MRSA is an occupational health

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Group 3: Antistaphylococcal Isoxazolyl Penicillins  135

Beta-­lactam Antibiotics: Penam Penicillins

hazard for veterinarians and veterinary staff, particularly for those who work with horses and swine (Jordan et al., 2011). Staphylococcus pseudintermedius, including MRSP, can also cause zoonotic infections (Somayaji et  al.,  2016) although the risk of transmission is generally regarded as low. Routine hand hygiene and use of protective clothing, as well as routine facility cleaning and disinfection, are critical in controlling the spread of multidrug pathogens such as MRSA and MRSP (Morris et al., 2017). Methicillin-­resistant (heteroresistant) S. aureus may be overlooked. While no single method is ideal and consensus is lacking, methicillin-­ resistant S. aureus and S. pseudintermedius are best detected using oxacillin disks, with S. aureus grown 18–24 hours at 30 °C or 35 °C. Confirmation by polymerase chain reaction (PCR) of the presence of the mecA gene is desirable. Heteroresistant S. aureus is often multidrug resistant, including to other beta-­ lactams, gentamicin, macrolides, and tetracyclines. Methicillin-­resistant S. pseudintermedius is considered to be resistant (MRSP) if the MIC to oxacillin is >0.5 μg/ml, whereas the breakpoint for MRSA is >4 μg/ml (Bemis et al., 2009). A new methicillin resistance gene, named mecC, was first described in 2011 in both humans and animals. Since then, this gene has been detected in different food producing and other animals and as an agent causing infections in some humans. Veterinary cases of  mecC-­MRSA infections have been reported across Europe and the UK, and in Malaysia and Japan. Isolates usually contain few resistance (except for beta-­lactams) and virulence genes, but the first isolates harboring important virulence genes or that are resistant to nonbeta-­ lactams have already been described. Moreover, severe and even fatal human infection cases have been detected.

­ roup 4: Extended-­spectrum G Penicillins: Aminobenzyl Penicillins: Ampicillin, Amoxicillin Ampicillin, amoxicillin, and the related esters bacampicillin, hetacillin, pivampicillin, and talampicillin have similar antimicrobial activity,

but amoxicillin and possibly pivampicillin have the advantage of achieving higher tissue concentrations because of better absorption from the intestine. The broad-­spectrum aminobenzyl penicillins are slightly less active than penicillin G against Gram-­positive and anaerobic bacteria and are equally susceptible to staphylococcal penicillinase. These broad-­spectrum drugs, however, have considerably greater activity against Gram-­negative bacteria such as E. coli, P. mirabilis, and Salmonella. Nevertheless, acquired resistance has considerably reduced the effectiveness of these drugs.

Antimicrobial Activity Antimicrobial susceptibility categories given below are guidelines only; laboratory breakpoints available in veterinary medicine will vary with dosage, sites of infection, route of administration, and other variables (Chapter 2). ●●

●●

●●

Good susceptibility (MIC 4 μg/ml, approximately). Bacteroides fragilis, B. bronchiseptica, Citrobacter spp., Enterobacter spp., Klebsiella spp., other Proteus spp., P. aeruginosa, Serratia spp., Y. enterocolitica.

Antimicrobial Resistance Plasmid-­ or integron-­mediated acquired resistance is common in Gram-­negative bacteria and is often multiple, such as that in most enterotoxigenic E. coli and S. typhimurium. Many E. coli associated with bovine intramammary infections are resistant. Aminobenzyl penicillins are susceptible to S. aureus beta-­lactamase.

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136

Pharmacokinetic Properties The basic pharmacokinetic properties of penicillins were described under General Considerations. The aminopenicillins can penetrate the outer layer of Gram-­negative bacteria better than penicillin G. However, resistance to the aminopenicillins is easily acquired by Gram-­negative bacteria. Both ampicillin and amoxicillin are relatively stable in acid. In dogs, the systemic availability of amoxicillin (60–70%) is about twice that of ampicillin (20–40%), so that peak blood concentrations are often twice or more those that occur after the same dose of ampicillin. The absorption of amoxicillin is unaffected by feeding, unlike ampicillin. Hetacillin and pivampicillin are esters of ampicillin developed to increase systemic availability, but it is questionable whether this occurs in dogs. Pivampicillin has significantly better bioavailability in horses than amoxicillin after oral administration. Ampicillin is available as a sodium salt that can be administered parenterally in a freshly prepared solution. The trihydrate salts are less soluble and therefore poorly absorbed from the intestine but form aqueous suspensions that can be injected either IM or SC. These preparations produce low peak concentrations in the serum but extend the dosing interval to 12 hours. Long-­acting preparations of amoxicillin and ampicillin trihydrate have been introduced which produce therapeutic serum concentrations for 48 hours against bacteria with good susceptibility. The lower peak plasma concentrations, however, may decrease penetration of the antibiotic to sites of infection.

Drug Interactions Aminobenzyl penicillins are commonly synergistic with aminoglycosides against Gram-­ positive bacteria and often also against Gram-­negative bacteria, but only if the latter are not resistant to both drugs. The broad-­spectrum beta-­lactamase inhibitors clavulanic acid and sulbactam show remarkable synergism with

aminobenzyl penicillins against beta-­lactamase-­ producing bacteria (Chapter 9).

Toxicities and Adverse Effects Toxic effects are similar to those described under General Considerations. One hazard with broad-­spectrum penicillins is the potential to disturb the normal intestinal flora. In dogs and cats, the effect may be less marked with amoxicillin, which is better absorbed. Ampicillin should not be administered to small rodents (guinea pigs, hamsters, gerbils) or to rabbits since it may produce clostridial colitis (C. difficile or, in rabbits, C. spiroforme). Moderate diarrhea has been described in calves after several days’ treatment with oral ampicillin, which appears to result from malabsorption caused by a direct effect on intestinal mucosa.

Administration and Dosage Recommended dosages are shown in Table 7.2. The soluble sodium salts can be administered parenterally and orally but the poorly soluble trihydrate form should only be administered IM. Reconstituted, aqueous sodium salts are unstable after more than a few hours. Because of their short half-­lives, preparations that are rapidly absorbed should be administered every six hours to maintain serum drug concentrations over 1 μg/ml for a significant length of time. Amoxicillin is preferred for oral administration because it is better absorbed than ampicillin, and its absorption is unaffected by feeding. Another advantage of oral amoxicillin over ampicillin is that it can be given twice daily to small animals. Long-­acting preparations of amoxicillin are available, but it is doubtful whether they maintain therapeutic serum concentrations for the 48-­hour recommended dosing interval.

Clinical Applications The aminobenzyl penicillins are bactericidal, relatively nontoxic drugs with a broader spectrum of activity than penicillin G and are better distributed in the body. Even with these

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­Group 4: Extended-­spectrum Penicillins: Aminobenzyl Penicillins: Ampicillin, Amoxicilli  137

Beta-­lactam Antibiotics: Penam Penicillins

advantages, relatively high doses are required to treat infections caused by Gram-­negative bacteria. The relatively high prevalence of acquired resistance has limited their effectiveness. Amoxicillin is the best penicillin for the treatment of urinary tract infections and enteric infections caused by susceptible organisms in dogs and cats and has similar activity to penicillin G in the treatment of anaerobic infections. Although amoxicillin offers pharmacokinetic advantages and is preferred over ampicillin, it has some of the same difficulty as ampicillin in attaining concentration in tissues approximating those of susceptible Gram-­negative bacteria. The main clinical applications are similar to those shown in Table 7.3. Amoxicillin is a drug of choice in the treatment of leptospirosis. Long-­acting amoxicillin administered twice at 15  mg/kg IM q48 hours was effective in removing the Leptospira hardjo kidney carrier state from the majority of experimentally infected cattle (Smith et al., 1997). Sodium ampicillin is used in combination with an aminoglycoside (e.g., amikacin or gentamicin) for surgical prophylaxis and the treatment of severe infections such as neonatal sepsis and bacterial pleuropneumonia. Ampicillin or amoxicillin are drugs of choice for mixed aerobic-­anaerobic infections such as cat-­bite infections. Amoxicillin is a good first-­line option for the treatment of sporadic bacterial cystitis in dogs and cats; the MIC breakpoint of 4 × MIC) may be a drawback of newer cephalosporins, since these characteristics are associated with resistant bacterial superinfection and gastrointestinal disturbance. Widespread use of third-­ generation cephalosporins in human and veterinary medicine may have been one of the important factors underlying the resistance crisis in medicine and has been associated with the striking emergence and dissemination of multiple forms of beta-­lactamases observed in recent years. This may also apply to the use of cefovecin in dogs. The fifth edition of this book stated that second-­ and third-­generation cephalosporins are not first-­choice antimicrobial agents in animals but rather should be reserved for use where susceptibility testing indicates that alternatives are not available. This remains the opinion of the authors, but these drugs are increasingly widely used in veterinary medicine as first-­choice antimicrobials. This use is under increasing scrutiny by the medical profession, public health officials, and governments. There has been a remarkable rise in

resistance through ESBLs in Enterobacterales from both food and companion animals (including food-­borne pathogens, such as Salmonella) associated with the increased use of later generation cephalosporins. The association between ceftiofur use in eggs or day-­old broiler chicks with CMY-­2 beta-­lactamase-­ producing Salmonella and E. coli, and the spread of resistant S. Heidelberg into the human population documented in Canada and the US, led to drug label changes and bans on such use. One response to the rise of ESBLs in the US was the prohibition in 2012 by the Food and Drug Administration of the extra-­label use of cephalosporins in food animals (Food and Drug Administration,  2012). This prohibition extends to use for disease prevention, use at unapproved doses, frequencies, durations, or routes of administration, and use of human or companion animal drugs. Ceftiofur can be used for an extra-­label disease treatment (e.g., Gram-­negative sepsis) as long as the label dosage regimen is followed. The ban does not extend to the extra-­label use of cephapirin products in food-­producing species or extra-­ label use in food-­producing minor species (e.g., ducks, rabbits). In Denmark, voluntary discontinuation of cephalosporin use in swine in 2010  was associated with a decline in ESBL-­ resistant E. coli in pigs at slaughter. There are severe restrictions on the use of third-­ and later generation cephalosporins in some countries in Europe, including Denmark, France, and The Netherlands. For example, in France there is a legal requirement for both clinical examination and susceptibility testing before using highest priority, critically important antimicrobials such as third-­g eneration cephalosporins (ANSES-­ANMV, 2018). This includes the demonstration that lower World Health Organization (2016) category antimicrobials are ineffective. This regulation has considerably reduced the use of these drugs by veterinarians. Stewardship issues are discussed further in Section 3 of this book.

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150

F ­ irst-­generation Cephalosporins: Cefacetrile, Cephaloridine, Cefazolin, Cephapirin, Cephradine, Cephalothin, Cefadroxil, Cephradine, Cephalexin, and Cephaloglycin First-­generation cephalosporins have high activity against Gram-­positive bacteria, including penicillinase-­producing S. aureus and S. pseudintermedius; moderate activity against certain nontransferable, beta-­lactamase-­ producing, Gram-­negative Enterobacterales and fastidious Gram negatives; and no activity against Enterobacter spp., P. aeruginosa, and Serratia spp., among others. Antimicrobial activity of oral first-­generation cephalosporins is similar to that of aminopenicillins with the addition of resistance to the beta-­lactamase of S. aureus. For susceptibility testing, cephalexin should be used as the class drug in dogs rather than cephalothin, which has been used in the past. Cefazolin may also be tested since it is more active against Gram-­negative bacteria. Acquired resistance is common in Gram negatives and is particularly important in Enterobacterales. Methicillin-­resistant S. aureus and methicillin-­resistant S. pseudintermedius (MRSP), discussed in Chapter  7, are resistant to all cephalosporins except those in the fifth generation. Apart from MRSA and MRSP, which are increasingly detected in companion animals, pigs, and horses, acquired resistance is rare in Gram-­positive bacteria. Canine isolates of MRSA and MRSP are commonly multidrug resistant (Morris et al., 2017). Antimicrobial susceptibility categories given below are guidelines only; bacterial susceptibility in veterinary medicine may vary by region and species, and over time (Chapter 2). ●●

Predictable susceptibility (32 μg/ml). Acinetobacter spp., Bacteroides fragilis, Bordetella bronchiseptica, Campylobacter spp., Citrobacter spp., Enterobacter spp., Nocardia spp., P. aeruginosa, Rhodococcus equi, Serratia spp., and Yersinia spp. MRSA and MRSP are resistant.

Pharmacokinetic Properties There is widespread distribution within extracellular fluids but poor penetration across biological membranes (including into the udder) and physiological barriers (such as the blood– brain barrier). Inflammation enhances passage across barriers. The majority of cephalosprorins are rapidly eliminated, unchanged, in the urine, and tubular secretion (but not glomerular filtration) can be inhibited by probenecid to reduce clearance from the body. Plasma protein binding is low. Parenteral formulations of first-­generation cephalosporins are rapidly and completely absorbed after IM or SC injection. Cephalothin and cephapirin are metabolized into less active desacetyl derivatives. The plasma elimination half-­life is less than one hour. Oral cephalosporins have pharmacokinetic properties similar to penicillin V and the aminobenzyl penicillins. Generally, they are rapidly and completely absorbed after oral administration in monogastrates, but not horses. These drugs are unaffected by the presence of food (except for cephradine). Plasma elimination half-­lives are short, usually less than one hour, although cefadroxil has a longer half-­life in dogs.

151

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­First-­generation Cephalosporins 

Beta-­lactam Antibiotics: Cephalosporins

administered to monogastrates three times daily, although cefadroxil may be administered twice daily at the higher dose. Oral cephalosporins should not be used in herbivores.

Drug Interactions First generation are synergistic with aminoglycosides. Beta-­lactams, as an inhibitor of cell wall synthesis, significantly enhance the uptake of aminoglycosides.

Clinical Applications

Toxicities and Adverse Effects

Oral cephalosporins have similar applications to penicillinase-­resistant penicillins and aminobenzyl penicillins in monogastrates, and are widely used in small animal medicine. The first-­generation cephalosporins are thus potentially useful in a variety of nonspecific infections caused by staphylococci, streptococci, Enterobacterales in some cases, and some anaerobic bacteria. The treatment of superficial bacterial folliculitis in dogs is one useful application, although this may better be treated with topical antiseptics, as well as deep pyoderma. Cephalexin has been described as the drug of choice for K. pneumoniae urinary tract infections, although a fluoroquinolone is now a better choice. Apart from skin and ­urinary tract infections caused by susceptible

Cephalosporins are among the safest of antimicrobial drugs. Allergic reactions, including acute anaphylactic hypersensitivity, are rare. In humans, the majority of allergic reactions are not cross-­reactive with penicillin. A small proportion of human patients may develop eosinophilia, rash, and drug-­associated fever. Vomiting and diarrhea may occur in a small proportion of dogs and cats given oral cephalosporins.

Administration and Dosage Recommended dosages are shown in Table 8.2. Because of the margin of safety, a range of dosages can be used depending on the MIC of susceptible bacteria. Oral cephalosporins should be Table 8.2  Dosage of first-­generation cephalosporins. Species

Dog, cat

Horse

Drug

Route

Dose (mg/kg)

Interval (h)

Cephradrine

IV, IM

22

6–­8

Cephalothin

IV

20–­40

6–­8

Cefazolin

IV, IM

15–­30

8

Cefadroxil

PO

22

12

Cephalexin

PO

30

12

Cephadrine

PO

10–­25

6–­8

Cephapirin

IV, IM

15–20

8

Cefazolin

IV, IM

15–­20

8

Comments

IV only (painful IM)

Highly susceptible, e.g., S. aureus

Horse (foals only)

Cephalexin

IV, IM

10

8–­12

Cefadroxil

PO

20–­40

8

Calves (pre-­ruminant)

Cephadrine

PO

7

12

Cefadroxil

PO

25

12

Cattle, sheep

Cefazolin

IV, IM

15–20

12

Poor udder penetration

Cephapirin

IV, IM

10

8–­12

As cefazolin

IM, intramuscular; IV, intravenous; PO, by mouth (per os).

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152

organisms, other applications include the treatment of nonaccessible abscesses (e.g., GI and lung abscess) and deep wound infections caused by susceptible organisms in dogs and cats. Resistance in canine S. aureus and S. pseudintermedius has become an increasing problem globally; canine isolates of MRSA and MRSP are commonly multidrug resistant (Morris et al., 2017). Reports suggested that oral cephalexin treatment in dogs might enhance nasal carriage of MRSP (Fungwithaya et al., 2017). Clinical applications of parenteral first-­ generation cephalosporins have become fewer with the development of beta-­lactamase-­ stable oral cephalosporins, with the exception of use for surgical prophylaxis. These drugs have been, and continue to be, used extensively in prophylaxis of surgical wound infections in human patients and are used for this purpose in dogs and cats. Cefazolin has been suggested for administration (22  mg/kg IV) 30–60  minutes prior to surgical incision, repeated every four hours for skin flora and every two hours for E. coli. In dogs and cats, cefazolin might be used to establish high tissue levels rapidly before using an oral cephalosporin. In the absence of susceptibility testing, their use in treating infections caused by Gram-­negative Enterobacterales is not recommended since activity is unpredictable (as is the case also for aminobenzyl penicillins). In cattle, different first-­generation cephalosporins are in widespread use for treatment and prevention (dry cow therapy) of intramammary infections caused by the Gram-­positive cocci, as alternatives to pirlimycin, cloxacillin, and penicillin-­novobiocin combination. Administration is by the intramammary route.

of beta-­lactamases. They are moderately active against Gram-­positive bacteria. Cephamycins (cefotetan, cefoxitin) are products of Streptomyces rather than of Cephalosporium species and differ from cephalosporins in the presence of a methoxy group in the 7 position of the cephalosporin nucleus. Cephamycins are stable to beta-­lactamases, including those of Bacteroides fragilis, although this is now less common, but like other second-­generation drugs are not active against P. aeruginosa.

Antimicrobial Activity Cefoxitin is resistant to most bacterial beta-­ lactamases, although it penetrates Gram-­negative bacteria relatively poorly. Antimicrobial activity is slightly broader and greater than that of cefazolin and other first-­generation cephalosporins for Gram-­negative bacteria and includes Enterobacter spp. and Serratia spp. Activity against Gram-­ positive bacteria is slightly less. Cefoxitin was at one time stable to the beta-­lactamase of B. fragilis and had good activity against this and other Bacteroides, Porphyromonas, and Prevotella spp., although this stability can no longer be relied on. Pseudomonas aeruginosa, enterococci, MRSA, MRSP, and some Enterobacterales are resistant. Cefotetan has the greatest activity of the 7-­methoxy cephalosporins against Gram-­negative bacteria but P. aeruginosa is resistant. A proportion of Citrobacter, Enterobacter, and Serratia spp. are resistant. Activity against anaerobes is similar to cefoxitin but a proportion of B. fragilis are resistant. Cefmetazole has a spectrum of activity similar to cefoxitin but it is more active against Enterobacterales.

Resistance

­Second-­generation Cephalosporins: Cefaclor, Cefoxitin, Cefmetazole, Cefotetan, and Cefuroxime Second-­generation parenteral cephalosporins have a wide spectrum of antibacterial activity largely because of their stability to a broad range

Stable depression of inducible beta-­lactamases associated with hyperproduction of AmpC beta-­ lactamases in certain Gram-­negative pathogens is an important mechanism of resistance. Cefoxitin is a powerful beta-­lactamase inducer and can therefore antagonize the effects of other beta-­lactams. As described earlier, in recent

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­Second-­generation Cephalosporins: Cefaclor, Cefoxitin, Cefmetazole, Cefotetan, and Cefuroxim  153

Beta-­lactam Antibiotics: Cephalosporins

years there has been increasing spread of a family of cephamycinase (CMY2)-­encoding plasmids in animals, noted not only in hospital-­ acquired E. coli infections in companion animals, but also in Salmonella. Certain strains of MRSP may be falsely reported as susceptible to cefoxitin by laboratories because of its poor induction of the mecA gene (Weese et al., 2009). This is why an oxacillin disk is preferable to cefoxitin in testing for methicillin resistance.

Pharmacokinetic Properties Pharmacokinetic properties and toxicities are similar to those of first-­generation parenteral cephalosporins. With one exception, they are not absorbed following oral administration. Cefuromine axetil is an ester of cefuroxime which is hydrolyzed in the intestinal mucosa and liver to yield active drug, producing good bioavailability after oral administration. Excretion, which can be delayed by probenecid, is largely renal. Plasma elimination half-­ lives in cattle and horses are about one hour.

Toxicities and Adverse Effects Second-­generation cephalosporins cause pain on IM injection and may cause thrombophlebitis when administered IV. Cefoxitin may cause hypoprothrombinemia and a tendency to bleed in human patients.

Administration and Dosage Administration is usually IV because of pain associated with IM administration. Dosage in animals, which in some cases is empirical, is shown in Table 8.3.

Clinical Applications Clinical applications in animals are limited by the expense of these drugs. In human medicine cefoxitin was at one time, and to some extent still is, valued particularly for its broad

activity against anaerobes, especially B. fragilis, as well as against Enterobacterales. Indications are thus treatment of severe mixed infections with anaerobes (aspiration pneumonia, severe bite infections, gangrene, peritonitis, pleuritis) and prophylaxis in colonic surgery or ruptured intestine. Cefuroxime is available and effective for short-­lasting dry cow therapy and for treatment of clinical intramammary infections in lactating cows. Cefuroxime axetil is used by the oral route in human medicine for the treatment of otitis media and upper respiratory infections caused by susceptible bacteria. The widespread use of cephalosporins for this purpose may have been largely responsible for the extensive emergence of penicillin resistance in Streptococcus pneumoniae, an important human pathogen, in recent years.

T ­ hird-­generation Cephalosporins: Cefmenoxime, Cefotaxime, Cefovecin, Ceftizoxime, Ceftriaxone, Ceftiofur, Latamoxef, Cefetamet, Cefixime, Cefpodoxime proxetil, Cefoperazone, Cefsulodin, and Ceftazidime Third-­generation cephalosporins are distinguished by their high antibacterial activity and their broad resistance to beta-­lactamases; they have particularly good activity against most Enterobacterales. Exceptions include Enterobacter and Serratia. Streptococci are highly susceptible, staphyloccci moderately susceptible, and enterococci are resistant. Latamoxef (moxalactam) is an oxacephem with an oxygen atom replacing the sulfur at the C1 position of the cephalosporin nucleus. Its wide anti-­Enterobacterales activity is similar to that of others in the group but latamoxef is more active against B. fragilis, Citrobacter spp., and Enterobacter spp., and less active against S. aureus. Some P. aeruginosa are resistant. Importantly, these drugs have become widely used in animals due to long-­acting

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154

Table 8.3  Dosage of second-­, third-­, and fourth-­generation cephalosporins in animals. Species

Drug

Route

Dose (mg/kg)

Interval (h)

Dog, cat

Cefotaxime

IM (SC)

20–­40

8 (SC 12)

Cefoperazone

IV, IM

20–­25

6–8

Cefovecin

SC

8

10–­14 days

Cefoxitin

IV, IM, SC

15–­30

6–­8

Cefiofur

IM

2.2

24

Cefixime

PO

5

12–­24

Cefpodoxime proxetil

PO

5–10

24

Ceftizoxime

IV, IM

25–40

8–­12

Ceftriaxone

IV, IM

25

12–­24

Cefuroxime

IV

10–­15

8–12

Cefoperazone

IM

20

6–­8

Ceftazidime

IM

25–­50

8–­12

Cattle, sheep, goats

Ceftiofur

IM, SC

1.1–2.2

24

Cattle

Ceftiofur crystalline free acid

Posterior ear

6.6

5 days

Horses

Cefoperazone

IM

30

6–­8

Cefquinome 2.5

IM

1 (calves 2)

24

Cefquinome LA

SC

2.5

48

Ceftazidime

IM

20–­40

12–24

Cefotaxime

IV

20–­30

6–­8

Cefoxitin

IV, IM

20

8

Cefquinome

IM

1

24 (foals 12 IV or IM)

Ceftiofur

IV, IM

2.2–­5

12–­24

Ceftiofur crystalline free acida IM (2 sites)

6.6

4 days

Ceftriaxone

IV, IM

25

12 (not adults)

Cefoperazone

IM

30

6–­8

Ceftazidimeb

IM

25–­50

8–­12

b

Horse (foals only)

Cefpodoxime proxetil

PO

10

6–­12

Swine

Cefquinome 2.5

IM

2

24

Ceftiofur

IM

3–­5

24

Ceftiofur crystalline free acid

IM

5

5 days

a

 Neonatal foals may require 13.2 mg/kg q48 hours.  Caution should be used as may cause colitis. IM, intramuscular; IV, intravenous; PO, by mouth (per os); SC, subcutaneous.

b

formulations without due consideration of their importance to human medicine and the risks of antimicrobial resistance. Voluntary restriction on third-­ and fourth-­generation cephalosporins in Danish pig and cattle production has abolished the use of ceftiofur in

Denmark, showing that other agents can be used effectively for managing disease. Following antimicrobial stewardship principles for veterinary medicine, these drugs should be reserved for pathogens where resistance to lower tier drugs has been documented.

155

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­Third-­generation Cephalosporins 

Beta-­lactam Antibiotics: Cephalosporins

Antimicrobial susceptibility categories given below are guidelines only; bacterial susceptibility in veterinary medicine may vary by region and species and over time (Chapter 2). ●●

●●

●●

Predictable susceptibility (MIC 8 μg/ml). Acinetobacter spp., Bordetella spp., some Enterobacter spp. and Serratia spp., some P. aeruginosa, enterococci, and MRSA and MRSP.

Other third-­generation cephalosporins only registered for humans, such as cefsulodin, ceftaxidime and cefoperazone, are distinguished by the high activity against P. aeruginosa.

Resistance Transferable resistance to third-­generation cephalosporins as a result of AmpC hyperproduction, extended spectrum beta-­lactamases, and to a lesser extent beta-­lactamase group 3 metallo-­beta-­lactamases has been discussed earlier. This is an important threat to the continued use of these cephalosporins in animals, particularly in food animals because of public health considerations. Ceftazidime-­ specific PER-­type extended-­spectrum beta-­ lactamases have been described (see Table 9.1). Multidrug resistance plasmids carrying the blaCMY2 encoding resistance to ceftiofur and ceftriaxone have been identified in Salmonella

enterica serovars, among others, and are often found in strains with concomitant resistance to ampicillin, chloramphenicol, streptomycin, sulfamethoxazole, and tetracycline. The cmy-­2 gene appears to have been mobilized into different plasmid backbones that have spread through E. coli and Salmonella through conjugation (Carattoli et  al.,  2002). More recently, blaCTX-­M variants have been observed in bacterial isolates recovered from a wide range of animal species, such as poultry, beef cattle, swine, horses, and dogs (Cormier et al., 2019), discussed further in Chapter 9. In human medicine, the breakpoint for resistance used to be 64 μg/ml, so that there was confusion between resistance reported for animal isolates as it relates to resistance in human isolates. For example, in the Canadian Integrated Program for Antimicrobial Resistance Surveillance report for 2003  data (CIPARS, 2005), ceftiofur resistance (breakpoint >8 μg/ml) was particularly high in Salmonella isolated from chicken in Québec but not as high for human Salmonella isolates tested for ceftriaxone (breakpoint >64 μg/ml); when the same 8 μg/ml breakpoint was applied to both drugs, percentage resistance was the same. As discussed further under beta-­ lactamases in Chapter  9, the human medical CLSI breakpoints for ceftriaxone for Enterobacterales were revised in 2010 to 4 μg/ml for resistance. The CLSI breakpoints for cefpodoxime for treating S. aureus and S. pseudintermedius soft tissue infections in dogs are: susceptible 8 μg/ml.

Pharmacokinetic Properties The pharmacokinetic properties of oral third-­ generation cephalosporins are typical of those of beta-­lactams generally. Cefpodoxime has a relatively long plasma elimination half-­life in dogs of about 5.6 hours, so that plasma concentrations exceed 1 μg/ml for about 24 hours after a dose of 10 mg/kg.

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Parenteral third-­generation cephalosporins are rapidly and well absorbed after IM or SC administration, giving peak serum concentrations in 0.5–1 hour. While data are often lacking, the plasma elimination half-­life is about one hour following IV injection. In cattle, the plasma elimination half-­life of ceftiofur is about 2.5 hours. Distribution into tissues in extracellular fluid is widespread but passage across membranes or physiological barriers is poor. Meningeal inflammation significantly enhances otherwise poor CSF penetration so that, because of exceptional antibacterial ­activity, these cephalosporins are drugs of choice for bacterial meningitis caused by Enterobacterales. Cefotaxime is metabolized in the body to the less active desacetyl-­ cefotaxime. Excretion is largely through the urinary tract, with cefotaxime being excreted through tubular mechanisms and the others through glomerular filtration. Probenecid administration delays tubular excretion. Biliary elimination also occurs, notably for ceftriaxone and latamoxef. These drugs should therefore be avoided in species with expanded large intestines. Ceftriaxone has a long elimination half-­life, giving this drug the advantage of twice-­daily dosing. Ceftiofur is very rapidly (almost instantly) metabolized in the body to an active metabolite called desfuroylceftiofur. Desfuroylceftiofur is very highly protein bound (99%) due to a sulfhydryl group in its chemical structure. The efficacy of ceftiofur is attributed to binding to acute-­phase proteins (alpha-­1-­antitrypsin). These act as a reservoir for active drug and carry the bound drug to the site of inflammation, where a new equilibrium is established between free and bound drug. Desfuroylceftiofur does not have the same activity as does ceftiofur against Proteus and Staphylococcus, such that MIC results from laboratories can be misleading since they are only based on ceftiofur disks (e.g., S. aureus is 4–8 times less susceptible to desfuroylceftiofur than ceftiofur). For ceftiofur, most of the antibacterial activity is attributed to desfuroylceftiofur. Ceftiofur and

desfuroylceftiofur distribute quite differently compared to other drugs because of the very high degree of protein binding. Reversible covalent bonding with plasma and tissue proteins produces lower than expected free concentrations following clinical doses. However, tissue chamber studies have shown that ceftiofur and desfuroylceftiofur concentrations are higher in infected compared to normal tissue. There is no interaction with other protein-­ bound drugs (e.g., phenylbutazone) since ceftiofur and phenylbutazone do not compete for the same protein-­binding site. The ceftiofur hydrochloride formulation is more stable than ceftiofur sodium. Drug distribution to tissues and residue concentrations are similar for both. A crystalline free acid formulation of ceftiofur has the advantage of delayed absorption, so that for highly susceptible bacteria, dosing frequency can be reduced to 96–120 hours apart, depending on the species and route of administration. For example, the crystalline free acid formulation administered as a single subcutaneous injection into the ear of cattle at 6.6 mg/kg is slowly absorbed and gives plasma concentrations exceeding the MIC of common respiratory tract bacterial pathogens for about six days. Similarly, this formulation administered intramuscularly in swine also has a long half-­life, with plasma concentrations after IM injection of 5 mg/kg exceeding the MIC of common respiratory tract pathogens for about five days. Maintaining concentrations above the MIC for as long as possible is important for ceftiofur because it is not expected to have a postantibiotic effect (PAE) against Gram-­negative bacteria. Cefoperazone has largely hepatic elimination, which therefore tends to be relatively often associated with gastrointestinal disturbance in humans. Thus, this drug is contraindicated in horses and other herbivores with an expanded large bowel. Cefoperazone, but not ceftazidime, elimination in urine is reduced by probenecid. There has been little study of the pharmacokinetic properties of cefoperazone, ceftazidime or cefsulodin in animals.

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­Third-­generation Cephalosporins 

Beta-­lactam Antibiotics: Cephalosporins

Drug Interactions Third-­generation cephalosporins are synergistic with aminoglycosides although the combination is rarely used in veterinary medicine.

Toxicities and Adverse Effects Toxicities and adverse effects are similar to those described for first-­and second-­generation cephalosporins, but the nephrotoxic potential is low. Due to the broad antibacterial activity of these cephalosporins, gastrointestinal disturbances and superinfection by resistant microorganisms, including yeasts, may be anticipated. In human medicine, there is a strong association between parenteral use of third-­generation cephalosporins and C. difficile diarrhea. Anecdotal reports suggest that there is a link between ceftiofur use in neonatal piglets and the development of C. difficile infection. Colitis has been reported following the use of cephalosporins in horses, with a recent study reporting that 79% of horses which developed C. difficile colitis had been administered cephalosporins (Nomura et  al.,  2020). Gastrointestinal disturbances have especially been noted in horses administered ceftriaxone IV, probably because of its biliary excretion, so this drug should be used cautiously, if at all, in adult horses. Cutaneous drug reactions to ceftiofur, characterized by hair loss and pruritus, have been described in a cow. Cefpodoxime administered orally to dogs has not been associated with serious adverse effects, but vomiting, diarrhea, and decreased appetite may occur.

Administration and Dosage Recommended dosages, which in some cases are empirical, are shown in Table 8.3. To some extent, dosage can be tailored to the susceptibility of the organism, with the aim of maintaining drug concentrations > MIC throughout the majority of the dosing interval. For example, dosage of ceftiofur sodium or hydrochloride for

highly susceptible organisms associated with lower respiratory disease is usually 1.1–2.2 mg/kg q24 hours, but for E. coli infections caused by susceptible organisms the dose might be as high as 2.2–4.4 mg/kg q12 hours and for Salmonella spp. 5 mg/kg q12 hours may be required. Dosage of the crystalline free acid formulation in food animals and horses, and of cefovecin in  companion animals, is less frequent. Ceftriaxone has the advantage that dosage is twice daily whereas dosage of other parenteral third-­ generation cephalosporins (other than ceftiofur) is usually q8 hours. Cefixime’s long elimination half-­life allows once-­daily administration in people. Dosage recommended for cefetamet in children is 20  mg/kg q12 hours. Cefpodoxime has been approved in the United States for dosage to dogs at 5–10 mg/kg administered once daily, with twice-­daily administration in refractory infections. The upper dose is preferable for susceptible S. aureus or S. pseudintermedius infections. A suggested dosage of cefpodoxime in foals was 10 mg/kg q6–12 hours (Carrillo et al., 2005). The antipseudomonal third-­generation drugs (cefoperazone, cefsulodin, ceftazidime) are largely reserved in human medicine for P. aeruginosa and other Gram-­negative septicemias in neutropenic human patients, in which efficacy is considerably enhanced by combination with an aminoglycoside. Cephalosporins have slow bactericidal activity compared to aminoglycosides. Subcutaneous injection of 30 mg/kg q4 hours or constant IV infusion of 4.1 mg/kg/hr were estimated to produce serum concentrations of cefoperazone exceeding the MIC of canine clinical isolates of P. aeruginosa (Moore et al., 2000).

Clinical Applications Because of expense, the availability of cheaper alternatives, and the potential to select for resistant bacteria, third-­generation cephalosporins should be reserved for serious, ­probably life-­threatening, infections caused by Gram-­ negative bacteria, especially Enterobacterales.

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They are listed as highest priority, critically important antimicrbials by the World Health Organization (WHO, 2024). Despite recommendations to reserve these drugs for serious infections, and only for infections where susceptibility testing indicates that alternatives are not available, there is an increasing and unfortunate tendency to use these drugs as first choice in animals. As noted earlier, because of resistance concerns, in 2012 there was a prohibition in the US by the FDA on the extra-­label use of cephalosporins  in food animals (Food and Drug Administration, 2012). They are recommended, in combination with an aminoglycoside, in severe infections caused by multiple resistant bacteria in compromised hosts, such as neutropenic hosts. These drugs have potential application in ­septicemia, osteomyelitis and septic arthritis,  some lower respiratory tract infections, intraabdominal infections caused by Enterobacterales, and some soft tissue infections where cheaper alternative drugs are not available. The poor activity of some of these cephalosporins against Gram-­negative anaerobes is a drawback; ceftiofur, however, has good activity against anaerobes in horses with the exception of members of the B. fragilis group. Activity against anaerobes in cattle may be reduced (Samitz et al., 1996), perhaps as a result of the common usage of this drug in this species. Although not well documented in many animal species, third-­generation cephalosporins have a tendency to select for C. difficile infections. Cefpodoxime has been approved for use in dogs for skin infections (wounds and abscesses) caused by susceptible organisms. It has the advantage of once-­daily administration for this purpose but should be reserved in line with antimicrobial stewardship principles. Cattle, Sheep, and Goats

Ceftiofur sodium and hydrochloride is used extensively for the treatment of, and prophylaxis for, acute, undifferentiated bovine ­pneumonia with the advantage of a low recommended dose (1.1–2.2  mg/kg, q24 hours) and

zero drug withdrawal time in milk. Treatment is for 3–5  days and has proved as effective as treatment with sulbactam-­ampicillin or potentiated sulfonamides for this purpose. In one study of treatment of relapse of undifferentiated fever/bovine respiratory ­disease in feedlot cattle, ceftiofur was less effective than enrofloxacin (Abutarbush et al., 2012). Dosage IM of 3  mg/kg q12 hours was inadequate for the parenteral treatment of intramammary infections caused by E. coli (Erskine et al., 1995). Treatment of severe coliform mastitis with ceftiofur was shown to reduce death or culling (Erskine et  al.,  2002), likely due to improved penetration into the udder with severe inflammation (otherwise penetration is negligible) and treatment of septicemia. Ceftiofur sodium and hydrochloride is also used in the treatment of acute bovine interdigital necrobacillosis and the hydrochloride form is approved in the US and elsewhere for the treatment of postparturient metritis. Overall, procaine penicillin is a better choice than ceftiofur for treatment of these latter infections, because of its equivalent or better antibacterial activity and narrower spectrum, with less likelihood of producing resistance in “by-­stander” bacteria. Ceftiofur has also been administered experimentally at the extra-­label dose of 5 mg/kg q24 hours in the treatment of Salmonella infection of calves (Fecteau et  al.,  2003). Multiple treatments with ceftiofur sodium have been used to eliminate Leptospira from the kidneys of cattle although tetracycline and tilmicosin were equally effective and are of lesser importance to human medicine than third-­generation cephalosporins. The crystalline free acid formulation of ceftiofur administered SC in the ear of cattle at 6.6 mg/kg gives plasma concentrations exceeding the MIC of respiratory tract pathogens (H. somni, M. haemolytica, P. multocida) for over five days. It has application for the treatment of respiratory disease caused by these highly susceptible bacteria, as well as for ­treatment of interdigital necrobacillosis. Administration by routes other than SC in the

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­Third-­generation Cephalosporins 

Beta-­lactam Antibiotics: Cephalosporins

ear can lead to violative residues, and should be avoided. Inadvertent intraarterial injection (usually into an auricular artery) has been associated with sudden death. The advantage of this formulation is that most animals with susceptible infections will respond within 3–5 days. Horses

Ceftiofur sodium and the crystalline free acid (CCFA) are suitable for use in horses in treating bacterial infections caused by susceptible bacteria. The crystalline free acid is indicated specifically for treatment of lower respiratory tract infections caused by S. zooepidemicus (MIC 2 μg/ml over 90% of the time (Pusterla et al., 2016), exceeding the MIC of susceptible Enterobacterales. Although doses of ceftiofur sodium exceeding 5 mg/kg have been administered to foals, evidence is lacking regarding the need in terms of

pharmacokinetics and efficacy. Extra-­label uses of ceftiofur sodium include treatment for pleuritis and peritonitis and, via an intrauterine route, treatment for endometritis caused by susceptible organisms. Ceftiofur has been investigated for use in regional limb perfusions but concentrations only exceed MIC for common pathogens for 12 hours, limiting the usefulness of this drug when compared to aminoglycosides (Cox et  al.,  2017). Following intraarticular injection, ceftiofur maintains concentrations above MIC for at least 24 hours (Mills et al., 2004). In addition, this route may avoid the metabolism of ceftiofur to desfuroylceftiofur and thereby be more effective in the treatment of joint sepsis caused by S. aureus. As noted earlier, colitis has been reported following the use of cephalosporins in horses, with a recent study reporting that 79% of horses which developed C. difficile colitis had been administered cephalosporins (Nomura et  al.,  2020). Seizures, collapse, and sudden death, typically within 30 minutes of injection, have been associated with intramuscular administration of crystalline free acid ceftiofur formulations in a small number of horses. Cefotaxime has been used effectively in the treatment of neonatal septicemia and meningitis caused by Acinetobacter spp., Enterobacter spp., and P. aeruginosa. Ceftriaxone may be particularly suitable for the treatment of meningitis in foals because it crosses the healthy blood–cerebrospinal fluid barrier. A dosage suggested for Gram-­negative bacterial meningitis is 25 mg/kg q12 hours. This drug should, however, be used with caution in adult horses because of its hepatic excretion and potential to cause severe colitis. Swine

Ceftiofur sodium is available for use in swine in the treatment of respiratory or systemic infections caused by susceptible bacteria such as P. multocida, beta-­lactamase-­producing Actinobacillus spp., Haemophilus parasuis, and Streptococcus suis. The crystalline free acid formulation of ceftiofur administered at 5 mg/kg

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gives plasma concentrations exceeding the MIC of respiratory tract pathogens (A. pleuropneumoniae, H. parasuis, P. multocida, S. suis) for over five days, so that it has the advantage of single-­dose treatment of infections caused by such susceptible bacterial infections. Ceftiofur has also been used in the control of Salmonella choleraesuis infections. It also has application for IM administration in the treatment of neonatal colibacillosis. Anecdotally, the practice of routine injection of neonatal pigs with ceftiofur may predispose them to infection with C. difficile, which has emerged as a significant problem in some swine farms in recent years. Narrower spectrum drugs are often effective and should be preferred for the clinical applications outlined above. Dogs and Cats

Cefovecin is a long-­acting SC formulation for dogs and cats, with the remarkable property that it produces serum concentrations >0.25 μg/ ml (MIC90 of S. pseudintermedius) for most of 14 days. It is thus used for the single treatment of infections caused by highly susceptible bacteria, including those commonly involved in skin infections, bite wounds, and abscesses (S. pseudintermedius, S. canis, P. multocida). Because of urinary excretion, it is also effective against enteric bacteria causing urinary tract infections. Treatment can be repeated at 14-­day intervals on 2–4 occasions in cats and dogs, respectively, depending on susceptibility and clinical considerations. Cefovecin is in widespread use as a first-­line  antimicrobial drug in cats (Buckland et  al.,  2016; Hardefeldt et  al.,  2020; Hur et al., 2020). The advantage claimed, notably in cats, is that administration by this route ensures compliance in comparison to owners trying to administer amoxicillin or amoxicillin-­clavulanic acid pills twice daily by mouth, thus enhancing the likelihood of cure. Cefovecin is purportedly used in an extra-­label manner for treating wildlife for compliance and to avoid difficulties with handling. Cefovecin has a ­similar spectrum of clinical efficacy to amoxicillin-­clavulanic acid (Stegemann et al., 2007). Common usage in cats

includes cat fight injuries, abscesses and to a lesser extent dermatitis, urinary tract disorders, and prophylaxis (Hardefeldt et  al.,  2020). Cefovecin is less commonly used in dogs, with indications being surgical prophylaxis, dermatitis and to a lesser extent oral cavity disorders and traumatic injuries (Hardefeldt et al., 2020). Cefovecin is used in cats and dogs in many circumstances in which antimicrobials are not indicated or in which an antimicrobial of lower importance in human health is recommended (Hardefeldt et al., 2020). Owner/animal compliance should not be the motivating decision to use cefovecin. Serious adverse clinical effects have not been reported, with any hypersensitivity effects lasting 3–5 days. Many companion animal practices use amoxicillin-­clavulanic acid as a “first-­line” antimicrobial (Murphy et  al.,  2012; Buckland et  al.,  2016; Hur et  al.,  2020). Cefovecin has a similar spectrum of activity although dosage might give slightly lower serum concentrations against Enterobacterales than amoxicillin-­ clavulanic acid. However, antimicrobials should be chosen that have the narrowest spectrum of activity. For example, most cat bite infections can be successfully treated with amoxicillin and staphylococcal skin infections with cephalexin, so that these antimicrobials should be preferred over a potentiated aminopenicillin or third-­ generation cephalosporin. In addition, many guidelines for these infections now recommend 3–5-­day courses of narrow-­spectrum antimicrobials, thereby reducing concerns around compliance. The rapid rise and dissemination of broad-­spectrum beta-­lactamase resistance in Enterobacterales of companion animals support the enhanced stewardship of these drugs. Poultry

Ceftiofur is administered SC to day-­old chicken and turkey poults for the control of E. coli infections and navel infections, and has been injected in ovo for the same purpose. As described earlier, as a result of extra-­label use for egg injection, CMY-­2 beta-­lactamase-­ resistant strains have developed in E. coli and

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­Third-­generation Cephalosporins 

Beta-­lactam Antibiotics: Cephalosporins

Salmonella in broilers, with spread of resistant Salmonella to humans to cause serious infections. This has been best documented in Canada (Dutil et  al.,  2010) but is also recognized in the US and other countries. E. coli carrying the blaCTX-­M-­1 is common in poultry in France (Casella et al., 2017) and in Salmonella in the US and elsewhere (Denagamage et  al., 2019). There is evidence that resistant E. coli are also reaching humans and causing disease or, if not themselves disease causing, that they may be a transmission source of resistance genes (Manges, 2015). The use of third-­generation cephalosporin drugs in poultry has important public health considerations that suggest that they should not be used. Approval has been rescinded in many regions.

F ­ ourth-­generation Cephalosporins: Cefepime, Cefpirome, Cefquinome The fourth-­generation cephalosporins have high activity against Enterobacterales, moderate activity against P. aeruginosa, and enhanced activity against staphylococci. They are stable to hydrolysis by many plasmid or chromosomally mediated beta-­lactamases and are poor inducers of group 1 beta-­lactamases.

Antimicrobial Activity Cefepime and cefquinome are enhanced-­ potency, extended-­spectrum cephalosporins, the zwitterionic nature of which gives them rapid ability to penetrate through the porins of Gram-­negative bacteria to the cell membrane. Cefepime, cefpirome, and cefquinome have higher affinity for essential PBPs and greater resistance to hydrolysis by beta-­ lactamases than other cephalosporins. In particular, they are resistant to, and poor inducers of, group  1 beta-­lactamases. No reports of activity against specific animal pathogens are available for cefapime and cefpirome.

Antimicrobial susceptibility categories given below are guidelines only; bacterial susceptibility in veterinary medicine may vary by region and species and over time (Chapter 2). ●●

●●

Predictable susceptibility (MIC 32 μg/ml): Enterococcus spp., L. monocytogenes, Bacteroides spp., C. difficile, MRSA and MRSP.

Pharmacokinetic Properties Pharmacokinetic properties of these parenterally administered cephalosporins are typical of those of other parenteral cephalosporins generally. Most of these drugs are excreted through the urine.

Drug Interactions Combination of cefepime with aztreonam is synergistic against P. aeruginosa with derepressed cephalosporinases, since aztreonam protects cefepime against these enzymes in the extracellular environment.

Toxicities and Adverse Effects Toxicities and adverse effects in people are those of cephalosporins generally, with the major effect being gastrointestinal disturbance. Treatment was withdrawn in about 5% of patients treated  with cefpirome and 1–3% of patients treated with cefepime because of adverse effects. Gastrointestinal effects must be anticipated if these drugs are used in animals and have been observed in horses administered cefepime by the oral or IM route (Guglick et al., 1998).

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Administration and Dosage These drugs are administered IV or IM twice daily to human patients; dosage can to some extent be tailored to the nature and severity of the infection. In horses, a dosage recommendation of cefepime was 2.2  mg/kg q8 hours (Guglick et al., 1998). This is a very low dosage based on extrapolation from the empirical dose of 50 mg/kg q8 hours in children. By contrast, an IV dose of cefepime estimated for treatment of susceptible bacteria in neonatal foals was 11 mg/kg q8 and 40 mg/kg q6 hours for dogs (Gardner and Papich,  2001). Recommended dosage of cefquinome in adult horses with S. zooepidemicus respiratory disease is 1 mg/kg once daily for 5–10  days and in foals with E. coli septicemia 1 mg/kg q12 hours. Although a dose of cefquinome of 0.5 mg/kg q24 hours has been suggested for the treatment of S. equi subsp. equi (Lee et al., 2020), procaine penicillin is a better choice of antimicrobial because of its narrower spectrum and lower priority in human medicine. In Europe, approved dosage of cefquinome for respiratory disease in cattle caused by susceptible bacteria, foot rot or acute E. coli intramammary infections is 1  mg/kg q24 hours, in calves with E. coli septicemia is 2 mg/ kg q24 hours, and for swine is 2  mg/kg for MMA syndrome and respiratory disease. A long-­acting (q48 hours) form is available for cattle and swine.

Clinical Applications Fourth-­generation cephalosporins are used in human medicine in the treatment of nosocomial or community-­acquired lower respiratory disease, bacterial meningitis, urinary tract infections, and uncomplicated skin or skin-­related infections. These drugs are valuable extended-­ spectrum cephalosporins for the treatment of serious infections in people. Cefquinome is used in Europe and Japan in treatment of bovine respiratory disease and, by intramammary or IM administration, in the treatment of coliform and

other bacterial intramammary infections. In general, its efficacy in field studies of treatment of infections in cattle and swine has been similar to, and slightly superior to, that of ceftiofur sodium (Lang et  al.,  2003). In swine, cefquinome has been associated with significant increase in resistance gene expression compared to ceftiofur (Rutjens et al., 2023). It is approved in Europe for the treatment of equine respiratory disease caused by S. zooepidemicus or foal septicemia caused by E. coli. However, third-­ and fourth-­generation cephalosporins should be reserved for use where susceptibility testing indicates that alternatives are not available.

F ­ ifth-­generation Cephalosporins: Ceftobiprole, Ceftraroline Antimicrobial Activity Ceftraroline is a parenteral oxyimino cephalosporin with bactericidal activity against MDR Gram-­positive bacteria including MRSA and vancomycin-­resistant Staphylococcus aureus and has been used in the treatment of community-­ acquired pneumonia and Gram-­positive osteomyelitis in people. It also exhibits broad-­spectrum activity against many of the important community-­ acquired Gram-­positive and Gram-­negative pathogens. It does not possess activity against extensively resistant Gram-­negative bacteria and has limited activity against most nonfermentative Gram-­negative bacilli (e.g., Acinetobacter spp., Pseudomonas aeruginosa) and many anaerobic species. Methicillin-­resistant S. pseudintermedius have been shown to have relatively high rates of resistance (33%), using human MRSA breakpoints (Scherer et al., 2018). Ceftobiprole has potent Gram-­positive and Gram-­negative activity and is approved for the treatment of community-­acquired and hospital-­ acquired pneumonia, but not ventilator-­associated pneumonia, in human adults. Ceftobiprole also exhibits activity against MRSA and has similar activity to cefepime when tested against Enterobacterales and Pseudomonas aeruginosa.

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­Fifth-­generation Cephalosporins: Ceftobiprole, Ceftrarolin 

Beta-­lactam Antibiotics: Cephalosporins

Pharmacokinetic Properties Pharmacokinetic properties of these parenterally administered cephalosporins are typical of parenteral cephalosporins generally. Most of these drugs are excreted through the urine.

Drug Interactions In vitro studies have shown that ceftobiprole can be synergistic with daptomycin against MSSA, MRSA, and methicillin-­resistant Staphylococcus epidermidis strains and with amikacin and levofloxacin against Pseudomonas aeruginosa.

Toxicities and Adverse Effects Toxicities and adverse effects in people are common, with 45% and 70% of people experiencing a side-­effect although only 3% and 6% resulted

in discontinuation of ceftraroline and ceftobiprole respectively. The major adverse effects were nausea, headache, and gastrointestinal effects. There are no reports of use in animals.

Administration and Dosage Ceftraroline is indicated for the treatment of community-­acquired pneumonia, complicated skin and soft tissue infections, and Gram-­ positive osteomyelitis in people. The dose in people is 600  mg q8-­12 hours by one-­hour intravenous infusion. Ceftobiprole is indicated for community-­ and hospital-­acquired pneumonia. The dose is 500mg q8 hours by two-­ hour intravenous infusion. There are no reports of either drug being used in animals. Fifth generation cephalosporins are listed by the World Health Organization (WHO, 2024) as authorized for human use only.

­References and Bibliography Abutarbush SM, et al. 2012. Comparison of enrofloxacin and ceftiofur sodium for the treatment of relapse of undifferentiated fever/ bovine respiratory disease in feedlot cattle. Can Vet J 53:57. Albarellos GA, et al. 2010. Pharmacokinetics of cefoxitin after intravenous and intramuscular administration in cats. J Vet Pharm Ther 33:619. Anderson REV, et al. 2023. Characterization of Escherichia coli and other Enterobacterales resistant to extended-­spectrum cephalosporins isolated from dairy manure in Ontario, Canada. Appl Environ Microbiol 89:e01 86922. ANSES-­ANMV, French Agency for Food, Environmental and Occupational Health and Safety (ANSES) and French Agency for Veterinary Medicinal Products (ANMV). 2018. Sales survey of veterinary medicinal products containing antimicrobials in France in 2017. www.anses.fr/en/system/files/ANMV-­Ra­Antibiotiques2017EN.pdf

Barradell LB, Bryson HM. 1994. Cefepime. Drugs 47:471. Buckland EL, et al. 2016. Characterisation of antimicrobial usage in cats and dogs attending UK primary care companion animal veterinary practices. Vet Rec 179:489. Bush K. 2018. Past and present perspective on beta-­lactamases. Antimicrob Agents Chemother 62:e01076–18. Bush K, Bradford PA. 2020. Epidemiology of β-­lactamase-­producing pathogens. Clin Microbiol Rev 33:e00047–19. Carattoli A, et al. 2002. Characterization of plasmids carrying CMY-­2 from expanded cephalosporin-­resistant Salmonella strains isolated in the United States between 1996 and 1998. Antimicrob Agents Chemother 46:1269. Carrillo NA, et al. 2005. Disposition of orally administered cefpodoxime proxetil in foals and adult horses and minimum inhibitory concentration of the drug against common bacterial pathogens of horses. Am J Vet Res 66:30.

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Casella T, et al. 2019. High prevalence of ESBLs in retail chicken meat despite reduced of antimicrobials in chicken production, France. Int J Food Microbiol 257:271. Chen Y, et al. 2019. Increasing prevalence of ESBL-­producing multidrug resistant Escherichia coli from diseased pets in Beijing, China from 2012 to 2017. Front Microbiol 10:2852. Chong Y, et al. 2018. Current epidemiology, genetic evolution and clinical impact of extended-­spectrum β-­lactamase-­producing Escherichia coli and Klebsiella pneumoniae. Infect Genet Evol 61:185. Collard WT, et al. 2011. Pharmacokinetics of ceftiofur crystalline-­free sterile suspension in the equine. J Vet Pharm Ther 34:476. Cormier A, et al. 2019. Diversity of CTX-­M-­ positive Escherichia coli recovered from animals in Canada. Vet Microbiol 231:71. Cox KS, et al. 2017. Plasma, subcutaneous tissue and bone concentration of ceftiofur sodium after regional limb perfusion in horses. Equine Vet J 49:341. De Jong A, et al. 2020. Antimicrobial susceptibility monitoring of canine and feline skin and ear pathogens isolated from European veterinary clinics results of the ComPath Surveillance program. Vet Dermatol 31:431–e114. Denagamage TM. 2019. Detection of CTX-­M-­1 extended-­spectrum beta-­lactamase among ceftiofur-­resistant Salmonella enterica clinical isolates of poultry. J Vet Diagn Invest 31:681. Duplessis C, Crum-­Cianflone NF. 2011. Ceftraroline: a new cephalosporin with activity against methicillin-­resistant Staphylococcus aureus (MRSA). Clin Med Rev Ther 3:a2466. Dutil L, et al. 2010. Ceftiofur resistance in Salmonella enterica serovar Heidelberg from chicken meat and humans. Emerg Infect Dis 16:48. Elias L, et al. 2020. The occurrence and characterization of extended-­spectrum-­beta-­ lactamase-­producing Escherichia coli isolated

from clinical diagnostic specimens of equine origin. Animals 10:28. Elnekave E, et al. 2019. Circulation of plasmids harboring resistance genes to quinolones and/or extended-­spectrum cephalosporins in multiple Salmonella enterica serotypes from swine in the United States. Antimicrob Agents Chemother 63:e02602–18. Erskine RJ, et al. 1995. Ceftiofur distribution in serum and milk from clinically normal cows and cows with experimental Escherichia coli-­induced mastitis. Am J Vet Res 56:481. Erskine RJ, et al. 2002. Efficacy of systemic ceftiofur as a therapy for severe clinical mastitis in dairy cattle. J Dairy Sci 85:2571. Fecteau ME, et al. 2003. Efficacy of ceftiofur for treatment of experimental salmonellosis in neonatal calves. Am J Vet Res 64:918. Food and Drug Administration. 2012. New animal drugs: Cephalosporin drugs; extralabel animal drug use; order of prohibition. Federal Register 77:735. Fungwithaya P, et al. 2017. Nasal carriage of methicillin-­resistant Staphylococcus pseudintermedius in dogs treated with cephalexin monohydrate. Can Vet J 58:73. Gardner SY, Papich MG. 2001. Comparison of cefepime pharmacokinetics in neonatal foals and adult dogs. J Vet Pharm Ther 24:187. Gelalcha BD, Dego OK. 2022. Extended-­ spectrum beta-­lactamase-­producing Enterobacteriaceae in the USA dairy cattle farms and implications for public health. Antibiotics 11:1313. Giacobbe DR, et al. 2019. Ceftobiprole: drug evaluation and place in therapy. Expert Rev Anti-­infect Ther 17:9. Goudah A, et al. 2009. Evaluation of single-­dose pharmacokinetics of cefepime in healthy bull camels (Camelus dromedarius). J Vet Pharm Ther 32:393. Guglick MA, et al. 1998. Pharmacokinetics of cefepime and comparison with those of ceftiofur in horses. Am J Vet Res 59:458. Hall TL, et al. 2010. Pharmacokinetics of ceftiofur sodium and ceftiofur crystalline free acid in neonatal foals. J Vet Pharm Ther 34:403.

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  ­References and Bibliograph

Beta-­lactam Antibiotics: Cephalosporins

Hardefeldt L, et al. 2020. Use of cefovecin in dogs and cats attending first-­opinion veterinary practices in Australia. Vet Rec 187:e95. Hayer SS, et al. 2020. Genetic determinants of resistance to extended-­spectrum cephalosporins and fluoroquinolone in Escherichia coli isolated from diseased pigs in the United States. mSphere 5:e00990–20. Hur BA, et al. 2020. Describing the antimicrobial usage patterns of companion animal veterinary practices: free text analysis of more than 4.4 million consultation records. PLoS One 15:e0230049. Isgren CM, et al. 2019. Emergence of carriage of CTX-­M-­15 in fecal Escherichia coli in horses at an equine hospital in the UK; increasing prevalence over a decade (2008–2017). BMC Vet Res 15:268. Lang I, et al. 2003. A field study of cefquinome for the treatment of pigs with respiratory disease. Rev Med Vet 153:575. Lee D-­H, et al. 2020. Pharmacokinetic and pharmacodynamics integration for optimal dosage of cequinome against Streptococcus equi subsp. equi in foals. Vet Res 51:13i. Lister PD, et al. 1998. Cefepime-­aztreonam: a unique double beta-­lactam combination for Pseudomonas aeruginosa. Antimicrob Agents Chemother 42:1610. Manges AR. 2016. Escherichia coli and urinary tract infections: the role of poultry meat. Clin Microbiol Rev 22:122. Marshall WF, Blair JE. 1999. The cephalosporins. Mayo Clin Proc 74:187. Meyer S, et al. 2008. Pharmacokinetics of intravenous ceftiofur sodium and concentration in body fluids of foals. J Vet Pharm Ther 32:309. Mills ML, et al. 2004. Determination of synovial fluid and serum concentrations, and morphologic effects of intraarticular ceftiofur sodium in horses. Vet Surg 29:398. Moffat J, et al. 2020. Resistance to extended-­ spectrum cephalosporins in Escherichia coli and other Enterobacterales from Canadian turkeys. PLoS One 15:e0236442. .

Moore KW, et al. 2000. Pharmacokinetics of ceftazidime in dogs following subcutaneous administration and continuous infection and the association with in vitro susceptibility of Pseudomonas aeruginosa. Am J Vet Res 61:1204. Morris DO, et al. 2017. Recommendations for approaches to meticillin-­resistant staphylococcal infections of small animals: diagnosis, therapeutic considerations and preventative measures: Clinical Consensus Guidelines of the World Association for Veterinary Dermatology. Vet Dermatol 28:304. Murphy CP, et al. 2012. Out-­patient antimicrobial drug use in dogs and cats for new disease events from community companion animal practices in Ontario. Can Vet J 53:291. Nobrega DB, et al. 2020. Critically important antimicrobials are generally not needed to treat nonsevere clnical mastitis in lactating dairy cows: results from a network meta-­ analysis. J Dairy Sci 11:10585. Nomura M, et al. 2020. Mortality, clinical findings, predisposing factors and treatment of Clostridiodes difficile colitis in Japanese thoroughbred racehorses. Vet Rec 187:e14. Papich MG, et al. 2010. Pharmacokinetic, protein binding, and tissue distribution of orally administered cefopodoxime proxetil and cephalexin in dogs. Am J Vet Res 71:1484. Passmore CA, et al. 2007. Efficacy and safety of cefovecin (ConveniaTM) for the treatment of urinary tract infections in dogs. J Small Anim Pract 48:139. Petersen SW, Rosin E. 1993. In vitro antibacterial activity of cefoxitin and pharmacokinetics in dogs. Am J Vet Res 54:1496. Primeau CA, et al. 2022. Integrated surveillance of extended-­spectrum beta-­lactamase (ESBL)-­producing Salmonella and Escherichia coli from humans and animal species raised for human consumption in Canada from 2012 to 2017. Epidemiol Infect 151:E14. Pusterla N, et al. 2016. Pharmacokinetic parameters for single-­and multi-­dose regimens for subcutaneous administration of

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a high-­dose ceftiofur crystalline-­free acid to neonatal foals. J Vet Pharmacol Ther 40:88. Ramirez-­Castillo FY, et al. 2023. An overview of carbapenem-­resistant organisms from food-­producing animals, seafood, aquaculture, companion animals, and wildlife. Front Vet Sci 10:1158588. Rutjens S, et al. 2023. Cefquinome shows a higher impact on the pig gut microbiome and resistome compared to ceftiofur. Vet Res 54:45. Samitz EM, et al. 1996. In vitro susceptibilities of selected obligate anaerobic bacteria obtained from bovine and equine sources to ceftiofur. J Vet Diagn Invest 8:121. Sanchez S, et al. 2002. Characterization of multidrug-­resistant Escherichia coli isolates associated with nosocomial infections in dogs. J Clin Microbiol 40:3586. Scherer CB, et al. 2018. Ceftaroline resistance in Staphylococcus pseudintermedius gene mecA carriers. Pesq Vet Bras 38:12. Shaheen BW, et al. 2010. Antimicrobial resistance profiles and clonal relatedness of canine and feline Escherichia coli pathogens expressing multidrug resistance in the United States. J Vet Intern Med 24:323. Stegemann MR, et al. 2007. Clinical efficacy and safety of cefovecin in the treatment of canine pyoderma and wound infection. J Small Anim Pract 48:378. Sun D, et al. 2020. Optimal regimens based on PK/PD cutoff evaluation of ceftiofur against

Actinobacillus pleuropneumoniae in swine. BMC Vet Res 16:366. Toombs-­Ruane LJ, et al. Carriage of extended-­ spectrum beta-­lactamase and AmpC beta-­ lactamase-­producing Escherichia coli strains from humans and pets in the same households. Appl Environ Microbiol 86:e01613-­20. Weese JS, et al. 2009. Infection with methicillin-­ resistant Staphylococcus pseudintermedius masquerading as cefoxitin susceptible in a dog. J Am Vet Med Assoc 235:1964. Weese JS, et al. 2022. Fecal shedding of extended-­spectrum beta-­lactamase-­producing Enterobacterales in cats admitted to an animal shelter. J Feline Med Surg 24:1301. Weldhagen GF. 2004. Integrons and β-­lactamases. Int J Antimicrob Agents 23:556. World Health Organization. 2024. List of Medically Important Antimicrobials. A risk management tool for mitigating antimicrobial resistance due to non-human use. Geneva: World Health Organization. Wu MT, et al. 2016. Evaluation of oxacillin and cefoxitin disk and MIC breakpoints for prediction of methicillin resistance in human and veterinary isolates of Staphylococcus intermedius group. J Clin Microbiol 54:535. Zogg AL, et al. 2018. High prevalence of extended-­spectrum β-­lactamase-­producing Enterobacteriaceae among clinical isolates from cats and dogs admitted to a veterinary hospital in Switzerland. Front Vet Sci 5:62.

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  ­References and Bibliograph

9 Other Beta-­lactam Antibiotics: Beta-­lactamase Inhibitors, Carbapenems, and Monobactams John F. Prescott and Laura Y. Hardefeldt

The continuing development of beta-­lactam antibiotics by changes of atoms within the basic beta-­lactam ring or its attachment to the thiazolidine ring has produced compounds with significantly different activity from penam penicillins and the cephalosporins and cephamycins. Carbapenem and monobactam class antibiotics have been introduced into human medicine but none have been approved for use in veterinary medicine. By contrast, some beta-­lactamase inhibitors (clavulanic acid, sulbactam) have been successfully introduced into veterinary medicine in combination with aminobenzyl penicillins, producing broad-­spectrum antibacterial drugs which overcome the limitations that some of the acquired resistance had placed on the older extended-­spectrum penicillins. Resistance increasingly continues to threaten the efficacy of these beta-­lactams, however, as beta-­ lactamase resistance genes evolve and then expand and spread globally through mobile genetic elements in Gram-­negative bacteria. This expansion is partly spearheaded by the expansion of certain “high-­risk” bacterial clones, with subsequent dissemination of their resistance genes into the broader enteric and other Gram-­negative bacterial populations.

­Beta-­lactamases and Beta-­lactamase Inhibitors: Clavulanic Acid, Sulbactam, and Tazobactam Beta-­lactamase production is a major factor in  constitutive or acquired resistance of bacteria to beta-­lactam antibiotics. The clinical importance of beta-­lactamases has been associated particularly with the rapid ability of plasmid-­mediated resistance to spread through bacterial populations. Such resistance has considerably reduced the value of once important drugs such as amoxicillin. Three beta-­lactamase inhibitors, clavulanic acid, sulbactam, and tazobactam (Figure  9.1), have considerably enhanced the activity of penicillins against bacteria with acquired plasmid-­ mediated resistance. Although possessing weak antibacterial activity on their own, their irreversible binding to susceptible beta-­lactamases (Table 9.1) allows the active beta-­lactam antibiotic with which they are combined to bind to the penicillin-­binding proteins (PBPs), resulting in lysis of the bacterial pathogen. Antibiotics combined for clinical use with clavulanic acid or sulbactam, which both have similar spectrum of

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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Other Beta-­lactam Antibiotics: Beta-­lactamase Inhibitors, Carbapenems, and Monobactams

(A)

O2 S

O CH O

Figure 9.1  Structural formulas of clavulanic acid (A) and sulbactam (B).

(B)

CH2OH

N COOH

O

N

CH3 CH3

COOH

Table 9.1  Functional and molecular characteristics of the major groups of beta-­lactamases. Inhibited by:

a

Bush–Jacoby group

Molecular class

Common namea and attributes of beta-­lactamases in functional group

Clavulanic acid

Avibactam

1

C

Cephalosporinase; often chromosomal enzymes in Gram-­negative bacteria. Confer resistance to all classes of beta-­lactams, except carbapenems. Plasmid-­encoded include LAT, MIR, ACT, FOX, CMY family beta-­lactamases, including FOX-­1, CMY-­2, MIR-­1

–

+

1e

C

Cephalosporinase; increased hydrolysis of ceftazidime (CM-­37)

–­

+

2a

A

Penicillinase; staphylococcal and enterococcal penicillinases included. High resistance to penicillins

+

+

2b

A

Penicillinase; broad-­spectrum beta-­lactamases, primarily Gram-­negative bacteria (TEM-­1, SHV-­1)

+

+

2be

A

ESBL; confers resistance to oxyimino-­ cephalosporins (cefotaxime, ceftiofur, ceftazidime) and monobactams (CTX-­M, includes CTX-­M15, PER, SHV, some OXA, TEM, VEB,)

+

+

2br

A

IRT TEM beta-­lactamases; one inhibitor-­resistant SHV-­derived enzyme (TEM-­30, SHV-­10)

+

+

2c

A

Carbenicillinase; carbenicillin-­hydrolyzing enzymes (PSE-­1)

+

+

2d

D

Oxacillinase; cloxacillin-­hydrolyzing enzymes OXA family

+

+

2de

D

ESBL cephalosporinases (OXA-­11, OXA-­15)

+

+

2df

D

Carbapenemases (OXA-­23, OXA-­48)

+

+

2f

A

Carbapenemase; hydrolysis of carbapenems, cephalosporins, cephamycins, penicillins, weak inhibition by clavulanic acid (KPC-­2, IMI-­1)

+

+

3a

B

MBL carbapenemase; broad spectrum of all beta-­lactams except monobactams (IMP-­1, IND-­1, NDM-­1, VIM-­1)

–­

–­

3b

B

MBL carbapenemase; preferential hydrolysis carbapenems (CphA, Sfh-­1)

–­

–­

 Common name: ESBL, extended-­spectrum beta-­lactamase; IRT, inhibitor-­resistant TEM; MBL, metallo-­beta-­lactamase. Source: Table adapted from Bush and Fisher (2011), and Bush and Bradford (2019, 2020); See Beta-­Lactamase DataBase (www.bldb.eu) for an updated list of beta-­lactamases.

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beta-­lactamase-­inhibiting activities, have included amoxicillin, ampicillin, and ticarcillin. Clavulanic acid and sulbactam are synergistic with a number of penicillins and cephalosporins that are readily hydrolyzed by plasmid-­mediated beta-­lactamases, including benzyl and aminobenzyl penicillins and third-­generation cephalosporins. The introduction of clavulanic acid and sulbactam was a significant advance in antimicrobial therapy of infections. The beta-­lactamase inhibitor combinations should be used with caution in herbivores with expanded large intestines because of potential for disrupting normal flora resulting in diarrheic illness.

Beta-­lactamases: Classification Beta-­lactamases are enzymes which degrade beta-­lactam drugs by opening the beta-­lactam ring (see Figure 7.2). As described in Chapter 8, there has been a remarkable evolution of these enzymes in response to antimicrobial selection and widespread dissemination of beta-­ lactamase resistance genes through Gram-­ negative bacterial populations by means of plasmids, transposons, and other mobile genetic elements. The beta-­lactamases of clinically important pathogens have been studied in exquisite detail. They consist of a wide variety of related proteins, many hundreds of which have been fully characterized. They may be chromosomally mediated (inducible or constitutive) or plasmid mediated, with transferable spread causing the greatest chaos and threat to the continued use of these drugs. Numbers appear to be rising almost exponentially (Bush and Fisher,  2011; Bush,  2018). Beta-­lactamases of Gram-­positive bacteria may be exported extracellularly whereas beta-­lactamases of Gram-­ negative bacteria are usually found in the periplasmic space but may be found extracellularly when the bacterium lyses (see Figures 7.3, 7.4). Certain “high-­risk” clones of E. coli, Klebsiella pneumoniae, Acinetobacter baumannii, and Pseudomonas aeruginosa have

had an important role globally in human medicine in spreading resistance because of their ability to thrive in humans in hospital settings and to flexibly accumulate and change resistance, thus acting as a reservoir of resistance genes for other bacteria. Classification into four molecular classes is based on a combination of structural molecular characterization (nucleotide, amino acid sequence) and further by functional characterization (substrate, inhibition profile) (Table 9.1), although these do not account for other changes that can affect the susceptibility of a bacterium. Although there is general correlation with molecular structural-­based typing approaches, a functional approach to classification is preferred because minimal differences in molecular character may cause dramatic differences in function. Functional groups are identified by relative hydrolysis of the substrates of penicillins, cephalosporins, carbapenems, and monobactams (e.g., benzyl penicillin, ceftazidime, cefotaxime, imipenem), and further by their inhibition by avibactam, clavulanic acid, tazobactam, and EDTA (Table  9.1) (Bush and Fisher,  2011; Bush,  2018; Bush and Bradford,  2019,  2020). Extended-­spectrum beta-­lactamases (ESBLs) and cephalosporinases, including the CTX-­M enzymes that hydrolyze extended-­spectrum cephalosporins, now form the largest group. However, carbapenemases such as the IMP, KPC, NDM, and OXA-­48 families are increasing rapidly as their genes spread on plasmids globally, often but not always into high-­risk clonal bacteria of a variety of Gram-­negative pathogenic species in human hospital settings (Bush, 2018; Bush and Bradford, 2020). These are still rare in animal pathogens. The classification of beta-­lactamases continues to evolve (Mack et  al.,  2020; Yoon and Jeong, 2020). The ready availability of inexpensive genome sequencing without phenotypic information is complicating identification of genuine beta-­lactamases. Part of the increasing complexity of beta-­ lactamase resistance is that individual bacteria

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­Beta-­lactamases and Beta-­lactamase Inhibitors: Clavulanic Acid, Sulbactam, and Tazobacta  171

Other Beta-­lactam Antibiotics: Beta-­lactamase Inhibitors, Carbapenems, and Monobactams

may not only acquire and maintain multiple distinct beta-­lactamase enzymes, but this form of resistance may add to resistance mediated through changes in porin function and efflux mechanisms. The genes for beta-­lactamases are found in the chromosome or on plasmids and may be moved from these sites by transposons. Transfer of some of these genes has been widespread within and between species, genera, and families. The evolution of beta-­lactamases has occurred at a dramatic rate among bacteria, probably in response to selection by the extensive use of beta-­lactam antibiotics, especially those with an increasing spectrum of activity. This spread may have been aided by use of antimicrobials such as the fluoroquinolones that encourage bacterial mutation. Plasmid-­ mediated beta-­lactamases are centrally important in beta-­lactamase resistance. For example, plasmid-­mediated TEM-­1 beta-­lactamase, which encodes ampicillin resistance, has become widespread in E. coli. More recently, plasmid-­mediated ESBLs, discussed below, have emerged among Enterobacterales, although many remain sensitive to cefoxitin and imipenem and usually also to the beta-­ lactamase inhibitors clavulanic acid and tazobactam. However, some TEM variants resistant to beta-­lactamase inhibitors have been described (Table 9.1). All Gram-­negative bacteria produce beta-­ lactamases, usually functional group  1, from genes located on their chromosomes. In some  genera (e.g., Acinetobacter, Citrobacter, Enterobacter, Serratia), as described in Chapter 8 under Resistance to cephalosporins, these AmpC hyperproducers are inducible, producing high concentrations of enzyme that overwhelm local concentrations of beta-­ lactamase inhibitors. In some cases, therefore, mutants with derepressed inducible beta-­ lactamases have emerged among the genera listed which are resistant to beta-­lactams that previously were effective. More seriously, as described in Chapter  8, AmpC hyperproduction may become plasmid encoded by high

copy number plasmids (CMY2, FOX, MIR, MOX). The dissemination of CMY2  AmpC beta-­lactamase plasmids among E. coli and Salmonella is a particular concern. The most rapidly expanding family of beta-­ lactamases in the group 2 serine beta-­lactamases are the ESBLs, which contain the functional groups 1e, 2be, 2ber, and 2de (Table 9.1) (Bush and Fisher,  2011). The expansion of the CTX family of ESBLs has displaced the early TEM-­ and SHV-­derived ESBLs, with CTX-­M15 being the globally most widely distributed ESBL in human medicine (Bush,  2018; Bush and Bradford, 2020). Numerous studies have shown that ESBL-­producing Enterobacterales continue to expand globally in intensively reared food animals (Elnekave et  al.,  2019; Hayer et  al.,  2020; Moffet et  al., 2020) and in horses (Elias et al., 2020), and in companion animals both inside hospitals and increasingly in primary care veterinary clinical settings (Timofte et al., 2016; Walter et al., 2016; Chong et al., 2018; Zogg et  al.,  2018; Bortolami et  al.,  2019; Chen et  al.,  2019). For example, extended-­spectrum cephalosporin-­resistant E. coli from clinical infections in companion animals rose from 7% of isolates in 2010–2011 (Timofte et al., 2016) to 21% in 2010–2016 (Botolami et  al., 2019). Companion animal veterinary clinics play an important role in the selection and transmission of multidrug-­resistant bacteria, both in animals and to their owners (Dazio et al., 2021; Schmitt et  al.,  2021). Some companion animal isolates belong to sequence types of pandemic high-­risk multidrug-­resistant human clones (Bortolami et al., 2019). Identification of ESBLs can be problematic. At one time, most guidelines recommended screening based on reduced susceptibility to extended-­spectrum cephalosporins as a primary screen, followed by a second test (such as use of double different cephalosporin + clavulanic acid disks spaced at specific distances) to confirm ESBL production, although the latter will not always be necessary if an obvious ESBL is identified. This approach has been generally, but not always, replaced by recommended

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172

breakpoints for specific drugs. In the Committee for Clinical Laboratory Standards Institute (CLSI) human guidelines (CLSI, 2010), breakpoints for Enterobacterales were changed to 64 μg/ml). Although not an approved indication, tulathromycin has been shown to clear Leptospira borg­ petersenii from the urine and kidneys of experimentally infected cattle (Cortese et  al.,  2007). Combination of tulathromycin with ketoprofen showed reduced pyrexia and more rapid improvement of BRD clinical symptoms (respiration, depression scores) compared to tulathromycin alone (de Koster et al., 2022).

double-­blinded controlled study showed no benefit of tulathromycin over ­controls in the IM treatment of foals with ultrasonographic evidence of lung abscesses (lung score 8.0–15 cm) (Venner et al., 2013b). Tulathromycin is poorly active against R. equi in vitro with an MIC90 >64 μg/ml. However, the prolonged half-­life and potential dosing interval of seven days may provide an advantage in treatment of other infections (Leventhal et al., 2020).

Swine

Gamithromycin is a semisynthetic azalide approved for the treatment and control of BRD. It differs from most other macrolides approved for veterinary use in its structural composition by having a 15-­membered semisynthetic lactone ring with a uniquely positioned alkylated nitrogen atom at the 7a-­position.

Tulathromycin is indicated for the prevention and treatment of SRD caused by A. pleuro­ pneumoniae, P. multocida, B. bronchiseptica, H. parasuis, or Mycoplasma hyopneumoniae, Mycoplasma hyorhinis, and M. hyosynoviae. A single dose of tulathromycin was as effective as three daily administrations of enrofloxacin for the treatment of pigs inoculated experimentally with M. hyopneumoniae (Nanjiani et al., 2005). Sheep and Goats

Tulathromycin is used in the systemic treatment of early footrot in sheep and in the early treatment of acute bacterial pneumonia in goats and sheep. In an uncontrolled study of sheep and goats with caseous lymphadenitis, closed-­system lavage in combination with either intralesional or SC administration of tulathromycin resulted in resolution of the abscesses in most cases (Washburn et al., 2009). However, the in vitro activity of tulathromycin against Corynebacterium pseudotuberculosis has not been studied. Horses

In a controlled, randomized, double-­blinded clinical trial, foals with R. equi lung abscesses diagnosed by ultrasound and treated with tulathromycin IM were significantly improved compared to untreated foals but the efficacy was less than that of foals administered an zithromycin and rifampin combination (Rutenberg et  al.,  2017). However, an earlier randomized,

Gamithromycin

Antimicrobial Activity

The antimicrobial activity of gamithromycin is similar to that of other azalides such as azithromycin and to tilmicosin and tulathromycin. The drug is active in vitro against M. haemolyt­ ica, P. multocida, H. somni, some Mycoplasma bovis, Streptococcus equi subsp. zooepidemicus, Streptococcus suis, and R. equi. Resistance in M. haemolytica is increasingly problematic and in M. bovis is now widespread. Mycoplasma hyopneumoniae and Mycoplasma mycoides subsp. mycoides are susceptible. Pharmacokinetic Properties

The pharmacokinetics of gamithromycin in cattle are characterized by rapid absorption from the injection site, extensive distribution into tissues, and slow elimination, which collectively contribute to high and sustained concentrations in pulmonary epithelial lining fluid, bronchoalveolar cells, and lung tissue (Giguère et  al.,  2011b). The bioavailability of gamithromycin after SC administration to cattle is nearly 100% (Huang et  al.,  2010). The apparent volume of distribution after IV

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administration is 25 l/kg (Huang et al., 2010). Peak lung concentrations are approximately 28 μg/g after administration of an SC dose of 6  mg/kg. Lung concentrations are 16–650 times higher than concurrent plasma concentrations. Lung elimination half-­life values for cattle are 6–7 days (Huang et al., 2010; Giguère et  al.,  2011b). Pharmacokinetic studies in swine showed similar advantages to those in cattle; the plasma half-­life after IM injection is 94 hours (Hamel et al., 2021a). Toxicity and Adverse Effects

Gamithromycin is safe to use in cattle. No serious adverse events were noted during the clinical development program of the drug. Transient discomfort and mild to moderate injection site swelling may be seen in some treated animals, so that larger doses are given in different sites. Safety has not been assessed in other species. Administration and Dosage

Administration and dosages are summarized in Table 12.1. Clinical Applications Cattle

Gamithromycin is approved for the treatment or metaphylaxis of BRD associated with M. haemolytica, P. multocida, or H. somni in beef and nonlactating dairy cattle. The efficacy of gamithromycin for the treatment and control of bovine respiratory disease has been documented in multiple studies (Abell et al., 2017). Swine

Gamithromycin has the same indications for use in swine as tulathromycin. It was as effective as tildipirosin against respiratory tract disease in swine (Hamel et al., 2021a). Sheep and Goats

Although not licensed for such use in small ruminants, gamithromycin would be a reasonable alternative for the treatment of respiratory disease in sheep and goats. Subcutaneous injection of gamithromycin at a dose of

6 mg/kg was effective in the treatment of footrot in sheep (Hamel et al., 2021b). Horses

Intramuscular administration of gamithromycin to foals at a dosage of 6  mg/kg maintains pulmonary epithelial lining fluid concentrations above the MIC90 for S. equi subsp. zooepidemicus and phagocytic cell concentrations above the MIC90 for R. equi for approximately seven days (Berghaus et  al.,  2012). Gamithromycin (IM, weekly) was not inferior to azithromycin-­ rifampin in a controlled, randomized, double-­ blinded clinical trial in foals with mild to moderate R. equi bronchopneumonia but had a higher frequency of adverse reactions (colic, hindlimb lameness because of painful injection site) (Hildebrand et  al.,  2015). Intravenous injection was found to be both well tolerated and to have pharmacokinetic parameters similar to those following IM injection (Berlin et al., 2017).

Tildipirosin Tildipirosin is a semisynthetic 16-­membered macrolide derived from the naturally occurring tylosin. The chemical structure of tildiprosin is characterized by two piperidine substituents on C20 and C23, and a basic mycaminose sugar moiety at C5 of the macrocyclic lactone ring. Owing to three nitrogen atoms accessible to protonation, tildipirosin is a tribasic molecule. Antimicrobial Activity

The antimicrobial activity of tildipirosin is similar to that of other azalides such as azithromycin, gamithromycin, and to tilmicosin and tulathromycin. Pharmacokinetic Properties

The pharmacokinetics of tildipirosin in cattle and swine are characterized by rapid absorption from the injection site, extensive distribution into tissues and slow elimination, which collectively contribute to high and sustained concentrations in bronchial fluid, and lung tissue (Rose et  al., 2012; Menge et  al.,  2012).

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The bioavailability of tildipirosin after SC administration to cattle is approximately 80% (Menge et  al.,  2012). The apparent volume of distribution after IV administration to cattle is 49  l/kg (Menge et  al.,  2012). Peak lung concentrations are approximately 15 (Menge et al., 2012; Rose et al., 2013) and 4 μg/g after administration of a dose of 4  mg/kg to cattle and swine, respectively (Menge et  al.,  2012; Rose et  al.,  2013). Concentrations achieved in the lung are remarkable; the lung half-­life in swine is about seven days and bronchial fluid in cattle about 11  days. Pharmacokinetic studies in dogs, goats, rabbits, and sheep confirm the long half-­lives and high bioavailability of tildipirosin (Wang et al., 2018; Xiong et al., 2020; Galecio et al., 2022a,b). Toxicity and Adverse Effects

Tildipirosin is safe to use in cattle. No serious adverse events were noted during the clinical development program of the drug. Mild to moderate injection site swelling and pain on palpation of the injection site are common in treated cattle and swine. During clinical trials in swine, treatment with tildipirosin caused shock symptoms in two of 1048 treated animals. The higher dose cattle form should not be used in swine. Tildipirosin also appears safe in dogs, dairy goats, horses, and rabbits. Administration and Dosage

Administration and dosages are summarized in Table 12.1. Clinical Applications Cattle

Tildipirosin is approved for the treatment or control of BRD associated with M. haemolyt­ ica, P. multocida, or H. somni in beef and nonlactating dairy cattle. Swine

In some countries, tildipirosin is approved for the treatment of respiratory disease associated with A. pleuropneumoniae, P. multocida, B. bronchiseptica, and G. parasuis.

Tylvalosin Tylvalosin (acetylisovaleryltylosin) is a 16-­ membered lactone ring macrolide antibiotic recently approved in some countries for use in swine and poultry. Antimicrobial Activity

Tylvalosin is highly active in  vitro against Mycoplasma synoviae, M. hyopneumoniae, and M. gallisepticum, and more potent than other veterinary macrolides including, at least for avian mycoplasma, strains resistant to tilmicosin or tylosin (Morrow et al., 2020). It is also active against some but not all isolates of B. hyodysente­ riae and B. pilosicoli. The drug is active against some obligate anaerobes including Clostridium spp. The activity of tylvalosin against many bacterial pathogens of veterinary importance has not been studied. It is active against Toxoplasma. Pharmacokinetic Properties

Tylvalosin tartrate is rapidly absorbed after oral administration to pigs and chicken. Tylvalosin is rapidly metabolized to 3-­O-­acetyltylosin, which possesses equivalent microbiological activity to the parent compound. In pigs, plasma concentrations are below the limit of quantification after administration of the recommended dose. In chickens, peak plasma concentrations are achieved approximately one hour after a single oral dose. Tylvalosin is rapidly distributed to the major organs. Tylvalosin concentrations in the lung are detected for up to 12 hours after administration. Part of the overall efficacy of the product might be due to the activity of the metabolites rather than to tylvalosin alone. Toxicity and Adverse Effects

No adverse reactions related to the drug were observed during clinical or target animal safety studies. Administration and Dosage

Administration and dosages are summarized in Table 12.1.

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Clinical Applications Poultry

In some countries, tylvalosin tartrate is approved for the prevention and treatment of different mycoplasmoses and Clostridium per­ fringens infections in chickens and turkeys. The drug is also indicated for prevention and treatment of mycoplasmosis in pheasants. Swine

In the United States, tylvalosin tartrate is approved for the control of porcine proliferative enteritis associated with L. intracellularis infection. In many other countries, the drug is also approved for the treatment and prevention of porcine proliferative enteropathy, swine enzootic pneumonia caused by susceptible strains of M. hyopneumoniae, and swine dysentery caused by B. hyodysenteriae, although resistance in the latter is common.

Other Classic Macrolides Uncommon macrolide antibiotics (oleandomycin, josamycin, kitasamycin, rosaramicin) have activity similar to erythromycin, spiramycin, and tylosin. There is little reported experience with their use in veterinary medicine, although kitasamycin is used in Japan. The agents appear to have nothing to offer over the commonly used classic macrolide antibiotics.

­Advanced-­generation Macrolide Antibiotics: Roxithromycin, Clarithromycin, and Azithromycin Interest in other macrolides has been stimulated by their activity against both traditional and emerging human pathogens, including Campylobacter spp., Helicobacter spp., Legionella spp., as well as against intracellular organisms that have emerged through the AIDS epidemic, such as Bartonella spp. and Mycobacterium spp. New erythromycin derivatives with enhanced pharmacokinetic and in some cases broader antibacterial activities

include roxithromycin, clarithromycin, and azithromycin, which are in widespread use in human medicine. Roxithromycin is an acid-­stable derivative of erythromycin with similar activity to erythromycin and complete cross-­resistance with erythromycin but differs by enhanced oral bioavailability and longer half-­life, allowing for once-­ or twice-­daily administration. It is a well-­tolerated alternative to erythromycin for daily oral administration. Clarithromycin, a 6-­0-­methyl derivative of erythromycin (Figure  12.2), is approximately twice as active as erythromycin against bacteria on a weight basis, has a half-­life about twice that of erythromycin, and good activity against Mycobacterium avium. Azithromycin, an acid-­ stable 15-­membered ring azalide (Figure 12.2), is more active than erythromycin against Gram-­negative bacteria and has a considerably lengthened half-­life relative to erythromycin. The application of these and other newer macrolides for veterinary use will likely take advantage of their long half-­lives, which may allow for a single administration in the treatment of infections caused by pathogens such as Campylobacter and Mycoplasma, and of infections caused by intracellular bacteria.

Antimicrobial Activity Bacteria with MIC ≤2 μg/ml are generally regarded as susceptible and ≥8 μg/ml as resistant to newer macrolides. All these macrolides approved for use in human medicine share similar antibacterial spectrum of activity against Gram-­positive isolates. Azithromycin has the broadest in  vitro spectrum against Gram-­negative bacteria, including moderate activity against Salmonella enterica, but the others also have activity against important human Gram-­negative upper respiratory tract pathogens (Bordetella pertussis, Haemophilus influenzae, and Moraxella catarrhalis). Other important antibacterial effects include excellent activity against the genera Bartonella, Borrelia, Brucella, Campylobacter, Chlamydia,

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Legionella, Leptospira, Mycoplasma, members of the Spirochetaceae, and Ureaplasma. Mycobacteria such as M. avium are often moderately susceptible. Activity against anaerobic bacteria is variable.

Pharmacokinetic Properties In comparison to erythromycin, from which they have been developed, newer macrolides are acid stable, produce fewer gastrointestinal adverse effects, have higher bioavailability following oral administration, have considerably lengthened serum half-­lives, and produce higher tissue concentrations, so that single or twice-­daily dosing is appropriate. Oral bioavailability of azithromycin is approximately 97% in dogs and about 50% in cats and foals. The oral bioavailability of clarithromycin in dogs is lower, ranging between 60% and 80%. The bioavailability of clarithromycin in dogs is not significantly influenced by feeding. Azithromycin, but not clarithromycin, is also available as an IV formulation. Serum elimination half-­lives are 20 hours and 35 hours for azithromycin in foals and cats, respectively. The elimination half-­life of clarithromycin in foals (4.8 hours) is shorter than that of azithromycin but longer than that of erythromycin (one hour). The long half-­lives of these newer drugs, which is particularly marked for azithromycin, apparently result from extensive uptake by, and slow release from, tissues rather than resulting from delayed metabolism. The major route of excretion is the bile and intestinal tract, although clarithromycin is more markedly excreted through the kidney. About half the administered azithromycin is excreted unchanged in the bile in dogs and cats. Tissue half-­lives in cats vary from 13 hours in adipose tissue to 72 hours in heart muscle (Hunter et  al.,  1995). Concentrations of azithromycin within the lung and spleen of cats exceeded 1  μg/ml 72 hours after a single oral dose of 5.4 mg/kg (Hunter et al., 1995). Tissue concentrations of azithromycin are generally 10–100 times those achieved in serum.

The extensive tissue distribution of azithromycin appears to result from its concentration within macrophages and neutrophils. The half-­ life of azithromycin monotherapy in foal neutrophils is 49 hours. Bronchoalveolar cells  and pulmonary epithelial lining fluid concentrations in foals are 15–170-­fold and 1–16-­fold higher than concurrent serum concen­trations, respectively (Jacks et al., 2001). In foals, clarithromycin achieves considerably greater concentrations in pulmonary epithelial lining fluid and alveolar macrophages than either erythromycin or azithromycin. However, the half-­life of clarithromycin at these sites is much shorter than that of azithromycin (Suarez-­Mier et al., 2007).

Toxicity and Adverse Effects In humans, newer macrolides are typically well tolerated and cause fewer gastrointestinal disturbances than erythromycin. Experience in dogs and cats suggests the same is true in these species. As with earlier macrolides, these drugs occasionally can induce enterocolitis in foals and possibly in accompanying mares. Adult horses appear to have a higher incidence of enterocolitis associated with administration of macrolides than foals. Clarithromycin may be fetotoxic and should not be administered to pregnant animals.

Administration and Dosage Dosage recommendations for dogs, cats, and foals are summarized in Table 12.1.

Clinical Applications There is limited experience with the use of newer macrolides in veterinary medicine, but these drugs offer the advantage for monogastrates of better oral bioavailability, potentially fewer adverse effects, and less frequent administration compared to erythromycin. Their marked efficacy against intracellular organisms is a considerable advantage. Potential

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applications include those described for erythromycin, for example, as an alternative to penicillin in penicillin-­allergic animals for the treatment of infections caused by susceptible Gram-­positive aerobes, an alternative to amoxicillin or doxycycline in the treatment of leptospirosis, and an alternative to tetracyclines in treatment of Rickettsia and Coxiella infections. Newer macrolides may have advantages in the treatment of intracellular infections in monogastrates, including Bartonella, Chlamydia, and atypical mycobacterial infections. Clarithro­ mycin is effective in the treatment of atypical Mycobacterium infections, when combined with other antibiotics. Other areas that need to be investigated are use against Mycoplasma infections in animals, since medically important Mycoplasma are highly susceptible to clarithromycin in vitro. Dogs and Cats

Azithromycin in combination with atovaquone or artesunate was effective in eliminating Babesia gibsoni from persistently infected dogs (Birkenheuer et al., 2004; Karasová et al., 2022). Administration of azithromycin to dogs with experimental Rocky Mountain spotted fever resulted in improvement of most of the clinical signs but was not as effective as doxycyline or trovafloxacin in decreasing vascular injury to the eye and clearing viable circulating rickettsiae (Breitschwerdt et al., 1999). Azithromycin prevented or resolved episodes of acute arthritis and reduced the bacterial load but failed to eliminate Borrelia burgdorferi in infected dogs (Straubinger,  2000). Azithromycin, given at a dose of 10–15 mg/kg daily for three days and then twice weekly, provided a similar rapid resolution of clinical signs when compared to doxycycline in cats with Chlamydia felis infections. In a prospective, randomized, placebo-­ controlled clinical trial, azithromycin at a dose of 10  mg/kg PO once daily was surprisingly found to be safe and effective for the treatment of papillomatosis in dogs (Yağci et al., 2008). In one study, dogs with persistent leptospiruria responded to clarithromycin following ineffective

treatment with doxycycline and/or enrofloxacin (Mauro and Harkin, 2018). Clarithromycin, in combination with amoxicillin and a proton pump inhibitor, has been used successfully for the treatment of gastric ulcers associated with Helicobacter spp. in dogs. Horses

The main extra-­label use of azithromycin or clarithromycin in the horse is for the treatment of Rhodococcus equi infections in foals, where there are pharmacokinetic advantages over erythromycin (Suarez-­Mier et  al.,  2007). The incidence of diarrhea in foals treated with clarithromycin is similar to that observed with erythromycin. In most cases, diarrhea is mild and self-­limiting but diarrheic foals should be monitored carefully because some may develop depression and severe diarrhea, leading to dehydration and electrolyte loss. Because of the potential for severe enterocolitis, clarithromycin and azithromycin should, like erythromycin, not be used in adult horses unless no alternatives are available. As discussed earlier under Drug interactions, although a combination of a macrolide with rifampin has been a mainstay of treatment of  R. equi pneumonia in foals, there is no evidence that the combination improves clinical response compared to macrolides alone. There is also evidence for the adverse effects of rifampin on the metabolism of macrolides. In  particular, concurrent administration of rifampin considerably reduces absorption of clarithromycin in foals (Peters et  al.,  2011). A combination of clarithromycin with doxycycline or minocycline reduced the emergence of resistance in R. equi in  vitro (Erol et  al., 2021).  The combination of doxycycline with azithromycin was shown to be as effective as the combination of rifampin and azithromycin (Wetzig et al., 2020). The transferable basis of resistance to macrolides and rifampin, which is now relatively common, has been well described (Álvarez-­ Narváez et al., 2020), as has the international spread of the multidrug-­resistant R. equi clone

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(Val-­Calvo et  al.,  2022). The development of resistance was associated with mass prophylactic use of azithromycin in foals on endemically infected horse-­breeding farms in the United States (Huber et  al.,  2021). Changing the policy of treatment of foals with small lung abscesses to treatment of those only with larger abscesses was shown to decrease antimicrobial usage without increasing mortality (Arnold-­ Lehna et al., 2020) and will likely reduce selection pressure for further development or spread of resistance.

­Ketolides Ketolides are members of a new semisynthetic 14-­membered ring macrolide, with a 3-­keto group instead of an alpha-­L-­cladinose on the erythronolide A ring. The mode of action of ketolide molecules involves inhibition of protein synthesis by binding to the 50S subunit of the bacterial ribosome and by blocking the translation of messenger RNA. Through such mechanisms, peptidyl tRNA accumulates within the bacterial cell, resulting in the depletion of free transfer tRNA needed for alpha-­ amino acid activation. The ketolides also inhibit the formation of the 50S and 30S bacterial ribosomal subunits. The two most widely studied ketolides are telithromycin and cethromycin. Both have been developed for oral use. Their spectrum of activity is similar to that of the newer

generation macrolides. However, they offer the advantage of overcoming some, but not all, of the current mechanisms of resistance to standard macrolides within Gram-­positive cocci. In  general, Staphylococcus aureus and Streptococcus pyogenes strains with inducible MLSB resistance are susceptible to ketolides whereas strains with constitutive expression of MLSB are resistant. In Streptococcus pneumo­ niae, Wolter et  al. (2008) confirmed that the mutant erm(B) gene conferring telithromycin resistance was expressed at a constitutively higher level than the inducible wild-­type gene, and that the elevated erm(B) expression resulted in a higher level of rRNA methylation that presumably hindered telithromycin binding to the bacterial ribosome. Ketolides are also active against most Gram-­ positive isolates that are resistant to macrolides because of macrolide efflux (mef) genes. The pharmacokinetics properties of ketolides include a long half-­life as well as extensive tissue distribution and uptake into respiratory tissues and fluids, allowing for once-­ daily dosing. Adverse effects of ketolides in humans are similar to those of macrolides and usually related to the gastrointestinal tract, with diarrhea, nausea, and abdominal pain being the most frequently reported. However, telithromycin was withdrawn from clinical use in humans in the United States after visual disturbances, myasthenia gravis, and hepatotoxicity were noted.

­References and Bibliography Abell KM , et al. 2017. A mixed treatment comparison meta-­analysis of metaphylaxis treatments for bovine respiratory disease in beef cattle. J Anim Sci 95:626. Álvarez-­Narváez S, et al. 2020. Horizontal spread of Rhodococcus equi macrolide resistance plasmid pRErm46 across environmental Actinobacteria. Appl Environ Micro 86:e00108-­20.

Anacleto TP, et al. 2011. Studies of distribution and recurrence of Helicobacter spp. gastric mucosa of dogs after triple therapy. Acta Cir Bras 26:82. Andrés-­Lasheras S, et al. 2022. Bovine respiratory disease: conventional culture-­ independent approaches to studying antimicrobial resistance in North America. Antibiotics 11:487.

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Apley MD. 2015. Treatment of calves with bovine respiratory disease. Duration of therapy and posttreatment intervals. Vet Clin Food Anim 31:441. Arnold M, et al. 2022. Distribution, genetic heterogeneity, and antimicrobial susceptibility of Brachyspira pilosicoli in Swiss pig herds. Vet Microbiol 269:e109421. Arnold-­Lehna D, et al. Changing policy to treat foals with Rhodococcus equi pneumona in the later course of disease decreases antimicrobial usage without increasing mortality rate. Equine Vet J 52:531. Baptiste KE, Kyvsgaard NC. 2017. Do antimicrobial mass medications work? A systematic review and meta-­analysis of randomized clinical trials investigating antimicrobial prophylaxis or metaphylaxis against naturally occurrnig bovine respiratory disease. Pathogen Dis 75:ftx083. Båverud V, et al. 1998. Clostridium difficile associated with acute colitis in mares when their foals are treated with erythromycin and rifampicin for Rhodococcus equi pneumonia. Equine Vet J 30:482. Berghaus LJ, et al. 2012. Plasma pharmacokinetics, pulmonary distribution, and in vitro activity of gamithromycin in foals. Vet Pharmacol Ther 35:59. Berlin S, et al. 2016. Pharmacokinetics and pulmonary distribution of clarithromycin and rifampicin after concomitant and consecutive administration in foals. Mol Pharm 13:1089. Berlin S, et al. 2017. Pharmacokinetics and pulmonary distribution of gamithromycin after intravenous administration in foals. J Vet Pharmacol Ther 40:406. Berlin S, et al. 2018. Intestinal and hepatic contributions to the pharmacokinetic interaction between gamithromycin and rifampicin after single-­dose and multiple-­dose administration in healthy foals. Equine Vet J 50:525. Birkenheuer AJ. 2004. Efficacy of combined atovaquone and azithromycin for therapy of chronic Babesia gibsoni (Asian genotype) infections in dogs. J Vet Intern Med 18:494.

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Mannheimia haemolytica in calves at high risk of developing bovine respiratory disease. J Anim Sci 96:1259. Davedow T, et al. 2020. Investigation of a reduction in tylosin on the prevalence of liver abscesses and antimicrobial resistance in Enterococci in feedlot cttle. Front Vet Sci 7:90. De Jong A, et al. 2021a. Antimicrobial susceptibility monitoring of Mycoplasma hyopneumoniae isolated from seven European countries during 2015–2016. Vet Microbiol 253:108973. De Jong A, et al. 2021b. Minimal inhibitory concentration of seven antimicrobials to Mycoplasma gallisepticum and Mycoplasma synoviae isolates from six European countries. Avian Pathol 50:161. De Koster J, et al. 2022. Treatment of bovine respiratory disease with a single administration of tulathromycin and ketoprofen. Vet Rec 190:e834. Erol E, et al. 2021. Synergistic combinations of clarithromycin with doxycycline or minocycline reduce the emergence of antimicrobial resistance in Rhodococcus equi. Equine Vet J 54:799. Fischer CD, et al. 2011. Anti-­inflammatory benefits of antibiotic-­indu ced neutrophil apoptosis: tulathromycin induces caspase-­3-­ dependent neutrophil programmed cell death and inhibits NF-­kappaB signaling and CXCL8 transcription. Antimicrob Agents Chemother 55:338. Foster DM, et al. 2016. Comparison of active drug concentrations in the pulmonary epithelial lining fluid and interstitial fluid of calves injected with enrofloxacin, florfenicol, ceftiofur, or tulathromycin. PLoS One 11:e0149100. Francois Watkins LK, et al. 2021. Ongoing outbreak of extensively drug-­resistant Campylobacter jejuni infections associated with US pet stores pupplies, 2016–2020. JAMA Netw Open 4:e2125203. Galecio JS, et al. 2022a. Pharmacokinetics of tildipirosin in horses after intravenous and intramuscular administration and its potential muscle damage. Res Vet Sci 152:20.

Galecio JS, et al. 2022b. Pharmacokinetics of tildipirosin in plasma, milk, and somatic cells following intravenous, intramuscular, and subcutaneous administration in dairy goats. Pharmaceutics 14:860. Gautier-­Bouchardon AV, et al. 2014. Overall decrease in the susceptibility of Mycoplasma bovis to antimicrobials over the past 30 years in France. PLoS One 9:e87672. Giguère S, et al. 2004. Retrospective comparison of azithromycin, clarithromycin, and erythromycin for the treatment of foals with Rhodococcus equi pneumonia. J Vet Intern Med 18:568. Giguère S, et al. 2011a. Diagnosis, treatment, control, and prevention of infections caused by Rhodococcus equi in foals. J Vet Intern Med 25:1209. Giguère S, et al. 2011b. Disposition of gamithromycin in plasma, pulmonary epithelial lining fluid, bronchoalveolar cells, and lung tissue in cattle. Am J Vet Res 72:326. Godinho KS, et al. 2005. Efficacy of tulathromycin in the treatment of bovine respiratory disease associated with induced Mycoplasma bovis infections in young dairy calves. Vet Ther 6:96. Hamel D, et al. 2021a. Gamithromycin in swine: pharmacokinetics and clinical evaluation against swine respiratory disease. Vet Med Sci 7:455. Hamel D, et al. 2021b. Gamithromycin in sheep: pharmacokinetics and clinical evaluation against footrot. Res Vet Sci 142:94. Hampson DJ, et al. 2019. Antimicrobial resistance in Brachyspira – an increasing problem for disease control. Vet Microbiol 229:59. Hildebrand F, et al. 2015. Efficacy of gamithromycin for the treatment of foals with mild to moderate bronchopneumonia. J Vet Intern Med 29:333. Holschbach C, et al. 2020. Prevalence and temporal trends in antimicrobial resistance of bovine respiratory disease pathogen isolates submitted to the Wisconsin Veterinary Diagnostic Laboratory: 2008–2017. J Dairy Sci 103:9464.

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 ­References and Bibliograph

Macrolides, Azalides, and Ketolides

Huang RA, et al. 2010. Pharmacokinetics of gamithromycin in cattle with comparison of plasma and lung tissue concentrations and plasma antibacterial activity. J Vet Pharmacol Ther 33:227. Huber L, et al. 2021. Association between antimicrobial treatment of subclinical pneumonia in foals and selection of macrolide-­ and rifampicin-­resistant Rhodococcus equi at horse-­breeding farms in central Kentucky. J Am Vet Med Assoc 258:648. Hunter RP, et al. 1995. Pharmacokinetics, oral bioavailability and tissue distribution of azithromycin in cats. J Vet Pharm Ther 18:38. Ives SE, Richeson JT. 2015. Use of antimicrobial metaphylaxis for the control of bovine respiratory disease in high-­risk cattle. Vet Clin Food Anim 31:341. Jacks S, et al. 2001. Pharmacokinetics of azithromycin and concentration in body fluids and bronchoalveolar cells in foals. Am J Vet Res 62:1870. Jelinski M, et al. 2020. Antimicrobial sensitivity testing of Mycoplasma bovis isolates derived from Western Canadian feedlot cattle. Microorg 8:124. Karasová M, et al. 2022. Clinical efficacy and safety of Malarone®, azithromycin and artesunate combination for treatment of Babesia gibsoni in naturally infected dogs. Animals 12:706. Kilpinen S, et al. 2011. Effect of tylosin on dogs with suspected tylosin-­responsive diarrhea: a placebo-­controlled, randomized, double-­ blinded, prospective clinical trial. Acta Vet Scand 53:26. Kilpinen S, et al. 2014. Efficacy of two low-­dose oral tylosin regimens in controlling the relapse of diarrhea in dogs with tylosin-­ responsive diarrhea: a prospective, single-­ blinded, two-­arm parallel clinical field trial. Acta Vet Scand 56:43. Kinnear A, et al. 2020. Investigation of macrolide resistant genotypes in Mycoplasma bovis isolates from Canadian feedlot cattle. Pathogens 9:622. Klein U, et al. 2022. Antimicrobial susceptibility profiles of Mycoplasma hyorhinis strains

isolated from five European countries between 2019 and 2021. PLoS One e0272093. Lakritz J, et al. 1997. Effect of treatment with erythromycin on bronchoalveolar lavage fluid cell populations in foals. Am J Vet Res 58:56. Lavoie JP, et al. 2000. Equine proliferative enteropathy: a cause of weight loss, colic, diarrhoea and hypoproteinaemia in foals on three breeding farms in Canada. Equine Vet J 32:418. Leventhal HR, et al. 2020. Pharmacokinetics and pulmonary distribution of Draxxin® (tulathromycin) in healthy adult horses. J Vet Pharmacol Ther 44:714. Liu Y, et al. 2020. Molecular characteristics and antibiotic susceptibility profiles of Mycoplasma bovis associated with mastitis on dairy farms in China. Prev Vet Med 182:105106. Lombardi KR, et al. 2011. Pharmacokinetics of tilmicosin in beef cattle following intravenous and subcutaneous administration. J Vet Pharmacol Ther 34:583. Luo W, et al. 2020. The dose regimen formulation of tilmicosin against Lawsonia intracellularis in pigs by pharmacokinetic-­ pharmacodynamic (PK-­PD) model. Microb Pathog 147:104389. Magstadt DR, et al. 2018. Treatment history and antimicrobial susceptibility results for Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni isolates from bovine respiratory disease cases submitted to Iowa State University Veterinary Diagnostic Laboratory from 2013 to 2015. J Vet Diagn Invest 30:99. Mainguy-­Seers S, et al. 2022. Effects of azithromycin on bronchial remodelling in the natural model of severe neutrophilic asthma in horses. Sci Rep 12:446. Manchester AC, et al. 2019. Long-­term impact of tylosin on fecal microbiota and fecal bile acids of healthy dogs. J Vet Intern Med 33:2605. Martinez-­Cortés I, et al. 2018. Tilmicosin modulates the innate immune response and preserves casein production in bovine mammary alveolar cells during

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Staphylococcus aureus infection. J Anim Sci 97:644. Mauro T, Harkins K. 2018. Persistent leptospiruria in five dogs despite antimicrobial treatment (2000–2017). J Am Anim Hosp Assoc 55:42. McKay SG, et al. 1996. Use of tilmicosin for treatment of pasteurellosis in rabbits. Am J Vet Res 57:1180. Menge M, et al. 2012. Pharmacokinetics of tildipirosin in bovine plasma, lung tissue, and bronchial fluid (from live, nonanesthetized cattle). J Vet Pharmacol Ther 35:550. Morrow CJ, et al. 2020. Antimicrobial susceptibility of pathogenic mycoplasmas in chickens in Asia. Vet Microbiol 250:e108840. Müller HC, et al. 2018. Effects of intermittent feeding of tylosin phosphate during the finishing period on feedlot performance, carcase characteristics, antimicrobial resistance, and incidence and severity of liver abscesses in steers. J Anim Sci 96:2877. Peters J, et al. 2011. Oral absorption of clarithromycin is nearly abolished by chronic comedication of rifampicin in foals. Drug Metab Dispos 39:1643. Rosales RS, et al. 2020. Antimicrobial susceptibility profiles of porcine mycoplasma isolated from samples collected in southern Europe. BMC Vet Res 16:324. Rose M, et al. 2013. Pharmacokinetics of tildipirosin in porcine plasma, lung tissue, and bronchial fluid and effects of test conditions on in vitro activity against reference strains and field isolates of Actinobacillus pleuropneumoniae. J Vet Pharmacol Ther 36:140. Rowan TG, et al. 2004. Efficacy of danofloxacin in the treatment of respiratory disease in European cattle. Vet Rec 154:585. Rutenberg D, et al. 2017. Efficacy of tulathromycin for the treatment of foals with mild to moderate bronchopneumonia. J Vet Intern Med 31:901. Schmidt JW, et al. 2020. In-­feed tylosin phosphate administration to feedlot cattle minimally affects antimicrobial resistance. J Food Protect 83:350.

Stieler AL, et al. 2016. Macrolide-­induced hyperthermia in foals: role of impaired sweat responses. Equine Vet J 48:590. Straubinger RK. 2000. PCR-­based quantification of Borrelia burgdorferi organisms in canine tissues over a 500-­day postinfection period. J Clin Microbiol 38:2191. Suarez-­Mier G, et al. 2007. Pulmonary disposition of erythromycin, azithromycin, and clarithromycin in foals. J Vet Pharmacol Ther 30:109. Trott DJ, et al. Comparative macrolide use in humans and animals: should macrolides be moved off the World Health Organization’s critically important antimicrobial list? J Antimicrob Chemother 76: 1955. Val-­Calvo J, et al. 2022. International spread of multidrug-­resistant Rhodococcus equi. Emerg Infect Dis 28:1899. Van Vliet AHM, et al. 2022. Genomic screening of antimicrobial resistance markers in the UK and US Campylobacter isolates highlights stability of resistance over an 18-­year period. Antimicrob Agents Chemother 66:e0168721. Venner M, et al. 2013a. Efficacy of mass antimicrobial treatment of foals with subclinical pulmonary abscesses associated with Rhodococcus equi. J Vet Intern Med 27:171. Venner M, et al. 2013b. Comparison of tulathromycin, azithromycin and azithromycin-­rifampin for the treatment of mild pneumonia associated with Rhodococcus equi. Vet Rec 173:397. Wang J, et al. 2018. Pharmacokinetics of tildipirosin in beagle dogs. J Vet Pharmacol Therap 41:e49-­e52. Washburn KE, et al. 2009. Comparison of three treatment regimens for sheep and goats with caseous lymphadenitis. J Am Vet Med Assoc 234:1162. Wennekamp TR, et al. 2022. Antimicrobial resistance in bovine respiratory disease: auction market-­and ranch-­raised calves. Can Vet J 63:47.

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 ­References and Bibliograph

Macrolides, Azalides, and Ketolides

Wetzig M, et al. 2020. Efficacy of the combination of doxycycline and azithromycin for the treatment of foals with mild to moderate bronchopneumonia. Equine Vet J 52:613. Wolter N, et al. 2008. Telithromycin resistance in Streptococcus pneumoniae is conferred by a deletion in the leader sequence of erm(B) that increases rRNA methylation. Antimicrob Agents Chemother 52:435. Woolums AR, et al. 2018. Multidrug resistant Mannheimia haemolytica isolated from high-­risk beef stocker cattle after antimicrobial metaphylaxis and treatment for bovine respiratory disease. Vet Microbiol 221:143. World Organisation for Animal Health. 2022. Annual Report on Antimicrobial Agents Intended for Use in Animals, 6th edn. www.woah.org/en/document/ annual-­report-­on-­antimicrobial-­agents-­ intended-­for-­use-­in-­animals/ Xiong J, et al. 2019. Tilmicosin enteric granules and premix to pigs: antimicrobial

susceptibility testing and comparative pharmacokinetics. J Vet Pharmacol Therap 42:336. Xiong J, et al. 2020. Pharmacokinetics and bioavailability of tildipirosin in rabbits following single-­dose intravenous and intramuscular administration. J Vet Pharmacol Ther 43:448. Yağci BB, et al. 2008. Azithromycin therapy of papillomatosis in dogs: a prospective, randomized, double-­blinded, placebo-­ controlled clinical trial. Vet Dermatol 19:194. Younes JA, et al. 2022. Changes in the phenotypic susceptibility of Mannheimia haemolytica isolates to macrolide antimicrobials uring the early feeding period following metaphylactic tulathromycin use in western Canadian feedlot calves. Can Vet J 63:920. Zaheer R, et al. 2020. Surveillance of Enterococcus spp. reveals distinct species and antimicrobial resistance diversity across a One-­Health continuum. Sci Rep 10:3937.

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13 Aminoglycosides and Aminocyclitols Patricia M. Dowling

G ­ eneral Considerations The aminoglycosides and aminocyclitols are bactericidal antibiotics primarily used to treat serious infections caused by aerobic Gram-­ negative bacteria and staphylococci. Amikacin and tobramycin have excellent activity against Pseudomonas aeruginosa. The use of aminoglycosides and aminocyclitols had been eclipsed by  the development of the fluoroquinolones, which have better safety profiles and better ­distribution kinetics. But in human and ­veterinary medicine, the increasing prevalence of multidrug-­resistant (MDR) Gram-­negative pathogens, including carbapenem-­resistant Enterobacterales, Pseudomonas aeruginosa, and Acinetobacter spp., renewed interest in aminoglycosides for use as monotherapy or typically in combination with other antimicrobials. However, the World Health Organization has reclassified the aminoglycosides (with the  exception of spectinomycin) as Critically Important Antibiotics (CIA) due to their use in the treatment of human infections. There is a high probability of transfer of aminoglycoside resistance determinants from nonhuman sources (van Duijkeren et  al.,  2019). Renal accumulation of aminoglycosides results in detectable drug residues for prolonged periods, so their extra-­label use in food animals is strongly discouraged.

Chemistry The aminoglycoside antibiotics – streptomycin, dihydrostreptomycin, kanamycin, gentamicin, tobramycin, amikacin, and neomycin  – are large molecules with numerous amino acid groups, making them basic polycations that are highly ionized at physiological pHs. Their polarity largely accounts for the pharmacokinetic properties that are shared by all members of the group. Chemically, they consist of a hexose nucleus to which amino sugars are attached by glycosidic linkages. This is why these molecules are also referred to as aminocyclitols or aminoglycosidic aminocyclitols. The aminoglycosides can be divided into four groups on the basis of the type and substitution pattern of their aminocyclitol molecule: derivatives containing the aminocyclitol streptidine (e.g., streptomycin and dihydrostreptomycin), derivatives containing the aminocyclitol streptamine (e.g., spectinomycin), derivatives containing a  4,5-­disubstituted deoxystreptamine moiety (e.g.,  neomycin), and derivatives containing a 4,6-­disubstituted deoxystreptamine moiety (e.g., gentamicin, kanamycin, amikacin, tobramycin).

Mechanism of Action Aminoglycosides must penetrate the bacteria to assert their effect. Penetration can be enhanced by the presence of a drug that

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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Aminoglycosides and Aminocyclitols

interferes with cell wall synthesis, such as a beta-­lactam antibiotic. Susceptible, aerobic Gram-­negative bacteria actively pump the aminoglycoside into the cell. This is initiated by an oxygen-­dependent interaction between the antibiotic cations and the negatively charged ions of the bacterial membrane lipopolysaccharides. This interaction displaces divalent cations (Ca++, Mg++), which affects membrane permeability. Once inside the bacterial cell, aminoglycosides bind to the 30S ribosomal subunit and cause a misreading of the genetic code, interrupting normal bacterial protein synthesis. This leads to changes in the cell membrane permeability, resulting in additional antibiotic uptake, further cell disruption, and, ultimately, cell death. The extent and types of misreading vary because different members of the group interact with different proteins. Streptomycin acts at a single site but the other drugs act at several sites. Other effects of aminoglycosides include interference with the cellular electron transport system, induction of RNA breakdown, inhibition of translation, interference with DNA metabolism, and damage to cell membranes. The bactericidal effect is through the formation of abnormal cell membrane channels by misread proteins. Aminoglycoside action is bactericidal and dose (concentration) dependent. For example, gentamicin concentrations in the range of 0.5–5.0 μg/ml are bactericidal for Gram-­ positive and some Gram-­negative bacteria. At  10–15 μg/ml, gentamicin is effective against the more resistant bacteria such as Pseudomonas aeruginosa, Klebsiella pneumoniae, and Proteus mirabilis. The clinical ­implication is that high initial doses increase ionic bonding, which enhances the initial concentration-­dependent phase of rapid antibiotic internalization and leads to greater immediate bactericidal activity. Human clinical studies demonstrate that proper initial therapeutic doses of aminoglycosides are critical in reducing mortality from Gram-­negative septicemia.

For antimicrobials whose efficacy is concentration dependent, high plasma concentration levels relative to the MIC of the pathogen (Cmax:MIC ratio, also known as the inhibitory quotient or IQ) and the area under the plasma concentration-­time curve that is above the bacterial MIC during the dosage interval (area under the inhibitory curve, AUIC = AUC/ MIC) are the major determinants of clinical efficacy. For the aminoglycosides, a Cmax:MIC ratio of 10  was suggested to achieve optimal efficacy (McKellar et  al.,  2004). Recent work suggests that the area under the plasma ­concentration–time curve (AUC)/MIC ratio may be a more reliable indicator of bacterial killing and clinical efficacy (Bland et al., 2018). This human study suggests that an AUC/MIC ratio of 30–50 for aminoglycoside therapy may be effective for noncritically ill/immunocompetent patients with less serious Gram-­negative infections such as urinary tract infections. An AUC/ MIC target of 80–100  may be prudent when treating patients with aminoglycoside monotherapy or in critically ill patients with serious Gram-­negative infections such as nosocomial pneumonia. Further evidence is needed in veterinary patients to determine the efficacy and safety of this PK/PD approach. The aminoglycosides have a significant postantibiotic effect (PAE) – the period of time where antimicrobial concentrations are below the bacterial MIC but the antimicrobial-­ damaged bacteria are more susceptible to host defenses. The duration of the PAE tends to increase as the initial aminoglycoside concentration increases.

Antimicrobial Activity The antimicrobial action of the aminoglycosides is directed primarily against aerobic, Gram-­negative bacteria. Because bacterial uptake is oxygen dependent, they are not active against facultative anaerobes or aerobic bacteria under anaerobic conditions. They are less potent in hyperosmolar environments or low pH environments. Purulent debris at the

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infection site binds ionically to aminoglycosides and inactivates them. They are active against some Gram-­positive bacteria, such as Staphylococcus spp. Emerging strains of methicillin-­resistant Staphylococcus aureus (MRSA) and Staphylococcus pseudintermedius (MRSP) typically retain susceptibility to gentamicin and/or amikacin. Enterococci are inherently resistant (Chapter  2), and therapy against streptococci is more effective when combined with a beta-­lactam antibiotic. Salmonella and Brucella spp. are intracellular pathogens and are often resistant. Some mycobacteria, spirochetes, and mycoplasma are susceptible. In potency, spectrum of activity, and stability to enzymes from plasmid-­mediated resistance, amikacin > tobramycin > gentamicin > ­neomycin = kanamycin > streptomycin. Amikacin was developed from kanamycin and has the broadest spectrum of activity of the aminoglycosides. It is effective against Gram-­ negative strains not susceptible to other aminoglycosides because it is more resistant to bacterial enzymatic inactivation. It is also considered the least nephrotoxic, but it is less efficacious against streptococci than gentamicin. Streptomycin and dihydrostreptomycin are the most active of these drugs against mycobacteria and Leptospira and the least active against other organisms. Paromomycin is effective against some protozoa and cestodes and is the only aminoglycoside with clinically important antileishmanial activity. The bactericidal action of the aminoglycosides on aerobic Gram-­negative bacteria is markedly influenced by pH, being most active in an alkaline environment. Increased local acidity secondary to tissue damage or bacterial destruction may explain the failure of aminoglycosides to kill usually susceptible pathogens. Another factor affecting activity is the presence of purulent debris, which ionically binds to aminoglycosides and inactivates them. When using an aminoglycoside to treat purulent infections (e.g., abscesses), surgical debridement and/or drainage increases efficacy.

Resistance to Aminoglycoside Antibiotics The most common mechanism of bacterial resistance to aminoglycosides is chemical modification by aminoglycoside-­modifying enzymes (AMEs) (Garneau-­Tsodikova and Labby, 2016). The AMEs originally had roles in normal cellular metabolism but have evolved to modify aminoglycosides with selective pressure from exposure to these antibiotics. The AMEs consist of three subclasses: aminoglycoside N-­acetyltransferases (AACs), amino­ glycoside O-­nucleotidyltransferases (ANTs), and aminoglycoside O-­phosphotransferases (APHs). Each AME modifies an aminoglycoside at a specific position, such as an exposed hydroxyl or amino groups to prevent ribosomal binding, and this information is included in the enzyme name. Bifunctional enzymes exist and are capable of multiple types of aminoglycoside modification. AAC(6′)-­Ib is the most prevalent and clinically relevant AME, with approximately 50 variants of AAC(6′)-­Ib in numerous Gram-­negative bacterial species. The AMEs are present in the periplasmic space of bacteria, so that extracellular inactivation of drug does not occur. The genes for AMEs are highly mobile; they are transferred on plasmids, integrons, transposons, and other transposable gene elements, often along with other resistance genes (such as beta-­lactamases). A single type of plasmid may confer cross-­resistance to multiple aminoglycosides and to other unrelated antimicrobials. A single bacterial isolate may have any one of a variety of combinations of resistance to different antibiotics conferred by the particular plasmid it carries. For example, one E. coli strain may be simultaneously resistant to ampicillin, apramycin, chloramphenicol, gentamicin, kanamycin, sulfonamide, streptomycin, tetracycline, and trimethoprim. Antimicrobial resistance in organisms such as E. coli and Salmonella species is a focus of international research due to potential transference of antimicrobial resistance from

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­General Consideration 

Aminoglycosides and Aminocyclitols

animal to human pathogens. Several strategies are being investigated to overcome resistance caused by AMEs, including regulating AME expression, designing new aminoglycosides that evade AMEs, and designing of AME inhibitors. Aminoglycosides target the A-­site of the bacterial ribosome to disrupt protein translation. Other acquired mechanisms of resistance include mutations of the ribosome or enzymatic modifications of the ribosome. The ­bacterial A-­site is located on the 16S RNA of the 30S bacterial ribosomal subunit. One known mechanism of aminoglycoside resistance occurs from mutations in the rrs gene, which codes for 16S rRNA, that hinders aminoglycoside binding. The aminoglycoside binding site may also be modified enzymatically by 16S ribosomal RNA methyltransferases (RMTases). The RMTases naturally occur in actinomycetes, the bacteria from which aminoglycosides were originally isolated. The RMTases are acquired by other bacteria by uptake of a plasmid containing the RMTase gene. The clinical prevalence of RMTases is low but increasing. This is a considerable threat to human and veterinary medicine because RMTases confer resistance to many clinically relevant aminoglycosides, including amikacin. In order to have their bactericidal activity, aminoglycosides must cross the bacterial cell wall. The structure of the cell wall of Gram-­ negative bacteria confers innate resistance to aminoglycosides. The outward facing half of the outer membrane of Gram-­negative bacteria consists of sugar-­functionalized lipopolysaccharides (LPSs), with a net negative charge, attracting cationic aminoglycosides. The most common LPS modification leading to reduced aminoglycoside uptake is the incorporation of the positively charged 4-­amino-­4-­deoxy-­L-­arabinose sugar, which effectively reduces the net negative charge of the LPS layer, decreasing affinity for aminoglycosides. Bacterial strains with reduced cell wall permeability and consequently 2–4-­fold increases in MIC may be selected during

treatment with aminoglycosides. Such strains show cross-­resistance to all other drugs within the group. If the aminoglycoside manages to penetrate the bacterial cell wall, active expulsion by efflux pumps may prevent effective intracellular concentrations. There are five families of efflux systems: the major facilitator super­ family, the ATP-­binding cassette family, the resistance-­nodulation division family (RND), the small MDR family, and the multidrug and toxic compound extrusion family. The majority of genes for efflux mechanisms are located on the chromosome, but members of the major facilitator superfamily are also located on plasmids. While the contribution of efflux pumps to aminoglycoside resistance is low overall, efflux pump expression may be used to monitor resistance to other classes of antimicrobials as a biomarker for determining how bacteria become multidrug resistant (Swick et al., 2011). Chromosomal mutation resulting in resistance is relatively unimportant except for streptomycin and dihydrostreptomycin, where it occurs readily as a result of a single-­step ­mutation to high-­level resistance. For the other drugs, chromosomal resistance develops slowly, because there are many 30S ribosomal binding sites. Both subinhibitory and inhibitory aminoglycoside concentrations produce resistance in bacterial cells surviving the initial ionic binding. This first-­exposure adaptive resistance is due to upregulation of aminoglycoside efflux pumps (Sidhu et  al.,  2012). Exposure to one dose of an aminoglycoside is sufficient to ­produce resistant variants of an organism with altered metabolism and impaired aminoglycoside uptake. In vitro, animal and clinical studies show that the resistance occurs within 1–2  hours of the first dose. The duration of adaptive resistance relates directly to the half-­ life of elimination of the aminoglycoside. With normal aminoglycoside pharmacokinetics, the resistance may be maximal for up to 16 hours after a single dose, followed by partial return of  bacterial susceptibility at 24 hours and

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complete recovery at 40 hours (Barclay and Begg,  2001). The clinical significance of this phenomenon is that frequent dosing or constant infusion of an aminoglycoside is less effective than high-­dose, once-­daily dosing.

Pharmacokinetic Properties Aminoglycosides are poorly absorbed from the normal gastrointestinal tract, but are well absorbed after IM or SC injection. Following parenteral administration, effective concentrations are obtained in synovial, perilymph, pleural, peritoneal, and pericardial fluid. When given to neonates or animals with enteritis, oral absorption may be significantly increased and result in violative tissue residues in food  animals. When given by intrauterine or in­tramam­mary infusion to cows, gentamicin is well absorbed and results in prolonged tissue residues. Aminoglycosides bind to a low extent to plasma proteins (less than 25%). As they are large molecules and highly ionized at physiological pH values, they are poorly lipid soluble and have limited capacity to enter cells and penetrate cellular barriers. These drugs do not readily attain therapeutic concentrations in transcellular fluids, particularly cerebrospinal and ocular fluid. The milk-­to-­plasma equilibrium concentration ratio is approximately 0.5. Their apparent volumes of distribution are relatively small (7–10 days), multiple doses per day, acidosis and electrolyte disturbances (hypokalemia, hyponatremia), volume depletion (shock, endotoxemia), concurrent nephrotoxic drug therapy, age (neonates, geriatrics), preexisting renal disease, and elevated trough concentrations (Mattie et al., 1989). Calcium supplementation reduces the risk of  nephrotoxicity. Nephrotoxicity can also be  decreased by feeding a high-­protein/

high-­calcium diet such as alfalfa to treated large animals and diets higher than 25% ­protein to small animals, as protein and calcium cations compete with aminoglycoside cations for binding to renal tubular epithelial cells (Behrend et al., 1994; Schumacher et al., 1991). High dietary protein also increases glomerular filtration rate and renal blood flow, thereby reducing aminoglycoside accumulation. Because nephrotoxicity is related to aminoglycoside accumulation in the renal proximal tubular cells, it is logical that peak concentrations are not related to toxicity and that longer dosage intervals result in less total drug ­contact with the renal brush border membrane. High-­ dose, once-­daily dosing of aminoglycosides is now common in human and veterinary medicine; it takes advantage of the concentration-­ dependent killing and long PAE of these drugs and avoids first-­exposure adaptive resistance and nephrotoxicity. Serum concentrations of aminoglycosides can be monitored to reduce toxicity and ­confirm therapeutic concentrations (Bartal et  al.,  2003). To allow for the distribution phase, blood sampling for the peak concentration is done at 0.5–1 hour after administration and the trough sample is usually taken prior to the next dose. The peak and trough concentrations can then be used to estimate

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Tubular lumen

Renal tubule epithelial cell

Brush border

Aminoglycoside cations

Ruptured lysosome

Lysosome

Tubular lumen with aminoglycoside cations Phospholipid anions Cell lumen

Figure 13.1  Aminoglycoside cations interact with phospholipid anions on the brush border of renal tubule epithelial cells. Then they are pinocytosed and accumulate in lysosomes until they cause the lysosome to rupture, which destroys the cell.

the elimination half-­life for the individual patient. An increase in the elimination half-­ life during therapy is a very sensitive indi­ cator of early tubular insult. If using a once-­daily regimen, a blood sample just prior to the next dose may be below the recommended trough concentrations and may even be below the limit of detection of the assay. For these patients, an eight-­hour postdose sample will provide a more accurate estimate of the elimination half-­life. Serum concentrations of drug should be 0.5–2 μg/ml before the next dose (gentamicin, tobramycin) or less than 6 μg/ml for amikacin. If therapeutic drug monitoring is unavailable, then nephrotoxicity is detected by an increase in urine gamma-­glutamyl transferase (GGT) enzyme and an increase in the urine GGT/urine creatinine (Cr) ratio (van der Harst et al., 2005). The UGGT/UCr may increase to 2–3 times baseline within three days of a nephrotoxic dose. If these tests are not

available, the development of proteinuria is the next best indicator of nephrotoxicity and it is easily determined in a practice setting. Elevations in serum urea nitrogen and Cr ­confirm nephrotoxicity, but are not seen for seven days after significant renal damage has occurred. Elimination half-­lives of 24–45 hours have been reported in horses with renal toxicity, further prolonging the toxic exposure to the drug. While peritoneal dialysis is useful in lowering creatinine and serum urea nitrates, it may not be effective in significantly increasing the elimination of the accumulating aminoglycoside. The animal’s ability to recover most likely depends on the type of medication exposure and the amount of healthy renal tissue remaining to compensate. Aminoglycoside ototoxicity occurs from the  same mechanisms as nephrotoxicity. The tendency to produce vestibular damage (streptomycin, gentamicin) or cochlear damage (amikacin, kanamycin, neomycin) varies with

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­General Consideration 

Aminoglycosides and Aminocyclitols

the drug. Tobramycin appears to affect both vestibular (balance) and cochlear (hearing) functions equally. This drug-­specific toxicity may be due to the distribution characteristics of each drug and concentration achieved in each sensory organ. The ototoxic effect of aminoglycosides is potentiated by the loop diuretics furosemide and ethacrynic acid and probably other diuretic agents. All aminoglycosides given rapidly IV cause bradycardia, reduce cardiac output, and lower blood pressure through an effect on calcium metabolism. These effects are of minor significance (Hague et  al.,  1997). Neuromuscular blockade is a rare effect, related to blockade of acetylcholine at the nicotinic cholinergic receptor. It is most often seen when anesthetic agents are administered concurrently with aminoglycosides. Affected patients should be treated promptly with parenteral calcium ­chloride at  10–20  mg/kg IV, calcium gluconate at 30–60 mg/kg IV or neostigmine at 100–200 μg/kg to reverse dyspnea from muscle response depression. Edrophonium at 0.5 mg/kg IV will also reverse neuromuscular blocking effects.

Dosage Considerations Aminoglycosides produce rapid, concentration-­ dependent killing of Gram-­negative aerobes and a prolonged PAE (McKellar et  al.,  2004). A  maximum plasma concentration (Cmax) to  MIC ratio is associated with efficacy. A Cmax:MIC ratio of 8–12/1 optimizes bactericidal activity. Higher initial serum concentrations may also be associated with a longer PAE. Traditionally, aminoglycosides were administered every 8–12 hours. If the aminoglycoside is dosed multiple times a day or the drug concentration remains constant, as with a continuous infusion, first-­exposure adaptive resistance persists and increases and the risks of nephrotoxicity and ototoxicity increase. Dose administration at 24-­hour intervals, or longer, may increase efficacy by allowing time for adaptive resistance to reverse. Some ­clinicians have expressed reservations about

once-­daily dosing when intestinal damage allows continued exposure to bacteria that may replicate during the prolonged periods of subtherapeutic aminoglycoside concentrations, but this has not been documented clinically. Studies in human and veterinary patients support high-­dose, once-­daily therapy of aminoglycosides (Bauquier et al., 2015; Albarellos et  al.,  2004; Godber et  al.,  1995; Magdesian et al. 1998; Nestaas et al., 2005). However, the optimal doses and the ideal therapeutic drug monitoring strategy are still unknown (Redpath et  al.,  2021). All dosage regimens should take into account the patient’s renal function, the exclusive renal excretion route of aminoglycosides, and their toxicity potential. Neonates typically have a higher percentage of extracellular water than adults; therefore, the volume of distribution of aminoglycosides is higher and they typically require higher dosages than adults.

Clinical Use The toxicity of aminoglycosides has largely restricted their use to the treatment of severe infections. Because there are few alternative treatment options, aminoglycosides are increasingly considered in the treatment of MRSA and  MRSP infections in companion animals (Papich,  2012). The more toxic aminoglycosides (e.g., neomycin) are largely restricted to topical or oral use for the treatment of infections caused by Enterobacterales. The less toxic aminoglycosides are usually reserved for the parenteral treatment of severe sepsis caused by Gram-­negative aerobes and, increasingly, the treatment of methicillin-­resistant staphylococcal infections. Of these, gentamicin is usually the first choice followed by amikacin, which due to expense is reserved for sepsis caused by organisms resistant to gentamicin. But even the expensive aminoglycosides can be used for local therapy of musculoskeletal infections. Antimicrobial-­impregnated polymethyl methacrylate beads, collagen sponges, and regional perfusion (intravenous or intraosseous) provide

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high local concentrations with less expense and reduced risk of systemic toxicity. Because aminoglycoside residues persist in renal tissues for prolonged periods, the extra-­ label use in food animals should be avoided. A voluntary resolution against the extra-­label administration of aminoglycosides has been adopted by the American Association of Bovine Practitioners, the Academy of Veterinary Consultants, the National Cattlemen’s Beef Association and the American Veterinary Medical Association.

S ­ treptomycin/ Dihydrostreptomycin Streptomycin and dihydrostreptomycin are members of the streptidine group. Dihydros­ treptomycin has very similar properties to streptomycin but is more likely to cause ototoxicity. Streptomycin was the earliest aminoglycoside introduced for clinical use.

Antimicrobial Activity Streptomycin and dihydrostreptomycin are active against mycobacteria, some mycoplasma, some Gram-­negative rods (including Brucella), and some Staphylococcus aureus. With the exception of mycobacteria, streptomycin is the least active of the aminoglycosides. Among ­susceptible ­bacteria are Leptospira, Francisella tularensis, Yersinia pestis, and most Campylobacter fetus ssp. venerealis. Organisms with MIC MIC and fAUC/MIC can be predictive of efficacy in vete­rinary species, where f describes the free (protein-­unbound) drug concentration (Toutain et  al.,  2021). The Clinical and Laboratory Standards Institute (CLSI) considers only fAUC/ MIC values of ≥25 to be predictive for the clinical efficacy of the tetracyclines. However, AUC/ MIC values ranging from 12 to 819 are reported

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depending on the species treated, the bacterial pathogens targeted, and the tetracycline used. Values also vary depending on the bacteriological effect and the body site targeted. Some studies suggest T>MIC is a more predictive index than AUC/MIC (Li et  al.,  2021; Zhang et  al., 2019). The fAUC/MIC is the preferred PK/PD index for long-­acting oxytetracycline injectables. The tetracyclines tend to have postantibiotic effects for 2–3 hours for both Gram-­positive and Gram-­negative pathogens.

Resistance to Tetracyclines The widespread use of tetracyclines in food animal production contributes to the worldwide spread of resistance in Gram-­positive and Gram-­negative bacteria and Mycoplasma spp. Resistance to tetracyclines is due to the ­acquisition of mobile genetic elements carrying tetracycline-­specific resistance genes, mutations within the ribosomal binding site, and/or chromosomal mutations leading to increased expression of intrinsic resistance mechanisms. Currently, over 1000 resistance genes are reported, including mosaic genes. Mosaic tetracycline resistance genes are a subgroup of the genes encoding ribosomal protection proteins (RPPs). They are formed when two or more RPP-­encoding genes recombine in a bacterium, resulting in a functional chimera. The following mechanisms of resistance are described, with the first two mechanisms being the most common: (1) energy-­dependent efflux pumps, most of which are antiporters that exchange an extracellular H+ for a cytoplasmic tetracycline-­Mg2+ complex; (2) synthesis of ribosomal protection proteins that dissociate the tetracycline from the binding site near the ribosomal AA-­tRNA docking site; (3) synthesis of antibiotic-­inactivating enzymes; (4) ribosomal 16S RNA mutation at the primary binding site of tetracyclines; (5) stress-­induced downregulation of the porins through which the drug crosses the outer Gram-­negative wall; and (6) multidrug transporters with similar mechanisms to efflux pumps.

Because of the absence of the dimethylamine group on carbon 4, the CMTs do not select for antimicrobial resistance (Figure 14.2). The oral administration of tetracyclines results in the fecal excretion of resistant bacteria as well as unchanged drug. The presence of tetracyclines in the environment selects for resistance in bacterial populations and alters microbial activities such as the decay process. While the tetracyclines are usually degraded with half-­lives ranging from less than 1 hour to 22 days, their persistence for 180 days in soil to six months in marine sediment has been reported. Composting contaminated manure may efficiently accelerate the degradation of the tetracyclines but the degradation products of some tetracyclines remain biologically active (Granados-­Chinchilla and Rodríguez, 2017).

Pharmacokinetic Properties In addition to the usual PK factors (e.g., molecular size, lipid solubility, degree of protein binding), the pharmacokinetics of tetracyclines can be greatly affected by exposure to multivalent cations (e.g., Ca2+, Mg2+, Zn2+, Cu2+, Fe2+, Fe3+, Al3+) and interactions with P-­glycoprotein (P-­gp) transporters. There are also wide species variations in the PK parameters. The oral bioavailability of the first-­generation tetracyclines is relatively low. If calcium, magnesium, iron or aluminum ions are present within the gastrointestinal tract, oral bioavailability is markedly reduced. Oral bioavailability is improved in fasted animals, but this is not a clinically feasible adjustment for sick animals. Large animal formulations are often administered by IM or SC injection. The type of salt, solvents or vehicles present in the formulation and the pH of the products influence the bioavailability. The extent of protein binding of the tetracyclines is highly variable and dependent on the tetracycline, the species treated, and the type of proteins present in the biological media. Several tetracyclines display an atypical nonlinear binding to serum protein, where the free

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­General Consideration 

Tetracyclines

fraction of the tetracycline decreases at higher total plasma concentrations, in contrast with what occurs with drugs for which binding is saturable (Toutain et  al.,  2021). Free tetracycline molecules circulate mostly as Ca2+ and Mg2+ chelates, which may facilitate the bacterial penetration. The ability to cross biological membranes varies between the tetracyclines. While a high degree of protein binding tends to limit tissue distribution, lipophilicity favors passage across cell membranes. Therefore, despite extensive binding to plasma proteins, the second-­ generation tetracyclines have more extensive tissue penetration than the first-­generation tetracyclines. The tetracyclines have generally good distribution in highly vascularized tissues without anatomical barriers (e.g., liver, gallbladder, salivary gland, seminal vesicles, kidney, heart, lungs, endometrium), reaching higher concentrations than in plasma. Diffusion into interstitial fluid such as pulmonary epithelial fluid, peritoneal and pleural fluids can be variable. The distribution in intermediately vascularized tissues, such as muscle and skin, is variable but can reach higher concentrations than in plasma. The concentrations reached in poorly perfused tissues and/or those with anatomical barriers (e.g., adipose tissue, joints, tendons, brain, CSF, prostate, ocular tissues) is generally low; however, moderate to high concentrations are achieved in milk. The passage of tetracyclines across the blood–brain barrier and placenta may be influenced by their affinity to P-­gp transporters. In infected tissues, the concentrations of tetracyclines can be higher than in healthy tissues, such as in infected lungs or in mastitic milk. The tetracyclines are among a limited number of osteotropic drugs and deposit in teeth or in sites of new bone formation. Tetracyclines are eliminated by glomerular filtration, biliary secretion, and intestinal excretion. As efflux transporters, P-­gp has a role in the elimination of some tetracyclines. The elimination of the second-­generation tetracyclines is

mainly by nonrenal routes, but the concentrations reached in urine and bile are higher than plasma concentrations for all tetracyclines. Enterohepatic recirculation contributes to the unusually long elimination half-­life for drugs mainly eliminated by renal route, and can cause a second plasma peak that has been detected in sheep, horses, and donkeys following parenteral administration (Chapuis et al., 2021a, b). Pharmacokinetic parameters of the tetracyclines can vary between species of a same genus, such as reported between adult horses, donkeys, and ponies. Young animals or neonates can have different PK parameters from adults, but the variation is species dependent. Comparing results of studies in ruminants, the stage of maturity of rumen function affects the PK of some tetracyclines even when administered parenterally.

Drug Interactions The oral absorption of the tetracyclines is impaired by antacids containing multivalent cations, such as magnesium and aluminum, by iron-­containing preparations such as ferrous sulfate, and by bismuth subsalicylate. Manipulation of the P-­gp transporters by tetracyclines can be done deliberately to change drug PK. For example, DXC was effective at inhibiting the efflux of ivermectin and doramectin in alpaca cells, which may improve CNS concentrations for more effective treatment of parelaphostrongylosis (Agbedanu et al., 2015). Evidence of synergism of tetracyclines with other antimicrobials is scant. Studies of synergistic effects are mainly performed in  vitro, using models, or evaluating the correlation of mass administration of medicated feed and ­animal performance. DXC and MIN showed in  vitro synergism when combined with clarithromycin against R. equi (Erol et al., 2022). As tetracyclines are typically bacteriostatic, they are considered antagonistic to bactericidal drugs that target actively growing cells (Ocampo et al., 2014).

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Toxicity and Adverse Effects The tetracyclines may impair urinary concentration function by inhibiting enzymes of the renal medulla, or by disturbing the response of antidiuretic hormone causing nephrogenic diabetes insipidus. The second-­generation tetracyclines are considered safer than the first-­generation drugs regarding nephrotoxicity, and some references suggest that it is unnecessary to adapt the dosage when these drugs are administered to patients with renal failure. The parenteral administration of tetracyclines disturbs the intestinal microbiota and is associated with gastrointestinal signs and hepatopathies. Tetracyclines form chelates with calcium in bone and teeth, resulting in discoloration of teeth, enamel defects, growth retardation of skeleton of fetuses and young animals, and changes in bone density. The tetracyclines may cause photosensitization in light-­pigmented animals.

C ­ hlortetracycline, Tetracycline, and Oxytetracycline Formulations Oral formulations of CTC, TTC, and OTC are available as premixes for medicated feeds for food animals and for milk replacer for calves. Water additives of TTC and OTC for food animals are also available. TTC is available as human and small animal tablets or capsules and as boluses and intramammary products for large animals. OTC is available in a number of formulations for parenteral use in large animals; the composition of the carrier solution determines the absorption and elimination from IM or SC injection sites (propylene ­glycol  – “short-­acting”, polyethylene glycol and 2-­pyrrolidone – “long-­acting”).

Pharmacokinetics The oral absorption of CTC, TTC, and OTC varies widely between species and is greatly affected by complexation with multivalent

cations that precipitate as pH increases (e.g., calcium in dairy products) and by food particles (Decundo et al., 2019). In pigs, feed did not affect the bioavailability of OTC (3% in both fasted and fed pigs); for TTC there was a significantly higher bioavailability in fasted (18%) than in fed (5%) pigs while for CTC the bioavailability was not significantly different between fasted (11%) and fed pigs (6%) (Nielsen and Gyrd-­Hansen, 1996). Therefore, therapeutic concentrations in plasma or tissues are not achieved after oral administration of any of the three tetracyclines to fed or fasted pigs and any benefit of oral administration is due to impact on gastrointestinal flora. Poor oral bioavailability also occurs in poultry and ruminants fed tetracyclines. Exposing the microbiota to high concentrations of the tetracyclines enhances development of drug-­resistant strains and the lack of absorption also results in tetracycline dissemination in the environment (Ricker et al., 2020). The IM or SC administration of long-­acting OTC formulations results in prolonged plasma and tissue concentrations. Vehicles such as polyethylene glycol, propylene glycol, povidone or pyrrolidone are nonionic surfactants that slow the absorption phase at the injection site and prolong the elimination rate. If injected IV, the long-­acting formulations show the same PK profile as the short-­acting formulations. When long-­acting formulations are injected IM or SC, the rate of absorption is initially high from vasodilation at the injection site. Then the rate of absorption decreases, from tissue damage and fibrosis. Therefore, the absorption phase controls the elimination phase, resulting in the “flip-­flop” phenomenon (Dowling and Russell,  2000). This increases the time between subsequent administrations, hence the term “long-­acting.” However, these formulations result in low concentrations in plasma and persistence of OTC at the injection site and in other tissues. Overall, this results in prolonged subtherapeutic concentrations and prolonged residues in food animals. Therefore, the use and development of long-­acting

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Tetracyclines

formulations are not consistent with good antimicrobial stewardship. The volume of distribution values range from 0.7 to 15 l/kg for CTC; from 1 to 5 l/kg for TTC; and from 0.6 to 7  l/kg for OTC. Plasma protein binding is moderate and varies between drug and species. Distribution of OTC in milk of dairy cattle is proportional to plasma concentrations, with a faster rate of distribution after IV administration compared to IM. The tetracyclines are well distributed to most tissues, except the CNS. Therapeutic levels may be achieved, however, with IV administration when the meninges are inflamed. OTC preferentially accumulates in the lungs of pneumonic calves. The multivalent cation-­chelating properties of the first-­generation tetracyclines cause deposition in egg shells, teeth, and bone. The tetracyclines undergo little hepatic metabolism and metabolites produced are inactive. Metabolites are mostly of concern in eggs from laying hens. Some jurisdictions set the maximum residue limit for tetracyclines based upon the sum of the concentrations of parent drug and its 4-­epimer. Unchanged drugs are excreted by renal and biliary routes. Renal clearance is related to glomerular filtration. Greater than 40% appears in the feces after biliary elimination and most drugs have some enterohepatic circulation. When administered orally, the mean residence times and elimination half-­lives range from three hours to 37 hours for CTC; from five to 16  hours for TTC; and from three hours to 28 hours for OTC. When administered IV, the elimination half-­life of OTC is typically 6–8 hours. Due to “flip-­flop” kinetics, the elimination half-­life is typically 22–24 hours after IM or SC administration of long-­acting OTC products.

Toxicity and Adverse Effects Anaphylactic reactions and injection site ­reactions may occur in cattle treated IM or SC with long-­acting OTC formulations. Rapid IV  administration of OTC causes a

dose-­dependent cardiotoxic effect in cattle, resulting in collapse, ataxia, and dyspnea. This is attributed to intravascular chelation of calcium, decreased blood pressure from the drug vehicle (propylene glycol), and the speed of administration. To prevent collapse, the IV injection of OTC should be administered over at least five minutes. Shivering, ataxia, dyspnea, and collapse were reported as acute reactions to OTC in horses; the vehicle used in the commercial preparation was also suspected to play a role in the adverse events. For the treatment of contracted tendons in foals, high doses of OTC were associated with cardiotoxicity, weakness, profuse sweating, diaphragmatic flutter, convulsion, rhabdomyolysis, and renal failure (Ellero et  al.,  2020). The rapid administration of OTC can cause rapid hypotension, bradycardia, and electrocardiographic abnormalities in cats and dogs, as well as dyspnea in cats. Nephrotoxicity is associated with dose-­ related functional changes in renal tubules, which may be exacerbated with the administration of high doses of OTC, or dehydration, hemoglobinuria, myoglobinuria, toxemia, or the co-­administration of nephrotoxic drugs (Ellero et al., 2020). Transient hemoglobinuria with trembling and subnormal temperatures is reported in cattle administered long-­acting OTC, but propylene glycol-­based products may also cause intravascular hemolysis. Fatal nephrotoxicity is reported in dogs administered an overdose of OTC. Immediate response included lacrimation, facial scratching, and depressed mentation; when the overdose was repeated, emesis, diarrhea, bradycardia, oliguria or anuria occurred. Dilution of OTC and slow administration prevented renal failure in dogs. Long-­term administration of tetracyclines may cause uroliths in dogs. In horses, acute colitis was reported following the parenteral administration of OTC or the consumption of feed contaminated with TTC. However, adverse effects of parenteral OTC were also associated with excessive dosage, concomitant use of other antimicrobials,

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and stressors such as surgery and transport. OTC is now the recommended treatment for colitis cause by Neorickettsia risticii (Potomac horse fever). In guinea pigs, CTC may cause clostridial enterotoxemia but its use is reported without adverse effects. Tetracyclines can rarely cause idiosyncratic hepatic necrosis in dogs and cats and augment hepatocellular lipid accumulation in many species. Esophageal stricture and gastric irritation are reported in cats receiving PO TTC tablets or capsules. TTC is associated with drug-­induced pyrexia in cats. The IM or SC administration of OTC is associated with mild to severe tissue irritation, depending on the species and the formulation administered, as solvents such as aqueous-­2-­ pyrrolidone, N-­methylpyrrolidone, glycerol formaldehyde, polyvinylpyrrolidone, propylene glycol, and dimethylacetamide are irritant substances. In general, long-­acting formulations of OTC should not be administered IM to (A)

horses; however, the formulation with polyethylene glycol vehicle is minimally irritating. Because of their retention in bone, TTC, CTC, and OTC cause brilliant yellow-­gold fluorescence under ultraviolet light in long bones and flat bones (Warner et  al.,  2022) (Figure 14.3). Egg shells of laying hens fed tetracyclines will also fluoresce and this can be used as a screening method to determine if eggs were laid during a medication period (Zurhelle et al., 2000). Administration of first-­ generation tetracyclines to growing puppies or pregnant females results in yellow discoloration of primary and permanent teeth of the puppies (Boy et al., 2016).

Dosage Considerations The tetracyclines are labelled for parenteral (IV, IM, SC) and topical (intrauterine, intramammary, cutaneous ointments or spray, ophthalmological ointments) administration. (B)

Figure 14.3  (A) Photograph under ultraviolet light of bones from a pig fed chlortetracycline. (B) Photograph under ultraviolet light of the bones from a pig not fed chlortetracycline.

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Intraperitoneal and subconjunctival administration of OTC are reported but may cause severe tissue irritation. The mass administration of tetracyclines through free-­choice medicated feed or water to food animals should be reconsidered even if products are labeled for such uses. Such use results in unpredictable and significant inter-­ and intraindividual variation in oral absorption. Reduced feed or water intake in sick animals may further contribute to the low plasma concentrations. Additionally, as milk decreases the bioavailability of oral tetracyclines, the reconstituting of tetracyclines in milk or milk replacer is not recommended despite being labeled for this in many countries.

Clinical Usage Ruminants

Tetracyclines are labeled for both the treatment and the prophylaxis/metaphylaxis of bovine respiratory disease, but antimicrobial resistance is very common in these pathogens. The efficacy of prophylactic/metaphylactic administration can be moderate and highly variable, and such use should be reconsidered from the perspective of antimicrobial stewardship (Baptiste and Kyvsgaard, 2017). In sheep and goats, OTC is used for the treatment of some respiratory diseases. The emergence of resistance, including in the enteric microbiota, is also reported in sheep receiving oral tetracyclines. While tetracyclines are labeled for the management of diarrhea in calves or lambs in some countries, there are high rates of antimicrobial resistance (Zhang et al., 2022). Labels exist for the treatment of enterotoxemia caused by Clostridium perfringens in lambs, but evidence of clinical efficacy is lacking. In many countries, OTC is labelled for the systemic and/or intramammary treatment of mastitis in cattle and small ruminants. Tetracyclines may be useful in the treatment of Brucella spp. and other reproductive diseases in ruminants. Tetracyclines fail to

eliminate the shedding of Chlamydophila abortus in sheep but are potentially efficacious in the prevention of enzootic abortion if started early in an outbreak. As PK/PD targets are not met, oral CTC is not likely efficacious for the prevention of Campylobacter-­ associated abortion in sheep even if higher than approved dosages are used (Washburn et al., 2018). Parenteral OTC may control leptospirosis in dairy cattle. Intravenous OTC may be efficacious in the treatment of bacterial meningitis in adult cattle or encephalitic listeriosis, with better success rates in cattle than in small ruminants. Tetracyclines can be used in the management of otitis media/interna in calves, especially from Mycoplasma spp. Oxytetracycline and CTC are labeled for the control of bovine anaplasmosis from A. marginale. They reduce bacteremia but do not eliminate the organism. Even if recommended, chronic administration increases selection for resistance in Anaplasma. OTC may be efficacious in the treatment of tick fever (A. phagocytophilum) in cattle and lambs. Tetracyclines may be effective in the treatment of heartwater disease (Ehrlichia ruminantium). The use of tetracyclines in the management of parasitic infection from Babesia bovis and B. bigemina (redwater) or East Coast fever (Theileria parva) reduces parasitemia and red blood cell destruction without inhibiting the development of immunity, but antiprotozoal drugs are preferred for treatment. The parenteral administration of OTC is labeled for the management of infectious keratoconjunctivitis (“pink eye”) in cattle with proven clinical efficacy (Angelos,  2015). Clinical efficacy is also reported in sheep but eradication of the infection can be incomplete. Tetracyclines are efficacious for treatment of bovine and ovine dermatophilosis. The tetracyclines are efficacious in the treatment of Fusobacterium necrophorum infections such as necrotic laryngitis in calves, interdigital necrobacillosis in ruminants, and abscesses. Their efficacy is not clear in the treatment of bovine

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papillomatous digital dermatitis or contagious ovine dermatitis (Apley,  2015; Duncan et  al., 2014). Milk residues are possible if tetracycline products are applied topically to the pasterns of lactating cows with digital dermatitis (Cramer et al., 2019). Camelids and Camels

Tetracyclines are widely used in camels and camelids for a variety of diseases. Camels and camelids show similarity in pharmacokinetics of these drugs. In some countries, OTC products are labeled for camels. OTC can be used to treat acute hemolysis from Candidatus Mycoplasma haemolamae in camelids, although treated animals remain chronic carriers. Swine

Despite their very poor oral bioavailability, the tetracyclines are widely used in many countries in the management of atrophic ­rhinitis and bacterial causes of porcine ­respiratory disease complex, but efficacy is difficult to determine (O’Connor et al., 2019; Sargeant et al., 2019). CTC in medicated feed is labeled in the control of porcine proliferative enteropathies (Lawsonia intracellularis) with reported efficacy in the literature; TTC and OTC are also reported as efficacious. CTC is also approved in some countries for the treatment of bacterial enteritis caused by Escherichia coli and Salmonella choleraesuis. However, antimicrobial resistance is common in these bacteria (Barton, 2014). Treatment and control of leptospirosis in swine should include a combination of vaccination, medication, and management changes. Successful use of oral tetracyclines to reduce and eliminate leptospire shedding in pigs has been reported. However, such approaches should be limited to outbreak situations and comply with regulatory requirements. Tetracyclines may decrease the incidence of acute hemolytic form of Mycoplasma suis infection, but pigs can become chronic carriers.

Poultry and Other Birds

Oral tetracyclines are labeled for chickens in many countries for the control of chronic respiratory disease and air sac infections caused by Mycoplasma gallisepticum and Escherichia coli, infectious synovitis caused by Mycoplasma synoviae, and in turkeys for the treatment infectious synovitis caused by Mycoplasma synoviae and secondary bacterial infection associated with transmissible enteritis (bluecomb). Oral tetracyclines are used to treat fowl cholera caused by Pasteurella multocida. Oral CTC is used to treat pleuropneumonia infection from Mycoplasma gallisepticum in chickens, while TTC and OTC are ineffective. CTC in medicated feed is used to treat avian chlamydiosis from Chlamydophila psittaci. In the EU and some other countries, tetracycline products are also labeled for homing pigeons, ornamental birds, common canaries, parrots, geese, and ducks. Horses

Despite historical concerns over inducing colitis in horses, IV OTC has become the treatment of choice for infection from Neorickettsia risticii (Potomac horse fever) and Anaplasma phagocytophilum (equine granulocytic anaplasmosis) (Oliver et  al.,  2023; Taylor,  2023). Parenteral OTC is one of several suggested treatments for Lawsonia intracellularis infection causing equine proliferative enteropathy in horses. Tetracyclines are commonly used to treat equine Lyme disease from Borrelia burgdorferi infection (Divers et al., 2018). Dogs and Cats

Tetracycline can be used for the treatment of bacterial urinary tract infection caused by E. coli and Staphylococcus spp. Tetracyclines are suggested as drugs of choice for the treatment of A. phagocytophilum, Ehrlichia canis, and Rickettsia rickettsii infections in dogs. While the treatments resolve the clinical signs, they may fail to clear the organisms. OTC is the drug of choice for the treatment of salmon poisoning (Neorickettsia helmintheca).

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Other Mammals

In the EU and some other countries, tetracycline products are labeled for rabbits, small rodents, mink, polecats, and other fur-­bearing animals. Indications can be vague and scant data supporting efficacy in these species are available. In mink, indications include the treatment of acute gastroenteritis, including from Salmonella spp. (Pyatnychko et al., 2021). Bees

Oxytetracycline is labeled in many countries for the management of American (Paenibacillus larvae) or European foulbrood (Melissococcus plutonius), but resistance is reported (Masood et al., 2022). The limitations of extended use of tetracyclines include lack of efficacy, drug residues in honey, and reduced longevity of bees. Fish and Crustaceans

The use of OTC HCl or dihydrate is approved in some countries for the treatment of ulcer disease (Haemophilus piscium), furunculosis (Aeromonas salmonicida), bacterial hemorrhagic septicemia and erythrodermatitis (Aeromonas hydrophila), columnaris disease (Flavobacterium columnare), cold-­water ­disease (Flavobacterium psychrophilum), enteric redmouth disease (Yersinia ruckeri), vibriosis (Vibrio anguillarum), piscirickettsiosis (Piscirickettsia salmonis), and Lactococcus garvieae infection in fish such as salmonids, cyprinids, catfish, seabass, sea bream, briss eel, and turbot. There are tetracycline products available in many countries and on the internet without a veterinary prescription for use in ornamental fish. Most concerning is that some of these products are diverted for human use. OTC is also used for skeletal marking to study fish populations, but this use is not consistent with antimicrobial stewardship. OTC is labeled for the management of gaffkemia (Aerococcus viridans) in lobsters. Reptiles

Dosages for OTC or CTC are reported in reptiles but studies are scarce. For the treatment of mycoplasmosis (M. alligatoris) in American alligators, pharmacokinetic studies suggest

that only IM or IV administrations could reach therapeutic concentration.

­Doxycycline and Minocycline Formulation In some countries, veterinary-­approved DXC is available as the hyclate salt as an injectable solution, water soluble or lactodispersable powders, tablets or capsules. In countries without veterinary-­approved products, human formulations are used either as is or in compounded drug products suitable for animal use. DXC gel is available in some countries formulated with a polymer delivery system made of N-­methyl-­2-­pyrrolidone and poly (DL-­lactide) that allows the slow release of DXC in the periodontal pocket of dogs with periodontal disease. Only human formulations of MIN as the hyclate salt are available for extra-­label use in veterinary patients.

Pharmacokinetics Compared to the first-­generation tetracyclines, DXC and MIN have greater oral bioavailability in most species. The oral bioavailability of feed-­administered doxycycline in swine is approximately 22%. Oral DXC is of limited usefulness in horses due to poor oral bioavailability. When doxycycline was combined with poloxamer gel, the oral bioavailability increased in horses but is greatly dependent on the compounding formulation used (Chapuis et  al.,  2021b). Oral DXC or MIN in nursing foals resulted in higher plasma peak and AUC than in adults (Giguère et al., 2017). Overnight fasting and delaying feeding of hay two hours after oral MIN administration improves drug bioavailability and increases plasma concentrations. In dogs, feeding is a significant source of variation in PK parameters so it is recommended to administer MIN without food. Concurrent administration of sucralfate

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tablets significantly decreases MIN absorption. For DXC, concurrent administration of tablets did not affect bioavailability but sucralfate suspension significantly reduced oral absorption. Doxycycline has an higher degree of protein binding (71–98%) than MIN (65–80%) in most species. Pigs have a relatively low degree of DXC protein binding (31%). Values for the volumes of distribution range from 0.3 to 25 l/kg for DXC and from 0.5 to 14 l/kg for MIN. Overall, the second-­generation tetracyclines have better intracellular and tissue penetration than the first-­generation drugs. In dogs, DXC reached higher concentrations in aqueous humor, vitreous fluid, and CSF than MIN, while the latter had higher distribution into the brain. DXC penetrates well into interstitial fluid in sheep and dogs but only achieved low concentrations in an infected tissue cage model in horses (Chapuis et  al.,  2021b). DXC accumulated in the synovial fluid in horses, with a penetration ratio of 4.6 between the AUC of plasma and the synovial fluid. The plasma elimination half-­life ranges from three to 24 hours for DXC and four to 20 hours for MIN. DXC and MIN are eliminated unchanged by both the renal and biliary routes. In rats, dogs, and humans with renal failure, DXC did not accumulate in plasma. While DXC does not undergo significant hepatic metabolism, MIN forms a variety of metabolites, some of which have antimicrobial activity.

Toxicity and Adverse Effects In sheep, the IV administration of DXC was associated with tachypnea, tremors, sialism, and hindlimb weakness. Intravenous administration of DXC caused fatal collapse in horses and ponies (Riond et  al.,  1992). This reaction appears to be due to chelation of intracellular calcium, resulting in neuromuscular blockade of the myocardium. The rapid IV administration of DXC can cause hypotension, bradycardia, and electrocardiographic abnormalities in cat and dogs, as well as collapse and dyspnea in

cats. However, no adverse events were reported with the IV administration of MIN in horses. Intravenous MIN caused severe hypotension and cardiovascular depression in dogs (Maaland et al., 2014). Overdose of oral DXC in calves is associated with anorexia, sialorrhea, dysphagia, arrhythmias, pulmonary distress, and sudden death. Postmortem findings include acute renal tubular necrosis, pulmonary edema, myocardial degeneration, and necrosis and myopathy of tongue and some striated muscles (Brihoum et  al.,  2010). Oral DXC was associated with fatal colitis when administered to fasted adult horses. This may be due to significant changes in fecal microbial diversity (Chapuis et al., 2022). DXC is associated with vomiting, diarrhea, anorexia, and an increase in hepatic enzymes in dogs and cats. Oral DXC and MIN tablets and capsules must be carefully administered to dogs and cats, as failure to pass in to the stomach may result in severe esophagitis, esophageal necrosis, and perforation or stricture formation. Oral MIN may cause hypersalivation and, at higher dosage, emesis in dogs. Despite historical concerns, DXC does not cause tooth discoloration in young children and there is no evidence that it does so in young animals (Stultz and Eiland,  2019). In humans, MIN is noted to cause blue-­green discoloration of teeth, bone, and skin.

Dosage Considerations In North America, DXC is only labeled in dogs as the subgingival gel. Compounded DXC from approved human formulations is convenient for use in small animals and horses, but the stability of compounded products is uncertain (Papich et  al.,  2013). In some countries, DXC anhydrate and hyclate products are labeled for oral administrations in multiple species, including cattle, sheep, goats, swine, poultry, homing pigeons, geese, rabbits, dogs, and cats. MIN is only available as human formulations and is frequently compounded to products suitable for use in small animals.

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­Doxycycline and Minocyclin  285

Tetracyclines

Clinical Applications Ruminants

In some countries, DXC is approved for use in drinking water or milk replacer for preruminant calves to treat bronchopneumonia and pleuropneumonia caused by Pasteurella spp., Streptococcus spp., Trueperella pyogenes, Histophilus somni, and Mycoplasma spp. DXC is used to treat bacterial infections such as pneumonia, skin and soft tissue infections, urinary tract infections, salmonellosis, and colibacillosis in sheep and goats. Swine

Doxycycline in medicated feed or water is used in many countries outside North America for the management of atrophic rhinitis caused by Pasteurella multocida and Bordetella bronchiseptica, bronchopneumonia caused by P. multocida, Streptococcus suis and Mycoplasma hyorhinis, and pleuropneumonia caused by Actinobacillus pleuropneumoniae. Horses

Based on PK studies in normal horses, oral DXC and MIN have been recommended for treatment of Gram-­positive bacterial infections. However, DXC treatment was ineffective in a tissue cage model of infection using a susceptible strain of S. zooepidemicus (Chapuis et al., 2021b). Oral DXC may be useful in the treatment of Lawsonia intracellularis infection (equine proliferative enteropathy) in foals. Oral DXC and MIN are potential options for the treatment of Borrelia burgdorferi infection (Lyme disease) in horses (Divers et al., 2018). Dogs and Cats

Doxycycline is labeled in some countries for the treatment of respiratory diseases; along with MIN, it is advocated as first-­line treatment of respiratory diseases in small animals (Lappin et  al.,  2017). DXC should be efficacious for the treatment of bacterial urinary tract infections in dogs and cats (Wilson et  al.,  2006) and is recommended for the

treatment of leptospirosis in dogs (Schuller et al., 2015). DXC and MIN are drugs of choice for the treatment of canine skin diseases from methicillin-­resistant Staphylococcus pseudintermedius (Hnot et al., 2015). DXC is recommended for the treatment of ehrlichiosis (Ehrlichia canis) and Rocky Mountain spotted fever (R. rickettsia), and may be more effective than OTC in the treatment of salmon poisoning (Neorickettsia helmintheca). Treatment of Lyme disease (Borrelia burgdorferi) in dogs with DXC or MIN is only recommended in clinically affected or proteinuric patients (Littman et al., 2018). Dogs clinically ill from plague (Yersinia pestis) recovered with DXC treatment (Nichols et  al.,  2014). DXC is the first drug of choice for treating Wolbachia, a member of the Rickettsiaceae, which has a symbiotic relationship with heartworms. Treatment with DXC reduces Wolbachia numbers in all stages of heartworms and improves outcomes and decreases microfilaremia in dogs treated for heartworm disease. MIN is a suitable alternative to DXC. Where approved, DXC hyclate 8.5% gel is efficacious for the local treatment of periodontal disease in the dog. MIN in a nanoparticle delivery system placed in periodontal pockets showed clinical efficacy in dogs (Yao et  al.,  2014). The administration of subantimicrobial doses of DXC was efficacious in the management of periodontal inflammation in dogs without altering subgingival microflora and or affecting antimicrobial susceptibility (Kim et al., 2016). In cats, DXC is suggested as the treatment of choice for anaplasmosis, ehrlichiosis, and rickettsiosis (Qurollo, 2019). DXC resolved clinical signs of feline hemoplasmosis but did not clear Mycoplasma haemofelis or Candidatus M. haemominutum in cats (Barker,  2019). In cats, DXC may be a treatment option for Lyme borreliosis (Qurollo,  2019). Despite the zoonotic potential, treatment of cats with Bartonella spp. bacteremia is only recommended in clinically ill cats or healthy bacteremic cats living in a household with immunocompromised people (Álvarez-­Fernández et al., 2018).

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Poultry

In some countries, DXC is approved for use in  drinking water of broilers, breeders, and replacement pullets for infections of the respiratory tract caused by Mycoplasma spp., Escherichia coli, Haemophilus paragallinarum and Bordetella avium, and enteritis caused by Clostridium perfringens and Clostridium colinum. However, a high resistance rate in E. coli isolated from chickens has been documented following DXC use. Reptiles

Dosages of DXC are reported for reptiles but studies are scarce. DXC PO was associated with clinical resolution of anaplasmosis in gopher tortoises (Raskin et al., 2020).

G ­ lycylcyclines The glycylcyclines are classified as third-­ generation tetracyclines approved for human use only (WHO, 2024). The glycylcycline in clinical use is tigecycline, a minocycline with a tert-­butylglycylamino group on carbon-­9. Tigecycline binds to the A site of the 30S subunit and interacts with residues of the H34 ribosomal subunit. This leads to prevention of protein synthesis by blocking incorporation of amino acid residues into the elongating peptide chain. Binding of tigecycline to ribosomes is five times higher than that of older tetracyclines. Tigecycline has a broad antimicrobial spectrum, with efficacy against many multidrug-­resistant bacteria such as extended-­spectrum beta-­lactamase-­producing

Enterobacterales (ESBL), carbapenem-­resistant Enterobacterales (CRE), methicillin-­resistant Staphylococcus aureus (MRSA), vancomycin-­ resistant Enterococcus (VRE) and multidrug-­ resistant Acinetobacter sp. but is not active against Pseudomonas spp. The AUC/MIC ratio appears to be the appropriate PK/PD target for tigecycline. Tigecycline is available only as an injectable formulation for the IV treatment of complicated skin and soft tissue and abdominal infections in humans. Despite a high degree of protein binding, tigecycline has a large volume of distribution (7–9 l/kg), which reflects extensive distribution into tissues. Tigecycline is excreted mainly unchanged via the feces and to a lesser extent via urine. With clinical use, tigecycline resistance has emerged and the most common mechanisms are overexpression of nonspecific active efflux pumps or mutations within the drug-­binding site in the ribosome. Mobile tigecycline-­ resistance genes, tet(X3) and tet(X4), have now been identified in numerous Enterobacterales and Acinetobacter spp. isolated from animals, meat for consumption, and humans (He et al., 2019). Tet(X3) and Tet(X4) inactivate all tetracyclines, including tigecycline. Tigecycline has been proposed as a treatment alternative for dogs and cats for infections caused by multidrug-­resistant bacteria, which are increasing in incidence (Kim and Kim, 2021; Papich, 2012). Such use should be carefully considered according to antimicrobial stewardship principles due to its importance in human medicine.

R ­ eferences Agbedanu PN, et al. 2015. Doxycycline as an inhibitor of p-­glycoprotein in the alpaca for the purpose of maintaining avermectins in the CNS during treatment for parelaphostrongylosis. Vet Parasitol 212:303. Álvarez-­Fernández A, Breitschwerdt EB, Solano-­Gallego L. 2018. Bartonella infections

in cats and dogs including zoonotic aspects. Parasit Vectors 11:624. Angelos JA. 2015. Infectious bovine keratoconjunctivitis (pinkeye). Vet Clin North Am: Food Anim Pract 31:61. Apley MD. 2015. Clinical evidence for individual animal therapy for papillomatous digital

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dermatitis (hairy heel wart) and infectious bovine pododermatitis (foot rot). Vet Clin North Am: Food Anim Pract 31:81. Baptiste KE, Kyvsgaard NC. 2017. Do antimicrobial mass medications work? A systematic review and meta-­analysis of randomised clinical trials investigating antimicrobial prophylaxis or metaphylaxis against naturally occurring bovine respiratory disease. Pathogens Dis 75:ftx083. Barker EN. 2019. Update on feline hemoplasmosis. Vet Clin North Am: Small Anim Pract 49:733. Barton MD. 2014. Impact of antibiotic use in the swine industry. Curr Opin Microbiol 19:9. Boy S, Crossley D, Steenkamp G. 2016. Developmental structural tooth defects in dogs – experience from veterinary dental referral practice and review of the literature. Front Vet Sci 3:9. Brihoum M, et al. 2010. Descriptive study of 32 cases of doxycycline-­overdosed calves. J Vet Intern Med 24:1203. Chapuis RJ, et al. 2021a. Nonlinear mixed-­effect pharmacokinetic modeling and distribution of doxycycline in healthy female donkeys after multiple intragastric dosing – preliminary investigation. Animals 11: 2047. Chapuis RJ, et al. 2021b. Pharmacokinetics and pharmacodynamics of doxycycline in a Streptococcus equi subsp. zooepidemicus infection model in horses. J Vet Pharmacol Ther 44:766. Chapuis RJ, et al. 2023. Characterisation of faecal microbiota in horses medicated with oral doxycycline hyclate. Equine Vet J 55:129. Cramer G, Solano L, Johnson R. 2019. Evaluation of tetracycline in milk following extra-­label administration of topical tetracycline for digital dermatitis in dairy cattle. J Dairy Sci 102:883. Decundo JM, et al. 2019. Impact of water hardness on oxytetracycline oral bioavailability in fed and fasted piglets. Vet Med Sci 5: 517. Divers T, et al. 2018. Borrelia burgdorferi infection and Lyme disease in North American

horses: a consensus statement. J Vet Intern Med 32:617. Dowling P, Russell A. 2000. Pharmacokinetics of a long-­acting oxytetracycline-­polyethylene glycol formulation in horses. J Vet Pharmacol Ther 23:107. Duncan J, et al. 2014. Contagious ovine digital dermatitis: an emerging disease. Vet J 201:265. Ellero N, et al. 2020. Rhabdomyolysis and acute renal failure associated with oxytetracycline administration in two neonatal foals affected by flexural limb deformity. Vet Sci 7:160. Erol E, Shaffer CL, Lubbers BV. 2022. Synergistic combinations of clarithromycin with doxycycline or minocycline reduce the emergence of antimicrobial resistance in Rhodococcus equi. Equine Vet J 54:799. Giguère S, et al. 2017. Comparative pharmacokinetics of minocycline in foals and adult horses. J Vet Pharmacol Ther 40:335. Granados-­Chinchilla F, Rodríguez C. 2017. Tetracyclines in food and feedingstuffs: from regulation to analytical methods, bacterial resistance, and environmental and health implications. J Anal Methods Chem 2017, 1315497. He T, et al. 2019. Emergence of plasmid-­ mediated high-­level tigecycline resistance genes in animals and humans. Nat Microbiol 4:1450. Hnot ML, et al. 2015. Effect of feeding on the pharmacokinetics of oral minocycline in healthy research dogs. Vet Dermatol 26:399. Kim DH, Kim JH. 2021. Efficacy of tigecycline and linezolid against pan-­drug-­resistant bacteria isolated from companion dogs in South Korea. Front Vet Sci 8:693506. Kim S, et al. 2016. Clinical and microbiological effects of a subantimicrobial dose of oral doxycycline on periodontitis in dogs. Vet J 208:55. Lappin M, et al. 2017. Antimicrobial use guidelines for treatment of respiratory tract disease in dogs and cats: Antimicrobial Guidelines Working Group of the International Society for Companion Animal Infectious Diseases. J Vet Intern Med 31:279.

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Li Z, et al. 2021. Pharmacokinetics and ex vivo pharmacodynamics of Minocycline against Salmonella abortus equi in donkey plasma and tissue cage fluid. Res Vet Sci 135:293. Littman MP, et al. 2018. ACVIM consensus update on Lyme borreliosis in dogs and cats. J Vet Intern Med 32:887. Maaland MG, Guardabassi L, Papich MG. 2014. Minocycline pharmacokinetics and pharmacodynamics in dogs: dosage recommendations for treatment of meticillin-­ resistant Staphylococcus pseudintermedius infections. Vet Dermatol 25:182. Masood F, et al. 2022. Evaluating approved and alternative treatments against an oxytetracycline-­resistant bacterium responsible for European foulbrood disease in honey bees. Sci Rep 12:1. Nichols MC, et al. 2014. Yersinia pestis infection in dogs: 62 cases (2003–2011). J Am Vet Med Assoc 244:1176. Nielsen P, Gyrd-­Hansen N. 1996. Bioavailability of oxytetracycline, tetracycline and chlortetracycline after oral administration to fed and fasted pigs. J Vet Pharmacol Ther 19:305. Ocampo PS, et al. 2014. Antagonism between bacteriostatic and bactericidal antibiotics is prevalent. Antimicrob Agents Chemother 58: 4573. O’Connor AM, Totton SC, Shane D. 2019. A systematic review and network meta-­analysis of injectable antibiotic treatment options for naturally occurring swine respiratory disease. J Swine Health Prod 27:133. Oliver A, Conrado FO, Nolen-­Walston R. 2023. Equine granulocytic anaplasmosis. Vet Clin North Am Equine Pract 39:133. Papich MG. 2012. Selection of antibiotics for meticillin-­resistant Staphylococcus pseudintermedius: time to revisit some old drugs? Vet Dermatol 23:352. Papich MG, Davidson GS, Fortier LA. 2013. Doxycycline concentration over time after storage in a compounded veterinary preparation. J Am Vet Med Assoc 242:1674.

Pyatnychko O, et al. 2021. Treatment of acute intestinal infection in minks. Scientific and Technical Bulletin оf State Scientific Research Control Institute of Veterinary Medical Products and Fodder Additives аnd Institute of Animal Biology, 22:310. Qurollo B. 2019. Feline vector-­borne diseases in North America. Vet Clin North Am: Small Anim Pract 49:687. Raskin RE, Crosby FL, Jacobson ER. 2020. Newly recognized Anaplasma sp. in erythrocytes from gopher tortoises (Gopherus polyphemus). Vet Clin Pathol 49:17. Ricker N, et al. 2020. Toward antibiotic stewardship: route of antibiotic administration impacts the microbiota and resistance gene diversity in swine feces. Front Vet Sci 7:255. Riond JL, et al. 1992. Cardiovascular effects and fatalities associated with intravenous administration of doxycycline to horses and ponies. Equine Vet J 24:41. Sargeant JM, et al. 2019. A systematic review of the efficacy of antibiotics for the prevention of swine respiratory disease. Anim Health Res Rev 20:291. Schuller S, et al. 2015. European consensus statement on leptospirosis in dogs and cats. J Small Anim Pract 56:159. Stultz JS, Eiland LS. 2019. Doxycycline and tooth discoloration in children: changing of recommendations based on evidence of safety. Ann Pharmacother 53:1162. Taylor SD. 2023. Potomac horse fever. Vet Clin North Am Equine Pract 39:37. Toutain PL, et al. 2021. The pharmacokinetic/ pharmacodynamic paradigm for antimicrobial drugs in veterinary medicine: recent advances and critical appraisal. J Vet Pharmacol Ther 44:172. Warner AJ, et al. 2022. Tetracyclines and bone: unclear actions with potentially lasting effects. Bone 159:116377. Washburn K, et al. 2018. Pharmacokinetics of chlortetracycline in maternal plasma and in fetal tissues following oral administration to pregnant ewes. J Vet Pharmacol Therapeut 41:218.

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Wenzel M, et al. 2021. A flat embedding method for transmission electron microscopy reveals an unknown mechanism of tetracycline. Commun Biol 4:306. Wilson B, et al. 2006. Susceptibility of bacteria from feline and canine urinary tract infections to doxycycline and tetracycline concentrations attained in urine four hours after oral dosage. Australian Vet J 84:8. World Health Organization, 2024. WHO List of Medically Important Antimicrobials. A risk management tool for mitigating antimicrobial resistance due to non-human use. Geneva: World Health Organization.

Yao W, et al. 2014. Local delivery of minocycline-­ loaded PEG-­PLA nanoparticles for the enhanced treatment of periodontitis in dogs. Int J Nanomed 9:3963. Zhang X, et al. 2022. Invited Review: Antimicrobial use and antimicrobial resistance in pathogens associated with diarrhea and pneumonia in dairy calves. Animals 12:771. Zurhelle G, et al. 2000. Metabolites of oxytetracycline, tetracycline, and chlortetracycline and their distribution in egg white, egg yolk, and hen plasma. J Agric Food Chem 48:6392.

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15 Chloramphenicol, Thiamphenicol, and Florfenicol Patricia M. Dowling and Hélène Lardé

G ­ eneral Considerations

Chemistry

The phenicol antimicrobials are used to treat Gram-­positive, Gram-­negative, and anaerobic infections. The phenicols are considered highly important antimicrobials in human medicine by the World Health Organization due to their use in human medicine, including treatment of infections transmitted from nonhuman sources. In particular, chloramphenicol is being reinvestigated for its activity against vancomycin-­resistant Enterococcus faecium and Staphylococcus aureus and against multidrug-­resistant (MDR) Gram-­negative pathogens in combination with colistin (Čivljak et al., 2014). Thiamphenicol is used in human medicine in some countries, while florfenicol is used exclusively in veterinary medicine.

Chloramphenicol is a stable, lipid-­soluble, ­neutral compound. It is a derivative of dichloracetic acid and contains a nitrobenzene ­moiety. This para-­nitro group is associated with idiosyncratic (nondose-­dependent) ­aplastic anemia in humans (Figure 15.1). Thiamphenicol has a similar antibacterial spectrum to chloramphenicol but differs from the parent compound in that the p-­nitro group attached to the benzene ring is replaced by a sulfomethyl group. Florfenicol is a structural analogue of thiamphenicol that also lacks the p-­nitro group, and it is more active than ­thiamphenicol against bacteria. Neither thiamphenicol nor florfenicol is associated with dose-­independent aplastic anemia in humans or any other species, but both are associated with dose-­dependent bone marrow suppression.

C ­ hloramphenicol Chloramphenicol a broad-­spectrum antibiotic that was derived from Streptomyces venezuelae initially but has also been produced synthetically. Chloramphenicol is a time-­dependent, bacteriostatic drug that can attain effective concentrations at sites of infection that are relatively inaccessible to other antimicrobials.

Mechanism of Action Chloramphenicol is a potent inhibitor of microbial protein synthesis. It binds irreversibly to a receptor site on the 50S subunit of the bacterial ribosome, inhibiting peptidyl transferase and preventing the amino acid transfer to growing peptide chains and subsequently inhibiting protein formation. Chloramphenicol

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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Chloramphenicol, Thiamphenicol, and Florfenicol O

O2N

H

NH

C

C

C

CHCl2

CH2OH

OH H Chloramphenicol O

H3C

SO2 Florfenicol

H3C

H

NH

C

C

OH

H

CHCl2

CH2F

O H

NH

C

C

OH Thiamphenicol

H

SO2

C

C

CHCl2

CH2OH

Figure 15.1  Chemical structure of chloramphenicol, florfenicol, and thiamphenicol.

●●

also inhibits mitochondrial protein synthesis in mammalian bone marrow cells in a dose-­ dependent manner.

Antimicrobial Activity Chloramphenicol is active against a wide range of Gram-­positive and many Gram-­negative and anaerobic bacteria. It is bacteriostatic at usual therapeutic concentrations. Chloramphenicol is active against Hemobartonella, Rickettsia, and Chlamydia. While mycoplasma often show susceptibility in vitro, chloramphenicol therapy of mycoplasma pulmonary infections is often ineffective. ●●

Susceptible organisms (MIC ≤8 μg/ml) are ­Gram-­positive aerobic bacteria, including Actinomyces spp., Trueperella pyogenes, Bacillus  anthracis, Corynebacterium spp., Erysipelothrix rhusiopathiae, Listeria monocytogenes, many Enterococcus spp., Staphylococcus spp., and Streptococcus spp. Methicillin-­ resistant  Staphylococcus aureus (MRSA) and Staphylococcus pseudintermedius (MRSP) have

●●

emerged as significant pathogens in companion animals. Two major clonal MRSP lineages have disseminated in Europe and North America. Isolates originating from North America are often susceptible to chloramphenicol, whereas isolates from Europe are often resistant to ­chloramphenicol (Perreten et  al.,  2010). Staphylococcus schleiferi, a coagulase-­negative staphylococcus and emerging zoonotic pathogen isolated from dogs, is typically susceptible (Lee et  al.,  2019). Typically susceptible Gram-­ negative aerobic bacteria include Actinobacillus spp., Bordetella bronchiseptica, Brucella canis, Enterobacterales (including many E. coli), Klebsiella spp., Proteus spp., and Salmonella spp., Haemophilus spp., Histophilus somni, Leptospira spp., Moraxella bovis, Mannheimia haemolytica, and Pasteurella spp. Anaerobes (Bacteroides spp., Clostridium spp., Prevotella spp., Porphyromonas spp.) are commonly susceptible, including penicillin-­resistant Bacteroides fragilis. Intermediately susceptible organisms (MIC = 16 μg/ml) include Rhodococcus equi. Resistant organisms (MIC ≥32 μg/ml) include Pseudomonas spp., Mycobacterium spp., and Nocardia spp. Resistance often emerges in Gram-­negative enteric bacteria such as E. coli.

Resistance to Chloramphenicol The most common mechanism of bacterial resistance to chloramphenicol is enzymatic inactivation by acetylation by chloramphenicol acetyltransferases (CATs). Acetylation of the hydroxyl groups on chloramphenicol prevents drug binding to the 50S ribosomal subunit. There are also reports of other mechanisms of resistance, such as efflux systems, inactivation by phosphotransferases, and mutations of the target site or permeability barriers (Schwarz et al., 2004). Many of the genes coding for the CAT genes or specific transporters are located on mobile genetic elements, such as plasmids, transposons or gene cassettes. The CAT genes are commonly found on plasmids in

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Enterobacterales and Pasteurellaceae, and most of these plasmids carry one or more additional resistance genes. The other mechanisms of chloramphenicol resistance include an efflux mechanism due to chloramphenicol/florfenicol exporter ( fexA) and the 23S rRNA methyl transferase (cfr) that also mediates resistance to linezolid. Increasing rates of chloramphenicol resistance in MRSA appear to be due to acquisition of novel MRSA clones carrying a fexA variant associated with florfenicol use in animals (Udo et al., 2021).

Pharmacokinetic Properties In monogastric animals and preruminant calves, chloramphenicol is well absorbed from the gastrointestinal tract. The oral bioavailability of chloramphenicol in foals is 83%, but only 40% after a single administration in mares, declining to 20% after five doses (Brumbaugh et al., 1983; Gronwall et al., 1986). Chloramphenicol palmitate is poorly absorbed in cats. In ruminants, orally administered chloramphenicol is inactivated in the rumen. The apparent volume of distribution of chloramphenicol is large (>1  l/kg) in all species. This can be attributed to widespread distribution, as partitioning of the drug is independent of pH and there is no evidence of selective tissue binding. Because of its lipid solubility and moderately low protein binding (30–46%), chloramphenicol attains effective concentrations in most tissues and body fluids, including cerebrospinal fluid (CSF) and the central nervous system. Chloramphenicol may achieve CSF concentrations up to 50% of plasma concentrations when the meninges are normal and more when inflammation is present. Topical ophthalmic formulations achieve therapeutic concentrations in the aqueous humor. Chloramphenicol readily diffuses into milk, and pleural and ascitic fluids. It readily crosses the placenta, achieving concentrations 75% of those in maternal plasma. This may be of clinical ­significance, as the fetal liver is deficient in

glucuronyl transferase activity. Penetration of the blood–prostate barrier is relatively poor unless inflammation is present. The elimination half-­life of chloramphenicol varies widely between species. Elimination is primarily by hepatic metabolism by conjugation with glucuronic acid. Its elimination is short in horses (one hour) and long in cats (5–6  hours) because of feline deficiencies in glucuronide conjugation. A fraction of the dose is excreted unchanged by glomerular filtration in the urine of dogs (10%) and cats (25%), while a negligible amount is eliminated by renal excretion in herbivores. The hepatic metabolites, which are inactive, are excreted in the urine and to a much lesser extent in the bile. The glucuronide conjugate excreted in bile can be hydrolyzed by intestinal flora to liberate the parent drug. In newborn animals, the elimination half-­ life of chloramphenicol is considerably longer than in adult animals of the same species. This is due mainly to immature glucuronide conjugation mechanisms. Glucuronide conjugation develops most rapidly in foals, so that the half-­ life in the one-­week-­old foal approaches that of the adult horse.

Drug Interactions Chloramphenicol should not be used concurrently with bactericidal antimicrobials in treating infections where host defenses are poor. Concurrent chloramphenicol and penicillin G are antagonistic in treating bacterial meningitis and endocarditis in humans. Chloramphenicol acts on the same ribosomal site as the macrolides. Chloramphenicol is antagonistic to the fluoroquinolones, as inhibition of protein synthesis by chloramphenicol interferes with the production of autolysins necessary for cell lysis after the fluoroquinolone interferes with bacterial DNA supercoiling. Chloramphenicol inhibits the oxidase activity of cytochrome P450, and decreases the clearance of drugs metabolized through the same pathway, resulting in prolonged

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pharmacological effect. For example, chloramphenicol markedly prolongs the effect of barbiturates, opioids, xylazine, and propofol.

Toxicity and Adverse Effects The main toxic effects of chloramphenicol in humans are bone marrow depression, which can be either an idiosyncratic, nondose-­dependent aplastic anemia or a dose-­dependent anemia from suppression of protein synthesis associated with the p-­nitro group (Yunis et  al.,  1980). Aplastic anemia appears to be a genetically determined idiosyncrasy of individual humans. The incidence of fatal aplastic anemia has been estimated as 1 in every 25 000–60 000 humans who use the drug. A few cases of aplastic anemia in humans have occurred following contact exposure (ophthalmic use, medicated sprays, handling), so veterinarians and owners should wear protective gloves and face masks when handling chloramphenicol products. A “gray baby” syndrome occurs in newborn infants because their deficiency in glucuronic acid conjugation causes a dose-­dependent anemia. In animals, chloramphenicol toxicity is related to both the dose and duration of treatment, and cats are more likely than dogs to develop toxicity. In cats, clinical signs of toxicity may be seen when the usual maintenance ­dosage of 25 mg/kg of base or palmitate ester is given twice daily for 21  days (Watson,  1991). Chloramphenicol causes changes in the peripheral blood and bone marrow due to reversible, dose-­related disturbances in red cell maturation. Administration for less than 10 days using the maintenance dose is not likely to cause toxicity in either dogs or cats, unless the animals have depressed hepatic microsomal enzyme activity or severely impaired renal function. Use in dogs is associated with frequent adverse gastrointestinal effects (e.g., vomiting, diarrhea, weight loss, nausea, anorexia, decreased appetite), as well as lethargy, shaking, and hindlimb weakness (especially in large dogs) (Short et al., 2014).

Dosage Considerations Therapeutic efficacy of chloramphenicol is maximized by maintaining an average steady-­ state plasma concentration of 5–10 μg/ml. Chloramphenicol is available for either oral (free base or palmitate ester) or parenteral (sodium succinate) administration. For local treatment of eye or ear infections caused by susceptible organisms, topical preparations are available. Because chloramphenicol is well absorbed from the gastrointestinal tract in small animals, it can be given orally as either the base or palmitate ester. The ester is hydrolyzed in the small intestine prior to absorption of the active free base. Subcutaneous injection of chloramphenicol sodium succinate is an alternative to oral administration. While both routes may provide equivalent concentrations, the oral route is preferable as injection of the parenteral preparation is painful. Intramuscular injection provides significantly lower plasma levels than subcutaneous and oral routes, and is not recommended. The total length of treatment should not exceed 10 days, especially in cats. Do not administer chloramphenicol to patients with evidence of bone marrow suppression or renal/hepatic insufficiency. The short half-­life of chloramphenicol in horses (one hour), together with its generally bacteriostatic action, makes IV administration impractical. Tablets of the free base drug can be administered orally or the sodium succinate formulation can be given by IM injection. After absorption from injection sites, the inactive succinate ester is rapidly hydrolyzed to the active drug. Because of the risks of idiosyncratic aplastic anemia in humans, chloramphenicol is banned for use in food animals in most countries. The drug should not be used in the early neonatal period unless plasma concentrations are monitored, and should be used with caution in pregnant animals because of the potential adverse effects on the fetus.

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Clinical Use The potential for idiosyncratic fatal aplastic anemia in humans has led to prohibition of chloramphenicol use in food animals in many parts of the world. However, because of its low price, broad spectrum of antimicrobial activity, and good pharmacokinetic properties, the illegal use of chloramphenicol still occurs in bees, poultry, and aquaculture, especially in developing countries. Florfenicol is the appropriate analogue to use in food animals. With the development of fluoroquinolones for companion animals, there were few primary indications for the use of chloramphenicol, but it was still considered for some anaerobic infections, serious ocular infections, prostatitis, otitis media/interna and salmonellosis in horses, dogs, and cats. Use in dogs and cats has been increasing in frequency due to the increase in MRSA and MRSP infections, but chloramphenicol is associated with more adverse effects (mainly gastrointestinal) than other treatment options such as doxycycline, clindamycin, and amikacin. Human toxicity from handling chloramphenicol should be discussed with the owner and appropriate precautions taken when prescribing chloramphenicol for veterinary use. In addition, the zoonotic potential of animal-­origin staphylococci should be discussed with owners (Carroll et al., 2021).

T ­ hiamphenicol Thiamphenicol is a derivative of chloramphenicol, in which the p-­nitro group has been replaced by a sulfomethxyl group. Because it lacks the p-­nitro group, it does not induce irreversible bone marrow aplasia in humans, although it can cause dose-­dependent bone marrow suppression. Thiamphenicol is generally less active than chloramphenicol against Enterobacterales, although it has equal activity against Haemophilus, B. fragilis, and streptococci. Cross-­ resistance with chloramphenicol is complete in

bacteria that possess CATs. Thiamphenicol glycinate, the ester prodrug of thiamphenicol, is sometimes used in water-­soluble parenteral formulations; the prodrug is cleaved quickly and completely by the tissue esterase to thiamphenicol (Yang et al., 2011). Absorption and distribution are similar to chloramphenicol, and it is also  equally well distributed into tissues. Bioavailability after intramuscular administration is high in most species: 76% in pigs (Castells et al., 1999), 96% in dogs (Castells et al., 1998), 88% in sheep (Abdennedi et al., 1994a), 84% in cattle (Abdennebi et al., 1994b). Oral bioavailability is 75–78% in geese and ducks (Tikhomirov et  al.,  2019,  2021), 60% in preruminant lambs and calves (Mengozzi et  al.,  2002), and 30% in pigs and adult sheep (Abdennebi et  al., 1994; Haritova et al., 2002). Thiamphenicol is not eliminated by hepatic glucuronide conjugation but excreted unchanged in the urine. Unlike chloramphenicol, its elimination is unaffected by liver disease and by the use of other drugs metabolized in the liver. The pharmacokinetic parameters of thiamphenicol follow allometric scaling, in that values for elimination half-­life and volume of distribution increase with body size from mice through rats, rabbits, dogs, pigs, sheep, and calves (Castells et  al.,  2001). Therapeutic concentrations are achieved in milk of lactating cows (Abdennebi et al., 1994b). Thiamphenicol is used in Europe, South America, China, and Japan but is not available in North America. Thiamphenicol appears underutilized in the treatment of many infections caused by susceptible organisms. While detailed dosage information is not available because of the lack of pharmacokinetic and clinical studies, suitable dosage in animals would appear to be similar to that of chloramphenicol.

­Florfenicol Florfenicol is a fluorinated derivative of thiamphenicol, in which the hydroxyl group at C-­3 is replaced with fluorine. Florfenicol is a potent

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inhibitor of microbial protein synthesis with the same mechanisms of action as chloramphenicol. Like thiamphenicol, florfenicol lacks the p-­nitro group and does not cause idiosyncratic aplastic anemia in humans but can cause dose-­dependent bone marrow suppression in animals.

Antimicrobial Activity Florfenicol is more active than chloramphenicol in its range of antimicrobial activity. Bordatella bronchiseptica, Streptococcus suis, Pasteurella multocida, and Actinobacillus pleuropneumoniae from swine and Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni from cattle are susceptible at ≤2 μg/ml. Salmonella enterica subsp. enterica serovar Choleraesuis from swine are susceptible at ≤4 μg/ml. Other enteric bacteria may have MIC values higher than 4 μg/ml and would not be considered susceptible. Fusobacterium necrophorum, Bacteroides melaninogenicus, and Moraxella bovis are highly susceptible. Florfenicol shows in  vitro activity against Mycoplasma hyopneumoniae in pigs and M. bovis in cattle but the lack of specific clinical and laboratory standards makes data interpretation and correlation to in  vivo efficacy difficult (Klein et al., 2017). Florfenicol is active against a number of important bacterial pathogens of fish including Aeromonas salmonicida, Vibrio salmonicida, Vibrio anguillarum, and Yersinia ruckeri in salmon and trout and Edwardsiella ictaluri in catfish.

Antimicrobial Resistance Because of the substitution of a hydroxyl group with a fluorine molecule, florfenicol is not susceptible to resistance from bacteria expressing CAT enzymes. But seven florfenicol resistance genes and their variants, floR, cfr, fexA, fexB, ramA, pexA, optrA, and estDL136, have been discovered. Mobile genetic elements play an important role in the replication of these

resistance genes and the horizontal resistance gene transfer. There is increasing concern over the use of florfenicol in food animals with environmental contamination leading to cross-­resistance to other classes of drugs important in human and veterinary medicine. Preexposure of enteric bacterial pathogens to subinhibitory concentrations of florfenicol, as would be found in manure of treated animals, substantially increased cross-­resistance to ampicillin, ­tetracycline, and nalidixic acid (Singh and Bhunia, 2019). Although oxazolidinones (e.g., linezolid) have never been approved for use in livestock, there is an abundance of oxazolidinone resistance genes in livestock and their manure that is associated with florfenicol residues (Nuesch-­Inderbinen et  al.,  2022; Wang et  al.,  2020). The gene cfr confers ­resistance to five classes of antimicrobial agents including phenicols, lincosamides, oxazolidinones, pleuromutilins, and streptogramin A. The second transferable oxazolidinone resistance gene, optrA, encodes a ribosomal protection protein of the ABC-­F family and confers resistance to oxazolidinones and phenicols. Another ribosomal protection ­protein of the ABC-­F family, poxtA, leads to decreased susceptibility to phenicols, oxazolidinones, and tetracyclines. These transmissible resistance genes are of concern as linezolid is a WHO Critically Important Antimicrobial in human medicine and they have been shown to easily persist in animal manure (Ma et al., 2022). Florfenicol resistance in Gram-­negative bacteria is frequently related to the floR gene carried by a transposon located on a plasmid. This gene codes for a membrane-­associated exporter protein that promotes efflux of chloramphenicol and florfenicol. In cases of neonatal calf diarrhea from E. coli, if floR was present, the MIC range was 16 to ≥256 μg/ml (White et al., 2000). After a single dose of florfenicol, feedlot cattle showed a shift in fecal flora to MDR E. coli, likely due to selection for plasmids containing the floR gene linked with

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other resistance genes. The antimicrobial resistance associated with florfenicol treatment declined over four weeks post treatment but a higher proportion of fecal E. coli was resistant than when the cattle entered the feedlot (Berge et al., 2005). In swine, oral or intramuscular use of florfenicol selected for florfenicol resistant (MIC ≥256 μg/mL) E. coli (De Smet et al., 2018). There is a clear trend of decreasing susceptibility of the three major Gram-­negative pathogens to the antimicrobials commonly used for treatment and control of bovine respiratory disease (BRD), including florfenicol (DeDonder and Apley, 2015; Depenbrock et al., 2021). A large number of studies have found that floR is the most common gene in poultry MDR Salmonella strains isolated from the chicken intestinal tract, chicken breeding environment, or slaughterhouse.

Pharmacokinetic Properties Florfenicol is lipid soluble and characterized by good extravascular absorption and extensive distribution. The oral bioavailability of florfenicol in horses is 83% and 81% after IM injection (McKellar and Varma, 1996). It is 89% in 2–5-­week-­old calves but decreases when administered with milk replacers (Varma et  al.,  1986). Florfenicol readily transfers into the milk of lactating dairy cows when administered IM or SC, and IMM administration results in significant plasma concentrations. While values of volume of distribution for florfenicol are slightly lower than for chloramphenicol, florfenicol is well distributed into many tissues including lungs, muscle, bile, kidney, and urine. With IV administration, cerebrospinal fluid concentrations are 46% of plasma concentrations, achieving potentially therapeutic concentrations for H. somni, but not Gram-­negative enteric bacteria (de Craene et  al.,  1997). With IM administration to beef calves, the serum concentration of florfenicol remains above 1 μg/ml for 22 hours (Lobell et al., 1994).

The commercially available formulation of florfenicol for cattle and swine is long-­acting, so that “flip-­flop” kinetics occurs, where elimination is prolonged due to slow absorption from the IM or SC injection site. While rapidly eliminated after IMM administration to dairy cows, florfenicol residues last longer in milk after IM administration and are greatly prolonged (>16.5  days) after SC administration (Kawalek et  al.,  2016). After SC administration to dairy goats, florfenicol residues could be detected with a rapid residue test for >30 days (Richards et al., 2022). In cattle, 64% of a dose is excreted as parent drug in the urine. Florfenicol amine is the slowest metabolite to deplete from the liver and is used as the marker residue for withdrawal times. Sheep show similar pharmacokinetics to cattle for florfenicol. Florfenicol is well absorbed orally in dogs, but is rapidly eliminated so never developed as a commercial product (Park et al., 2008). While not approved, florfenicol is used extra-­ label in a number of species. Pharmacokinetics have been described in goats, camels, North American elk, rabbits, alpacas, llamas, dogs, and many species of birds and fish (Alcorn et al., 2004; Ali et al., 2003; Park et al., 2007, 2008; Pentecost et al., 2013, 2015).

Drug Interactions Florfenicol shows synergistic activity in combination with thiamphenicol or aminoglycosides as florfenicol acts as an initiating modulator of membrane permeability that allows increased uptake of multiple antibiotics by susceptible Gram-­negative bacteria with the potential to reduce the overall usage, drug residues, and withdrawal times (Assane et al., 2019; Kim et al., 2020; Rattanapanadda et al., 2019; Wei et al., 2016). Polyether ionophore antibiotics, such as salinomycin, monensin, and maduramycin, increase the absorption, distribution, and elimination of florfenicol, resulting in reduced plasma concentrations.

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Toxicity and Adverse Effects Transient diarrhea or inappetence may occur in cattle treated with florfenicol, but resolves within a few days of discontinuing treatment. In swine, perianal inflammation and/or rectal eversion may occur in treated animals, but should resolve completely within one week. The injectable florfenicol formulations for cattle and swine are only labeled for a maximum of two doses, so bone marrow suppression has not been reported with injectable use in these species. In mice, florfenicol induced cell cycle arrest and apoptosis of hematopoietic cells, and changed the bone marrow hematopoietic microenvironment by decreasing the number of peripheral blood cells. It also induced hypoplasia and atrophy of the spleen and thymus, induced cell cycle arrest, splenocyte apoptosis, and decreased the proliferation and viability of lymphocytes and humoral and cellular immunity (Hu et al., 2016). When fed at therapeutic dosages, florfenicol induced temporary toxicity in the hematopoietic and lymphoid organs of piglets and affected hematopoiesis and immune function (Hu et al., 2014). Potentially fatal bone marrow suppression, from suppression of protein synthesis in erythroid cells, has been documented with overdose or prolonged florfenicol administration (Holmes et  al.,  2012; Tuttle et al., 2006). In goats, high doses of florfenicol produced reversible dose-­dependent effects on functional indicators of kidney and liver such as urea, creatinine, TP, ALP, SGOT, SGPT, GGT, and bilirubin (Shah et al., 2016). In dogs, the florfenicol combination otic products are safe when administered carefully. But contact of the product with the eyes of either the dog or the person applying the treatment can result in corneal ulcers, eye irritation, conjunctivitis, squinting, and eye pain. At high florfenicol doses in water, florfenicol induced hepatotoxicity in young chicks by inhibiting enzyme activity in the Nrf2-­ARE

pathway to increase oxidative stress and promoting apoptotic protein expression to accelerate hepatocyte apoptosis (Han et al., 2020). Florfenicol induces nephrotoxicity in young broiler chicks by causing intense lipid peroxidation reactions and abnormal cell apoptosis in the kidney, resulting in reduced the growth performance (Wang et  al.,  2021). The toxicity can be alleviated by herbal products such as Salvia miltiorrhiza polysaccharides (Wang et  al.,  2022). At high doses, florfenicol caused early embryonic death in broilers at approximately five days of development (Al-­Shahrani and Naidoo, 2015). The use of florfenicol in horses is not recommended. Despite a high oral bioavailability and good tissue distribution, florfenicol administration to horses altered fecal consistency with single doses administered IV, PO, or IM (McKellar and Varma, 1996).

Administration Florfenicol injectable solution is approved in numerous countries for beef cattle and nonlactating dairy cattle at 20  mg/kg IM twice at a 48-­hour interval or 40  mg/kg SC once. The products combined with meloxicam or flunixin meglumine are only labeled for SC administration. In some countries, injectable florfenicol is approved for sheep at 20  mg/kg IM daily for three consecutive days and for swine at 15 mg/kg IM twice at a 48-­hour interval. In swine it should be injected into the neck at no more than 5 ml per site. In the US, the EU, and other countries, florfenicol is approved as a swine premix or a water additive at 10 mg/kg fed for five days. In some countries, florfenicol is approved for use in poultry at 10–20 mg/kg in water for three days. Florfenicol is poorly soluble in water, so administration by this route may lead to variability in individual exposure. In fish, the florfenicol premix is mixed in unmedicated feed prior to pelleting or used to surface coat pelleted feed and fed to deliver 10–15  mg/kg per day for 10 consecutive days.

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Florfenicol is available in combination products for the treatment of canine otitis.

Clinical Use Cattle, Sheep and Goats

Florfenicol is used for metaphylaxis and for treatment of BRD caused by highly susceptible bacteria such as Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni (MIC ≤2 μg/ml). The same dosage regimen will treat infectious pododermatitis caused by Fusobacterium necrophorum and Bacteroides melaninogenicus and infectious bovine keratoconjunctivitis caused by Morexella bovis, but penicillin or oxytetracycline are less expensive and narrower in antimicrobial spectrum and should be used as first-­line treatments for these infections. The label dosage does not result in systemic concentrations that would be effective against Gram-­negative enteric pathogens. There are approved formulations that combine florfenicol with the nonsteroidal antiinflammatory drugs flunixin meglumine or meloxicam to control pyrexia in cattle with BRD. Florfenicol/flunixin meglumine was equivalent to tildipirosin in the treatment of pneumonia due to Mannheimia, Pasteurella, and Moraxella in preweaned dairy heifers, but neither drug was effective against Mycoplasma spp. (Bringhenti et al., 2021). In the early phase of a natural outbreak of M. bovis in calves, potentially complicated by Pasteurellaceae infection, florfenicol treatment resulted in a more rapid cure rate and less antimicrobial usage than oxytetracycline (Jourquin et  al., 2022). After SC administration of florfenicol, synovial fluid concentrations greater than the MIC90 for F. necrophorum were achieved in the metatarsophalangeal joint for at least six days (Jones et al., 2015). While it has high systemic bioavailability, IMM administration of florfenicol for the treatment of bovine mastitis caused by a variety of pathogens had no advantage over cloxacillin (Wilson et al., 1996). Milk residues in lactating dairy cows are prolonged. In some countries,

florfenicol is approved in sheep for the treatment of ovine respiratory tract infections due to Mannheimia haemolytica and Pasteurella multocida. Florfenicol distributes into the preocular tear fluid of sheep, but doses >20  mg/kg would be needed to achieve concentrations greater than the MICs for most strains of Mycoplasma spp. that cause infectious keratoconjunctivitis (Regnier et  al.,  2013). Florfenicol is used extra-­label in meat and dairy goats, but milk residues can persist for >30 days. Poultry

Florfenicol is used in poultry in many countries in the treatment of respiratory diseases due to its activity on Haemophilus spp., Salmonella typhi, Klebsiella pneumoniae, Staphylococcus aureus, Pasteurella multocida, and E. coli. It is effective in broiler chickens for the treatment and control of air sacculitis associated with E. coli susceptible to florfenicol. In China, florfenicol is the most commonly used antimicrobial on layer and broiler chicken farms. Such widespread use is associated with increasing rates of transmissible antimicrobial resistance (Wang et al., 2018). Swine

Florfenicol is approved for the control of swine respiratory disease associated with Actinobacillus pleuropneumoniae, Pasteurella multocida, Streptococcus suis, and Bordetella bronchiseptica. Florfenicol in feed or by injection reduces illness due to Actinobacillus pleuropneumoniae and M. hyopneumoniae in pigs (Del Pozo Sacristan et al., 2012). Florfenicol was equally effective as gamithromycin in treating acute Actinobacillus pleuropneumoniae-­infected pigs (Papatsiros et al., 2019). As in poultry, extensive use of florfenicol in swine production is associated with increasing rates of transmissible antimicrobial resistance (Nuesch-­Inderbinen et al., 2022). Dogs

Florfenicol is not commonly used systemically in dogs. When the injectable cattle product was used in the treatment of chronic pyoderma in

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two dogs with MDR Staphylococcus pseudintermedius, the isolates became resistant due to acquisition of the fexA gene that codes for a phenicol-­specific efflux pump (Couto et  al.,  2016). One isolate also carried the cfr gene, which confers multidrug resistance including to CIA such as linezolid. Florfenicol has been developed as canine otic products in combination with a corticosteroid (mometasone or betamethasone) and antifungal (terbinafine)in long-­lasting gel formulations. The veterinarian-­administered otic gels provide equivalent efficacy and higher quality of life to dogs with otitis externa compared to traditional daily owner-­administered topical otic therapy (Noli et al., 2017). Fish

Florfenicol is used in the treatment of susceptible bacterial diseases of fish, including furunculosis in salmon and vibriosis in salmon and cod, pseudotuberculosis in Japanese yellowtail,

enteric septicemia in channel catfish, and enteric redmouth in trout. In Canada, florfenicol is approved for the treatment of furunculosis caused by susceptible strains of Aeromonas salmonicida in salmon. In the US, it is approved for the control of mortality due to furunculosis associated with Aeromonas salmonicida and coldwater disease associated with Flavobacterium psychrophilum in salmonids, for the control of mortality due to columnaris disease associated with Flavobacterium columnare in freshwater-­ reared finfish, for the control of mortality due to enteric septicemia associated with Edwardsiella ictaluri in catfish, and for the control of mortality due to streptococcal septicemia associated with Streptococcus iniae in freshwater-­reared warmwater finfish. In Japan, florfenicol is labeled for the treatment of pseudotuberculosis and streptococcosis in Perciformes (yellowtail, amberjack, red sea bream, tilapia, etc.) and for  the treatment of edwardsiellosis disease in eel.

References Abdennebi EH, et al. 1994a. Thiamphenicol pharmacokinetics in sheep. J Vet Pharmacol Ther 17:12. Abdennebi EH, et al. 1994b. Thiamphenicol pharmacokinetics in beef and dairy cattle. J Vet Pharmacol Ther 17:365. Alcorn J, et al. 2004. Pharmacokinetics of florfenicol in North American elk (Cervus elaphus). J Vet Pharmacol Ther 27:289. Ali BH, Al-­Qarawi AA, Hashaad M. 2003. Comparative plasma pharmacokinetics and tolerance of florfenicol following intramuscular and intravenous administration to camels, sheep and goats. Vet Res Commun 27:475. Al-­Shahrani S, Naidoo V. 2015. Florfenicol induces early embryonic death in eggs collected from treated hens. BMC Vet Res 11:213. Assane IM, et al. 2019. Combination of antimicrobials as an approach to reduce their

application in aquaculture: emphasis on the use of thiamphenicol/florfenicol against Aeromonas hydrophila. Aquaculture 507:238. Berge AC, Epperson WB, Pritchard RH. 2005. Assessing the effect of a single dose florfenicol treatment in feedlot cattle on the antimicrobial resistance patterns in faecal Escherichia coli. Vet Res 36:723. Bringhenti L, et al. 2021. Effect of treatment of pneumonia and otitis media with tildipirosin or florfenicol + flunixin meglumine on health and upper respiratory tract microbiota of preweaned Holstein dairy heifers. J Dairy Sci 104:10291. Brumbaugh GW, et al. 1983. Pharmacokinetics of chloramphenicol in the neonatal horse. J Vet Pharmacol Ther 6:219. Carroll KC, Burnham CD, Westblade LF. 2021. From canines to humans: clinical importance of Staphylococcus pseudintermedius. PLoS Pathog 17:e1009961.

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Castells G, et al. 1998. Pharmacokinetics of thiamphenicol in dogs. Am J Vet Res 59:1473. Castells G, et al. 1999. Thiamphenicol disposition in pigs. Res Vet Sci 66:219. Castells G, et al. 2001. Allometric analysis of thiamphenicol disposition among seven mammalian species. J Vet Pharmacol Ther 24:193. Čivljak R, et al. 2014. Could chloramphenicol be used against ESKAPE pathogens? A review of in vitro data in the literature from the 21st century. Exp Rev Anti-­infect Ther 12:249. Couto N, et al. 2016. Acquisition of the fexA and cfr genes in Staphylococcus pseudintermedius during florfenicol treatment of canine pyoderma. J Glob Antimicrob Resist 7:126. De Craene BA, et al. 1997. Pharmacokinetics of florfenicol in cerebrospinal fluid and plasma of calves. Antimicrob Agents Chemother 41:1991. DeDonder KD, Apley MD. 2015. A literature review of antimicrobial resistance in pathogens associated with bovine respiratory disease. Animal Health Res Rev 16:125. Del Pozo Sacristan R, et al. 2012. Efficacy of florfenicol injection in the treatment of Mycoplasma hyopneumoniae induced respiratory disease in pigs. Vet J 194:420. Depenbrock S, et al. 2021. In-­vitro antibiotic resistance phenotypes of respiratory and enteric bacterial isolates from weaned dairy heifers in California. PLoS One 16:e0260292. De Smet J, et al. 2018. Similar gastro-­intestinal exposure to florfenicol after oral or intramuscular administration in pigs, leading to resistance selection in commensal Escherichia coli. Front Pharmacol 9:1265. Gronwall R, et al. 1986. Body fluid concentrations and pharmacokinetics of chloramphenicol given to mares intravenously or by repeated gavage. Am J Vet Res 47 :2591. Han C, et al. 2020. Florfenicol induces oxidative stress and hepatocyte apoptosis in broilers via Nrf2 pathway. Ecotoxicol Environ Saf 191: 110239. Haritova A, et al. 2002. Pharmacokinetics of thiamphenicol in pigs. J Vet Pharmacol Ther 25:464.

Holmes K, et al. 2012. Florfenicol pharmacokinetics in healthy adult alpacas after subcutaneous and intramuscular injection. J Vet Pharmacol Ther 35:382. Hu D, et al. 2014. Toxicity to the hematopoietic and lymphoid organs of piglets treated with a therapeutic dose of florfenicol. Vet Immunol Immunopathol 162:122. Hu D, et al. 2016. Florfenicol induces more severe hemotoxicity and immunotoxicity than equal doses of chloramphenicol and thiamphenicol in Kunming mice. Immunopharmacol Immunotoxicol 38:472. Jones ML, et al. 2015. Synovial fluid pharmacokinetics of tulathromycin, gamithromycin and florfenicol after a single subcutaneous dose in cattle. BMC Vet Res 11:26. Jourquin S, et al. 2022. Randomized field trial comparing the efficacy of florfenicol and oxytetracycline in a natural outbreak of calf pneumonia using lung reaeration as a cure criterion. J Vet Intern Med 36:820. Kawalek JC, et al. 2016. Depletion of florfenicol in lactating dairy cows after intramammary and subcutaneous administration. J Vet Pharmacol Ther 39:602. Kim S, et al. 2020. Synergy between florfenicol and aminoglycosides against multidrug-­ resistant Escherichia coli isolates from livestock. Antibiotics 9:185. Klein U, et al. 2017. Antimicrobial susceptibility monitoring of Mycoplasma hyopneumoniae and Mycoplasma bovis isolated in Europe. Vet Microbiol 204:188. Lee GY, et al. 2019. Carriage of Staphylococcus schleiferi from canine otitis externa: antimicrobial resistance profiles and virulence factors associated with skin infection. J Vet Sci 20:e6. Lobell RD, et al. 1994. Pharmacokinetics of florfenicol following intravenous and intramuscular doses to cattle. J Vet Pharmacol Ther 17:253. Ma W, et al. 2022. Fate and exposure risk of florfenicol, thiamphenicol and antibiotic resistance genes during composting of swine manure. Sci Total Environ 839:156243.

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Mckellar QA, Varma KJ. 1996. Pharmacokinetics and tolerance of florfenicol in Equidae. Equine Vet J 28:209. Mengozzi G, et al. 2002. A comparative kinetic study of thiamphenicol in pre-­ruminant lambs and calves. Res Vet Sci 73:291. Noli C, Sartori R, Cena T. 2017. Impact of a terbinafine-­florfenicol-­betamethasone acetate otic gel on the quality of life of dogs with acute otitis externa and their owners. Vet Dermatol 28:386. Nuesch-­Inderbinen M, et al. 2022. Fattening pigs are a reservoir of florfenicol-­resistant enterococci harboring oxazolidinone resistance genes. J Food Prot 85:740. Papatsiros V, et al. 2019. In vivo effectiveness of injectable antibiotics on the recovery of acute Actinobacillus pleuropneumoniae-­infected pigs. Microb Drug Resist 25:603. Park BK, et al. 2007. Pharmacokinetics of florfenicol and its major metabolite, florfenicol amine, in rabbits. J Vet Pharmacol Ther 30:32. Park BK, et al. 2008. Pharmacokinetics of florfenicol and its metabolite, florfenicol amine, in dogs. Res Vet Sci 84:85. Pentecost RL, et al. 2013. Pharmacokinetics of florfenicol after intravenous and intramuscular dosing in llamas. Res Vet Sci 95:594. Pentecost RL, et al. 2015. Absorption and disposition of florfenicol after intravenous, intramuscular and subcutaneous dosing in alpacas. Res Vet Sci 99:199. Perreten V, et al. 2010. Clonal spread of methicillin-­resistant Staphylococcus pseudintermedius in Europe and North America: an international multicentre study. J Antimicrob Chemother 65:1145. Rattanapanadda P, et al. 2019. In vitro and in vivo synergistic effects of florfenicol and thiamphenicol in combination against swine Actinobacillus pleuropneumoniae and Pasteurella multocida. Front Microbiol 10:2430. Regnier A, et al. 2013. Florfenicol concentrations in ovine tear fluid following intramuscular

and subcutaneous administration and comparison with the minimum inhibitory concentrations against mycoplasmal strains potentially involved in infectious keratoconjunctivitis. Am J Vet Res 74:268. Richards ED, et al. 2022. Comparison of florfenicol depletion in dairy goat milk using ultra-­performance liquid chromatography with tandem mass spectrometry and a commercial on-­farm test. Front Vet Sci 9:991772. Schwarz S, et al. 2004. Molecular basis of bacterial resistance to chloramphenicol and florfenicol. FEMS Microbiol Rev 28:519. Shah JM, et al. 2016. Impact of therapeutic and high doses of florfenicol on kidney and liver functional indicators in goat. Vet World 9:1135. Short J, et al. 2014. Adverse events associated with chloramphenicol use in dogs: a retrospective study (2007–2013). Vet Rec 175:537. Singh AK, Bhunia AK. 2019. Animal-­use antibiotics induce cross-­resistance in bacterial pathogens to human therapeutic antibiotics. Curr Microbiol 76:1112. Tikhomirov M, et al. 2019. Pharmacokinetics of florfenicol and thiamphenicol in ducks. J Vet Pharmacol Ther 42:116. Tikhomirov M, et al. 2021. Pharmacokinetics of florfenicol and thiamphenicol after single oral and intravenous, as well as multiple oral administrations to geese. Br Poult Sci 62:25. Tuttle AD, et al. 2006. Bone marrow hypoplasia secondary to florfenicol toxicity in a Thomson’s gazelle (Gazella thomsonii). J Vet Pharmacol Ther 29:317. Udo EE, et al. 2021. Resurgence of chloramphenicol resistance in methicillin-­ resistant Staphylococcus aureus due to the acquisition of a variant florfenicol exporter (fexAv)-­mediated chloramphenicol Resistance in Kuwait Hospitals. Antibiotics 10:1250. Varma KJ, et al. 1986. Pharmacokinetics of florfenicol in veal calves. J Vet Pharmacol Ther 9:412. Wang M, et al. 2018. Variations of antibiotic resistance profiles in chickens during

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administration of amoxicillin, chlortetracycline and florfenicol. J Appl Microbiol doi: 10.1111/jam.14065. Wang X, et al. 2021. Florfenicol induces renal toxicity in chicks by promoting oxidative stress and apoptosis. Environ Sci Pollut Res Int 28:936. Wang X, et al. 2022. Salvia miltiorrhiza polysaccharides alleviates florfenicol induced kidney injury in chicks via inhibiting oxidative stress and apoptosis. Ecotoxicol Environ Saf 233:113339. Wang Y, et al. 2020. Association of florfenicol residues with the abundance of oxazolidinone resistance genes in livestock manures. J Hazard Mater 399:123059. Watson AD. 1991. Chloramphenicol 2. Clinical pharmacology in dogs and cats. Aust Vet J 68:2. Wei CF, et al. 2016. Florfenicol as a modulator enhancing antimicrobial activity: example

using combination with thiamphenicol against Pasteurella multocida. Front Microbiol 7:389. White DG, et al. 2000. Characterization of chloramphenicol and florfenicol resistance in Escherichia coli associated with bovine diarrhea. J Clin Microbiol 38:4593. Wilson DJ, et al. 1996. Efficacy of florfenicol for treatment of clinical and subclinical bovine mastitis. Am J Vet Res 57:526. Yang B, et al. 2011. Pharmacokinetics of the prodrug thiamphenicol glycinate and its active parent compound thiamphenicol in beagle dogs following intravenous administration. Xenobiotica 41:226. Yunis AA, et al. 1980. Chloramphenicol toxicity: pathogenetic mechanisms and the role of the p-­NO2 in aplastic anemia. Clin Toxicol 17:359.

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References  303

16 Sulfonamides, Diaminopyrimidines, and Their Combinations Jennifer M. Reinhart and John F. Prescott

The value of the sulfonamides, as single antimicrobial agents, has been greatly diminished by widespread acquired resistance. However, when combined with diaminopyrimidines such as trimethoprim, resistance occurs less frequently and thus their usefulness is enhanced.

S ­ ulfonamides Chemistry The sulfonamide antimicrobials are derivatives of sulfanilamide, which contains the structural prerequisites for antibacterial activity. The sulfonamides differ in the radical (R) attached to the sulfonamido (-­SO2NHR) group or occasionally in the substituent on the amino (-­NH2) group (Figure 16.1). It should be noted that other drug classes can also contain a sulfonamido group (e.g., carbonic anhydrase inhibitors, loop diuretics, certain coxibs) and thus may be classified as sulfonamides. However, these drugs lack the arylamine ring that the antimicrobials contain, which has important implications for mechanism of action and potential for toxicity. The various derivatives differ in physicochemical and pharmacokinetic properties as well as the degree of antimicrobial activity. As a group, sulfonamides are quite insoluble; they are more soluble at an alkaline pH than at an

acidic pH and the degree of solubility is dependent on the individual drug moiety. Specifically, sulfamethoxazole and sulfadimethoxine are more soluble than older sulfonamides such as sulfadiazine (Cribb et al., 1996). The sodium salts of sulfonamides are readily soluble in water, and parenteral preparations are available for IV injection. These solutions are highly alkaline in reaction, with the notable exception of sodium sulfacetamide, which is nearly neutral and is available as an ophthalmic preparation.

Mechanism of Action Sulfonamides interfere with the biosynthesis of folic acid in bacterial cells by competitively preventing paraaminobenzoic acid (PABA) from incorporation into the folic (pteroylglutamic) acid molecule (Figure 16.2). Specifically, sulfonamides compete with PABA for the enzyme dihydropteroate synthetase. Their selective bacteriostatic action depends on the difference between bacterial and mammalian cells in the source of folic acid. Susceptible microorganisms must synthesize folic acid, whereas mammalian cells use preformed folic acid. The bacteriostatic action can be reversed by an excess of PABA, so that any tissue exudates and necrotic tissue should be removed if animals are to be treated with sulfonamides.

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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Sulfonamides, Diaminopyrimidines, and Their Combinations

SO2NH2

SO2

NH N

N CH

CH3 NH2

NH2

Sulfanilamide

Sulfamethazine

SO2 NH

NH

SO2

N

N N

NH2

NH

S

CO

Sulfadiazine

COOH SO2 NH

N

Phthalylsulfathiazole CH3

O

NH2 Sulfamethoxazole

Figure 16.1  Structural formulas of some sulfonamide antimicrobials. Figure 16.2  Pathway for folic acid synthesis and inhibition by sulfonamide and diaminopyridine antimicrobials.

Dihydropteroate diphosphate + PABA Dihydropteroate synthetase

Dihydropteric acid

Diaminopyrimidines

Dihydrofolic acid

Sulfonamides

Dihydrofolate reductase

Tetrahydrofolic acid (active form)

Antimicrobial Activity Sulfonamides are antimicrobial agents with intrinsic activity against many bacteria, Toxoplasma, and other protozoal agents such as coccidia. However, their antibacterial activity

as a single agent is significantly limited by the extensive resistance that has developed over 90  years of use. Different sulfonamides may show quantitative but not necessarily qualitative differences in activity.

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The minimum inhibitory concentration (MIC) of sulfonamides is markedly affected by the composition of the medium and the bacterial inoculum concentration. Because of this, in  vitro tests may sometimes falsely report a bacterium to be resistant. This will not be the case with proper quality controls employing a thymidine-­sensitive strain of Enterococcus faecalis. In agar diffusion tests, Mueller Hinton agar containing lyzed horse blood is the ideal medium because it contains thymidine phosphorylase that decreases the quantity of thymidine in the medium. Veterinary-­specific breakpoints for sulfonamide antimicrobials have not been established, so human breakpoints are commonly reported (CLSI Group C agent). An MIC of 8–32 μg/ml is a reasonable definition of susceptibility for short-­acting systemic sulfonamides; an MIC of >64–128 μg/ml can be interpreted as evidence of resistance, although the CLSI (or other standardizing organization) should be consulted for the most up-­to-­date recommendations. Sulfonamide antimicrobials achieve high concentrations in the urine (>100 μg/ml), so different breakpoints are used. The CLSI criteria for people describe susceptibility in bacteria for urinary tract infections as those having an MIC of 80%) protein binding increases the elimination half-­life. In any one species, the extent of protein binding, apparent volume of distribution, and elimination half-­life vary widely among individual sulfonamides. This information, together with designating 100  μg/ml as the desired steady-­state plasma sulfonamide concentration, facilitates calculation of dosages. Sulfonamides are eliminated by a combination of renal excretion and biotransformation. This combination contributes to species variations in the half-­lives of individual drugs. Sulfadimethoxine, for example, has elimination half-­lives of 12.5 hours in cattle, 8.6 hours in goats, 11.3 hours in horses, 15.5 hours in swine, 13.2 hours in dogs, and 10.2 hours in cats. These relatively long elimination half-­lives have been attributed to extensive binding to plasma albumin and pH-­dependent passive reabsorption of the drug from acidic distal renal tubule fluid. Sulfonamides undergo metabolic alterations to a variable extent in the tissues, especially the liver. N4-­acetylation of the paraamino group is the principal metabolic pathway for most sulfonamides and takes place in humans and all domestic animals except the dog, which lacks functional genes for the N-­acetyltransferase enzymes (Trepanier,  2004). Acetylation takes place in the reticuloendothelial rather than the

parenchymal cells of the liver and other tissues such as the lungs. Aromatic hydroxylation of the methyl group on the pyrimidine ring, which may be the principal metabolic pathway for sulfonamides in ruminants, and glucuronide conjugation are microsomal-­mediated metabolic reactions. The glucuronide conjugates are highly water soluble and are rapidly excreted. Renal excretion mechanisms include glomerular filtration of free (unbound) drug in the plasma, active carrier-­mediated proximal tubular excretion of ionized unchanged drug and metabolites, and passive reabsorption of nonionized drug from distal tubular fluid. The extent of reabsorption is determined by the pKa of the sulfonamide and the pH of the fluid in the distal tubules. Urinary alkalinization increases both the fraction of the dose that is eliminated by renal excretion (unchanged in urine) and the solubility of sulfonamides in the urine.

Drug Interactions The important synergistic interaction of ­sulfonamides with antibacterial diaminopyrimidines such as trimethoprim and baquiloprim is  ­discussed below under the Antibacterial Diaminopyrimidine–Sulfonamide Combinations section. The agents appear not to antagonize the bactericidal effect of penicillins, but the procaine of procaine penicillin is an analogue of PABA that will antagonize sulfonamides. Thus, sulfonamides should not be combined with procaine penicillin therapeutically.

Toxicity and Adverse Effects The sulfonamides can produce a wide variety of usually reversible adverse effects, which may be divided into dose-­dependent (Type A) and idiosyncratic (Type B) reactions.  Of the dose-­ dependent reactions, ­gastrointestinal upset occurs fairly commonly. Sulfonamides have been associated with Clostridioides (Clostridium) difficile enterocolitis in horses, which may be fatal (Diab et  al.,  2013). Renal tubular necrosis can

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also occur as a result of drug precipitation and tubular crystalluria. However, newer sulfonamides such as sulfamethoxazole and sulfadimethoxine have higher solubility in normal urine pH and so are less likely to precipitate. Interestingly, acetylated sulfonamide metabolites have lower aqueous solubility so dogs, which lack the N-­acetyltransferase enzymes, may be at lower risk for nephrotoxicity. Sulfonamide antimicrobials also have dose-­dependent antithyroid effects via reversible inhibition of thyroid peroxidase. In dogs, sulfonamides reliably decrease circulating thyroid concentrations and can result in clinical hypothyroidism in a dose-­ and time-­dependent manner. In late gestational sows, ormetoprim-­ sulfadimethoxine has been documented to cause goitrous disease, increasing the number of stillborn or weak piglets (Blackwell et al., 1989). In a small proportion of animals (approximately 0.25% of dogs), sulfonamide therapy can produce idiosyncratic drug reactions, which are unpredictable and rare events occurring five days to several weeks after first exposure. These adverse events are thought to be due to a hypersensitivity reaction to the drug haptenized to endogenous proteins via its arylamine ring. The syndrome in dogs may include fever, ­polyarthropathy, blood dyscrasias, hepatopathy, epistaxis, skin eruptions, uveitis, retinitis, lymphadenopathy, proteinuria, edema, pneumonitis, pancreatitis, and neurological complications (Trepanier, 2003). Isolated cases of skin eruptions, uveitis, blood dyscrasias, and neurological signs have also been reported in horses. However, among the veterinary species, dogs appear to be at increased risk for sulfonamide hypersensitivity, possibly because they lack the ability to acetylate the arylamine ring, protecting it from oxidation and haptenization. Doberman Pinschers appear to be particularly susceptible, which may have a genetic basis (Reinhart et al., 2018). In contrast to most other sulfonamide-­ related morbidities, keratoconjunctivitis sicca (KCS) is a fairly common occurrence in dogs, with a reported incidence of 15% (Berger et al., 1995). Sulfonamide-­associated KCS also

appears to be more common in small breeds and is delayed in onset relative to other manifestations, occurring months after initiation of antimicrobial therapy. There is also evidence that some sulfonamide antimicrobials may be  directly toxic to the lacrimal gland mediated  by a pyrimidine or pyridine ring in the R-­group structure. In these ways, sulfonamide-­ associated KCS may be a separate reaction with a distinct pathogenesis, rather than part of the classic idiosyncratic hypersensitivity. However, KCS can occur in combination with other classic signs of sulfonamide hypersensitivity, so the classification of this adverse reaction remains unclear. Accidental overdose of sulfaquinoxaline in a juvenile broiler flock resulted in hemorrhage and renal tubular injury (Fulton and Buchweitz, 2023). Unlike other sulfonamides, sulfaquinoxaline is an inhibitor of the dithiothreitol-­dependent reduction of both vitamin K epoxide and vitamin K quinone, similar to coumarins and hydroxyquinones.

Administration and Dosage Dosages for sulfonamide antimicrobials used in veterinary medicine are presented in Table 16.1 Although it has previously been recommended to double the dosage on the first day and some veterinary-­approved product labels include this recommendation, loading doses of sulfonamide antimicrobials are no longer commonly used in veterinary practice. Although a large number of sulfonamide preparations are available for use in veterinary medicine, many of these are different dosage forms of sulfamethazine. This sulfonamide is most widely used in food-­producing animals and can attain effective plasma concentrations when administered either orally or parenterally. Because of their alkalinity, most parenteral preparations should be administered only by IV injection. Rapid IV injection of high doses of sulfonamide preparations should be avoided due to concerns for hypotension. At least one prolonged-­release

309

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­Sulfonamide 

Sulfonamides, Diaminopyrimidines, and Their Combinations

Table 16.1  Examples of usual dosages of sulfonamides in animals. Dosing interval (hrs)

Comment

Drug

Route

Dose (mg/kg)

Short-­acting sulfadiazine, sulfamethazine

IV, PO

50–­60

12

Double first dose

IV, IM, SC, PO

27.5

24

Double first dose

PO

137.5

96

Intermediate-­acting sulfadimethoxine sustained release, cattle sulfadiazine

PO

50

8

Double first dose

Sulfisoxazole

PO

50

8

Urinary tract infections

Gut-­active phthalylsulfathiazole

PO

100

12

sulfasalazine

PO

25

12

Silver sulfadiazine

Topical

Special-­use See text

IM, intramuscular; IV, intravenous; PO, by mouth (per os); SC, subcutaneous.

oral preparation of sulfamethazine is available for use in calves and could be administered to sheep and goats. This is a convenient form of maintenance therapy in that a single dose provides an effective level for 36–48 hours. Different oral forms have different systemic availability. Sulfadimethoxine preparations are used in small animals in addition to cattle. In dogs and cats, sulfadimethoxine is administered as an oral tablet or suspension. A 40% parenteral solution as well as oral bolus forms are available for use in cattle. In these species, ­sulfadimethoxine is usually administered at a dosage of 27.5 mg/kg daily. Also, a sustained-­ release oral formulation is available for cattle in some countries. Unlike the sodium salts of other sulfonamides, sodium sulfacetamide is nearly neutral. It is the only sulfonamide available for topical ophthalmic use. When a 30% solution is applied to the conjunctivae, it penetrates well and attains high concentrations in ocular fluids and tissues.

Clinical Use Widespread resistance greatly limits the effectiveness of sulfonamides in treating bacterial diseases of animals, so that indications for primary use are few. Trimethoprim– or other antibacterial diaminopyrimidine–sulfonamide combinations have largely replaced sulfonamides as therapeutic agents used in companion animals, although resistance also increasingly limits their use. Purulent material must always be removed, since free purines neutralize the effect of sulfonamides. Primary uses include treatment of toxoplasmosis (when combined with pyrimethamine), of chlamydiosis, of Pneumocystis spp., and possibly of nocardiosis (combined with minocycline), and the use of sulfasalazine in the treatment of chronic colitis. Cattle, Sheep, and Goats

Widespread resistance limits the use of sulfonamides in these animals, and it is best to give these  agents in combination with ­trimethoprim. Orally administered, long-­acting, sustained-­release

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dosage forms result in effective plasma concentrations for 3–5  days. Such a preparation has been effective in clinical trials assessing prevention and treatment of feedlot pneumonia, an unexpected result in view of the resistance reported in bovine Mannheimia and Pasteurella spp. Sulfonamides are used successfully to treat bovine interdigital necrobacillosis and coccidiosis. Sulfadimethoxine is the only sulfonamide approved for use in dairy cows over 20 months of age in the United States; extra-­label use of all sulfonamides in dairy cows is prohibited. Sustained-­release oral sulfamethazine and orally administered pyrimethamine, 0.5 mg/ kg once daily, might be drugs of choice in preventing outbreaks of Toxoplasma abortion in sheep. Sulfonamides have been used with chlortetracycline in feedlot lambs to prevent clostridial enterotoxemias. Swine

Sulfonamides have been used to control group E streptococcal infections and atrophic rhinitis caused by Bordetella bronchiseptica in swine. The sulfonamides are often combined with chlortetracycline. Horses

Sulfonamides are used commonly in horses in combination with antibacterial diaminopyrimidines. For the treatment of equine protozoal myeloencephalitis, sulfadiazine (20  mg/kg PO SID or BID, for up to 12 weeks or longer) is combined with pyrimethamine (1.0 mg/kg PO SID, for up to 120 days or longer) (Reed et al., 2016). Dogs and Cats

Sulfadimethoxine is commonly used for the treatment of isosporosis (coccidiosis) in dogs and cats at a dosage of 50–60 mg/kg/day orally for 5–20  days. Other than this indication, single-­agent sulfonamides have largely fallen  out of systemic use in small animals. Sulfonamides are one of the drugs of choice in the treatment of Nocardia infections, but combination products with diaminopyrimidines are more often used. Silver sulfadiazine cream has been used as a treatment in chronic otitis

externa caused by multidrug-­resistant P. aeruginosa, as the drug acts as a broad-­spectrum antimicrobial antiseptic. This preparation has been effective in controlling bacteria that infect burn wounds in human patients; activity is almost certainly the result only of the silver component. Sulfasalazine (salicylsulfapyridine) has been recommended in the treatment of canine chronic enteropathy. It is hydrolyzed by intestinal bacteria to yield sulfapyridine and 5-­aminosalicylate; it is likely that the antiinflammatory effect of the latter is responsible for the therapeutic effect. Comparably high concentrations of salicylate cannot be achieved in the colon by oral administration. The dosage of sulfasalazine for the dog is 25  mg/kg PO three times daily. The same dose in cats may induce salicylate poisoning. Poultry

Sulfonamides have been used in the prevention and treatment of coccidiosis, infectious coryza, pullorum disease, and fowl typhoid.

Environmental Impact As the oldest synthetic antimicrobial class currently in use, sulfonamides have been around for almost 90 years. This prolonged and widespread use in human and veterinary medicine has led to significant environmental contamination. Consequences include alterations in environmental bacterial populations; adverse effects on aquatic organisms including algae, plants, and fish; and drug accumulation in foods intended for human consumption including honey, plants, seafood, and farmed animals. These impacts emphasize the need for judicious use of sulfonamides by veterinarians, particularly in production medicine where mass medication practices are common. These practices have high potential for environmental contamination through livestock waste waters and manure (Nunes et al., 2020).

311

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­Sulfonamide 

Sulfonamides, Diaminopyrimidines, and Their Combinations

­ ntibacterial Diaminopyrimidines: A Aditoprim, Baquiloprim, Ormetoprim, and Trimethoprim Diaminopyrimidines interfere with folic acid production by inhibition of dihydrofolate reductase. Some diaminopyrimidines have marked specificity for bacterial dihydrofolate reductases (aditoprim, baquiloprim, ormetoprim, trimethoprim), others for protozoal enzymes (pyrimethamine), and others for mammalian enzymes (methyltrexate). The earliest antibacterial diaminopyrimidine introduced for clinical use was trimethoprim (Figure  16.3), a synthetic drug that is widely used in combination with

NH2 OCH3 CH2

N

N

NH2

OCH2 OCH3

Trimethoprim NH2 OCH3 CH2

N

N

NH2

OCH2

Baquiloprim

N

sulfonamides. It is a weak base with a pKa of about 7.6 and is poorly soluble in water. Other antibacterial diaminopyrimidines have similar antibacterial activities to trimethoprim but offer significant pharmacokinetic advantages, particularly those of greater half-­lives and tissue distribution.

Mechanism of Action Diaminopyrimidines interfere with the synthesis of tetrahydrofolic acid from dihydrofolate by combining with the enzyme dihydrofolate reductase (Figure  16.2). Selective antibacterial activity occurs because of greater affinity for the bacterial  rather than the mammalian enzyme. Diaminopyrimidines thus inhibit the same metabolic sequence as the sulfonamides, preventing bacterial synthesis of purines and thus of DNA. A synergistic and bactericidal effect occurs when the diaminopyrimidines are combined with sulfonamides (see Antibacterial Diaminopyrimidine– Sulfonamide Combinations), and for this reason these drugs are invariably used with a sulfonamide in veterinary medicine. Interestingly, in the United Kingdom trimethoprim alone rather than the combination is recommended for first-­line therapy of uncomplicated urinary tract infections in adult women (SIGN, 2012). This recommendation primarily is derived from antimicrobial stewardship concerns. However, other reasons supporting the use of single-­agent trimethoprim include the following. ●●

NH2 OCH3 CH2

N

NH2

N

N OCH3

CH2

●●

CH2

Aditoprim

Figure 16.3  Structural formulas of some diaminopyridines.

●●

Bactericidal synergy between trimethoprim and sulfonamides is only demonstrable when the concentration of each drug is less than bacteriostatic, but the bacteriostatic effect of trimethoprim in urinary tract infections is often detectable in urine for several days. Diaminopyrimidines are more widely distributed into tissues than sulfonamides, reaching sites, such as inside cells, which sulfonamides do not penetrate well. Most of the adverse effects of the combination are the result of the sulfonamide component.

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●●

The original claim that the combination prevented the emergence of resistance is dubious because sulfonamide resistance is widespread and because plasmids conferring sulfonamide resistance often also confer resistance to trimethoprim.

Antimicrobial Activity Antibacterial diaminopyrimidines are generally bacteriostatic and active against Gram-­positive and Gram-­negative aerobic bacteria, but not ­usually against anaerobes. Bacteria with an MIC 0.5/9.5 μg/ ml) is shown among the following Gram-­ positive aerobes: S. aureus, streptococci, Arcanobacterium spp., Corynebacterium spp., E. rhusiopathiae, L. monocytogenes. Gram-­negative aerobes: Acinetobacter spp.,  Actinobacillus spp., Bordetella spp., Burkholderia cepacia, Brucella spp., Dermatophilus congolense, Enterobacterales (E. coli, Klebsiella spp., Proteus spp., Salmonella spp., Yersinia spp.), Histophilus spp., Pasteurella spp., Stenotrophomonas maltophila. Anaerobes: Actinomyces spp., Bacteroides spp., Fusobacterium spp., some Clostridium spp., and Chlamydia spp. Intermediate susceptibility (MIC >2/38 μg/ ml) includes some Mycobacterium spp. and some Nocardia spp. Intrinsic resistance (MIC >4/76 μg/ml) is shown by Rickettsia, Leptospira spp., P. ­aeruginosa, enterococci, and Mycoplasma spp.

Resistance Mechanisms of resistance were discussed under the individual components of the combination. Resistance to the combination has developed progressively with use. Multiple integron-­ associated resistance, which includes both sulfonamide and trimethoprim resistance, has

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­Antibacterial Diaminopyrimidine–Sulfonamide Combinations (Potentiated Sulfonamides  315

Sulfonamides, Diaminopyrimidines, and Their Combinations

been described in some Salmonella serovars and in pathogenic E. coli isolated from animals.

Pharmacokinetic Properties In people, the half-­lives of trimethoprim and sulfamethoxazole are similar, and maintenance dosages provide continuous, therapeutic concentrations of both drugs in plasma. In animals, the half-­lives of the drugs are not similar, but the combination is often clinically effective because of the relatively broad range of drug ratio over which synergism occurs. The diaminopyrimidine component is concentrated in tissues whereas the sulfonamide component moves only slowly from plasma into tissues. The longer half-­lives of newer diaminopyrimidines (baquiloprim, ormetoprim) give the advantage of better maintenance of the 1:20 ratio said to be desirable, and of less frequent dosing. Following SC injection in cattle, trimethoprim seems to deposit in a slow-­release form, so that serum concentrations remain below MIC (Kaartinen et al., 2000). Because of this, the SC route cannot be recommended in cattle and perhaps in other species.

Drug Interactions Trimethoprim–sulfonamide has sometimes been used in conjunction with ampicillin to provide “broad-­spectrum bactericidal antimicrobial coverage” before microbiology data are available. However, one study showed that addition of ampicillin to trimethoprim–­sulfonamide dosing regimens only marginally increased the spectrum of activity. There is no known mechanism to suggest that such a combination might be synergistic. Rather, such a combination may be effective in treating polymicrobial infections involving aerobic bacteria susceptible to the ­trimethoprim–sulfonamide combination and anaerobic bacteria susceptible to ampicillin. Sulfonamide antimicrobials are cytochrome P450 (CYP) substrates and both sulfonamides and trimethoprim are known CYP inhibitors. Therefore, drug–drug interactions with other

CYP substrates, inducers, or inhibitors are possible and should be considered in multidrug protocols.

Toxicity and Adverse Effects The combination has a wide margin of safety, and adverse effects can mainly be attributed to the sulfonamide. These effects are discussed in the general description of the adverse effects of each drug class. In horses, minor tissue damage and pain may occur after IM injection; transient pruritus has been reported to follow the first but not subsequent doses. In isolated incidents a fatal adverse reaction followed IV injection of the combination preparation in horses, in some cases in anesthetized horses. In vitro investigations suggest that these incidents could be caused by potassium channel blockade by the trimethoprim component and subsequent fatal arrhythmias (Trachsel et al., 2018). A 7% incidence of diarrhea was observed in a study of the effect of twice-­daily administration of oral 30  mg/kg trimethoprim–sulfadiazine in horses. The prevalence of diarrhea noted following trimethoprim–­ sulfonamide use in horses in another study was not significantly different from that observed in horses receiving other antimicrobials, including penicillin (Wilson et al., 1996). Neurological abnormalities in horses characterized by reversible hypermetric gait, agitation, and erratic behavior have been described as an unusual adverse reaction (Stack et al., 2011). Sulfonamides cross the placenta and are tetratogenic at very high doses. Abortions can occur if given to pregnant mares due to placentitis. Direct infusion of the combination into the uterus may cause endometrial inflammation.

Administration and Dosage Usual dosages are shown in Table 16.2. Dogs and cats can be given the oral form (tablets or suspension) at the same dosage. Higher doses may be ­recommended for the treatment of certain ­protozoal infections. In horses, 30  mg/kg

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Table 16.2  Usual dosages of potentiated sulfonamide combinations in animals. Dose (mg/kg)

Dosing interval (hrs)

PO, IV, IM

30

24

May be divided q12 h Not IM in horses

PO

27.5

24

Double first dose May be divided q12 h

Drug (species)

Route

Trimethoprim-­ sulfonamide Ormethoprim-­ sulfadimethoxine Baquiloprim-­ sulfadimethoxine Dogs Cats Cattle, swine

Comment

Not available in the United States PO PO IM

30 30 10

48 24 24

IM, intramuscular; IV, intravenous; PO, by mouth (per os).

trimethoprim–sulfadiazine should be administered twice daily rather than once daily due to rapid elimination, particularly of the trimethoprim component, and the need to maintain therapeutic concentrations above the MIC for the majority of the dosing interval (van Duijkeren et  al.,  1994). These considerations may be further complicated by the biphasic absorption of the sulfonamide component observed in horses (see Sulfonamides, Administration and Dosage). Equine oral formulation options include a combination paste and a powder; availability varies by region. Sulfonamide clearance is apparently greater in the donkey compared to the horse, so dosage adjustments for this species may be required.

Clinical Applications Diaminopyrimidine–sulfonamide combinations have the advantage of good distribution into tissues, safety, a relatively broad-­spectrum bactericidal activity, and oral administration. A disadvantage is antagonism of action by infected tissue debris. The combination can be recommended in the treatment of urinary tract infections caused by common opportunist pathogens and has a particular place in the treatment of bacterial prostatitis because of good tissue penetration. Other indications include the treatment of enteric infections (E. coli,

Salmonella, Y. enterocolitica). The drug is of value in the treatment of canine brucellosis, often in combination with rifampin or doxycycline. The combination is a drug of choice in the treatment of Nocardia infections, but high oral dosage (3  mg trimethoprim ­equivalent/kg q6 h) must be used for prolonged periods. Other indications include the treatment of Neospora caninum, Pneumocystis spp., Toxoplasma gondii, Chlamydia and Chlamydophila infections, listeriosis, certain fast-­growing mycobacterial infections (M. kansasii, M. marinum), and Coxiella infections. In human medicine, the combination is used for the treatment of otherwise resistant infections caused by Acinetobacter, Burkholderia, and Stenotrophomonas spp., as well as methicillin-­ resistant S. aureus (MRSA) (Goldberg and Bishara,  2012). Livestock-­associated MRSA has, however, been associated with multiple drug resistance, including a novel trimethoprim resistance gene (dfrK) (Kadlec et al., 2012). The drug is also used in the treatment of acute upper and lower respiratory tract infections caused by susceptible organisms, as well as in infections in other sites. Cattle, Sheep, and Goats

The drug combination has been widely used in dairy and beef cattle in the treatment of salmonellosis in calves, as well as in undifferentiated ­diarrhea, bacterial pneumonia, footrot, and

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Sulfonamides, Diaminopyrimidines, and Their Combinations

s­ epticemic colibacillosis. It has largely been replaced by other antimicrobial drugs for which resistance is less of a problem. Baquiloprim– sulfadimidine was not as efficacious as danofloxacin in the treatment of experimentally induced E. coli diarrhea in calves (White et al., 1998), presumably because the organism is less susceptible to the combination drug. Although trimethoprim–sulfonamides have been used in the past for the prevention and treatment of bovine respiratory disease complex, several recent metaanalyses have demonstrated that the drug combination is largely ineffective for this indication (O’Connor et  al.,  2016,  2019). Other options, particularly macrolide antibiotics, are more efficacious and should be used instead. The potential for ­diaminopyrimidine–sulfonamide combination use in coliform septicemia and meningitis seems excellent but is increasingly limited by resistance. In meningitis the drug should be administered IV 3–4 times daily at the usual dosage. The potential for use in the treatment of Listeria meningoencephalitis appears excellent. Experimental studies have confirmed the antagonistic effect of infected tissue debris on the action of the combination and so use should be avoided in highly purulent disease or after debridement (Greko et al., 2002). When used to treat acute mastitis, the drug should be given IV at high dose because of poor bioavailability after IM injection and relatively poor udder penetration; a dosage of 48–50 mg/kg every 12 hours is appropriate for acute mastitis. A beneficial effect of ­trimethoprim–sulfonamide on the treatment of coliform mastitis has been noted, particularly when combined with nonsteroidal antiinflammatory drugs (Shpigel et al., 1998). Other uses in cattle include the treatment of urinary tract infections. The drug has potential but unproven use for the treatment of L. monocytogenes encephalitis in ruminants. A special application in goats and sheep is in preventing Toxoplasma abortion; the drug is also potentially useful in preventing chlamydial abortion in sheep. In experimental Toxoplasma infections in mice, protection by trimethoprim–sulfonamide

was inferior to pyrimethamine–sulfadiazine, but clinical results in naturally occurring infections in humans have been excellent. Swine

Trimethoprim–sulfonamide combinations have been used successfully in controlling a wide variety of conditions in pigs, including neonatal and postweaning colibacillosis, salmonellosis, atrophic rhinitis, greasy pig disease, Streptococcus suis-­associated septicemia, and pneumonia, but it has largely been replaced by other antimicrobial drugs for which resistance is less a problem. Atrophic rhinitis may be controlled by incorporating the drug in feed or water, or by injecting piglets at various times such as the third day of life and again in the third and sixth weeks. The mastitis-­metritis-­agalactia syndrome has been controlled by the prophylactic administration of 15 mg/kg PO for three days before and day days after parturition. The combination has been used in the eradication of A. pleuropneumoniae infection from herds by treating adults through the water for three weeks in combination with removal of serologically positive animals. Isolates of MRSA from clinical infections in Dutch swine were all found to be susceptible to the combination (Wolf et al., 2012), in marked contrast to nasal isolates from swine in Belgium (Crombé et al., 2012); the ST398 strain found in swine appears to be able readily to acquire multiple resistance genes (Argudin et al., 2011). Other diaminopyrimidine–sulfonamide combinations are available for swine for similar purposes to trimethoprim–sulfonamide combinations (Table  16.2). Susceptibility testing is required before instituting treatment in view of the variable reports of resistance of common swine pathogens to the combination, including bacteria such as H. parasuis that used to be highly susceptible. Horses

Trimethoprim–sulfonamide combinations are the most popular antimicrobials used in equine practice (Tallon et al., 2023), likely because they can be administered as an oral formulation to horses with few adverse effects. They are painful when administered IM. They are therefore

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used orally to treat acute respiratory infections including strangles, acute urinary tract infections, and wounds and abscesses, and may be a drug of choice in salmonellosis. In recent years, however, resistance has apparently increased in Streptococcus equi subsp. zooepidemicus, so that in some studies less than 90% of isolates are susceptible in  vitro (Peyrou et  al.,  2003). However, Feary et  al. (2005) have shown that reports of resistance may represent laboratory error, which is still quite frequent. The combination was ineffective in eradicating S. equi subsp. zooepidemicus in a tissue chamber model of infection despite in vitro susceptibility of the isolate and high concentrations of the drugs in the tissue chamber fluid (Ensink et  al.,  2003). For these reasons, and because it can be partially antagonized by tissue debris, it is a less desirable choice than procaine penicillin G for treatment of streptococcal infections. The combination of sulfadiazine with pyrimethamine is a drug of choice in the treatment of protozoal encephalomyelitis (see Antiprotozoal diaminopyrimidines). It is a drug of choice for Pneumocystis spp. infections in foals. Dogs and Cats

Trimethoprim–sulfonamide or ormetoprim–­ sulfadimethoxine combinations have wide application in dogs and cats against specific and nonspecific infections. The combination is highly effective against many opportunist bacteria present in canine urinary tract, skin, and ear infections (S. pseudintermedius, streptococci, and Enterobacterales including E. coli and Proteus). It is considered a first-­line, empirical treatment for uncomplicated lower urinary tract infections (Weese et  al.,  2019). In female dogs without recurrent signs, a short, three-­day course of ­trimethoprim–sulfamethoxazole (15  mg/kg PO q12 h) has been shown to have similar cure rates to a 10-­day course of a beta-­lactam (Clare et al., 2014). This short-­duration protocol, similar to what is used in women, may decrease the development of antimicrobial resistance and the likelihood for adverse effects to the drug. In recent reports, susceptibility to potentiated sulfonamides in urinary isolates from

small animals has been generally high (~90%), supporting their use in empirical therapy (Wong et  al.,  2015; Scarborough et  al.,  2020). However, susceptibility can vary between pathogens, host species, and geographic locations (Marques et  al.,  2016), so clinicians should be aware of current trends in their regions. Furthermore, susceptibility to potentiated sulfonamides is significantly lower in recurrent urinary tract infections, so culture should always be performed in these cases even if empirical therapy is instituted prior to receiving results (Wong et  al.,  2015). In particular, methicillin-­resistant S. pseudintermedius (MRSP) are often resistant to trimethoprim, as part of their common multidrug resistance (Perreten et al., 2010). Potentiated sulfonamides are also considered a first-­tier therapy for superficial bacterial folliculitis in the dog. However, if treating empirically, regional susceptibility patterns for S. pseudintermedius should be taken into account (Hillier et al., 2014). The combination drug is effective against Bordetella bronchiseptica, although relapses after treatment with ­trimethoprim–sulfadiazine for five days were common in experimental kennel cough. In one study, a significant number of isolates were found to be resistant to the combination drug (Speakman et  al., 2000). In general, potentiated sulfonamides are not considered a first-­ line therapy for respiratory infections in dogs and cats and are primarily indicated based on culture and susceptibility results (Lappin et al., 2017). The drug has been used successfully in the treatment of canine actinomycosis, although penicillins are the drug of choice. The combination has been effective in treating coccidiosis in dogs and cats, although single-­ agent sulfadimethoxine is generally used. The excellent penetration of diaminopyridines into the prostate makes the combination product a treatment of choice in prostatic infections in dogs, although fluoroquinolones may be a better empirical choice while awaiting culture results due to the possibility of Brucella canis infection (Weese et  al.,  2019). Similarly, the excellent penetration (50% of

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serum concentrations) of the aqueous and vitreous humors of the eyes by both drugs makes the combination suitable in the parenteral treatment of panophthalmitis caused by Gram-­ negative bacteria. The combination is used together with clindamycin and pyrimethamine in the initial treatment of Hepatozoon americanum infections in dogs. The combination is also used with clindamycin in the treatment of Neospora caninum infection and may be considered for toxoplasmosis in both dogs and cats. Poultry

Trimethoprim–sulfaquinoxaline and ­sulfamethoxazole–ormetoprim are used in the prophylaxis and treatment of E. coli, Haemophilus, and Pasteurella infections, as well as coccidiosis in poultry species. The combination has been used successfully in the treatment of Plasmodium gallinaceum malaria in chickens (Williams, 2005). Depending on the extent of use in different countries, which varies, resistance can be widespread among E. coli isolated from broilers. The combination has also been used to treat Riemerella anatipestifer infection in ducks.

­Antiprotozoal Diaminopyrimidines Some diaminopyrimidines such as pyrimethamine have high activity against protozoa by inhibiting dihydrofolate reductase and thus preventing purine synthesis. These drugs are used in the treatment of systemic protozoal infections such as toxoplasmosis, neosporosis, and equine protozoal myelitis. They are also highly active against Pneumocystis spp. Pyrimethamine and sulfadiazine are the most effective drugs in the treatment of toxoplasmosis

in humans and are generally preferred over alternatives such as azithromycin and trimethoprim– sulfamethoxazole. The adult human dosage is 75 mg pyrimethamine and 4 g sulfadiazine PO/ day in four divided doses, administered for up to four weeks. Dapsone combined with pyrimethamine has good activity experimentally against Toxoplasma. Combination of pyrimethamine (1  mg/kg) and sulfonamide antimicrobial (20 mg/kg) is a standard treatment for equine protozoal myeloencephalitis (EPM) (Reed et  al.,  2016). A pyrimethamine/sulfadiazine oral suspension is available, but the components may be administered separately. If pyrimethamine is combined with a potentiated sulfonamide, the trimethoprim component is unnecessary. A small proportion of horses may develop progressive anemia during treatment. Alternate drugs for the treatment of EPM in pregnancy are required, since pyrimethamine is teratogenic for animals and may lead to myeloid, erythroid or lymphoid hypoplasia with epithelial dysplasia and renal hypoplasia or nephrosis in newborn foals. Such effects may be exacerbated by administering folic acid to mares being treated for EPM (Toribio et  al.,  1998). About 60% of horses with moderate to severe EPM will improve with any of the FDA-­approved treatments (sulfadiazine–pyrimethamine, ponazuril or diclazuril) (Reed et al., 2016). Pyrimethamine and diaveridine are commonly combined with sulfaquinoxaline for their synergistic effect against coccidia. Pyrimethamine (1 mg/kg daily) combined with a sulfadoxine (20 mg/kg daily) or ­trimethoprim– sulfadiazine has been used successfully in the treatment of Neospora caninum infection in dogs (Thate and Laanen, 1998).

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Trepanier LA. 2004. Idiosyncratic toxicity associated with potentiated sulfonamides in the dog. J Vet Pharmacol Ther 27:129. Twedt DC, et al. 1997. Association of hepatic necrosis with trimethoprim-­sulfonamide administration in 4 dogs. J Vet Intern Med 11:20. Van Duijkeren E, et al. 1994. A comparative study of the pharamacokinetics of intravenous and oral trimethoprim/sulphadiazine formulations in the horse. J Vet Pharmacol Ther 17:440. Van Duijkeren E, et al. 1995. Pharmacokinetics and therapeutic potential for repeated oral doses of trimethoprim/sulphachlorpyridazine in horses. Vet Rec 137:483. Van Duijkeren E, et al. 1996. In vitro and in vivo binding of trimethoprim and sulphachlorpyridazine to equine food and digesta and their stability in caecal contents. J Vet Pharmacol Ther 19:281. Van Miert ASJPAM. 1994. The sulfonamide-­ diaminopyrimidine story. J Vet Pharmacol Ther 17:309. Weese JS, et al. 2019. International Society for Companion Animal Infectious Diseases (ISCAID) guidelines for the diagnosis and management of bacterial urinary tract infections in dogs and cats. Vet J 247:8.

White DG, et al. 1998. Comparison of danofloxacin with baquiloprim/ sulphadimidine for the treatment of experimentally induced Escherichia coli diarrhoea in calves. Vet Rec 143:273. Williams RB. 2005. The efficacy of a mixture of trimethoprim and sulphaquinoxaline against Plasmodium gallinaceum malaria in the domesticated fowl Gallus gallus. Vet Parasitol 129:193. Wilson RC, et al. 1989. Bioavailability and pharmacokinetics of sulfamethazine in the pony. J Vet Pharmacol Ther 12:99. Wilson WD, et al. 1996. Case control and historical cohort study of diarrhea associated with administration of trimethoprim-­ potentiated sulfonamides to horses and ponies. J Vet Intern Med 10:258. Wolf PJ, et al. 2012. Staphylococcus aureus (MSSA) and MRSA (CC398) isolated from post-­mortem samples from pigs. Vet Microbiol 158:136. Wong C, et al. 2015. Antimicrobial susceptibility patterns in urinary tract infections in dogs (2010–2013). J Vet Intern Med 29:1045. Wu S, et al. 2010. Prevalence and characterization of plasmids carrying sulfonamide resistance genes among Escherichia coli from pigs, pig carcasses and human. Acta Vet Scand 52:47.

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 ­Reference

17 Fluoroquinolones Patricia M. Dowling

I­ ntroduction The fluoroquinolones, also known as quinolones, 4-­quinolones, pyridine-­beta-­carboxylic acids, and quinolone carboxylic acids, are a large and expanding group of synthetic antimicrobial agents. The first of these compounds, nalidixic acid, was initially described in 1962, introduced into clinical practice in 1963, and then approved for clinical use in 1965. Nalidixic acid had limited clinical application because of its poor absorption following oral administration, its moderate antibacterial activity (MICs of 4–16 μg/ml for Enterobacterales), high protein binding (92–97%), and poor patient tolerance. Between the mid-­1960s and the early 1980s several other quinolones were approved for clinical use in humans, for example, oxolinic acid, pipemidic acid, piromidic acid, and flumaquine. These drugs exhibited increased antibacterial activity but still had limited absorption and distribution. In the 1980s, the addition of both a fluorine molecule at the 6 position of the basic quinolone structure and a piperazine substitution at the 7 position enhanced the antibacterial activity of these compounds, with activity against Pseudomonas aeruginosa and staphylococci. These modifications also increased the

oral absorption and tissue distribution. The ­ uinolone nucleus with the fluorine molecule q gave the group the name “fluoroquinolones.” The first fluoroquinolone approved for use in human clinical medicine was norfloxacin, followed shortly thereafter by ciprofloxacin. The first fluoroquinolone approved for use in animals was enrofloxacin, which was approved for use in the United States in companion animals in 1988. The fluoroquinolones and their current clinical uses in veterinary medicine are listed in Table 17.1. The World Health Organization (WHO) classifies the fluoroquinolones as Critically Important Antimicrobials for human medicine as they are needed for treatment for Campylobacter spp. infections, invasive disease due to Salmonella spp., and multidrug-­resistant Shigella spp. infections. The clear evidence that treatment of entire groups of animals with fluoroquinolones selects for resistance in important zoonotic pathogens mandates that these drugs should be used in ­veterinary medicine only when supporting ­laboratory data demonstrate that no suitable alternatives of less human health importance are  available and use as mass medications should be restricted (Kenyon, 2021; McEwen and Collignon, 2018).

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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325

Fluoroquinolones

Table 17.1  Fluoroquinolones used in veterinary medicine.a Fluoroquinolone

Comments

Enrofloxacin

Available as tablets and injectable formulation for dogs and cats and as an injectable solution for cattle. Only approved for treatment and respiratory diseases in beef and nonlactating cattle and swine in the United States and Canada.a Approved uses vary widely between countries, with some approvals for lactating dairy cows, swine, rabbits, fin fish, and poultry. Canine otic formulation available in some countries. Used extra-­label in horses and exotic animals.

Ciprofloxacin

In Europe and North America, injectable solutions, tablets, and ophthalmic formulations only approved for humans but used extra-­label in small animals. Available in veterinary formulations in some countries.

Danofloxacin

Only approved for treatment of respiratory disease in cattle in the United States and Canada, but approved for use in cattle, swine, and poultry in Europe.

Difloxacin

Injectable formulations for ruminants, camelids, poultry, swine, and dogs are available in some countries. Approved tablets for dogs have been discontinued. Oral poultry formulations also available in some countries.

Marbofloxacin

Available as small animal oral formulations in the United States and Canada. Large animal injectable formulations are available in Canada, Europe and other countries. Canine otic formulation available in some countries. Used extra-­label in horses.

Pradofloxacin

Oral formulations for use in dogs and cats.

Orbifloxacin

Oral formulations for use in dogs and cats. Canine otic formulation available in some countries. Used extra-­label in horses.

a

 Extra-­label use of fluoroquinolones in food-­producing animal species is prohibited in the United States.

C ­ hemistry The fluoroquinolones, like sulfonamide and nitrofurans, are synthetic compounds so they are antimicrobials but not antibiotics. The first clinically approved 4-­quinolone-­type compound was nalidixic acid. Since the discovery of nalidixic acid’s antibacterial activities, more than 10 000 compounds have been designed from the parent bicyclic 4-­quinolone molecule. Clinically, nalidixic acid had several limitations, including a narrow spectrum of activity, poor pharmacokinetic properties, toxic effects, and a tendency to select for resistant organisms. Replacing the hydrogen atom at position 6 of the 4-­quinolone molecule with a fluorine atom resulted in increased activity against both Gram-­positive and Gram-­negative bacteria. The increased activity is attributed to increased penetration of the bacterial cell membrane. Substituting a piperazinyl ring for the methyl

group at position 7  increased Gram-­negative activity, including antipseudomonal activity. These modifications led to the development of the first broad-­spectrum fluoroquinolone, norfloxacin. Additional studies demonstrated that substantial changes in potency could be obtained by variations at the N-­1 and C-­7 ­positions. For example, ciprofloxacin is similar in structure to norfloxacin but has a cyclopropyl group in place of the ethyl group at N-­1. This substitution enhances ciprofloxacin’s Gram-­positive and Gram-­negative activity. This cyclopropyl group is also found on enrofloxacin, danofloxacin, pradofloxacin, and orbifloxacin (Figure  17.1). Difloxacin has a phenyl ring at position N-­1 that gives it enhanced activity against Gram-­positive bacteria, relative to enrofloxacin. Difloxacin also has a second ­fluorine atom in its structure, whereas orbifloxacin has a total of three fluorine atoms. These additional fluorine atoms do not appear

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326

O

O H

5 7

8

1

N

COOH

F

2

COOH

F

N

N N

C2H5 Nalidixic Acid

N

N

N

COOH

N

Enrofloxacin

O COOH

F N

N

N

F CH3

C2H5

Ciprofloxacin

CH3 N

N

O

O

F

COOH

F N

N

CH3

Orbifloxacin

N

N CH3

Danofloxacin O

O COOH

F N

N

N

N

C2H5 Norfloxacin

F

COOH

F

O

O

3

6

CH3

COOH

4

F Difloxacin

N

N

HN

N HCL

COOH

F

CH3

N O

N CH3

Marbofloxacin F Sarafloxacin

Figure 17.1  Structures of fluoroquinolones used in veterinary medicine.

to influence the antibacterial activity of these compounds. Overall, there have been several chemical modifications at each of the eight positions in the 4-­quinolone molecule. Some increase absorption, some increase antibacterial activity, and others increase toxicity. For example, ciprofloxacin and enrofloxacin are similar molecules except for the ethyl group on the piperazinyl ring of enrofloxacin. This ethyl group enhances the oral absorption of enrofloxacin over ciprofloxacin but decreases its antipseudomonal activity. Biological classification places the 4-­quinolones in four groups or generations (Pham et  al.,  2019). First-­generation quinolones are those with antibacterial activity restricted to the Enterobacterales (e.g., nalidixic acid and flumequine). Second-­generation quinolones have an extended spectrum of antibacterial activity. Most fluoroquinolones

approved for use in people (including ciprofloxacin, norfloxacin, and ofloxacin) and all but one of the fluoroquinolones approved for use in veterinary medicine are second-­generation fluoroquinolones. Third-­generation fluoroquinolones have considerably improved activity against streptococci and obligate anaerobes. Examples of third-­generation fluoroquinolones approved for human use are sparfloxacin, grepafloxacin, and gatifloxacin. Pradofloxacin is the only third-­generation fluoroquinolone approved for use in animals. Fourth-­generation fluoroquinolones have all the activities of third-­ generation drugs and extra anaerobic activity. Examples of fourth-­generation fluoroquinolones approved for human use are trovafloxacin and moxifloxacin. Newer compounds are being explored that optimize the various substitutions and allow for the fluorine atom at position 6 to be replaced, which may reduce adverse effects, decrease metabolism, and decrease

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­Chemistr 

Fluoroquinolones

interactions with other drugs. However, the emergence of resistance to the entire class of fluoroquinolones remains problematic.

­Mechanism of Action The bacterial chromosome is a continuous, circular, double-­stranded DNA molecule approximately 1000 times longer than the bacterium in which it is contained. In order for such a long molecule to fit into the cell, it is densely packed in a negative supercoil, twisted in the opposite direction to the right-­ handed double helix of DNA. This supercoiled configuration is so highly strained that to improve function, the chromosome is divided into approximately 50 topologically independent domains. Fluoroquinolones target two essential bacterial topoisomerase enzymes: DNA gyrase and DNA topoisomerase IV. Topoisomerase I is characterized by reactions involving single-­stranded DNA, whereas topoisomerase II is involved in reactions with double-­stranded DNA. Topoisomerase II, also known as DNA gyrase, consists of two subunits, GyrA and GyrB. The gyrA gene encodes two alpha-­subunits while the gyrB gene encodes two beta-­subunits; the active DNA gyrase is an A2B2 complex. DNA gyrase binds to DNA; a segment of approximately 130 nucleotide wraps around the DNA gyrase. This wrapped DNA is cleaved in both strands, forming a DNA–protein covalent bond between the GyrA subunit and the 5’-­phosphates of the DNA molecule. Another segment of DNA is passed through this double-­ stranded break, which may then be resealed. The alpha-­subunit of the DNA gyrase is important in the breakage and reunion that allow for this ­relaxation of the DNA molecule. In multiple ­species of bacteria, it has been shown that the  4-­quinolone molecule interrupts the DNA ­breakage–reunion step by binding to the DNA gyrase–DNA complex at the interface between protein and DNA near the active site tyrosine and thus leads to defects in the negative supercoiling (Hooper and Jacoby, 2016).

Fluoroquinolones have a second intracellular target, DNA topoisomerase IV (Topo IV). This is a bacterial type II DNA topoisomerase and is also a multimeric protein composed of two ParC subunits and two ParE subunits, which exhibit sequence homology to GyrA and GyrB, respectively. This enzyme mediates relaxation of duplex DNA and the unlinking of daughter chromosomes following replication. However, unlike the DNA gyrase, Topo IV cannot supercoil DNA. Instead it is involved in the ATP-­dependent relaxation of DNA. It is a more potent decatenase than DNA gyrase. Fluoroquinolones can differ in their potency for the two enzymes, with a general pattern among the drugs in clinical use that there is greater activity against DNA gyrase in Gram-­ negative bacteria and greater activity against topoisomerase IV in Gram-­positive bacteria; but exceptions occur, and some quinolones have similar potency against both enzymes. The effect of fluoroquinolones on bacterial proliferation suggests three mechanisms of cell killing (Martinez et al., 2006). ●●

●●

●●

Mechanism A is common to all fluoroquinolones. It requires RNA and protein synthesis and only acts on dividing bacteria. Mechanism A appears to involve the blocking of replication by the gyrase–quinolone complex on DNA. Mechanism B does not require RNA and protein synthesis and can act on bacteria that are not multiplying. Mechanism B correlates with dislocation of the gyrase subunits that constrain the ternary complex. Mechanism C requires RNA and protein synthesis but does not require cell division. Mechanism C may correlate with trapping of Topo IV complexes on DNA.

­Antimicrobial Activity The important features of the antimicrobial activity of the fluoroquinolones is their rapid bactericidal and concentration-­dependent killing. Targeting fluoroquinolone dosage to the

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328

MIC of the pathogen, as discussed below under Pharmacodynamic Properties, not only increases clinical efficacy but reduces the emergence of resistance. The fluoroquinolones have excellent activity in  vitro against a wide range of aerobic Gram-­ negative bacteria, including the Enterobacterales, Actinobacillus pleuropneumoniae, Histophilus somni, Mannheimia haemolytica, and Pasteurella spp. including P. multocida. They are also active against Bordetella bronchiseptica, Brucella spp., Chlamydia/Chlamydophila spp., Mycoplasma spp., and Ureaplasma spp. Fluoroquinolones are active against rapidly growing mycobacteria isolated from dogs and cats. In general, pradofloxacin tends to be more active (i.e., lower MICs) against Gram-­negative bacteria than other veterinary fluoroquinolones (Silley et  al.,  2012). Activity against Pseudomonas aeruginosa is dependent on the fluoroquinolone, with ciprofloxacin being the most potent agent against this bacterium. For the most part, the first-­ and second-­generation fluoroquinolones are less active against Gram-­positive bacteria, especially enterococci, and have poor activity against anaerobic bacteria. Newer (third-­ and fourth-­generation) fluoroquinolones target this deficiency. For example, trovafloxacin, moxifloxacin, and gatifloxacin are newer human fluoroquinolones with good in vitro activity against obligate anaerobes. Pradofloxacin is active against anaerobic bacteria from dogs in cats including Clostridium spp., Bacteroides spp., Fusobacterium spp., and Prevotella spp. In general, fluoroquinolones have poor efficacy against streptococci and monotherapy with enrofloxacin has been associated with streptococcal shock syndrome and necrotizing fasciitis in dogs (Ingrey et  al.,  2003). While most staphylococci initially show susceptibility to fluoroquinolones, methicillin-­resistant Staphylococcus spp. frequently show resistance to all fluoroquinolones (Kizerwetter-­Świda et al., 2016). Fluoroquinolones exhibit a biphasic dose response curve (the Eagle effect) in that there is an optimum bactericidal concentration (OBC)

above the pathogen’s MIC and beyond which the bactericidal activity decreases (Lewin et  al.,  1991). As the ratio of fluoroquinolone concentration to MIC increases from ≤1:1 to the OBC (usually shown to be approximately 10:1–12:1 but is drug-­bacterium dependent), bacterial killing increases and is usually very rapid. As illustrated in Figure 17.2, when a strain of M. haemolytica is exposed to a fluoroquinolone at concentrations that are 25% of its MIC, the drug exhibits a slight stationary effect but then the bacterium resumes growth at a rate similar to that of the untreated control. As the concentration of the drug is increased above the MIC, there is a decrease in the number of viable organisms. For drug concentrations that are equivalent to the MIC, there is a slight decrease in the number of viable organisms but after 24  hours of exposure, the number of viable organisms has increased to more than what was in the starting suspension. This is without  an increase in MICs. This suggests that this  fluoroquinolone, at concentrations that are equal to the MIC, has a static effect on M. haemolytica. When the concentration of this fluoroquinolone is increased to four times the MIC, there is a nearly 4 log10 reduction in the number of viable organisms within four hours of exposure. However, this killing effect stabilizes and then the organisms begin to proliferate, again without an increase in MIC. This is in contrast to the growth rate when the concentration of the fluoroquinolone is eight times the MIC. Under this circumstance there is a very rapid bactericidal effect, 7 log10 reduction in viable organisms, and after a 24-­hour exposure there was no detectable regrowth of the bacterium. This suggests that at this concentration to MIC ratio, there was a 100% bactericidal effect. The concentration-­dependent killing effect may plateau when the ratio of fluoroquinolone concentration to MIC reaches 15:1–20:1 and at ratios greater than 20:1 the fluoroquinolones may become bacteriostatic. The decrease in antibacterial activity at high drug concentrations is

329

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­Antimicrobial Activit 

Fluoroquinolones 1000000000 100000000 10000000

CFU/mL

1000000 100000 10000 1000 100 10 1

0

1

2

CONTROL

3 4 Time (hours) 1/4×

5

1×

4×

6

24

8×

Figure 17.2  Concentration-­dependent killing effect of a fluoroquinolone tested against Mannheimia (Pasteurella) haemolytica.

thought to be caused by the inhibition of RNA and protein. This implies that protein synthesis may be required for quinolone-­mediated cell death. Supporting this, protein synthesis inhibitors (such as chloramphenicol) and RNA synthesis inhibitors (such as rifampin) reduce fluoroquinolone effectiveness in bacterial killing in vitro.

Antimicrobial Resistance Resistance to the fluoroquinolone occurs by target modification, decreased permeability, efflux, and/or target protection. Each of these fluoroquinolone resistance mechanisms can occur simultaneously within the same cell, thereby leading to very high resistance levels. To date, no mechanisms based on enzymatic inactivation/modification of fluoroquinolones have been discovered. Because fluoroquinolones are synthetic antimicrobials with no known natural analogues, it is less likely that this type of mechanism will emerge. Selection of resistant mutants with decreased permeability or efflux mechanisms generally means a 2–8-­fold

increase in MIC, whereas alteration of the DNA gyrase binding site or target protection may result in high-­level resistance. Resistance to one fluoroquinolone frequently results in resistance to all. This is especially true for the older drugs and for high-­level resistance. Fluoroquinolone resistance due to target mutations typically results in decreased susceptibility or resistance to other fluoroquinolones. Resistance due to alterations in permeability or activation of the efflux pump can confer resistance to other antimicrobial agents such as the cephalosporins, carbapenems, and tetracyclines even if these drugs have not been used in the patient (Hooper and Jacoby, 2016). Because fluoroquinolones mediate DNA damage by binding to susceptible enzymes, fluoroquinolone resistance mutations are recessive. For topoisomerase-­mediated fluoroquinolone resistance to be transferred horizontally, an acquired mutated gene has to supplant the wild-­type gene. The development of fluoroquinolone resistance via mutations in topoisomerases has been ­studied  extensively. Resistance is mediated primarily by target mutations in DNA gyrase

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330

(topoisomerase II), with secondary mutations in topoisomerase IV contributing to higher levels of resistance. Amino acid substitutions that result in bacterial resistance have been localized to a specific topoisomerase subdomain termed the quinolone resistance-­determining region (QRDR) within gyrA and parC. In E. coli, most mutations associated with quinolone resistance occur in the QRDR at serine 83 (Ser83) and aspartate 87 of gyrA, and at serine 79 and aspartate 83 of parC and at analogous sites in other species (Gebru et al., 2012). DNA sequence analysis of S. aureus and Streptococcus genes shows that the situation can be reversed in Gram-­positive bacteria, where topoisomerase IV (encoded by grlA and grlB) is the primary fluoroquinolone target. In both cases, mutations decrease the fluoroquinolone affinity for the enzyme/DNA complex and allow DNA replication to continue in the presence of fluoroquinolone concentrations that are inhibitory to wild-­type cell growth. In Gram-­negative organisms, fluoroquinolone resistance typically develops in a stepwise manner. A single QRDR mutation, usually at Ser83, confers resistance to nalidixic acid and decreases susceptibility to fluoroquinolones (ciprofloxacin MICs may go from a wild-­type baseline of 0.015–0.03 μg/ml to 0.125–1 μg/ml). Secondary mutations in the gyrA QRDR lead to overt fluoroquinolone resistance (ciprofloxacin MICs ≥4 μg/ml). However, this does not hold true for all Gram-­negative bacteria. In Campylobacter spp., which lack topoisomerase IV, a single mutation in gyrA is sufficient to impart high-­level ciprofloxacin MICs (32 μg/ml) (Griggs et  al.,  2005). This feature helps explain  the higher prevalence of resistance in Campylobacter, compared to E. coli, from food animals exposed to fluoroquinolones. As indicated above, fluoroquinolone resistance may also be mediated by decreased permeability of the bacterial cell wall through altered outer membrane porins (OmpF) and by the activity of energy-­dependent efflux pumps. Most fluoroquinolones cross the Gram-­negative outer membrane through protein channels called porins, although some may diffuse directly across

the lipid bilayer. Resistance due to decreased fluoroquinolone influx is generally reflected in low-­level changes in susceptibility and may explain differences in potency among different fluoroquinolone derivatives. Porin deficiency has been associated with quinolone resistance in E. coli and Pseudomonas. For example, mutations of the E. coli porin OmpF produce about a two-­fold increase in quinolone MICs. However, it is difficult to experimentally assess the role of porins without also accounting for effects due to efflux. Permeability changes mediated by altered porins are often part of a coordinated cellular response to the presence of numerous toxic agents, which includes simultaneous upregulation of efflux. In E. coli, de-­repression in regulatory loci such as marA or soxS leads to decreased fluoroquinolone susceptibility via simultaneous upregulation of the AcrAB-­TolC efflux pump and downregulation of the OmpF porin. This mechanism confers decreased susceptibility to a large number of other antimicrobial agents in addition to fluoroquinolones. Analogous regulatory loci exist among other species of bacteria. In antimicrobial efflux systems, membrane-­ localized proteins actively pump drug from the cell before it can diffuse to its primary target within the active site of DNA gyrase. Because they are driven by the proton motive force, energy uncouplers can be used to study their role in resistance. The E. coli genome carries as many as 30 potential efflux pumps, many of which mediate antimicrobial efflux. Some are effective for specific agents, whereas others protect against a variety of structurally diverse compounds. In addition, a single bacterium may contain multiple efflux pumps (e.g., AcrAB and CmlA) that are capable of extruding the same antimicrobial agent. Constitutive and inducible efflux is a known mechanism of fluoroquinolone resistance in both Gram-­negative and Gram-­positive bacteria, and may be more important than secondary mutations in topoisomerase IV genes. For example, it has been shown that deletion of the gene encoding the inducible AcrAB efflux pump reduces ciprofloxacin MICs

331

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­Antimicrobial Activit 

Fluoroquinolones

to near wild-­type levels in cells carrying topoisomerase mutations. In Campylobacter, where efflux mediated by CmeAB is constitutive, fluoroquinolone MICs in wild-­type cells are 3–4-­fold higher than those typical of E. coli. Insertional inactivation of CmeAB in C. jejuni reduces ciprofloxacin MICs to levels near that of wild-­type E. coli (0.003 μg/ml). Bacterial fluoroquinolone resistance was once thought to disseminate exclusively via clonal expansion under selective pressure. A plasmid-­ mediated quinolone resistance gene (qnrA) was first described in clinical isolates of Klebsiella pneumoniae in 1998 and was soon found worldwide in Gram-­negative bacteria, indicating that plasmid acquisition of qnrA and other fluoroquinolone resistance determinants has occurred independently multiple times. The qnrA gene is located near sequences (qacEA¨ 1, sulI) typically associated with class I integrons: the qnrA gene encodes a 218 amino acid protein belonging to the pentapeptide repeat family. In a concentration-­ dependent manner, qnrA functions by protecting E. coli DNA gyrase, but not topoisomerase IV, from inhibition by ciprofloxacin. The qnrA gene confers a small decrease in fluoroquinolone susceptibility such that qnrA + strains are still considered clinically susceptible. A second mechanism of fluoroquinolone resistance is modification of certain quinolones by a variant of the common aminoglycoside-­ modifying acetyltransferase AAC(60)-­Ib. A third mechanism for fluoroquinolone resistance is from plasmid-­encoded quinolone efflux pumps: OqxAB and QepA (Hooper and Jacoby, 2016). The qnr genes are usually found in multiresistance plasmids linked to other resistance determinants, including beta-­ lactamase genes (including genes for ESBLs), AmpC enzymes, and carbapenemases.

P ­ harmacokinetic Properties The fluoroquinolones are rapidly and well absorbed from the gastrointestinal tract of monogastric animals and preruminant calves.

Enrofloxacin is more lipid soluble than ciprofloxacin and has a higher oral bioavailability than ciprofloxacin in horses and small animals. All the oral veterinary products typically have high bioavailability in dogs and cats, but enrofloxacin bioavailability is poor in neonatal kittens. The oral bioavailability of enrofloxacin is approximately 60% in adult horses and 42% in foals. While it is extremely low in adult cattle, it is surprisingly good in sheep (80%). The pharmacokinetic parameters of fluoroquinolones administered to dogs, cats, cattle, horses, and pigs are given in Table  17.2. Ingestion with food may delay the time to peak serum concentrations without affecting total serum concentrations, unless the food is rich in magnesium or aluminum ions. Increases in oral dose usually produce linear increases in serum concentrations. Following absorption, fluoroquinolones exhibit rapid and extensive tissue distribution because of their lipophilic nature and low (1  l/kg). In general, fluoroquinolone concentrations in interstitial fluid, skin, and bones are 35–100% of those obtained in the serum, whereas bronchial secretions and prostatic concentrations may be 2–3 times the corresponding serum concentrations. Penetration into cerebrospinal fluid is ­approximately 25% of serum concentration. Therapeutic concentrations for Gram-­negative bacteria may be achieved in the CSF and ocular fluids. High concentrations are found in the bile and organs of excretion (liver, intestine, and urinary tract). The fluoroquinolones are concentrated within phagocytic cells. Uptake occurs by simple diffusion, and intracellular concentrations may be several times greater than plasma concentrations. Intracellular drug is microbiologically active; in vitro studies indicate that ciprofloxacin reduces survival of intracellular pathogens such as Brucella spp., Mycoplasma spp., and Mycobacterium spp. The fluoroquinolones are predominantly excreted as unchanged drug in the urine by glomerular filtration and active tubular

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332

Table 17.2  Comparative pharmacokinetic parameters of selected fluoroquinolones administered orally to cats, dogs, horses, cattle, and pigs.

Fluoroquinolone

Animal species

Route

Dose (mg/kg)

Ciprofloxacin

Cats

IV

10

PO

10

Dogs

Enrofloxacin

CMAX (μg/ml)

3.9 1.26

AUC0–­>24 (μg · hr/ml)

4.5

17

3.7

11

Bioavailability (%)

33

10

3.1

PO

10

1.55

Ponies

IV

5

3.45

2.5

Cats

IV

5

2.37

6.7

18.6

Kittens

IV

5

1.8

4.2

16.7

PO

5

4.8

5.7

33.7

8.74

83

Horses Foals Cattle Pigs

IV

5

PO

5

IV

5

PO

5

IV

5

PO

10

IV

5

SC

8

4.9

3.7 1.41 2.3 2.47 2.12 4.0 0.81

IV

5

PO

10

1.4

6.11

Cattle

SC

8

2.4

Dogs

PO

5

1.1

PO

15

6.04

Marbofloxacin

Cats

IV

2

PO

2

IV

2

6

2.4 4.1

5.4

Difloxacin

Dogs

2.2

0.5

Danofloxacin

4.4 6.1

35.6

17.1

48.54

18.4

58.47

2.6

4.4

7.3

7.51

10.5

63 42

11.2 83

4.7

3.8

14.76

6.9

9.34

3.4

21.28

1.01

7.9

21.26

7.8

24.73

100

1.37

12.4 94

2.34

69.1

PO

2

1.47

9.1

13.07

Cattle

IM

2

1.98

6.3

7.65

Cats

IV

2.5

PO

2.5

IV

2.5

PO

2.5

1.37

Cats

PO

3

1.2

4

8

6

70

Dogs

PO

3

1.6

2

7

13

100

Dogs Pradofloxacin

T1/2ß (h)

IV

Dogs

Orbifloxacin

Vd (l/kg)

1.3 2.06 1.2

4.5

10.6

5.5

10.82

5.4

14.3

7.1

12.72

100 100

IM, intramuscular; IV, intravenous; PO, by mouth (per os); SC, subcutaneous.

secretion. The exception is difloxacin, where 80% is excreted in the feces. Metabolites and the parent compound may be excreted in an active form in the bile and urine. The major

metabolite of enrofloxacin is ciprofloxacin. The amount of ciprofloxacin produced varies with different species, with some producing ciprofloxacin concentrations that exceed the

333

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­Pharmacokinetic Propertie 

Fluoroquinolones

MIC of some pathogens. The plasma elimination half-­lives of the fluoroquinolones depend on the drug and the animal species, but are relatively long. These long plasma elimination half-­lives make the fluoroquinolones ideal for once-­daily dosing, which improves compliance with the treatment regimen.

P ­ harmacodynamic Properties

Probability of remaining susceptible

With ideal pharmacokinetic parameters but a potential to select for resistant bacteria, ­optimal therapeutic dosage regimens for fluoroquinolones require pharmacokinetic/­ pharmacodynamic (PK/PD) integration (Chapter 4). Pharmacodynamic indices describe the interaction of drug concentration, which is dependent on dose and pharmacokinetic properties, with the bacterial killing of the drug. The fluoroquinolones show concentration-­ dependent killing with persistent postantibiotic effects (PAE), and the PK/PD parameters best associated with efficacy are fAUC0–­24/MIC (the ratio of the area under the plasma ­concentration–time curve of free drug divided by MIC) or fCmax/MIC (the ratio of maximum free plasma concentration over MIC) ratios. Studies with ciprofloxacin in critically ill people have shown that an AUC0–24/MIC of ≥125 is linked with favorable clinical and microbiological outcomes, whereas an AUC0–24/MIC of 100 90 80 70 60 50 40 30 20 10 0

+

+ + + + +

+ +

2 hours) surgery with much tissue manipulation.

Escherichia coli, the microbes most likely to cause postoperative wound infections in dogs and cats. Cefazolin usually has excellent activity against susceptible staphylococci and E. coli and has low toxicity. It is also active against many obligate anaerobes, which might be preferred if anaerobes are of particular concern, as in colonic or rectal surgery. Where available, injectable ampicillin-­sulbactam preparations are economical alternatives to cephalosporins and may have improved activity against anaerobes. If used, these antimicrobials should be administered every two hours during surgery and treatment should not be continued beyond the perioperative period. Unfortunately, the emergence of methicillin-­ resistant staphylococci (and especially Staphylococcus pseudintermedius) in some locations (North America and Europe) has meant that prophylactic treatment with beta-­ lactam drugs such as cephalosporins has the potential to select for these bacteria. These drugs have no efficacy against methicillin-­ resistant bacteria or most multidrug-­resistant Gram-­negative bacteria. Thus, antimicrobial drugs should never be used as a substitute for

careful infection control measures, which should include proper patient preparation, proper use of scrubbing and sterile drapes, attention to hemostasis, and minimization of surgical time.

Antimicrobial Prophylaxis in Nonsurgical Patients Prophylactic use of antimicrobial drugs in nonsurgical patients is controversial and veterinary data are limited, but it is not generally warranted. Chemoprophylaxis might be effective if the period of risk is brief (a few hours or days), as with some chemotherapy-­induced myelosuppression, or the target is a single drug-­susceptible species. For example, trimethoprim-­sulfonamides have been used successfully for prophylaxis in dogs treated with chemotherapeutics such as doxorubicin (Chretin et  al.,  2007). However, attempts at long-­term chemoprophylaxis are liable to simply select for bacteria that are resistant, especially if host defenses remain compromised. Prophylactic antimicrobial drug treatment of animals with indwelling urinary catheters is strongly discouraged, as it increases the risk of infection with resistant bacteria. If indwelling urinary catheterization is required, a closed sterile collection system should be used, and the catheter should be removed as soon as it is no longer required, because the risk of ascending infection increases with every day the catheter is left in place. Instead of using prophylactic antimicrobials, a better approach is to carefully monitor individuals at risk for signs of infection and to treat promptly and appropriately if infection occurs, as well as resolving underlying causes of compromised host defenses whenever possible. Routine culture of catheters at the time of removal is not recommended, because they become contaminated with ascending bacteria (which does not equate to infection). If infection is suspected (based on the presence of pyuria or hematuria) and a catheter is in place, the catheter should be removed and the urine collected for culture

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608

by cystocentesis or through a newly placed catheter, which should then be withdrawn if possible. Urine culture should never use specimens from the collection bag, and there is no indication to culture the catheter tip (Weese et al., 2019).

Routine treatment of dogs and cats with antimicrobials after removal of a catheter is controversial but could be considered if the consequences of infection are likely to be severe (such as reobstruction in cats with urethral obstructions).

­References and Bibliography Bakken JS. 2004. The fluoroquinolones: how long will their utility last? Scand J Infect Dis 36:85. Chretin JD, et al. 2007. Prophylactic trimethoprim-­sulfadiazine during chemotherapy in dogs with lymphoma and osteosarcoma: a double-­blind, placebo-­ controlled study. J Vet Intern Med 21:141. Clare S, et al. 2014. Short-­and long-­term cure rates of short-­duration trimethoprim-­ sulfamethoxazole treatment in female dogs with uncomplicated bacterial cystitis. J Vet Int Med 28:818. Hillier A, et al. 2014. Guidelines for the diagnosis and antimicrobial therapy of canine superficial bacterial folliculitis (Antimicrobial Guidelines Working Group of the International Society for Companion Animal Infectious Diseases) Vet Dermatol 25(3):163. Lappin MR, et al. 2019. Antimicrobial use guidelines for treatment of respiratory tract disease in dogs and cats: antimicrobial guidelines working group of the International Society for Companion Animal Infectious Diseases (ISCAID). J Vet Intern Med 31:279.

Piscitelli SC, et al. 1992. Clarithromycin and azithromycin: new macrolide antibiotics. Clin Pharm 11:137. Sivapalasingham S, Steigbigel N. 200911. Macrolides, clindamycin and ketolides. In: Mandell G, Bennett J, Dolin R (eds) Mandell, Douglas, and Bennett’s Principles and Practice of Infectious Diseases, 7th edn. Philadelphia: Churchill Livingstone Elsevier, Philadelphia, pp. 427–448. Weese JS. 2006. Investigation of antimicrobial use and the impact of antimicrobial use guidelines in a small animal veterinary teaching hospital: 1995–2004. J Am Vet Med Assoc 228:553. Weese JS, et al. 2019. International Society for Companion Animal Infectious Diseases (ISCAID) guidelines for the diagnosis and management of bacterial urinary tract infections in dogs and cats. Vet J 247:8. Westropp JL, et al. 2012. Evaluation of the efficacy and safety of high-­dose short duration enrofloxacin treatment regimen for uncomplicated urinary tract infections in dogs. J Vet Int Med 26:506.

609

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  ­References and Bibliograph

29 Antimicrobial Therapy in Beef Cattle Michael D. Apley, Brian V. Lubbers, and Nora D. Schrag

Antimicrobial options for cattle have changed dramatically in the past 50 years. Novel properties of new drug groups, changes in route of administration, and advances in drug formulations have significantly altered characteristics of treatment regimens. Many antimicrobials are single-­injection products, which minimizes the need for animal handling and greatly improves regimen compliance. These drugs are used in an environment of increasing regulatory and political pressure, an expanding array of branded food product lines, and increased scrutiny by consumer and special interest groups. Antimicrobial resistance in both veterinary and human bacterial pathogens threatens the effectiveness and access to these drugs in cattle. This chapter addresses important areas of consideration in constructing antimicrobial regimens in cattle within this context, including reasonable antimicrobials for selected diseases and an extended discussion of some common therapeutic challenges.

obligated to provide written treatment g­ uidelines. The treatment guidelines should be constructed to contain the following information, where appropriate. ●● ●●

●●

●● ●●

●●

●●

●●

●●

G ­ eneral Considerations of Antimicrobial Use in Beef Cattle When giving treatment instructions to clients, especially in large-­scale production facilities where lay personnel will be identifying and treating ill animals, the veterinarian is

Case definition for initial treatment. Initial regimen: drug(s), dose, route, ­duration, frequency, slaughter withdrawal. Specific administration instructions: injection site, volume per site, needle size, injection technique. Safety precautions or warnings. Environmental management during treatment: housing, water, feed. Case definitions for treatment success and failure and the time at which animals become eligible for retreatment (posttreatment interval, PTI). Secondary regimen for treatment of animals failing the initial treatment regimen. Any additional regimens for animals not responding after the first and second regimens. Disposition of animals not responding to therapy.

It is essential that the treatment protocols not be altered except after agreement by all parties involved. Consistency of protocol application is an absolute necessity in order to evaluate therapeutic and preventive programs in production systems.

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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611

Antimicrobial Therapy in Beef Cattle

In constructing these regimens, the ­veterinarian must make several key decisions.

Monotherapy or Combination Antimicrobial Therapy in Each Regimen The search for antimicrobial synergy is ­prevalent in all branches of medicine. Clinical studies and metaanalyses in human medicine evaluating combination versus single antimicrobial therapy are equivocal, with some studies showing no statistical reduction in patient mortality with combination therapy (Bowers et al., 2013; Heffernan et al., 2020) and others demonstrating a reduced mortality risk with combination antimicrobial therapy (Schmid et al., 2019). There is little evidence to either support or refute the routine use of combination therapy in cattle. An in  vitro study of M. haemolytica and P. multocida failed to demonstrate antimicrobial synergism with antimicrobials commonly used to treat BRD (Sweeney et al., 2008). One clinical study evaluating the concurrent use of ceftiofur and tulathromycin versus a single dose of florfenicol demonstrated a reduction in BRD mortality and relapse rates for the combination regimen; however, the study authors concluded that “the relative in  vivo effects of concomitant therapy compared to monotherapy with these antimicrobials cannot be determined” as neither drug in the combination treatment was studied individually (Booker et al., 2017). Anecdotal reports often claim that the preferred combination reduces relapses or improves initial treatment response. Arguments that combination therapy will suppress resistance development must be evaluated considering that the bacterial population will also be exposed to a wider variety of antimicrobials.

Same or Different Second-­line Therapy If the animal did not respond to the initial antimicrobial regimen, was it because of antimicrobial resistance in the pathogen or other factors that lead to therapeutic failure? By convention, many treatment protocols specify that subsequent

treatments consist of an ­antimicrobial from a different class, with the rationale being that if antimicrobial resistance contributed to drug failure, a drug with another mechanism of action might circumvent the particular resistance mechanism. There is not strong evidence to support or refute the practice of changing drug classes for follow­up antimicrobial therapy. While switching to a different drug class seems logical, the reality is that there are many reasons why an antimicrobial may fail to elicit a clinical response (improper diagnosis, advanced disease states, disease-­induced alterations in pharmacokinetics, etc.) that will not be resolved by changing drug classes. In the lead author’s personal experience with randomized, controlled respiratory disease trials, repeating the first drug regimen in first treatment failures resulted in a similar second treatment response as trials where a therapeutic from a different antimicrobial class was selected. This is dependent on the first treatment providing satisfactory treatment response and isolates with similar susceptibility profiles being present in all cases. Furthermore, recent reports of integrative-­ conjugative elements (ICE) that confer resistance to multiple drug classes would appear to make class switching a relatively futile exercise. As a counterpoint, while there are many reasons why antimicrobials may not cure an infection, the most significant improvement in clinical outcomes is probably realized when an effective antimicrobial is used. If antimicrobial resistance truly contributed to the clinical failure (something that is very rarely known at the time subsequent treatment is administered), it would be of benefit to change to a drug with a different mechanism as cross-­resistance is not uncommon for antimicrobials used to treat bovine respiratory disease.

Timing of Second-­line Therapy Administration In the diagnosis of undifferentiated fever/respiratory disease, the animal may have from three to 10 days to respond to the initial regimen

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612

before being classified as a treatment failure. Extended durations of antimicrobial coverage are commonplace among current antimicrobials, such as approximately seven days for ceftiofur crystalline free acid and 300  mg/ml long-­acting oxytetracycline, up to approximately two weeks for tulathromycin and gamithromycin, and a claimed 28 days for tildipirosin. These longer durations of therapy bring forth the challenge of deciding when concentrations are low enough that nonresponders should receive additional therapy. Trials evaluating the effect of a longer PTI for bovine respiratory disease demonstrate that longer PTIs did not negatively impact mortality or treatment success rates. These limited data suggest that a 5–­7-­day PTI is reasonable for most antimicrobial products. In the authors’ opinion, the success of longer PTIs is likely less about the prolonged concentration of antimicrobial and more about the time needed for the drug and host immune system to resolve the disease process. The current evidence for extended PTIs is notable; however, the successful implementation of PTIs in treatment protocols should consider the impact of “withholding treatment” on animal welfare and employee moral distress.

Extra-­label Drug Use (ELDU) In the United States, regulations for ELDU were promulgated as directed by the Animal Medicinal Drug Use Clarification Act (AMDUCA,  1996). The regulations should be consulted for actual guidance, but the overall order of expected use may be summarized as follows. 1) Use of an antimicrobial according to label directions. 2) Use of a drug labeled in that food animal species but in an extra-­label manner. 3) Use of a drug labeled for use in another food animal species. 4) Use of a veterinary nonfood animal-­labeled drug or human-­labeled drug. 5) Use of a compounded product meeting the requirements of the AMDUCA regulations.

One component of the AMDUCA ­regulations is that the veterinarian must determine an extended slaughter withdrawal time for animals subjected to ELDU. In the United States and Canada, this information may be obtained by consulting with the US Food Animal Residue Avoidance Databank (FARAD) or Canadian gFARAD (see Chapter  26). If adequate information for construction of an extra-­ label slaughter withdrawal time is not available, then the drug may not be used in food animals. Extra-­label drug use regulations vary in other countries/regions. The European Union has very strict regulations regarding the ELDU of antimicrobials (Regulation 2019/6). The AMDUCA regulations also specify certain pharmaceutical products for which ELDU is prohibited or restricted. The following antimicrobials are prohibited from being used in any extra-­label manner in food animals: chloramphenicol, fluoroquinolones, nitroimidazoles, nitrofurans, and glycopeptides. These regulations also prohibit the extra-­label use of sulfonamides in lactating dairy cows. In 2012, these regulations were amended to also prohibit the extra-­label use of cephalosporins for disease prevention, at unapproved dosages, frequencies, durations, or routes of administration in cattle, swine, chickens or turkeys (21 CFR Part 530,  2012). Furthermore, use of cephalosporins that are not approved for use in one of these species, and for extra-­label disease prevention is also prohibited. The extra-­label use of cephapirin in food-­producing animals is exempt from these restrictions. The extra-­label use of cephalosporins in these species is permitted for the treatment or control of an extra-­ label disease indication provided this use adheres to a labeled dosage regimen approved for that particular species and production class. Extra-­label use of a cephalosporin in a food-­producing minor species, such as ducks or rabbits, is also allowed. Veterinarians should be familiar with regulations in their respective countries in order to protect the interests of their clients and the consuming public.

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­General Considerations of Antimicrobial Use in Beef Cattl  613

Antimicrobial Therapy in Beef Cattle

Antimicrobial Stewardship Guidelines Concerns about the proper use of antimicrobials in food animals, especially related to the development of antimicrobial resistance, have prompted development of antimicrobial stewardship guidelines. For example, the American Veterinary Medical Association (AVMA) has made antimicrobial stewardship definition and core principles available to veterinary ­professionals (AVMA, 2021). These principles include a commitment to stewardship, continuously improving disease prevention practices, judicious selection and use of antimicrobials, evaluation of antimicrobial use practices, and encouraging the development of expertise in antimicrobial stewardship. While these guidelines do not give specific recommendations for antimicrobial applications, they do highlight some of the key considerations that should guide veterinarians as they design antimicrobial regimens for cattle. Some antimicrobials have specific limitations in cattle that either preclude their use or that require special consideration. The extra-­ label, systemic use of aminoglycosides in cattle has been the subject of resolutions or policy statements by the AVMA, American Association of Bovine Practitioners (AABP), Academy of Veterinary Consultants (AVC), and the National Cattlemen’s Beef Association (NCBA). In general, these statements discourage the extra-­label use of aminoglycosides in cattle due to the prolonged slaughter withdrawal potential due to renal accumulation. Veterinarians should pay special attention to these statements, especially when a producer organization joins with veterinary organizations in discouraging the extra-­label use of a drug in cattle. Some antimicrobials have significant potential for tissue damage when injected intramuscularly. These include the macrolides (e.g., tilmicosin, tylosin, erythromycin) and some oxytetracycline formulations. Although a visible lesion is not necessary for an adverse effect on tenderness, persistent visible lesions add to

trim loss when primal cuts are fabricated into retail cuts. Intravenous use of tylosin and erythromycin is a possibility, but the nonwater-­ soluble properties of these drugs in commercially available forms combined with the propylene glycol carriers make adverse ­reactions a possibility. In addition, repeated intravenous injections have become less attractive in light of effective alternatives with less frequent, subcutaneous administration routes.

Is Susceptibility Testing Useful in Selecting Antimicrobials for Use in Beef Cattle? The answer to this question depends on both the testing methods used in the laboratory and which interpretive criteria were used. Standardized antimicrobial susceptibility test methods are approved by the Clinical and Laboratory Standards Institute (CLSI)  –­ Veterinary Antimicrobial Susceptibility Testing (VAST) for bacteria isolated from animals to promote harmonization and reduce inter-­ and intralaboratory variation. The approved methods are prescriptive in terms of bacterial inoculum, media, incubation conditions, and routine use of quality control testing. Alterations to the approved methods may lead to a test result that is not correlated to clinical outcome (see Chapter 2 for additional discussion of antimicrobial susceptibility testing). Interpretive criteria (“susceptible”, “intermediate,” and “resistant” breakpoints) are approved by the CLSI-­VAST through the evaluation of microbiological, pharmacological, and clinical data and are specific to the antimicrobial, bacterial pathogen, host species, drug regimen, and disease process. The CLSI-­VAST has approved veterinary-­specific breakpoints for bovine respiratory disease (BRD), metritis, and mastitis for common bacterial pathogens  and commonly used antimicrobials (CLSI,  2024a). Approved BRD-­specific breakpoints have been established for ampicillin, ceftiofur (sodium, hydrochloride, and crystalline free acid formulations), enrofloxacin,

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614

florfenicol, gamithromycin, penicillin G, ­spectinomycin sulfate, tetracycline, tildipirosin, and tulathromycin for Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni. Danofloxacin breakpoints are approved for M. haemolytica and Pasteurella multocida, while tilmicosin breakpoints are only approved for M. haemolytica. There are no approved test methods or breakpoints for any antimicrobials for Mycoplasma bovis. For other antimicrobials, laboratories may extrapolate the breakpoints developed from other animal species, including humans. Examples of this include breakpoints reported for potentiated sulfonamides, aminoglycosides, and erythromycin. It should be noted that there are no veterinary approved breakpoints for enteric disease in any species. For in-­depth information on the conduct and interpretation of susceptibility testing in cattle, and all veterinary species, the reader is referred to the most recent editions of the Clinical and Laboratory Standards Institute publications VET01, VET01S, and VET09 (CLSI, 2024a,b). Arguments that susceptibility testing results have no utility in antimicrobial selection are often based on the fact that animals with “susceptible” organisms have failed to resolve infections and animals with “resistant” pathogens have recovered. It is important to realize that antimicrobial susceptibility testing does not guarantee a specific clinical result in an individual animal. Rather, for veterinary approved breakpoints, it places the animal/ drug regimen/pathogen combination in a population where clinical resolution is more (for susceptible isolates) or less (for resistant isolates) likely compared to other categories. The veterinarian must determine when susceptibility testing may be of use in monitoring a population of animals and pathogens. Veterinarians should also consider the utility of cumulative antimicrobial susceptibility data (“antibiograms”) for guiding empiric therapy of future cases. By monitoring susceptibility test results over time, veterinarians may detect

changing patterns of antimicrobial resistance that would necessitate a change in treatment protocols.

­Antimicrobial Use in Cattle Specific therapeutic antimicrobial application suggestions in cattle are reported in Table 29.1. Where appropriate, justifications for drug recommendations are presented in a referenced narration. These suggestions should be considered as starting points for application of evidence-­based therapeutic decision processes. Additional discussions for Mycoplasma bovis, enteric Salmonella spp. and E. coli, and Cryptosporidium parvum are provided in the text.

Mycoplasma bovis There has been debate as to whether M. bovis is a primary respiratory pathogen in cattle. However, in the United States, M. bovis is now listed as a label respiratory pathogen for tulathromycin, gamithromycin, enrofloxacin, and florfenicol. Standardized susceptibility testing methods and interpretive criteria have not yet been established for M. bovis for any indication. Variations in methods may contribute to variations in minimal inhibitory concentrations (MIC) results reported in Table  29.2. It is apparent in this table that there is a wide range of MICs determined for each drug, suggesting that some isolates will be refractive to therapy, although the maximum MIC correlated with therapeutic efficacy has not been established. No clinical trials evaluating antimicrobial therapy of arthritis or tenosynovitis due to M. bovis are available. The antimicrobial selected by the veterinarian should have at least the potential for effectiveness; beta-­ lactam antimicrobials (penicillins and cephalosporins) are not reasonable choices as Mycoplasma spp. lack the cell wall that these drugs target. It is reasonable to begin

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­Antimicrobial Use in Cattl  615

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Table 29.1  Specific antimicrobial use suggestions. Individual product labels should be consulted for indications and complete instructions for use. Label applications are US labels except for those in bold print which are EU labels.

Drugs for which this disease is a label application (therapy and/or prevention)

Category

Disease/Pathogen(s)

Respiratory disease

Pneumonia: Mannheimia haemolytica, Pasteurella multocida, Histophilus somni

Amoxicillin trihydrate, ampicillin trihydrate, ceftiofur (sodium, hydrochloride, and crystalline free acid salts), chlortetracycline, danofloxacin, enrofloxacin, erythromycin, florfenicol, gamithromycin, oxytetracycline, procaine penicillin G, penicillin G benzathine/penicillin G procaine, spectinomycin sulfate, sulfabromomethazine, sulfadimethoxine, sulfaethoxypyridazine, sulfamethazine, tetracycline, tildipirosin, tilmicosin, tulathromycin, tylosin, cefquinome, trimethoprim/sulfadiazine. trimethoprim/sulfadoxine, procaine penicillin/dihydrostreptomycin, amoxicillin trihydrate, amoxicillin/ clavulanic acid

Respiratory disease

Pneumonia –­ Mycoplasma bovis

Enrofloxacin, gamithromycin, florfenicol, tulathromycin, tylosin

Respiratory disease

Diphtheria (necrotic laryngitis) Fusobacterium necrophorum

Oxytetracycline, sulfabromomethazine, sulfadimethoxine, Sulfamethazine, tylosin

0005858924.indd 616

Reasonable extra-­label antimicrobial choices

Unreasonable extra-­ label antimicrobial selections for this disease

Comments

Gentamicin due to potential for toxicity in dehydrated animals and prolonged renal residues in cattle.

Antimicrobials with bovine respiratory disease on the label may be indicated for one or all of these pathogens. The italicized antimicrobials are the authors’ primary choices for cattle in the US in advanced stages of the disease or which have experienced extensive stress. Antimicrobials in bold are available in other countries.

Oxytetracycline, spectinomycin, tildipirosin, fluoroquinolones*

Any beta-­lactam (penicillins, cephalosporins) due to lack of a cell wall.

*In the US, fluoroquinolones would only be legal when used for the purpose of respiratory disease due to the primary label pathogens.

Ampicillin, ceftiofur, florfenicol, penicillin G, other macrolides

Enrofloxacin/ danofloxacin. Fluoroquinolones would be prohibited for ELDU in the US and anaerobic bacteria are not

Extra-­label recommendations are made based on the expected spectrum of activity (anaerobes) of the antimicrobial and/or label inclusion of foot rot due to Fusobacterium necrophorum.

10/25/2024 10:22:36 AM

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within the spectrum The nature of the site of necrotic of activity for these laryngitis may make therapy with less lipid-­soluble antimicrobials more of a drugs. challenge. Infectious enteric disease

Scours, neonatal diarrhea due to E. coli

Amoxicillin trihydrate, ampicillin trihydrate, chlortetracycline, neomycin, oxytetracycline, streptomycin sulfate, sulfabromomethazine, sulfachlorpyridazine, sulfaethoxypyridazine, sulfamethazine, tetracycline (all these antimicrobials display consistently high MICs that suggest the drugs would be ineffective), amoxicillin/clavulanic acid bolus, apramycin, cefquinome (septicemia), danofloxacin, enrofloxacin (septicemia and colibacillosis), marbofloxacin bolus, trimethoprim/sulfadiazine, trimethoprim/sulfadoxine

Ceftiofur, potentiated sulfonamides (all only after susceptibility testing)

(These extra-­label indications demonstrated very high MICs to most isolates.) Macrolides penicillin, ampicillin, florfenicol.   Fluoroquinolones are likely active against the pathogen, but ELDU is prohibited in the US.

Recommended extra-­label antimicrobials are based on susceptibility data and serum pharmacokinetics and should therefore be interpreted as relating to septicemia associated with enteric disease. See text for additional discussion.

Infectious enteric disease

Scours, neonatal diarrhea due to Salmonella spp.

Chlortetracycline, oxytetracycline, streptomycin, tetracycline (these antimicrobials display consistently high MICs that suggest the drugs would be ineffective), apramycin, enrofloxacin, trimethoprim/ sulfadiazine, trimethoprim/ sulfadoxine, procaine penicillin/ dihydrostreptomycin

Ceftiofur, potentiated sulfonamides (all only after susceptibility testing)

Gentamicin will cause extended withdrawal times that will compromise the ability to slaughter an animal that recovers from the acute disease but does not return to satisfactory production.

Recommended extra-­label antimicrobials are based on susceptibility data and serum pharmacokinetics and should therefore be interpreted as relating to septicemia associated with enteric disease. See text for additional discussion.

Infectious enteric disease

Enterotoxemia, overeating disease –­ Clostridium perfringens type C, D

Amoxicillin, ampicillin, penicillin G, ceftiofur, florfenicol, tetracyclines

Fluoroquinolones, aminoglycosides (lack of spectrum and illegal ELDU / prohibitive WD times)

Antiserum therapy is more likely related to therapeutic success. Septicemia resulting from enterotoxemia may involve multiple gut-­related bacteria. Antimicrobial selection should reflect this possibility (see septicemia related to neonatal diarrhea above).

(Continued)

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Drugs for which this disease is a label application (therapy and/or prevention)

Reasonable extra-­label antimicrobial choices

Unreasonable extra-­ label antimicrobial selections for this disease

Category

Disease/Pathogen(s)

Infectious enteric disease

Hemorrhagic bowel disease –­Clostridium perfringens type A

Infectious enteric disease

Cryptosporidiosis –­ Cryptosporidium parvum

Infectious enteric disease

Giardia

Albendazole, fenbendazole, metronidazole (see comments)

The extra-­label use of nitroimidazoles (e.g., metronidazole) in food animals is prohibited in the US. Fenbendazole regimens of 5 mg/kg q12h for 3 days or 5 mg/kg q24h for 5 days, PO, have been suggested (Rings and Rings, 1996). Fenbendazole liquid is labeled for Giardia in puppies and kittens in the EU.

Infectious enteric disease

Coccidiosis –­Eimeria Prevention/control: monensin, bovis, E. zeurnii lasalocid, amprolium, decoquinate, sulfaquinoxaline Therapy of acute disease: sulfaquinoxaline, sulfamethazine, amprolium

Sulfadimethoxine, sulfadimidine

Amprolium and sulfadimidine were found superior to halofuginone in an induced Eimeria bareillyi calf model (Sanyal et al., 1985). Toltrazuril was found effective in a dose-­dependent manner against an induced Eimeria bovis model in calves (Mundt et al., 2003).

0005858924.indd 618

Halofuginone lactate, parmomycin

Comments

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Table 29.1  (Continued)

Amoxicillin, ampicillin, penicillin G, ceftiofur, florfenicol, tetracyclines

Fluoroquinolones, aminoglycosides (lack of spectrum and illegal ELDU / prohibitive WD times_

Prognosis of hemorrhagic bowel disease is very guarded, with surgery necessary for resolution in many cases (Dennison et al., 2002). There is no published evidence that antimicrobial intervention changes the clinical outcome. While there are no published data to support florfenicol efficacy in this disease, the general activity against anaerobes makes it a reasonable consideration.

For prevention: lasalocid in calves ≥1 week old (toxic in neonates at effective doses!)

Amprolium, sulfas

See text for comments on clinical trial data for cryptosporidiosis. Affected calves have severe acid–­base and hydration insults.

10/25/2024 10:22:36 AM

Leptospirosis

Oxytetracycline, dihydrostreptomycin, tylosin (spirochetes on label)

Genitourinary

Metritis/endometritis Ceftiofur ( hydrochloride, crystalline free acid), ampicillin, oxytetracycline, sulfabromomethazine, sulfamethazine, sulfaethoxypyridazine (septicemia), tylosin

Genitourinary

Seminal vesiculitis –­ Trueperella pyogenes, Brucella abortus., E. coli, Pseudomonas spp., Actinobacillus seminis, Actinomyces bovis, Histophilus somni (Haemophilus somnus), Salmonella spp., Chlamydia spp.

Amoxicillin (does not eliminate renal carrier state), penicillin/ dihydrostreptomycin, ceftiofur

Antimicrobial therapy has not been shown to make a difference in clinical outcome. Oxytetracycline in the feed at various doses has been used for prevention. Tilmicosin phosphate (likely other macrolides as well), long-­acting oxytetracycline, and florfenicol have been used in therapeutic attempts.

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Genitourinary

Fluoroquinolones are likely active against the pathogen, but ELDU is prohibited in the US.

Ceftiofur was effective in clearing induced leptospirosis (hardjo) in cows at 2.2 and 5.0 mg/kg q24h for 5 days. These regimens were not effective when administered for 3 days. Long-­acting 200 mg/ml oxytetracycline (20 mg/kg) and penicillin/dihydrostreptomycin (25 mg/kg) were effective after single doses (Alt et al., 2001).

Intrauterine administration of penicillins, aminoglycosides, and sulfonamides is questionable, as these may undergo enzymatic cleavage, operate poorly in an anaerobic environment, or lose activity in the presence of pus.

Chenault et al. (2004) reported 14-­day cure rates of 77%, 65%, and 62% for cows suffering from acute postpartum metritis treated with 2.2 mg/kg IM/SQ ceftiofur HCl (CE) q24h for 5 days, 1.1 mg/kg CE q24h for 5 days, and controls, respectively. Konigsson et al. (2000) reported that cows treated with 10mg/kg IM oxytetracycline SID for 5 days demonstrated a shorter time to eradication of intrauterine T. pyogenes and F. necrophorum than untreated controls (p < 0.05). Trueperella pyogenes is the most common agent in the US. Brucella abortus is the most common in countries with this disease. There is debate as to the role of bacterial or viral pathogens in the pathogenesis of seminal vesiculitis (Larson, 1997).

(Continued)

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Drugs for which this disease is a label application (therapy and/or prevention)

Reasonable extra-­label antimicrobial choices

Category

Disease/Pathogen(s)

Genitourinary

Nephritis/ pyelonephritis –­ Corynebacterium renale, Trueperella pyogenes, E. coli

Trimethoprim/sulfadiazine, trimethoprim/sulfadoxine

For C. renale, Trueperella pyogenes –­penicillin G, ampicillin, florfenicol, tetracycline; for E. coli –­ceftiofur, fluoroquinolones (where legal)

Genitourinary

Cystitis

Amoxicillin, trimethoprim/ sulfadiazine, trimethoprim/ sulfadioxine

Amoxicillin, ampicilllin, ceftiofur, oxytetracycline, florfenicol, fluroquinolones (where legal), penicillin G, trimethoprim/sulfa

Enrofloxacin –­mycoplasma arthritis in cattle less than 2 years old

Oxytetracycline, florfenicol, fluoroquinolones (where allowed by law), tulathromycin, spectinomycin, gamithromycin, lincomycin (given due consideration to potential rumen flora alterations).

Musculoskeletal Adult arthritis –­ Histophilus somni, Mycoplasma bovis

0005858924.indd 620

Unreasonable extra-­ label antimicrobial selections for this disease

Comments

E. coli –­ aminoglycosides have a prohibitively long withdrawal time. MIC distributions for florfenicol and tetracyclines show many isolates with high MICs (EUCAST). Antimicrobials for cystitis have traditionally been chosen for their urine concentrations. However, the infection of concern is in the wall of the bladder, not the urine. Therefore, while urine concentrations may be of benefit, lack of significant urine concentrations does not necessarily preclude selection for cystitis. If M. bovis is suspected, any beta-­lactam would be an unreasonable choice. If another organism is confirmed, then ceftiofur and ampicillin may be considered.

Other pathogens may be present as listed for neonatal arthritis. However, therapy of adult bovine arthritis should include consideration of these organisms unless ruled out by culture. An extended duration of therapy (1–­2 weeks) and a prolonged recovery period are necessary.

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Table 29.1  (Continued)

10/25/2024 10:22:36 AM

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Musculoskeletal Neonatal arthritis –­ E. coli, Trueperella pyogenes, Staphylococcus spp., Streptococcus spp.

Amoxicillin trihydrate, amoxicilin/ clavulanic acid, procaine penicillin/ dihydrostreptomycin, procaine penicillin G

Potentiated sulfonamides, flouroquinolones (where allowed by law)

The potential presence of E. coli and the varied susceptibility results of ampicillin, florfenicol, and oxytetracycline suggest they are not primary considerations for this disease. The primary metabolite of ceftiofur has a greatly elevated MIC90 value for Staphylococcus spp. compared to the parent compound (Salmon et al., 1996), indicating it is not a primary choice where Staph. spp. may be part of the infection.

Central nervous system disease

Listeriosis –­Listeria monocytogenes

Procaine penicillin/ dihydrostreptomycin, procaine penicillin G

Penicillin G, oxytetracycline, enrofloxacin (depending on legal status). Therapy durations of 1–­2 weeks may be necessary.

Varying results are reported for the recomended drugs. Five of 6 bulls in a case report survived after therapy with oxytetracycline and dexamethasone (Ayars et al., 1999). A sheep and goat case report indicated poor response to chloramphenicol and oxytetracycline, but 6 of 9 animals recovered when treated with penicillin and gentamicin (Braun et al., 2002). Enrofloxacin has been reported as effective (Tripathi et al., 2001) but is illegal in countries with a ban on extra-­label use of fluoroquinolones in food animals (e.g., United States).

Central nervous system disease

Thromboembolic meningoencephalitis (TEME), Histophilus somni (Haemophilus somnus)

Ceftiofur, oxytetracycline, florfenicol

Oxytetracycline is a standard drug of choice for this application. Florfenicol is also suggested due to low MICs for H. somni combined with high lipid solubility.

(Continued)

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Drugs for which this disease is a label application (therapy and/or prevention)

Reasonable extra-­label antimicrobial choices

Unreasonable extra-­ label antimicrobial selections for this disease

Category

Disease/Pathogen(s)

Central nervous system disease

Meningitis –­E. coli in neonates, multiple other pathogens possible

Procaine penicillin/ dihydrostreptomycin

Ceftiofur, florfenicol, fluoroquinolones (where legal), trimethoprim/sulfa

Due to inconsistent coverage of the potential Enterobacterales component: penicillin G, first-­generation cephalosporins, macrolides, tetracyclines, florfenicol.

While consideration of blood–­brain barrier penetration is valid, it is likely that this barrier is disrupted in meningitis, allowing greater penetration of water-­soluble compounds. Doxycycline is a lipid-­ soluble tetracycline but the high protein binding in serum limits the amount available to the diffusionary pool, and therefore CNS penetration.

Central nervous system disease

Otitis media and interna –­potential pathogens include respiratory (all ages) and enteric pathogens (neonates). Mycoplasma bovis should be suspected in dairy calves where M. bovis mastitis is present in the herd.

Trimethoprim/sulfadiazine, tylosin

In cattle where respiratory pathogens are suspected: macrolides, florfenicol, fluoroquinolones (where legal). Beta-­lactams might be expected to have lower concentrations in otic tissues and no activity against Mycoplasma spp.

Aminoglycosides may be expected to have extensive binding to protein debris at the site of infection and are less active in areas with lowered pH.

Without adequate trial data, extra-­ label recommendations are made on the basis of reported pathogen population MICs and lipid solubility of the compound. Many of the extra-­label recommendations would not have activity against at least one possible pathogen (e.g., enrofloxacin –­Strep. spp., ceftiofur –­Staph. spp. and M. bovis, macrolides and florfenicol –­ inconsistent against Enterobacterales, penicillin G and ampicillin –­ Enterobacterales and M. bovis).

Tissue/ integumentary disease

Infectious bovine keratoconjunctivitis (pinkeye) –­ Moraxella bovis

Oxytetracycline, topical gentamicin, tulathromycin Florfenicol (Canada)

Penicillin G, tilmicosin, topical benzathine cloxacillin, ceftiofur

0005858924.indd 622

Comments

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Table 29.1  (Continued)

Florfenicol was found to be effective against IBK at either of the label dose regimens (Angelos et al., 2000; Dueger et al., 1999). Topical benzathine cloxacillin, 250 or 375 mg/eye, has been shown to be effective in naturally occurring and induced pinkeye models

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Tissue/ integumentary disease

Infectious pododermatitis (foot rot) –­Fusobacterium necrophorum, Bacteroides melaninogenicus (Prevotella melaninogenica), Porphyromonas levii

Procaine penicillin G, Amoxicillin, ceftiofur (sodium, ampicillin trihydrate hydrochloride, crystalline free acid), erythromycin, florfenicol, oxytetracycline, sulfabromomethazine, sulfadimethoxine, sulfaethoxypyridazine, sulfamethazine, tulathromycin, tylosin, cefquinome, tilmicosin, sulfadiazine/ trimethoprim chlortetracycline –­prevention of footrot (Canada)

Different labels will have different pathogens. Severe tissue reactions result from intramuscular use of tylosin and erythromycin.

Tissue/ integumentary disease

Actinobacillosis, “wooden tongue” –­ Actinobacillus lignieresii

Oxytetracycline, amoxicillin trihydrate, amoxicillin/clavulanic acid, cephalexin, dihydrostreptomycin, trimethoprim/ sulfadiazine (Actinobacilli on label)

Streptomycin, sodium iodide combined with antimicrobial therapy for effect on granulomatous tissue

A case report indicated that cattle receiving IV sodium iodide and intralesional streptomycin regressed lesions faster than negative controls or penicillin-­treated cattle (Campbell et al., 1975). No clinical trials are available.

Tissue/ integumentary disease

Actinomycosis, “lumpy jaw” –­ Actinomyces bovis

Amoxicillin trihydrate, amoxicilin/ clavulanic acid, dihydrostreptomycin, cefalexin, trimethoprim/ sulfadiazine (Actinomycae on label)

Penicillin G, ampicillin trihydrate, oxytetracycline. Sodium iodide may be combined with antimicrobial therapy for effect on granulomatous tissue

No clinical trials are available to confirm efficacy of these antimicrobials. Prolonged therapy is recommended with surgical debridement of the lesion if possible.

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(Daigneault and George, 1990). Tilmicosin was shown to be effective at both 5 and 10 mg/kg (Zielinski et al., 1999). Although local penicillin G is a standard treatment, one report indicated no difference in healing of naturally occurring IBK after subconjunctival administration (Allen et al., 1995).

(Continued)

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Category

Disease/Pathogen(s)

Tissue/ integumentary disease

Blackleg –­C. chauvoei; malignant edema –­C. sordellii, C. septicum; tetanus –­Clostridium tetani; bacillary hemoglobinuria –­ Clostridium hemolyticum; Black disease –­C. novyi

Tissue/ integumentary disease

Peritonitis –­ Escherichia coli, Trueperella pyogenes, Clostridium perfringens, multiple Gram-­positive and Gram-­negative aerobes and anaerobes. Isolate reports in other species include organisms in all 4 quadrants.

Tissue/ integumentary disease

Omphalophlebitis (navel ill)

0005858924.indd 624

Drugs for which this disease is a label application (therapy and/or prevention)

Reasonable extra-­label antimicrobial choices

Amoxicillin trihydrate, amoxicillin/ clavulanic acid, cefalexin, procaine penicillin G (C. chauvoei), procaine/ benzathine penicillin G (C. chauvoei), tylosin

Penicillin G, florfenicol

Trimethoprim/sulfa (probably the most consistent for E. coli), florfenicol, oxytetracycline (both inconsistent on E. coli), ceftiofur for short withdrawal but may not cover Staph. spp.   Combination therapy may be appropriate to increase spectrum of coverage. Amoxicillin trihydrate, amoxicilin/ clavulanic acid, procaine penicillin/ dihydrostreptomycin, procaine penicillin G

Unreasonable extra-­ label antimicrobial selections for this disease

Comments

All the approved drugs have “clostridia” on the label without indications for specific clostridial diseases unless indicated. Japanese isolates of C. perfringens, C. septicum, and C. sordellii displayed phenotypic resistance to oxytetracycline and were confirmed to carry oxytetracycline resistance genes (Sasaki et al., 2001). Penicillin/ gentamicin is reasonable as to spectrum but gentamicin engenders an extreme withdrawal that precludes salvage slaughter attempts in recovered animals. Fluoroquinolones lack activity against important anaerobic pathogens and would be prohibited for use in the US.

No clinical trials are available in cattle. Recommendations are based on activity against likely pathogens, lipid solubility, and duration of activity. An extended duration of therapy (≥1 week) is necessary. Prognosis is extremely poor in advanced cases. Note that the MIC90 of the ceftiofur metabolite against Staph. spp is approximately 8 times that of the parent compound.

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Table 29.1  (Continued)

10/25/2024 10:22:37 AM

Trichophytosis (ringworm)

Tissue/ integumentary disease

Rainrot (dermatophilosis) –­ Dermatophilus congolensis

Cardiovascular/ systemic

Anaplasmosis

Cardiovascular/ systemic

Endocarditis –­ Trueperella pyogenes and Streptococcus spp. are most common. Escherichia coli, other organisms also possible. Enterococcus spp.?

Benzalkonium chloride (0.15% topical solution), enilconazole

Chlortetracycline in the feed for control of active infection; oxytetracycline, enrofloxacin (US conditionally approved)

Topical iodine solution/scrub

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Tissue/ integumentary disease

Penicillin G, oxytetracycline, florfenicol

Penicillin G and oxytetracycline are often cited for therapy of dermatophilosis. A paper evaluating MIC and MBC concentrations, in vitro data, and unbound serum concentrations also recommended erythromycin, ampicillin, streptomycin, amoxicillin, and chloramphenicol (Hermoso-­de Mendoza et al., 1994).

Imidocarb diproprionate

Prevention or amelioration of clinical signs with oxytetraycline are well established. However, there are reports citing both successful and unsuccesful clearance of carriers with oxytetracycline. More recent work has documented unsuccessful clearance of induced anaplasmosis carrier status with the OIE regimen of 22 mg/kg oxytetracycline, IV, q24h, for 5 days (Coetzee et al., 2005). Clearance of the carrier state with imidocarb has been documented (Roby, 1972).

Penicillin G, presence of a Gram-­negative on blood culture indicates ampicillin, amoxicillin, or cefiofur

Prolonged therapy is necessary; 4–­6 weeks has been suggested as an appropriate duration (Dowling and Tyler, 1994; McGuirk, 1991). Lack of clinical efficacy may be due to lack of antimicrobial penetration into vegetative lesions. Florfenicol would be appropriate for pathogens with appropriate MICs (variable on E. coli).

(Continued)

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Category

Disease/Pathogen(s)

Drugs for which this disease is a label application (therapy and/or prevention)

Reasonable extra-­label antimicrobial choices

Unreasonable extra-­ label antimicrobial selections for this disease

Comments

In cases where the law and economics permit, fluoroquinolones would be appropriate if an organism other than a Strep. spp. was confirmed. Cardiovascular/ systemic

0005858924.indd 626

Anthrax –­Bacillus anthracis

Amoxicillin, amoxicillin/ clavulanic acid, tylosin (Bacillus on label) Oxytetracycline

Penicillin G, fluoroquinolones (where legal) doxycycline, first-­generation cephalosporins. Chloramphenicol results suggest florfenicol may be an option.

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Table 29.1  (Continued)

A study evaluating the MICs of 25 genetically diverse B. anthracis isolates from multiple countries reported MIC90 values as follows: ciprofloxacin 0.09 μg/ml, penicillin 0.2 μg/ml, doxycycline 0.34 μg/ml, cefuroxime 32 μg/ml, cephalexin 0.25 μg/ml, cefaclor 1.65 μg/ml, and tobramycin 0.97 μg/ml (Coker et al., 2002). Except for cefuroxime and possibly cefachlor, these MIC90 values are in a range where efficacy might be expected with typically used doses. Universally “susceptible” disk diffusion results with unvalidated interpretive criteria have been reported for tetracycline, ampicillin, streptomycin, chloramphenicol, and erythromycin in South African isolates (Odendaal et al., 1990).

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Rosenbusch et al. (2005) 223 US isolates

Antimicrobial

MIC90 Range MIC50 (μg/ml) (μg/ml) (μg/ml)

Ayling et al. (2000) 62 British isolates MIC50 MIC90 Range (μg/ml) (μg/ml) (μg/ml)

Klein et al. (2019) 232 European isolates

Jelinski et al.(2020) 211 Canadian isolates

MIC50 MIC90 Range (μg/ml) (μg/ml) (μg/ml)

MIC50 MIC90 Range (μg/ml) (μg/ml) (μg/ml)

0.25

Enrofloxacin

0.25

0.5

0.03–­4

NA

NA

NA

0.5

8

0.125–­32

NA

NA

NA

0.5

0.5

0.25–­8

0.25

1

0.064–­8

Florfenicol

1

4

0.06–­8

4

16

4–­128

4

8

0.5–­32

2

2

≤0.25–­8

1.0a

1.0a

0.5–­1.0a

16

0.25 to > 32

NA

NA

NA

NA

NA

NA

8

16

≤1–­32

NA

NA

NA

Gamithromycin

>64

>64

0.5 to >64

>256

>256

1 to >256

Marbofloxacin

1

4

0.25 –­>64 2

4

0.5–­16

NA

NA

NA

NA

NA

NA

>128

>128

8 to >128

>256

>256

2 to >256

NA

NA

Oxytetracycline

2

16

0.125 to >32

32

64

2 to >128

8

32

0.25 to >64

Spectinomycin

2

4

1 to >16

4

>128

2 to >128

NA

NA

NA

Tildipirosin Tilmicosin Tulathromycin

64 NA

>128 NA

0.5 to >128 NA

Tylosin a

 Nuflor® Gold label (2009), 59 US isolates. b  Draxxin® label (2009), 43 US isolates. NA, data not available.

0005858924.indd 627

>128 NA

>128 NA

16 to >128 NA

>64

>64

0.25 to >64

≤0.12–­16

MIC50 MIC90 Range (μg/ml) (μg/ml) (μg/ml)

Danofloxacin

Chlortetracycline 4

4

Label data as indicated by footnote

>64

>64

0.032 to >64

>256

>256

≤0.25 to >256

64

>64

0.5 to >64

>128

>128

1 to >128

NA

NA

NA

NA

NA

NA

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Table 29.2  Mycoplasma bovis susceptibility data.

NA 0.125

b

b

1

≤0.063 to >64b

10/25/2024 10:22:37 AM

Antimicrobial Therapy in Beef Cattle

consideration of those antimicrobials with M. bovis on the label for respiratory disease, even though the site of infection is different. For some cases, the tetracyclines may be appropriate, although it is important to recognize that the pharmacokinetics of injectable and oral tetracyclines are markedly different. The MICs for tilmicosin reported in Table 29.2 are considerably higher than those reported for respiratory pathogens against which this antimicrobial is effective, making effectiveness questionable for Mycoplasma spp. While the fluoroquinolones display good in vitro activity against M. bovis, and one of the compounds has M. bovis on the label for respiratory disease, extra-­label use of this drug class in food animals is illegal in the US. Therefore, use of any fluoroquinolone for therapy of arthritis, otitis media or tenosynovitis would be illegal in the US. In countries without this restriction, the fluoroquinolones are a reasonable consideration for extra-­label therapy of these disease conditions due to M. bovis. However, the pharmacodynamic or clinical justification for use of a fluoroquinolone for this specific pathogen in a nonrespiratory scenario has not been confirmed. Evaluating the impact of drug selection for Mycoplasma infections can be problematic as many of the clinical signs for nonrespiratory infections (e.g., arthritis) persist well beyond the infectious stage of the disease due to local inflammatory reactions.

Enteric Disease and Septicemia Associated with E. coli and Salmonella spp. A previous review of the literature showed that there is a paucity of data to support the efficacy of antimicrobial therapy for bacterial enteric disease in calves (Constable,  2004). Little has changed between the time of this review and the writing of this chapter (2023). The practitioner is hampered by two ­obstacles: the previously mentioned lack of clinical data from prospective controlled and

randomized clinical trials, and the lack of ­validated susceptibility testing breakpoints for classification of enteric pathogens as ­susceptible or resistant. However, the authors’ discussions with practitioners indicate that few would be willing to forego antimicrobial therapy considering that a proportion of calves with enteric disease are likely septicemic. Also, the potential for septicemia in adult cattle with coliform masitits and salmonellosis calls for guidance in reasonable antimicrobial selection. From an empirical approach, reasonable initial considerations include third-­generation cephalosporins, potentiated aminopenicillins, fluoroquinolones (not legal in the US), ­potentiated sulfonamides, and florfenicol. The confirmation of these initial selections would depend heavily on clinical response to therapy and on susceptibility testing as presently available. Susceptibility testing for enteric disease is not based on CLSI-­approved breakpoints but rather on breakpoints developed for another veterinary indication or adapted from human medicine. Approved CLSI breakpoints are developed based on a combination of in  vivo efficacy data coupled with isolate MIC distributions and pharmacokinetic/ pharmacodynamic (PK/PD) data. When applying these breakpoints to other indications, such as enteric disease, it is hoped that the PK/PD indices and the changes in MIC due to a resistance gene are at least similar. Therefore, we might more accurately refer to the process for enteric disease as “resistance testing,” where resistant isolates would be considered more likely to possess resistance genes, rendering the antimicrobial incapable of having an effect on the pathogen. Therefore, a reasonable approach is to first rule out any of the potential enteric therapeutics based on legal issues relevant to the practice area and also eliminate antimicrobials that lack activity against the suspected pathogen(s). For example, aminoglycosides would be a poor choice for clostridial infections as they lack activity against anaerobes. Next, empirical therapy may be guided by accessing enteric

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628

culture susceptibility summaries available from your diagnostic laboratory, or by monitoring susceptibility trends within specific production units. A preponderance of resistant classifications for a potential antimicrobial would indicate that the population of pathogens being submitted to that laboratory likely carry some resistance mechanism. This antimicrobial could therefore be moved down on the list of potential selections. A diagnostic laboratory finding of “susceptible” does not have any validated correlation with the likelihood of therapeutic success in the patient for enteric disease. However, we would assume that in the absence of phenotypic resistance (lack of resistance genes), the antimicrobial has some potential to contribute to the clinical recovery to the extent possible with the regimen and site of infection. There are obviously many assumptions in this discussion, highlighting the need for controlled clinical trials addressing the antimicrobial treatment component of neonatal enteric disease in calves.

Cryptosporidium parvum Multiple antimicrobials have been evaluated in C. parvum calf disease models. Antimicrobials reported as ineffective in calves up to 14 days of age when administered in the milk replacer during a 10-­day challenge model include amprolium, sulfadimidine, dimetridazole, metronidazole, ipronidazole, quinacrine, and monensin. Trimethoprim/sulfadiazine was also ineffective when administered daily as a bolus. Lasalocid was ineffective at 0.8  mg/kg per day. At 8 mg lasalocid/kg per day, six of the 10 treated calves died, with one of the four surviving calves becoming infected (Moon et  al.,  1982). Sulfadimethoxine has also been shown to be ineffective against C. parvum in a 1–­7-­day-­old calf challenge model (Fayer, 1992). Lasalocid has been used as a preventive or therapeutic agent for Cryptosporidium in calves based on anecdotal reports. This use in neonatal calves has resulted in reported

toxicities after 100  mg twice daily in milk replacer or 200 mg oral once-­daily doses starting at birth, with death occurring after 1–­3 administrations (Benson et al., 1998). The lead author confirmed this toxicity experimentally by dosing neonatal calves once at 5 mg/kg. In other studies, lasalocid has been tolerated in calves ≥7 days old with cessation of oocyst shedding three days after the last of three daily doses of 15  mg/kg (Gobel,  1987). These data would suggest that effective doses of lasalocid are toxic in neonatal calves but may be used in calves of at least one week of age. A trial evaluating decoquinate in a calf C. parvum challenge model found a significant decrease in number of days with abnormal stool scores in the treated groups given 875 or 1750 mg (10 times label dose) decoquinate per day, but no difference in oocyst shedding or weight gain (Redman and Fox, 1994). Another challenge study found no difference in days to diarrhea, days to shedding, or duration of diarrhea or oocyst shedding in calves given 2 mg/ kg decoquinate in milk replacer (Moore et al., 2003). In naturally occurring C. parvum infections, halofuginone lactate administered in milk replacer at 60 μg/kg per day cleared all shedding of oocysts within six days after the start of treatment in 98% of the treated animals. It should be noted that 93% of the untreated controls in this study also cleared the organism within 10  days of arrival at the facility (Villacorta et  al.,  1991). In a natural disease model, calves receiving 5 mg of halofuginone lactate daily in milk replacer were 70% less likely to shed C. parvum oocysts compared to untreated controls. Weight gain and milk and starter intakes were not significantly different between groups (Jarvie et al., 2005). In a challenge study, halofuginone reduced disease at 60 and 120 μg/kg per day but was ineffective at 30 μg/kg per day (Naciri et al., 1993). Other antimicrobials such as paromomycin, azithromycin, and clarithromycin have been demonstrated to be efficacious in murine ­models or human therapy (Fichtenbaum

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­Antimicrobial Use in Cattl  629

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et al., 1993; Rehg, 1991, Holmberg et al., 1998). The prophylactic potential of paromomycin was also demonstrated in a calf model (Fayer and William, 1993). Azithromycin significantly suppressed oocyst shedding and resulted in significant clinical improvement and weight gain

in naturally infected calves (Elitok et al., 2005; Nasir et al., 2013). However, such use of human antimicrobials is questionable for antimicrobial stewardship and the costs of these three agents are prohibitive for food animals and have mainly prevented their use.

­References and Bibliography 21 CFR Part 530. 2012. New Animal Drugs; Cephalosporin Drugs; Extralabel Animal Drug Use; Order of Prohibition. www.gpo.gov/ fdsys/pkg/FR-­2012-­01-­06/pdf/2012-­35.pdf Allen LJ, et al. 1995. Effect of penicillin or penicillin and dexamethasone on cattle with infectious keratoconjunctivitis, J Am Vet Med Assoc 206:1200. Alt DP, et al. 2001. Evaluation of antibiotics for treatment of cattle infected with Leptospira borgpetersenii serovar hardjo, J Am Vet Med Assoc 219:636. AMDUCA. 1996. 21 CFR Part 530, Extralabel Drug Use in Animals; Final Rule, Department of Health And Human Services, Food and Drug Administration, Docket No. 96N-­0081, RIN 0910-­AA47, ACTION: Final rule. www. ecfr.gov/current/title-­21/chapter-­I/ subchapter-­E/part-­530?toc=1 Angelos JA, et al. 2000. Efficacy of florfenicol for treatment of naturally occurring infectious bovine keratoconjunctivitis. J Am Vet Med Assoc 216:62. Apley MD. 2003. Susceptibility testing for bovine respiratory and enteric disease. Vet Clin North Am Food Anim Pract 19:3. AVMA. 2003. Judicious Therapeutic Use of Antimicrobials. www.avma.org/resources-­ tools/avma-­policies/judicious-­therapeutic­use-­antimicrobials AVMA. 2021. Antimicrobial stewardship definition and core principles. www.avma.org/ resources-­tools/avma-­policies/antimicrobial-­ stewardship-­definition-­and-­core-­principles Ayars WHJ, et al. 1999. An outbreak of encephalitic listeriosis in Holstein bulls. Bovine Practitioner 33:138.

Ayling RD, et al. 2000. Comparison of in vitro activity of danofloxacin, florfenicol, oxytetracycline, spectinomycin and tilmicosin against recent field isolates of Mycoplasma bovis. Vet Rec 146:745. Baba E, et al. 1989. Antibiotic susceptibility of Fusobacterium necrophorum from bovine hepatic abscesses. Br Vet J 145:195. Benson JE, et al. 1998. Lasalocid toxicosis in neonatal calves. J Vet Diagn Invest 10:210. Berg JN, Scanlan CM. 1982 Studies of Fusobacterium necrophorum from bovine hepatic abscesses: biotypes, quantitation, virulence, and antibiotic susceptibility. Am J Vet Res 43:1580. Booker CW, et al. 2017. An evaluation of concomitant therapy for the treatment of arrival fever in feedlot calves at ultra-­high risk of developing undifferentiated fever/bovine respiratory disease. Int J Appl Res Vet Med 15:85. Bowers DR, et al. 2013. Outcomes of appropriate empiric combination versus monotherapy for Pseudomonas aeruginosa bacteremia. Antimicrob Agents Chemother 57:1270. Braun U, et al. 2002. Clinical findings and treatment of listeriosis in 67 sheep and goats. Vet Rec 150:38. Campbell SG, et al. 1975. An unusual epizootic of actinobacillosis in dairy heifers. J Am Vet Med Assoc 166:604. Chenault JR, et al. 2001 Efficacy of ceftiofur hydrochloride administered parenterally for five consecutive days for treatment of acute post-­partum metritis in dairy cows. 34th Annual Convention of the AABP, pp. 137–­138.

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CLSI. 2008. Publication M31-­A3: Performance Standards for Antimicrobial Disk and Dilution Susceptibility Tests for Bacteria Isolated form Animals; Approved Standard, 3rd edn. www. clsi.org CLSI. 2024a. Performance Standards for Antimicrobial Disk and Dilution Susceptibility Tests for Bacteria Isolated from Animals, 7th edn. CLSI supplement VET01S. Clinical and Laboratory Standards Institute. CLSI. 2024b. Understanding Susceptibility Test Data as a Component of Antimicrobial Stewardship in Veterinary Settings, 2nd edn. CLSI report VET09. Clinical and Laboratory Standards Institute. Coetzee JF, et al. 2005. Comparison of three oxytetracycline regimens for the treatment of persistent Anaplasma marginale infections in beef cattle. J Vet Parasitol 127:61. Coker PR, et al. 2002. Antimicrobial susceptibilities of diverse Bacillus anthracis isolates. Antimicrob Agents Chemother 46:3843. Constable PD. 2004. Antimicrobial use in the treatment of calf diarrhea. J Vet Intern Med 18:8. Daigneault J, George LW. 1990. Topically applied benzathine cloxacillin for treatment of experimentally induced infectious bovine keratoconjunctivitis. Am J Vet Res 51:376. Daigneault J, George LW. 1990. Topically applied benzathine cloxacillin for treatment of experimentally induced infectious bovine keratoconjunctivitis. Am J Vet Res 51:376. Dennison AC, et al. 2002. Hemorrhagic bowel syndrome in dairy cattle: 22 cases (1997–­2000). J Am Vet Med Assoc 221:686. Dowling PM, Tyler, JW. 1994. Diagnosis and treatment of bacterial endocarditis in cattle. J Am Vet Med Assoc 204:1013. Dueger EL, et al. 1999. Efficacy of florfenicol in the treatment of experimentally induced infectious bovine keratoconjunctivitis. Am J Vet Res 60:960. Duran SP, et al. 1991. Comparative in-­vitro susceptibility of Bacteriodes and Fusobacterium isolated from footrot in sheep to 28 antimicrobial agents. J Vet Pharmacol Ther 14:185.

Elitok B, et al. 2005. Efficacy of azithromycin dihydrate in treatment of cryptosporidiosis in naturally infected dairy calves. J Vet Intern Med 19(4):590. Fayer R. 1992. Activity of sulfadimethoxine against cryptosporidiosis. J Parasitol 78:534. Fayer R, William E. 1993 Paromomycin is effective as prophylaxis for cryptosporidiosis in dairy calves. J Parisitol 79:771. FDA/CVM. 2005. www.fda.gov/cvm. Search “prohibited extralabel use.” Fichtenbaum CJ, et al. 1993. Use of paromomycin for treament of cryptosporidiosis in patients with AIDS. Clin Infect Dis 16:298. Gobel E. 1987. Diagnose und therapie der akuten kryptosporidiose beim kalb [Diagnosis and treatment of acute cryptosporidiosis in the calf]. Tierarztliche-­ Umschar 42:863. Heffernan AJ, et al. 2020. β-­lactam antibiotic versus combined β-­lactam antibiotics and single daily dosing regimens of aminoglycosides for treating serious infections: a meta-­analysis. Int J Antimicrob 55:105839. Hermoso-­de Mendoza J, et al. 1994. In vitro studies of Dermatophilus congolensis antimicrobial susceptibility by determining minimal inhibitory and bacteriocidal concentrations. Br Vet J 150:189. Holmberg SD, et al. 1998. Possible effectiveness of clarithromycin and rifabutin for cryptosporidiosis chemoprophylaxis in HIV disease. J Am Med Assoc 279:384. Jang S, Hirsh DC. 1994. Characterization, distribution, and microbiological associations of Fusobacterium spp. in clinical specimens of animal origin. J Clin Microbiol 32:384. Jarvie BD, et al. 2005. Effect of halofuginone lactate on the occurrence of Cryptosporidium parvum and growth of neonatal dairy calves. J Dairy Sci 88:1801. Jelinski M, et al. 2020. Antimicrobial sensitivity testing of Mycoplama bovis isolates derived from western Canadian feedlot cattle. Microorganisms 8:124. Jousimies-­Somer H, et al. 1996. Susceptibilities of bovine summer mastitis bacteria to

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  ­References and Bibliograph

Antimicrobial Therapy in Beef Cattle

antimicrobial agents. Antimicrob Agents Chemother 40:157. Klein U, et al. 2019. New antimicrobial susceptibility data from monitoring of Mycoplasma bovis isolated in Europe. Vet Microbiol 238:108432. Konigsson K, et al. 2000. Effects of NSAIDs in the treatment of postpartum endometritis in the cow. Reprod Domest Anim 35:186. Larson RL. 1997 Diagnosing and controlling seminal vesiculitis in bulls. Vet Med 12:1073. Lechtenberg KF, et al. 1998. Antimicrobial susceptibility of Fusobacterium necrophorum insolated from bovine hepatic abcesses. Am J Vet Res 59:44. Mateos E, et al. 1997. Minimum inhibitory concentrations for selected antimicrobial agents against Fusobacterium necrophorum isolated from hepatic abscesses in cattle. J Vet Pharmacol Ther 20:21. McGuirk SM. 1991. Treatment of cardiovascular disease in cattle. Appl Pharmacol Ther 7:729. Moon HW, et al. 1982 Attempted chemoprophylaxis of cryptosporidiosis in calves. Vet Rec 110:181. Moore DA, et al. 2003. Prophylactic use of decoquinate for infections with Cryptosporidium parvum in experimentally challenged neonatal calves. J Am Vet Med Assoc 223:839. Mundt HC, et al. 2003. Efficacy of toltrazuril against artificial infections with Eimeria bovis in calves. Parasitol Res 90(3): S166. Naciri MR, et al. 1993. The effect of halofuginone lactate on experimental Cryptosporidium parvum infections in calves. Vet Parasitol 45:199. Nasir A, et al. 2013. Treating Cryptosporidium parvum infection in calves. J Parasitol 99(4):715. Odendaal MW, et al. 1990. The antibiotic sensitivity patterns of Bacillus anthracis isolated from the Kruger National Park. J Vet Res 58:17. Piriz Duran S, et al. 1990 Susceptibilities of Bacteroides and Fusobacterium spp. from foot rot in goats to 10 beta-­lactam antibiotics. Antimicrob Agents Chemother 34:657.

Redman DR, Fox JE. 1994. The effect of varying levels of DECCOX on experimental cryptosporidia infections in Holstein bull calves. Proceedings of the 26th Annual American Association of Bovine Practitioners Conference, p. 157. Rehg JE. 1991. Activity of azithromycin against Cryptosporidia in immunosuppressed rats. J Infect Dis 163:1293. Rings DM, Rings MB. 1996. Managing Cryptosporidium and Giardia infections in domestic ruminants. Vet Med 12:1125. Roby TO. 1972. Elimination of the carrier state of bovine anaplasmosis with imidocarb. Am J Vet Res 33:1931. Rosenbusch RF. 1998. Antibiotic susceptibility of Mycoplasma bovis strains recovered from mycoplasmal pneumonia and arthritis in feedlot cattle. AS leaflet R1548. 1998 Beef Research Report, Iowa State University. Rosenbusch RF, et al. 2005. In vitro antimicrobial inhibition profiles of Mycoplasma bovis isolates recently recovered from various regions of the United States from 2002 to 2003. J Vet Diagn Invest 17:436. Salmon SA, et al. 1996. In vitro activity of ceftiofur and its primary metabolite, desfuroylceftiofur, against organisms of veterinary importance. J Vet Diagn Invest 8:332. Samitz EM, et al. 1996. In vitro susceptibilities of selected obligate anaerobic bacteria obtained from bovine and equine sources to ceftiofur. J Vet Diagn Invest 8:121. Sanyal PK, et al. 1985. Chemotherapeutic effects of sulphadimidine, amprolium, halofuginone and chloroquine phosphate in experimental Eimeria bareillyi coccidiosis of buffaloes. Vet Parasitol 17:117. Sasaki Y, et al. 2001. Tetracycline-­resistance genes of Clostridium perfringens, Clostridium septicum and Clostridium sordellii isolated from cattle affected with malignant edema. Vet Microbiol 83:61. Schmid A, et al. 2019. Monotherapy versus combination therapy for multidrug-­resistant Gram-­negative infections: systematic review and meta-­analysis. Sci Rep 9:15290.

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Sweeney MT, et al. 2008. In vitro activities of tulathromycin and ceftiofur combined with other antimicrobial agents using bovine Pasteurella multocida and Mannheimia haemolytica isolates. Vet Ther 9:212. Tripathi D, et al. 2001. Serodiagnosis and treatment of listeriosis in repeat breeder cattle. Indian J Anim Sci 71:3. Villacorta I, et al. 1991. Efficacy of halofuginone lactate against Cryptosporidium

parvum in calves. Antimicrob Agents Chemother 35:283. Zielinski GC, et al. 1999. Efficacy of different dosage levels and routes of inoculation of tilmicosin in a natural outbreak of infectious bovine keratoconjunctivitis. Proceedings of the 32nd Annual Convention of the American Association of Bovine Practitioners, p.261.

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  ­References and Bibliograph

30 Antimicrobial Therapy in Dairy Cattle Sarah Wagner and Sarah Depenbrock

In dairy cattle production, good antimicrobial stewardship requires changes in the way bacterial infections are identified and mitigated to reduce antimicrobial interventions. This chapter provides evidence-­based information on antimicrobial use decisions for the more common diseases of dairy animals: intramammary infections (IMI, also commonly called mastitis), metritis, bovine respiratory disease (BRD), and calf diarrhea. The reader is encouraged to consider that herd-­wide disease risk identification and mitigation improve overall animal health outcomes more than the use of antimicrobials after disease prevention efforts have failed.

D ­ airy Animal Use Considerations The intended use of the dairy animal is very important when making antimicrobial use decisions. Consideration of residue avoidance to protect the food supply and avoid costly mistakes such as contamination of a bulk tank is imperative in dairy practice. Ideally, the antimicrobial selected for use should be one that is labeled for the specific production group(s) indicated on the label. Specific dairy age groups that are commonly defined on antimicrobial labels include preruminating calves, ruminating calves, and adult female dairy cattle (20 months of age or older).

Male calves that are used for veal production include bob veal  –­ calves slaughtered by 21 days of age; milk-­fed veal –­ calves fed only milk or milk replacer that are slaughtered at 18–­20 weeks of age; red veal –­ calves that get additional grain and hay and are slaughtered at 18–­20 weeks; and free raised veal –­calves that remain with their dams on pasture and are slaughtered at 24  weeks. In veal production, antimicrobial use is limited as many antimicrobials carry the disclaimer “not to be used in veal calves” due to physiological differences in very young calves compared to older animals. Residue depletion is complicated by the common disease conditions being treated (e.g., dehydration from diarrhea) and short lifespan for drug withdrawal prior to processing for human consumption. Adult female dairy cattle are considered lactating animals, even if they are not actually lactating, because these animals are at risk for lactation. For example, an animal may inadvertently get bred early and calve early, or abort prior to the anticipated due date but late enough to come into milk, and thus produce milk unexpectedly early. Dairy replacement heifers may have more flexible antimicrobial treatment as long as animals are maintained until adulthood prior to entering the food chain. When label dose, route, volume per site, frequency, duration, indication, and production

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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group/age are followed, the label withdrawal time (WDT) can be used. Any time any one of these conditions is altered, an extended withdrawal interval (WDI) must be provided by the prescribing veterinarian. When an appropriate antimicrobial is not available for the desired age group or lactation status, extra-­ label drug use (ELDU) may be considered, and the jurisdictional regulations should be followed (see Chapter 26). Commercial tests validated for antimicrobials in milk can be purchased for on-­farm use. When antimicrobials are used in severely compromised animals in which drug elimination may be delayed, it is prudent to prolong the withdrawal recommendation past the label WDT for on-­label use or greatly extend the recommended WDI for ELDU.

I­ ntramammary Infection The most common use of antimicrobials on dairy farms is to treat mastitis, which is caused by IMI. The expenses associated with IMI (decreased milk production, decreased milk quality, drug treatment costs, and discarded milk) are considerable, and require dairy producers to implement management programs focused on IMI prevention and treatment. A farm that is experiencing high somatic cell counts, frequent occurrence of clinical IMI, a high prevalence of subclinical IMI, or all of these should investigate the reason and develop a program to mitigate the cause and prevent new occurrences. Money spent on effective prevention of IMI is likely to provide an overall financial benefit to the dairy, and treatment alone is not an effective way to resolve herd-­ level IMI problems. Even on well-­managed farms with IMI prevention protocols in place, treatment of clinical or subclinical IMI may be desirable when there is a good chance that treatment will improve the milk quality, productivity, well-­ being, and longevity of the cow. Subclinical IMI may be detected through a combination

of individual cow somatic cell counts (SCCs) as measured by dairy herd improvement (DHI) testing, or estimated using the California Mastitis Test (CMT) and confirmed by microbial culture of milk samples. Clinical IMI is described as mild when abnormal milk is the only sign; moderate when abnormal milk and udder swelling are present; or severe when milk and udder abnormalities are accompanied by signs of systemic illness such as fever, hypothermia, recumbency, or depression. The discussion presented here focuses on clinical cases of IMI, but the principles described are also generally applicable to the treatment of subclinical IMI.

Cow Factors In some cases, it is more rational not to treat IMI, either because treatment is unnecessary or because treatment is unlikely to result in resolution of clinical signs. Risk factors that have been found to decrease therapeutic efficacy include increasing cow age, high SCC before treatment, long duration of infection, multiple infected quarters, and infections caused by Staphylococcus aureus. Questions to ask about the cow before deciding to treat mastitis are presented in Table 30.1.

Herd-­based Therapeutic Protocols Many dairies implement a standardized approach to IMI prevention and therapy. Key benefits to standardizing IMI therapy are that treatment decisions are made in advance instead of “cow-­side” and that a consistent approach is developed. Less time is spent deciding whether or not to treat a case of IMI, selecting a drug and treatment regimen, and assigning an appropriate WDT for milk and meat from treated cows. Moreover, when treatments are standardized and good records are kept, it is simple to evaluate whether or not a given treatment is successful on the farm. A key component of a standardized approach to

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Table 30.1  Questions to ask about the cow before deciding to treat mastitis. Question

1)  Is this a new case of IMI or a relapse? 2)  How severe is it?

3)  How many quarters are affected? 4)  What is the cow’s stage of lactation? 5)  Does the cow have other health problems or risk factors?

Relevant information ●● ●●

●●

●●

●●

●●

●●

Repeated treatment of a recurrent case of IMI is frequently unrewarding. If a recurrent IMI is to be treated, the therapeutic regimen should be more extensive than what would be used for a mild, acute case. An IMI in a cow that has become systemically ill (septic/toxic) will require a therapeutic protocol that includes systemic antimicrobial therapy, supportive therapy (such as intravenous fluids and nonsteroidal antiinflammatory drugs) and closer monitoring than a case in which clinical signs are limited to the udder and milk. The expense and the likelihood of treatment failure increase as the number of affected quarters increases. For a cow in late lactation, economic and therapeutic advantages may be gained by treating the cow simultaneously with drying off. Older cows and cows with higher SCCs have a diminished likelihood of cure. Treating a cow for IMI may not provide benefit relative to cost if she has other conditions such as chronic lameness or low fertility that negatively affect her welfare, value to the herd, or probable longevity in the herd.

IMI treatment is regular veterinary review of protocols to ensure that they are being followed and continuing to generate expected outcomes.

On-­farm/Rapid Result Microbial Culture and Treatment Simple on-­farm microbial culture systems (or local laboratories that provide rapid milk culture results) allow culture results from each case of IMI to be incorporated into treatment protocols. The simplest and most common approach is to perform aerobic milk culture on agar gel plates with multiple selective media types used to categorize results as Gram-­positive pathogen growth, Gram-­negative pathogen growth, or no pathogen growth. In a typical protocol, no treatment of mild or moderate cases of IMI is initiated for 18–­24 hours while culture results are pending. Intramammary (IMM) antimicrobial treatment is reserved for cows with Gram-­ positive pathogens or mixed pathogens identified on microbial milk culture. Multiple research studies have compared the IMM treatment of all cows with mild or moderate

IMI to the treatment of only those with Gram-­ positive pathogens on milk culture, using outcomes such as bacteriological cure, clinical cure, rate and rapidity of recurrence, and culling. Selective, culture-­based treatment has comparable short-­term and long-­term outcomes to treatment of all clinical cases of IMI. This is because many cases of IMI that yield no growth or Gram-­ negative pathogen growth on culture resolve spontaneously and antimicrobial treatment is of no benefit. Decreasing the number of treated cows on farms that have previously treated every case of IMI results in financial benefit, even after the cost of conducting microbial culture of each new case is factored in. In addition, these culture-­ based selective treatment programs can reduce the number of cows treated with IMM antimicrobials by 50% or more, thereby decreasing the selection pressure for drug-­resistant bacterial populations. If a culture-­based approach is taken, it is necessary to understand that Gram-­negative IMI can develop into chronic infection or severe systemic illness. In addition, Mycoplasma spp. require special culture media and more stringent incubation conditions than this simple

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­Intramammary Infectio 

Antimicrobial Therapy in Dairy Cattle

approach provides, so they will not be identified by typical on-­farm culture programs. Retaining milk samples in the freezer for a period of time after the initial treatment is a useful practice. If the treatment outcome is undesirable, the retained sample can be submitted to a diagnostic laboratory for detailed microbiological diagnosis. By saving the pretreatment sample of cows treated with antimicrobials, there is no need to wait for the drugs to be eliminated from the mammary gland prior to sampling for repeat culture. For cows left untreated that do not improve, a repeat culture may be taken at any time to obtain results indicative of their current microbiological status. Culture is also an irreplaceable aid to determining whether a chronic case of IMI might be resolved by extended antimicrobial therapy or if the pathogen causing the IMI is not amenable to therapy and the greatest financial benefit to the farm would be not to treat it again. An example of a culture-­based herd mastitis treatment protocol is given in Figure 30.1.

Pathogen Identification and Antimicrobial Susceptibility Testing Antimicrobial susceptibility testing is a way of quantifying the interaction between microbes and antimicrobials in the laboratory (see Chapter 2). Validated veterinary breakpoints are used to determine whether a particular laboratory outcome indicates susceptibility for a specific drug, dosing regimen, pathogen, affected species, and disease condition. Currently, validated veterinary breakpoints are available for two US Food and Drug Administration (FDA)-­ approved preparations for IMM treatment of lactating cows (ceftiofur, pirlimycin) and one FDA-­approved intramammary preparation for treatment of dry cows (a combination of penicillin and novobiocin). Susceptibility testing appears to have limited value as an aid to therapeutic decision making for bovine IMI. The relationship between ­susceptibility determined by laboratory susceptibility testing and the outcome of clinical cases of IMI is inconsistent.

The issue is further complicated by the use of variable outcomes in trials assessing resolution of clinical IMI; the achievement of a cure may be defined as resolution of clinical signs, one or more negative microbial cultures, or some combination of these outcomes, and the amount of time between treatment and the assessment of treatment outcomes is not standardized. A more practical approach to assessing whether an IMI therapy works is to design farm protocols for treatment of clinical IMI with selected antimicrobials, then periodically evaluate the protocols for the efficacy of the selected drugs in achieving the farm’s therapeutic objectives. Common therapeutic objectives include clinically normal milk and/or a reduction in SCCs.

Pathogen Factors Microbial culture of IMI is a valuable aid in determining whether to initiate drug therapy and if so, what approach to use. Intramammary infections with certain pathogens are likely to respond to antimicrobial therapy, while other pathogens have a more variable response. As mentioned above, some IMI are likely to resolve without any treatment. Common IMI pathogens that are unlikely to be responsive to antimicrobial therapy are listed in Table 30.2. A brief overview of some other commonly encountered pathogens follows. Streptococcus agalactiae is a contagious IMI pathogen. It is typically responsive to therapy with antimicrobials with activity against Gram-­positive pathogens, such as beta-­lactams and lincosamides. Staphylococcus aureus is also a contagious IMI pathogen. Chronicity decreases the responsiveness of S. aureus infection to antimicrobial therapy. A new case in one quarter of a young cow, caused by a S. aureus strain that is susceptible to penicillin, is more likely to respond to appropriate therapy than one or more quarters chronically infecting an older cow, particularly with a penicillin-­resistant strain. Extending the duration of therapy improves the likelihood of therapeutic success when treating IMI caused by S. aureus. If treatment duration is extended

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Does the cow have abnormal milk? No Yes

Stop here.

Take a milk sample from the affected quarter using aseptic technique and write the cow number, date, AM or PM, and quarter on the sample container.

Is the cow acting sick/depressed? Are her eyes sunken? Does she have diarrhea? Is she down? Does she have a temperature over 104° F? Yes to any

No to all

Treat with intramammary and systemic antibiotics (farm choice), oral and intravenous fluids, and anti-inflammatory drugs. Monitor closely.

Begin microbial culture of milk sample. Incubate for 24 hours.

Check culture after 24 hours and record result. Begin microbial culture of milk sample. Incubate for 24 hours. No growth or contaminated: Do not treat. Check culture in 24 hours, modify treatment if necessary. Continue close monitoring and treatment.

Gram-positive: IMM treatment.

Gram-negative: IMM treatment or not, depending on farm protocol.

Recheck milk culture for microbial pathogen growth after 48 hours. Record milk culture results, treatment administered, if any, and treatment outcome, so that patterns of microbial etiology and treatment efficacy can be determined.

Figure 30.1  Example herd mastitis treatment protocol incorporating on-­farm culture.

beyond the label dosage, additional withdrawal time is required and milk testing for drug residues is recommended. Streptococci other than S. agalactiae and staphylococci other than S. aureus are the most

commonly isolated Gram-­positive pathogens found on microbial milk culture. IMM treatment with amoxicillin or ceftiofur resulted in  similar outcomes including clinical cure (return to normal milk and udder appearance)

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Antimicrobial Therapy in Dairy Cattle

Table 30.2  Mastitis pathogens unlikely to respond to antimicrobial drug treatment. Bacillus spp. Mycobacterium spp. Mycoplasma bovis Nocardia spp.

culture methods (i.e., blood agar in an aerobic culture environment), and it is associated with diseases other than IMI, such as otitis media and joint infections. Mastitis caused by Mycoplasma may occasionally resolve without treatment, but antimicrobial therapy does not affect the outcome (Gelgie et al., 2022).

Pasteurella spp. Proteus spp. Prototheca spp. (algae) Pseudomonas spp. Serratia spp. Trueperella pyogenes Yeasts (e.g., Candida spp.; IMM treatment will delay spontaneous cure)

and bacterial cure (no growth on milk culture) rates among cows affected with mild or moderate mastitis due to a variety of Gram-­positive pathogens, with clinical cure rates higher than bacterial cure rates (Tomazi et al., 2021). Both types of cure were less common in cows affected with Streptococcus uberis than other pathogens. Cows with Trueperella pyogenes or S. aureus infections were excluded from the study because these pathogens are not responsive to treatment, and no cows had a positive culture for S. agalactiae. For IMI with Gram-­negative pathogens that do not spontaneously resolve or that recur, treatment with antimicrobials may be warranted. Multiple studies have failed to produce solid evidence that treatment of a new case of IMI caused by a Gram-­negative pathogen will improve clinical or bacteriological outcomes when compared with no treatment. Unfortunately, what to do with Gram-­negative IMI that does not resolve without treatment or that recurs has not been established. If treatment of such cases is undertaken, selection of a drug with an appropriate spectrum of activity (an aminopenicillin or cephalosporin, not a lincosamide) is indicated, and an extended duration of therapy should be considered. Unlike other mastitis pathogens, Mycoplasma bovis cannot be grown using typical on-­farm

Intramammary Antimicrobial Use After cow factors, culture results, and pathogen factors have been weighed up and the decision has been made to treat IMI, a suitable therapeutic regimen must be designed. Components of a therapeutic regimen include the drug to be used, drug dose, route of administration, frequency of administration, duration of use, and meat and milk withdrawal times. For mild to moderate IMI, antimicrobial therapy is usually administered by the IMM route. There are eight antimicrobials approved by the US FDA for IMM use in lactating cows: amoxicillin, ceftiofur, cephapirin, cloxacillin, erythromycin, hetacillin, penicillin, and pirlimycin. Although they remain approved, products containing erythromycin, cloxacillin or pirlimycin are no longer marketed in the US. In Canada, only ceftiofur and cephapirin are available for IMM treatment in lactating cows. In the European Union (EU) and other countries, available IMM formulations include  amoxicillin/clavulanic acid, cephalexin/­kanamycin, novobiocin/ dihydrostreptomycin/neomycin, neomycin/ streptomycin/penicillin G, cephalexin/­ kanamycin, cefquinome, ­dihydrostreptomycin/ framycetin/penethamate, tetracycline/­ neomycin/bacitracin, rifaximin, lincomycin/ neomycin, cefuroxime, cefoperazone, and ­cloxacillin/neomycin. Intramammary use of drug preparations not specifically manufactured for IMM administration is not recommended; such substances may be irritating to udder tissues and promote inflammation. In addition, compounded preparations are at risk for ­contamination with infectious pathogens,

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and milk and meat withdrawal times recommended for other routes of administration are likely to be inaccurate. Different antimicrobial preparations should not be used simultaneously in one quarter, since interactions between the two drug formulations may decrease efficacy. Antimicrobial spectrum of activity is a key consideration when selecting an antimicrobial for IMM therapy of mastitis. One of the earliest beta-­lactam drugs to be developed, penicillin G, is available for IMM administration alone or in combination formulations. Penethamate is the diethylaminoethyl ester of penicillin G. Penicillin is active against many streptococci and nonpenicillinase-­producing staphylococci. The drug is inactive against the Enterobacterales, and staphylococcal resistance is common. Amoxicillin and hetacillin are aminopenicillins that share similar spectra of activity. The aminopenicillins are active against bacteria susceptible to penicillin G, as well as some Enterobacterales such as E. coli. Many E. coli isolates are now resistant to the aminopenicillins through beta-­lactamase production. Amoxicillin in combination with the beta-­ lactamase inhibitor clavulanic acid is available for IMM administration in the EU; this combination is more effective than aminopenicillins alone against beta-­lactamase-­producing bacterial strains. Cloxacillin is a penicillinase-­resistant penicillin active against penicillinase-­producing S. aureus strains that are resistant to the natural penicillins and aminopenicillins. It is less active against other penicillin-­sensitive organisms. Cephapirin and cephalexin are first-­generation cephalosporin drugs generally active against staphylococci and streptococci and sometimes active against coliforms such as E. coli and Klebsiella spp. All Enterococcus spp. are inherently resistant to cephalosporins. Cefuroxime is a second-­generation cephalosporin with increased activity against coliforms. The third-­ generation cephalosporins ceftiofur and cefoperazone and fourth-­generation cephalosporin

cefquinome have further increased activity against coliforms. Dihydrostreptomycin, streptomycin, neomycin, kanamycin, and framycetin are aminoglycosides available in combination IMM formulations in the EU and other countries. These antimicrobials have activity against aerobic, Gram-­negative bacteria and against Staphylococcus spp. Pirlimycin, a lincosamide, was the only drug available as an IMM preparation in the US and Canada that was not a member of the beta-­ lactam drug class. Lincosamide is available in IMM formulations in some countries. Lincosamides have a primarily Gram-­positive antimicrobial spectrum and are not active against coliform IMI pathogens. A recent metaanalysis evaluated 30 clinical trials of IMI treatment in cows using various drugs against various pathogens, comparing outcomes based on classifications of drug importance in human medicine (third-­ and fourth-­generation cephalosporins, highest priority; penicillins, high priority; and first-­ and second-­generation cephalosporins, highly important) (Nobrega et  al.,  2020). For nonsevere IMI, treatment of infection due to E. coli showed no benefit over nontreatment, and among the other most common pathogens, using an WHO critically important antimicrobial (CIA) (e.g., ceftiofur, cefquinome) did not improve outcomes compared to using noncritically important antimicrobials. In the interests of antimicrobial stewardship, these findings should be considered when selecting an antimicrobial for the treatment of IMI. When possible, preference should be given to the use of antimicrobials of least importance to human medicine. The drugs currently available in the US and Canada as IMM preparations are time-­ dependent inhibitors of bacterial growth. From a pharmacodynamic standpoint, efficacy is maximized by keeping the concentration of drug at the site of infection above the level ­necessary to inhibit microbial growth (minimum inhibitory concentration, MIC) as long as

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Antimicrobial Therapy in Dairy Cattle

possible between doses of the drug. Once the MIC of the drug is achieved at the site of infection, increased drug concentrations above the MIC are unlikely to improve efficacy. Consequently, for an IMI that is unresponsive to the label dosing regimen, extending the duration of therapy will be more effective than giving a higher dose at each treatment time without extending the duration of therapy. By contrast, some of the antimicrobials available in the EU for IMM administration to lactating cows are aminoglycosides, a drug class whose efficacy is maximized by administering high doses intermittently, with the drug concentration allowed to drop below the MIC between doses. Regardless of the approach, it is critical that ELDU of any antimicrobial in dairy cattle be in accordance with regulations governing such use and accompanied by extended milk and meat withdrawal periods (see Chapter 26).

Systemic Antimicrobials Used to Treat Intramammary Infections For acute mild-­to-­moderate IMI, systemic antimicrobial therapy is not indicated. For severe cases that involve systemic clinical signs (such as fever or depression) in addition to abnormal milk and udder swelling, systemic administration of antimicrobials is appropriate. Supportive care by administration of fluids and antiinflammatory drugs is also critical in such cases, and is discussed elsewhere (Suojala et al., 2013). Although systemic illness due to IMI is frequently caused by Gram-­negative pathogens, it may also be caused by Gram-­positive pathogens such as Streptococcus uberis and S. aureus. Because microbial culture generally takes 24 hours to yield a preliminary result, empirical systemic therapy of severe IMI must initially be broad spectrum. Because administration of IMM antimicrobials is not beneficial when the IMI is caused by Gram-­negative pathogens, a rational approach to therapy of severe acute IMI would be to treat suspected bacteremia by  using a systemic antimicrobial with a

spectrum of activity including Gram-­negative pathogens (which have been isolated from the blood of cows with severe IMI), combined with an IMM preparation that is active against Gram-­positive pathogens. In the US and many other countries, any systemic use of an antimicrobial as a therapy for IMI is an ELDU. The Animal Medicinal Drug Use Clarification Act specifically prohibits ELDU of sulfonamides in lactating dairy cows, prohibits any ELDU of fluoroquinolones and only permits EDLU of cephalosporins according to label dosing regimens. In Canada, short-­acting oxytetracycline ­formulations are approved for IMI while ­trimethoprim/sulfadoxine is only approved for treatment of septicemia but carries milk withdrawal times for lactating cows. In some countries, penethamate, cefquinome, lincomycin, danofloxacin, enrofloxacin, marbofloxacin, and gentamicin are approved for systemic treatment of IMI. As third-­ and fourth-­generation cephalosporins, aminoglycosides, and fluoroquinolones are considered CIA, their systemic use in dairy cattle should be based on evidence of efficacy. For IMM treatments, preference should be given to the use of antimicrobials of least importance to human medicine. When used in combination with IMM antimicrobials, antiinflammatory drugs, and other supportive therapy, the addition of intramuscular ceftiofur to the treatment regimen for severe acute IMI decreased the likelihood of a cow subsequently dying or being culled (Erskine et  al.,  2002). There is less evidence of benefit from systemic administration of a fluoroquinolone (Persson et  al.,  2015). Other drugs approved for bovine respiratory disease (BRD), such as florfenicol, tulathromycin, and tilmicosin, are poor choices for Gram-­negative bacteremia associated with severe IMI and require extremely long milk withdrawal periods. Although the Gram-­negative respiratory pathogens Mannheimia and Pasteurella may be susceptible to these drugs, the Gram-­negative coliform organisms that commonly cause IMI

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are either resistant to these drugs or the drug concentration required to inhibit their growth would not be reached in the udder at any reasonable dosage.

Antimicrobial Therapy of Dry Cows The dry (nonlactating) period of the lactation cycle is a critical time for dairy cattle. The periparturient period and the early dry period constitute the times of greatest risk for new IMI in the lactation cycle of the cow. Once involution of the mammary gland is complete, 10–­14 days after the start of the dry period, the immune environment for bacterial pathogens is more hostile. The most important defense against IMI in dry cows, as with lactating cows, is the teat canal. This barrier is enhanced during the dry period by the formation of a keratin plug. Additionally, during the dry period the mammary gland contains increased numbers of macrophages and lymphocytes and higher concentrations of complement and immunoglobulins that can help orchestrate more efficient phagocytosis. Lactoferrin, a potent iron-­chelating protein, also markedly increases in dry cow secretions, helping to inhibit growth of Gram-­negative bacteria, particularly E. coli. Consequently, the dry period is an ideal time to attain synergy between antimicrobial therapy and immune function to eliminate pathogens from the gland, without incurring the extensive milk withdrawal costs typical of lactating cow therapy. Intramammary administration of antimicrobials at the end of lactation has been a standard practice in dairy mastitis management for over 35 years. Because of concern regarding increased antimicrobial resistance, selective dry cow therapy (treatment of infected cows only) versus total or blanket dry cow therapy (treatment of all cows) is now recommended. Reviews and meta-analyses of relevant research indicate that if all cows –­drug treated or not –­ are treated with an internal teat sealant at dry-­ off, outcomes with selective dry cow therapy are not different from blanket therapy in the

rate of bacterial cure, prevention of new infections in the dry period, milk SCC, milk production, and rate of new IMI in early lactation (Kabera et al., 2021). Among the studies evaluated, a variety of inclusion criteria for drug treatment were used, including high SCC and/ or clinical IMI or microbial growth on milk culture. The studies used either ceftiofur hydrochloride or penicillin G procaine/novobiocin in cows selected for drug treatment. This evidence supports both the benefit of drug treatment for cows that have evidence of IMI and the viability of using only a teat sealant at dry-­off in qualified cows. Across the studies that implemented selective dry cow therapy, antimicrobial use was reduced by 66% compared to blanket dry cow therapy. Under the Veterinary Medicines Regulation (Regulation (EU) 2019/6), blanket dry cow treatment is no longer acceptable in EU member states. A variety of antimicrobials are marketed in dry cow products. Which drug is used does not seem to be as important as the use of any dry cow treatment for cows with evidence of IMI at dry-­off. A meta-analysis published in 2009 reviewed studies of IMM dry cow treatment from North America, Europe, New Zealand, South Africa, and Israel, and found that there were no studies demonstrating differences in efficacy among different IMM antimicrobials for resolving IMI during the dry period (Halasa et al., 2009).

M ­ etritis Acute puerperal metritis occurs within the first three weeks after calving and may include systemic signs of illness such as fever or depression in addition to abnormal vaginal discharge. E. coli and Trueperella pyogenes are especially important pathogens in bovine puerperal metritis. When cows have systemic signs of illness, treatment of metritis is indicated. Although metritis research sometimes reports cure rates, the definition of cure is not consistent among studies. Unlike research into

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­Metriti 

Antimicrobial Therapy in Dairy Cattle

the treatment of mastitis, microbial culture and the evaluation of “bacterial cure” are rare in metritis research. Despite the relatively frequent incidence of puerperal metritis in dairy cows, the scope of randomized controlled clinical trials of treatments for metritis is limited. A review reported that of 17 studies evaluating the use of systemic ceftiofur for the treatment of acute puerperal metritis, seven found that ceftiofur improved clinical parameters, but none reported improved reproductive performance (Haimerl and Heuwieser,  2014). Systemic treatment with ampicillin has been shown to be as effective as ceftiofur for the improvement of clinical signs in cows with puerperal metritis (Lima et al., 2014). However, it is worth considering that by one estimate, over half of untreated cases may resolve within 14  days (Sannmann et al., 2013). After 21 days post calving, uterine inflammation with or without vaginal discharge is called endometritis. Postpartum uterine abnormalities such as retained fetal membranes and endometritis have been treated using intrauterine infusion of tetracycline class drugs, but this is no longer recommended. Intrauterine infusion of oxytetracycline to treat endometritis produces drug residues in the milk for an average of five days, up to a maximum observed duration of eight days (Tan et al., 2007). It has been reported that four treatments of intrauterine chlortetracycline over a period of two weeks prevented milk loss and decreased infertility in dairy cows with acute puerperal metritis (Goshen and Shpigel, 2006). Such a protocol, four intrauterine treatments with chlortetracycline over two weeks, would produce a long duration of milk residues after treatment and is not an approved use of chlortetracycline in most countries. Treatment of retained fetal membranes with intrauterine oxytetracycline and fenprostaline increased the risk of pyometra and did not decrease the interval from calving to conception; that interval was also unaffected by the use of intrauterine oxytetracycline treatment without fenprostaline (Stevens et al., 1995).

For cows with clinical endometritis, treatment prior to 26 days in milk was not found to be beneficial (Leblanc et  al.,  2002). Cows treated for endometritis with intrauterine cephapirin between 27 and 33 days in milk were found to have a shorter time to pregnancy compared to untreated cows. The specificity of this limited time frame makes validation of this finding critically important, but no validating study has yet been published. There is no FDA-­approved cephalosporin product for intrauterine administration to cattle in the US, but a cephapirin product is approved for intrauterine use in Canada, the EU, and other countries. Subclinical endometritis occurs at least three weeks after parturition. Research into the treatment of subclinical endometritis has not provided consistent evidence of positive outcomes for any treatment.

­ ovine Respiratory Disease B in Dairy Cattle The bovine respiratory disease (BRD) complex is a common and serious health problem in the dairy industry, particularly in calf-­rearing facilities; hutch calves are most affected. BRD and diarrheal diseases are the most common diseases reported in dairy calves, and they are the most common reasons antimicrobials are used in hutched dairy calves and in calves that have recently been weaned off milk and moved to group pens (National Animal Health Monitoring System,  2018). The BRD complex also occurs in older calves and adult dairy cattle, particularly around times of stress such as during the transition period, shipping, or extreme weather events. This section is intended to present additional considerations specifically for the management of BRD in dairy cattle. The BRD complex is described generally in Chapter 29. For more information about the host, pathogen, and environmental factors associated with BRD in dairy cattle, the  reader is referred to the book chapter,

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“Preventing bacterial disease in dairy cattle” (Aly and Depenbrock, 2021). When using antimicrobials for BRD treatment in dairy calves, early diagnosis and treatment is associated with decreased progression of disease and improved outcomes (Binversie et  al.,  2020). Using antimicrobials to prevent disease or treat disease that could have been prevented is not good antimicrobial stewardship and may promote increased antimicrobial resistance (AMR). The BRD complex can have long-­term effects on dairy animal health and productivity, even after antimicrobial treatment (Adams and Buczinski,  2016; Schaffer et al., 2013). Because BRD is frequently subclinical in dairy calves (Binversie et  al.,  2020), defining protocols for BRD identification and diagnostic parameters for antimicrobial treatment is important to herd health management and antimicrobial stewardship planning. Clinical disease can be diagnosed on physical exam or using a scoring system validated for either preweaned or weaned dairy calves (Maier et al., 2019, 2020). However, physical examination and scoring systems lack perfect sensitivity and specificity; the physical exam often fails to identify subclinical cases and some non-­ BRD diseases such as heart disease or acid–­ base disturbances may mimic BRD signs. A literature review of clinical signs of BRD could not identify a clinically repeatable and validated definition for the clinical signs of BRD (Ferraro et al., 2021). Thoracic imaging such as ultrasound can provide evidence of lung consolidation. Thoracic ultrasound for BRD diagnosis is gaining popularity among livestock practitioners due to the relative portability of equipment and the ability to use a linear rectal probe which many livestock veterinarians already carry for reproductive work. More information on the use of thoracic ultrasound to diagnose, characterize, and monitor BRD in dairy cattle can be found in a review article (Ollivett and Buczinski,  2016). Airway sampling for cytological evaluation and bacterial culture and susceptibility can be used to

determine bacterial species and antimicrobial susceptibility associated with individual cases or groups of animals.

Selection of Antimicrobials for Treatment of BRD in Dairy Cattle The BRD complex in young, typically hutch-­ housed and milk-­fed dairy calves is often called “enzootic calf pneumonia” and is most frequently associated with P. multocida. The BRD complex that follows stressful events such as shipping is often called “shipping fever” and is most commonly associated with M. ­haemolytica. Although this form of the BRD complex is classically associated with feedlot cattle, M. ­haemolytica can also be involved in pneumonia of dairy cattle. Investigations into the microbiome of the upper and lower respiratory tracts of cattle have demonstrated the common presence of the respiratory Pasteurellaceae and Mycoplasma spp. classically associated with the BRD complex, as well as many other bacterial species. The composition of the microbiome changes with disease status as well as with many other host and environmental factors. Despite the complex nature of the bovine respiratory microbiome, antimicrobial therapy is still typically narrowly marketed for treatment of M. haemolytica, P. multocida, H. somni, and sometimes M. bovis infections. These bacteria can be found in the upper respiratory tract of both healthy and diseased cattle and are common invaders of the respiratory tissues in disease. Mycoplasma spp. are in a different class of bacteria from the Pasteurellaceae, characterized by lack of a cell wall. The following classes of antimicrobials contain drugs that are approved in various ­countries for treatment of BRD: macrolides, phenicols, fluoroquinolones, beta-­lactams (including amoxicillin/clavulanic acid), tetracyclines, aminoglycosides, trimethoprim/sulfonamides, and sulfonamides. Not all antimicrobials that are effective against the respiratory Pasteurellaceae are effective against Mycoplasma spp. The organism

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Antimicrobial Therapy in Dairy Cattle

M. bovis is a serious infectious disease concern in dairy cattle and has been commonly associated with otitis, arthritis, and mastitis in addition to its role in BRD. Spread of M. bovis from lactating animals through milk or colostrum to calves is a major mechanism by which this organism is maintained in a herd, and it can cause serious disease in calves. The antimicrobials that inhibit cell wall synthesis, such as beta-­lactams, are inherently ineffective against Mycoplasma spp. because these bacteria have no cell wall around their cell membranes. There are significant herd health risks of M. bovis shedding in a dairy facility and treatment of an affected animal is unlikely to permanently stop shedding of this pathogen. Thus, culling is preferable to antimicrobial treatment of dairy cattle with evidence of pneumonia or other diseases associated with M. bovis. The antimicrobials that are more likely to be effective against Mycoplasma spp. include several drugs labeled for use in BRD in the ­macrolide (e.g., tilmicosin, gamithromycin, ­tildipirosin, and tulathromycin), phenicol (e.g., florfenicol), fluoroquinolone (e.g., marbofloxacin, enrofloxacin and danofloxacin) or tetracycline classes. These drug classes have relatively high lipid solubility and act on intracellular structures. High lipid solubility is therapeutically desirable when treating BRD, but problematic in lactating dairy cattle because of prolonged milk residues or legal use restrictions. In the US, all fluoroquinolones are explicitly prohibited from use in female dairy cattle over 20 months of age. They are not prohibited for ELDU in Canada and there are approved products with milk withdrawal times in many other countries. The long-­acting macrolides and florfenicol have specific label restrictions against use in adult female dairy cattle. The ELDU of macrolides or florfenicol in adult female dairy cattle represents a serious risk to food safety due to prolonged and uncertain residues in milk. Chronic or abscessed cases of BRD may contain bacteria such as Trueperella pyogenes.

Severe chronic BRD and cases of BRD with significant abscessation are unlikely to resolve regardless of the drug selected. Even highly lipid-­soluble drugs may not reach therapeutic concentrations in large abscesses or severely compromised lung tissue; additionally, the contents of abscesses contain biological factors that inhibit antimicrobial drug function. Such cases are challenging to treat and may warrant euthanasia or advanced ­veterinary hospital care if economically warranted. Medicated feeds are used in dairy calf rearing but are ineffective in treatment of a BRD outbreak. Oral antimicrobial administration is more closely associated with enteric bacteria AMR than injectable antimicrobial administration (Checkley et  al.,  2010; Zhang et  al.,  2013). Clinically affected calves should be treated individually with injectable antimicrobials. Animals at risk of developing BRD should be managed to mitigate that risk by decreasing stressors, providing an appropriate plane of nutrition, and use of appropriate vaccination strategies for respiratory pathogens prior to known disease risk periods.

Antimicrobial Resistance in Bacteria Associated with BRD in Dairy Animals The existence of AMR in BRD isolates in dairy cattle is of growing concern. In a US study of 341 upper respiratory bacterial samples obtained from weaned dairy heifers, P. multocida and M. haemolytica isolates were not considered empirically susceptible to tetracycline, florfenicol, gamithromycin, tildipirosin, tilmicosin, danofloxacin, and enrofloxacin (Depenbrock et al., 2021). Empirical susceptibility was based on the general recommendation that greater than 80% of samples in a population should be susceptible. A 2021 Canadian study of 2834 deep nasopharyngeal swabs from cattle at arrival to a feed yard found that arrivals from dairy sources (mostly weaned calves) contained greater AMR in respiratory isolates, as well as more frequent

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multidrug resistance (>3 antimicrobial classes), than arrivals from beef sources. This study reported the most frequent resistance classifications to oxytetracycline, tulathromycin, and tilmicosin; each was classified as resistant in >20% of isolates (Andrés-­Lasheras et al., 2021). These findings raise concern for the frequent lack of susceptibility in respiratory isolates to antimicrobials commonly used to treat BRD in weaned dairy calves. Studies including respiratory isolates from preweaned dairy calves also report AMR to antimicrobials commonly used in the treatment of BRD (Maynou et al., 2017; Rérat et  al.,  2012). AMR appears to be more commonly reported in weaned heifers than hutched heifers, with the exception of veal calf rearing. This may be due to the relative high exposure of calves to antimicrobials, and more opportunities or time to collect AMR genes in weaned pens compared to younger hutched heifers. Multidrug resistance that includes resistance to tetracycline is common in respiratory pathogens of cattle, including dairy calves (Depenbrock et  al., 2021; Andrés-­Lasheras et  al.,  2021) , and the use of tetracycline in dairy calves may co-­select for AMR to other classes of antimicrobials needed to treat BRD (Holman et al., 2019). Although further investigation into the effect of drug use, resulting AMR patterns, and treatment outcomes in dairy calves is needed, it is prudent to avoid the use of antimicrobials to which there is frequent AMR or multidrug resistance in a given group, facility or region. Respiratory disease prevention in dairy calves is thus imperative to limit reliance on antimicrobials that may have poor efficacy in some populations of dairy cattle, particularly weaned calves. It is recommended to prevent BRD when calves are entering weaned pens, and ensure that calves are not being weaned with subclinical pneumonia that may become clinical with the stress of weaning and transition to group pens. For more information on the prevention of BRD in dairy operations, the reader

is referred to the cited reviews on the topic (Aly and Depenbrock,  2021; Lehenbauer,  2014; Lombard et al., 2020).

D ­ iarrhea Neonatal calf diarrhea is one of the leading causes of death in dairy calf-­rearing operations. The primary focus of this section is antimicrobial use relevant to calf rearing. The continual use of antimicrobials as mass medication for the prevention of diarrhea in neonatal or adult dairy animals is increasingly discouraged, restricted, or illegal in many countries. For information regarding diarrhea in adult cattle, the reader is referred to Holschbach and Peek (2018). There are many different causes of diarrhea in dairy calves that ultimately result in fecal water loss and varying degrees of dehydration, bacterial translocation, systemic illness, or death. Nutritional causes of neonatal calf diarrhea are common in bottle-­fed or bucket-­fed calves and may frequently be the underlying cause of indigestion and dysbiosis associated with diarrhea, regardless of the concurrent presence of infectious agents. The most common infectious agents associated with calf diarrhea at less than 21  days of age are ­bacteria  including E. coli (enterotoxogenic) and Salmonella spp., and viruses including ­rotavirus, coronavirus, and the protozoa Cryptosporidium parvum. Other agents associated with calf diarrhea include Clostridium perfringens, Giardia, bovine viral diarrhea (BVD) virus, other more recently reported viruses such as bovine torovirus and caliciviruses, and Eimeria spp. in older calves. Toxin ingestion can result in diarrhea in both calves and adult dairy cattle. Other systemic diseases may result in diarrhea in cattle of any age, such as heart disease, liver disease, peritonitis, endotoxemia or some rare congenital defects. Many causes of diarrhea in cattle are not caused by a bacterial infection and therefore will not be improved by antimicrobial therapy.

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­Diarrhe 

Antimicrobial Therapy in Dairy Cattle

Obtaining a diagnosis for the underlying causes of diarrhea on a dairy facility is important for planning treatment as well as disease prevention. Underlying management problems that allow for the enteric proliferation of undesirable microbial populations, such as feeding errors that result in indigestion or dysbiosis, sanitation errors that increase the spread of infectious agents, improper colostrum or nutritional management that results in decreased immunity, environmental stressors, toxin ingestion, or other animal management factors that may impact gastrointestinal health, should also be investigated. For further information on how to diagnose neonatal and adult cattle diarrhea, the reader is referred to reviews on the topic (Chigerwe and Heller, 2018; Holschbach and Peek, 2018). In general, interventions should be directed at prevention of diarrhea through modifying management practices, focusing on maximizing animal health and disease resistance, and minimizing feeding errors and stressors. When preventive methods have failed, sick calves with diarrhea are commonly treated with antimicrobials in addition to nonantimicrobial treatments including fluid therapy, antiinflammatory drugs, and enteral nutrition. To maintain judicious use of antimicrobials while protecting animal health, the primary goal of antimicrobial therapy should target bacteremia caused by translocation of enteric bacteria. Calves that are not otherwise systemically ill should be managed with supportive therapy that does not include antimicrobials. For more information on treatment of calves with diarrhea who are otherwise apparently healthy, the reader is referred to reviews for information on fluid therapy and supportive care of calves with diarrhea (Berchtold, 2009). Antimicrobial therapy should be reserved for calves that show signs of bacteremia. Targeted therapy for bacteremia decreased the total days of diarrhea in calves, and decreased the overall cost of treatment (Berge et  al., 2009). Conversely, some investigations have ­associated the use of therapeutic doses of

antimicrobials in calves with increased diarrhea incidence. The oral administration of neomycin sulfate, ampicillin trihydrate, and tetracycline hydrochloride was associated with evidence of malabsorption and diarrhea (Mero et al., 1985). Few antimicrobials are approved for treatment of diarrhea in dairy calves; in most countries, they include parenteral formulations of trimethoprim/sulfonamides and oxytetracycline and as feed or water additives containing chlortetracycline, tetracycline, oxytetracycline, sulfamethazine, and neomycin. In some countries, cefquinome is approved for E. coli septicemia in calves. The antimicrobials or formulations labeled for treatment of diarrhea in calves may not be effective for treating bacteremic calves and thus ELDU of other antimicrobials may be warranted and use regulations must be followed. When bacteremia is suspected or known, therapy should consist of antimicrobials with a spectrum of activity covering Gram-­negative enteric bacteria. The most common bacteria identified in the blood of scouring calves is E. coli. Parenteral administration, ideally coupled with supportive fluid therapy, is preferred over oral antimicrobial administration. Diarrheic calves are often dehydrated and/or hypovolemic, and drug absorption may be impaired from nonintravenous routes. For dehydrated and/or hypovolemic calves, fluid therapy should be instituted regardless of antimicrobial choice. While approved in many countries for the treatment of diarrhea in calves, oxytetracycline is not an ideal choice for the treatment of bacteremia, due to widespread coliform resistance to tetracyclines identified in samples from dairy cattle, especially dairy calves (Sawant et  al.,  2007). Oxytetracycline may cause nephrotoxicity, with a higher risk in dehydrated/ hypovolemic animals. In many countries, injectable trimethoprim/sulfadoxine formulations are approved for treatment of enteritis. Susceptibility of coliforms to sulfonamides has  been reported as poor in some studies

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(Smith,  2015), and sulfonamides may cause nephrotoxicity in dehydrated animals. Other antimicrobials that are approved for use in calves, but not specifically for the treatment of diarrhea, may have empirical efficacy against Gram-­negative enteric bacteria. They include ampicillin, amoxicillin/clavulanic acid, sulfadimethoxine, cephalexin, and ceftiofur. There are bacteriological and pharmacokinetic advantages to the use of ceftiofur or cefquinome for the treatment of bacteremic calves, including the drugs’ resistance to beta-­ lactamase and pharmacokinetic studies in calves demonstrating therapeutic concentrations against E. coli in the blood and intestine. However, third-­ and fourth-­generation cephalosporins are CIA and should not be used as first-­line treatments in calves due to use linked to AMR in zoonotic enteric pathogens (Awosile et al., 2018; Foster et al., 2019). Classes of antimicrobials that should be avoided when treating dairy calves with diarrhea include the fluoroquinolones and parenteral aminoglycosides. The fluoroquinolones are restricted from ELDU in some countries as they are CIA, and there are concerns that use in livestock perpetuates AMR, particularly in enteric zoonotic pathogens shed from calves (McEwen and Fedorka-­Cray, 2002). Aminoglycosides have empirical efficacy against Gram-­negative enteric bacteria, but there is a voluntary ban on the use of parenteral aminoglycosides in food animals in the US due to prolonged tissue residues. The use of the oral formulations of neomycin for the control of calf diarrhea is common on some calf-­ rearing operations. This practice assumes minimal absorption of this relatively polar drug from the GI tract. Because neomycin is poorly absorbed, the antimicrobial effects are primarily within the GI lumen, and thus it is not an effective treatment for bacteremia. There is widespread resistance to neomycin in Gram-­negative enteric bacteria. Additionally, if the barrier between the GI tract and the blood is disrupted due to pathology, unpredictable quantities of neomycin may be absorbed and cause violative tissue residues. Additionally, aminoglycosides

can cause nephrotoxicity, with an increased risk in dehydrated, hypovolemic diarrheic calves. Some causes of diarrhea associated with specific enteric pathogens require specific antimicrobial considerations, beyond treatment of bacteremia or intestinal coliform overgrowth. Examples include clostridial enteric diseases and the protozoal diseases coccidiosis and cryptosporidiosis. Clostridial enteritis or abomasitis in dairy calves is associated with abnormal nursing or feeding patterns, particularly intermittent feeding of large volumes of colostrum or milk replacer or feeding of milk replacers high in carbohydrates or protein. The oral ELDU of penicillin procaine G may be desirable when the cause of diarrhea is associated with enteric Clostridium spp., due to the drug’s efficacy against clostridial organisms and need for rapid efficacy at the enteric luminal site of clostridial toxin production (Simpson et al., 2018). An ELDU individual animal oral dose of 22,000 IU/kg orally every 24 hours for 3–­5  days has been suggested; for more information on prevention and treatment of clostridial abomasitis and enteritis, the reader is referred to a review on the topic (Simpson et al., 2018). A common protozoal cause of diarrhea in older calves is coccidiosis from pathogenic species of Eimeria. Calves are usually at least one month of age when they become clinically affected. Fecal shedding relative to exposure varies by Eimeria species and ranges from about six to 23  days (Bangoura and Bardsley,  2020). Calves typically acquire age-­ related immunity by one year and rarely show clinical signs beyond this time. Coccidiosis can present in a range of clinical signs from poor weight gain to diarrhea, to inflammatory diarrhea with variable amounts of blood, fibrin, or sloughed mucosa in the feces. Nonpharmaceutical management interventions, primarily centered on manure management and hygiene, are the preferred methods for control of Eimeria spp. The ionophore class of antimicrobials is not considered medically important and is often used as feed

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­Diarrhe 

Antimicrobial Therapy in Dairy Cattle

additives for the control of coccidiosis in dairy calves. Antimicrobial therapy of severely affected animals may include the oral administration of sulfonamides labeled for the treatment of coccidiosis, including sulfamethazine and sulfaquinoxaline. Nonantibiotic drugs that are also labeled for the treatment of coccidiosis in cattle in some countries include amprolium and toltrazuril. However, by the time oocysts are detected in the feces, the intestinal portion of this protozoal parasite’s life cycle has been completed and the intestinal damage has been done. Additional supportive therapy is necessary for severely affected animals and may include fluid therapy, plus systemic antimicrobials in some very severe cases where bacteremia is known or strongly suspected. For more information on coccidiosis in ruminants, the reader is referred to a review on the topic (Bangoura and Bardsley, 2020). Cryptosporidiosis, caused by the protozoa Cryptosporidium parvum, has been reportedly treated with the use of oral antimicrobials in calves. However, there are no antimicrobials considered effective or approved for this purpose in the US. Control of C. parvum in US dairy calves is focused on hygiene and improvement of calf health and resistance to infection. Treatment consists of supportive therapy, including fluid and nutritional support and decreasing stressors. The drug halofuginone is approved as an aid in reducing clinical signs of cryptosporidiosis in Canada and other countries. For severely affected calves showing signs of bacteremia, antimicrobial treatment directed against coliform bacteria may be beneficial as part of supportive therapy. For more information on cryptosporidiosis in cattle, the reader is referred to a review on the topic (Adkins, 2022).

Alternatives to Antimicrobial Use in the Management of Calf Diarrhea Many preventive methods, nonantimicrobial treatments, and supportive care methods have

been described for the management of diarrhea in dairy calves. Preventive methods are more likely to positively impact animal health than novel treatments applied after prevention has failed. Excellent colostrum management, provision of a high plane of nutrition, avoidance of feed errors or inconsistencies, appropriate housing hygiene, and minimization of environmental stressors are all crucially important for maintenance of healthy calves, particularly for prevention of diarrhea. For more information on calf management relative to the prevention of diarrhea, the reader is referred to reviews on this topic (Barrington et  al.,  2002; Lombard et al., 2020; McGuirk, 2008).

Antimicrobial Resistance in Enteric Pathogens The use of antimicrobials in neonatal calves has been shown to be a key driver of AMR on dairy operations (Catry et al., 2016; Formenti et al., 2021; Gonggrijp et al., 2016), and AMR in zoonotic pathogens has been linked to food animal production (Pérez-­Rodríguez and Mercanoglu Taban, 2019). The use of antimicrobials for disease treatment and prevention, and exposure of calves to subtherapeutic doses of antimicrobials through waste milk have all been associated with AMR on dairies (Duse et  al.,  2015; Firth et  al.,  2021; Formenti et al., 2021). It is thus necessary to maintain antimicrobial stewardship in prescribing practices for dairy cattle. For more information on making an antibiotic stewardship plan for dairy cattle, the reader is referred to guidelines published by the American Association of Bovine Practitioners (AABP, 2017). Monitoring AMR in enteric pathogens of dairy cattle can be challenging. Records of culture and susceptibility testing should be kept and used to tailor antimicrobial use according to recent susceptibility trends, when needed for severely affected animals. Producers are encouraged to work with their attending ­veterinarian and diagnostic laboratory to

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determine which bacterial isolates or strains are actually associated with disease or translocation into the blood to track AMR in the pathogens for which antimicrobial therapy is targeted. A lack of species-­ and disease-­appropriate susceptibility breakpoints presents a challenge of monitoring AMR in enteric bacteria of dairy cattle. Most breakpoints for enteric bacteria that are reported for cattle are actually intended  for use in humans with zoonotic enteric pathogens, and have not been developed specifically for cattle. Although E. coli has a breakpoint specifically for cattle, that breakpoint was designed for metritis (uterine infection), not diarrhea or bacterial translocation from the gastrointestinal tract (CLSI, 2018). This means that when an enteric isolate from cattle is deemed susceptible using breakpoints

extrapolated from other species or clinical sites, the likelihood that the isolate is actually going to be cured at the site of infection (presumably the bloodstream and intestinal wall) is uncertain; the pharmacokinetics of the drug used to determine breakpoints in susceptibility testing are likely not the same as in humans or other body sites of cattle. Due to the demonstrated contribution to AMR, particularly in zoonotic pathogens, as well as potential negative animal health impacts, the use of antimicrobials in neonatal calves should be limited. When antimicrobials are necessary to protect animal health, treatment regimen considerations should include clear disease diagnostic criteria to warrant antimicrobial use and discrete antimicrobial treatment protocols specifying dose, route, frequency, and duration of therapy.

­References and Bibliography Adams EA, Buczinski S. 2016. Short communication: Ultrasonographic assessment of lung consolidation postweaning and survival to the first lactation in dairy heifers. J Dairy Sci 99:1465. Adkins PR. 2022. Cryptosporidiosis. Vet Clin North Am Food Anim Pract 38:121. Aly SS, Depenbrock SM. 2021. Preventing bacterial disease in dairy cattle. In: Bouchard E (ed.) Improving Dairy Herd Health. Burleigh Dodds, London. American Association of Bovine Practitioners (AABP). 2017. Key elements for implementing antimicrobial stewardship plans in bovine veterinary practices working with beef and dairy operations. www.aabp.org/resources/ AABP_Guidelines/ antimicrobialstewardship-­7.27.17.pdf Andrés-­Lasheras S, et al. 2021. Prevalence and risk factors associated with antimicrobial resistance in bacteria related to bovine respiratory disease –­a broad cross-­sectional study of beef cattle at entry into Canadian feedlots. Front Vet Sci 8 :692646.

Awosile B, et al. 2018. Salmonella enterica and extended-­spectrum cephalosporin-­resistant Escherichia coli recovered from Holstein dairy calves from 8 farms in New Brunswick, Canada. J Dairy Sci 101:3271. Bangoura B, Bardsley KD. 2020. Ruminant coccidiosis. Vet Clin North Am Food Anim Pract 36:187. Barrington GM, et al. 2002. Biosecurity for neonatal gastrointestinal diseases. Vet Clin North Am Food Anim Pract 18:7. Berchtold J. 2009. Treatment of calf diarrhea: intravenous fluid therapy. Vet Clin North Am Food Anim Pract 25:73. Berge ACB, et al. 2009. Targeting therapy to minimize antimicrobial use in preweaned calves: effects on health, growth, and treatment costs. J Dairy Sci 92:4707. Binversie ES, et al. 2020. Randomized clinical trial to assess the effect of antibiotic therapy on health and growth of preweaned dairy calves diagnosed with respiratory disease using respiratory scoring and lung ultrasound. J Dairy Sci 103:11723.

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 ­References and Bibliograph

Antimicrobial Therapy in Dairy Cattle

Catry B, et al. 2016. Effect of antimicrobial consumption and production type on antibacterial resistance in the bovine respiratory and digestive tract. PLoS One 11:e0146488. Checkley SL, et al. 2010. Associations between antimicrobial use and the prevalence of antimicrobial resistance in fecal Escherichia coli from feedlot cattle in western Canada. Can Vet J 51:853. Chigerwe M, Heller MC. 2018. Diagnosis and treatment of infectious enteritis in adult ruminants. Vet Clin North Am Food Anim Pract 34:119. Clinical Laboratory Standards Institute (CLSI). 2018. Performance standards for antimicrobial disk and dilution susceptibility tests for bacteria isolated from animals. CLSI Supplement VET08, 4th ed. CLSI, Wayne, PA. Depenbrock S, et al. 2021. In-­vitro antibiotic resistance phenotypes of respiratory and enteric bacterial isolates from weaned dairy heifers in California. PLoS One 16 :e026 0292. Duse A, et al. 2015. Risk factors for antimicrobial resistance in fecal Escherichia coli from preweaned dairy calves. J Dairy Sci 98:500. Erskine RJ, et al. 2002. Efficacy of systemic ceftiofur as a therapy for severe clinical mastitis in dairy cattle. J Dairy Sci 85:2571. Ferraro S, et al. 2021. Scoping review on clinical definition of bovine respiratory disease complex and related clinical signs in dairy cows. J Dairy Sci 104:7095. Firth CLL, et al. 2021. The effects of feeding waste milk containing antimicrobial residues on dairy calf health. Pathogens 10:112. Formenti N, et al. 2021. Antimicrobial resistance of Escherichia coli in dairy calves: a 15-­year retrospective analysis and comparison of treated and untreated animals. Animals 11:2328. Foster DM, et al. 2019. Ceftiofur formulation differentially affects the intestinal drug concentration, resistance of fecal Escherichia coli, and the microbiome of steers. PLoS One 14:e0223378.

Gelgie AE, et al. 2022. Mycoplasma bovis mastitis. Curr Res Microb Sci 3:100123. Godden SM, et al. 2019. Colostrum management for dairy calves. Vet Clin North Am Food Anim Pract 35:535. Gonggrijp MA, et al. 2016. Prevalence and risk factors for extended-­spectrum β-­lactamase-­ and AmpC-­producing Escherichia coli in dairy farms. J Dairy Sci 99:9001–­9013. Goshen T, Shpigel NY. 2006. Evaluation of intrauterine antibiotic treatment of clinical metritis and retained fetal membranes in dairy cows. Theriogenology 66(9):2210. Haimerl P, Heuwieser W. 2014. Invited review: Antibiotic treatment of metritis in dairy cows: a systematic approach. J Dairy Sci 97(11):6649. Halasa T, et al. 2009. Meta-­analysis of dry cow management for dairy cattle. Part 1. Protection against new intramammary infections. J Dairy Sci 92(7):3134. Holman DB, et al. 2019. Antibiotic treatment in feedlot cattle: a longitudinal study of the effect of oxytetracycline and tulathromycin on the fecal and nasopharyngeal microbiota. Microbiome 7:86. Holschbach CL, Peek SF. 2018. Salmonella in dairy cattle. Vet Clin North Am Food Anim Pract 34(1):133. Kabera F, et al. 2021. Comparing blanket vs. selective dry cow treatment approaches for elimination and prevention of intramammary infections during the dry period: a systematic review and meta-­analysis. Front Vet Sci 8:688450. LeBlanc SJ, et al. 2002. The effect of treatment of clinical endometritis on reproductive performance in dairy cows. J Dairy Sci 85(9):2237. Lehenbauer TW. 2014. Control of BRD in large dairy calf populations. Anim Health Res Rev 15:184. Lima FS, et al. 2014. Efficacy of ampicillin trihydrate or ceftiofur hydrochloride for treatment of metritis and subsequent fertility in dairy cows. J Dairy Sci 97(9):5401. Lombard J, et al. 2020. Consensus recommendations on calf-­and herd-­level

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passive immunity in dairy calves in the United States. J Dairy Sci 103:7611. Maier GU, et al. 2019. Development of a clinical scoring system for bovine respiratory disease in weaned dairy calves. J Dairy Sci 102:7329. Maier GU, et al. 2020. A novel risk assessment tool for bovine respiratory disease in preweaned dairy calves. J Dairy Sci 103:9301. Maynou G, et al. 2017. Feeding of waste milk to Holstein calves affects antimicrobial resistance of Escherichia coli and Pasteurella multocida isolated from fecal and nasal swabs. J Dairy Sci 100:2682. McEwen SA, Fedorka-­Cray PJ. 2002 Antimicrobial use and resistance in animals. Clin Infect Dis 34(3):S93. McGuirk SM. 2008. Disease management of dairy calves and heifers. Vet Clin North Food Anim Pract 24(1):139. Mero KN, et al. 1985. Malabsorption due to selected oral antibiotics. Vet Clin North Food Anim Pract 1(3):581. National Animal Health Monitoring System. 2018. Dairy 2014 Health and Management Practices on U.S. Dairy Operations. www. aphis.usda.gov/animal_health/nahms/dairy/ downloads/dairy14/Dairy14_dr_PartIII.pdf Nobrega DB, et al. 2020. Critically important antimicrobials are generally not needed to treat nonsevere clinical mastitis in lactating dairy cows: results from a network meta-­ analysis. J Dairy Sci 103(11):10585. Ollivett TL, Buczinski S. 2016. On-­farm use of ultrasonography for bovine respiratory disease. Vet Clin North Am Food Anim Pract 32:19. Pérez-­Rodríguez F, Mercanoglu Taban B. 2019. A state-­of-­art review on multi-­drug resistant pathogens in foods of animal origin: risk factors and mitigation strategies. Review. Front Microbiol 10:2091. Persson Y, et al. 2015. Efficacy of enrofloxacin for the treatment of acute clinical mastitis caused by Escherichia coli in dairy cows. Vet Rec 176(26):673. Rérat M, et al. 2012. Bovine respiratory disease: efficacy of different prophylactic treatments in

veal calves and antimicrobial resistance of isolated Pasteurellaceae. Prev Vet Med 103:265. Sannmann I, et al. 2013.Comparison of two monitoring and treatment strategies for cows with acute puerperal metritis. Theriogenology 79:961. Sawant AA, et al. 2007. Antimicrobial-­resistant enteric bacteria from dairy cattle. Appl Environ Microbiol 73(1):156. Schaffer AP, et al. 2013. The effects of BRD in Holstein dairy calves during the first 120 days of life on subsequent production, longevity, and reproductive performance as cows. Proceedings of the American Association of Bovine Practitioners Annual Conference, p.143. Simpson K, et al. 2018. Clostridial abomasitis and enteritis in ruminants. Vet Clin North Food Anim Pract 34:155. Smith G. 2015. Antimicrobial decision making for enteric diseases of cattle. Vet Clin North Am Food Anim Pract 31:47. Stevens RD, et al. 1995. Evaluation of the use of intrauterine infusions of oxytetracycline, subcutaneous injections of fenprostalene, or a combination of both, for the treatment of retained fetal membranes in dairy cows. J Am Vet Med Assoc 207(12):1612. Suojala L, et al. 2013. Treatment for bovine Escherichia coli mastitis –­an evidence-­based approach. J Vet Pharmacol Therapeut 36(6):521. Tan X, et al. 2007. Persistence of oxytetracycline residues in milk after the intrauterine treatment of lactating cows for endometritis. Vet Rec 161(17):585. Tomazi T, et al. 2021. Negatively controlled, randomized clinical trial comparing different antimicrobial interventions for treatment of clinical mastitis caused by gram-­positive pathogens. J Dairy Sci 104(3):3364. Zhang, L, et al. 2013. Antibiotic administration routes significantly influence the levels of antibiotic resistance in gut microbiota. Antimicrob Agents Chemother 57:3659.

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 ­References and Bibliograph

31 Antimicrobial Therapy in Sheep and Goats Joe S. Smith and Amanda J. Kreuder

Despite their worldwide population and ­agricultural importance, sheep and goats are minor species within North America, and many veterinarians will have had limited exposure to them. In the United States, Canada, the European Union, and worldwide, there are few licensed veterinary pharmaceutical products available for sheep and goats. For example, in the United States there are nine antimicrobials labeled for sheep and only five labeled for goats (only one of which is parenteral). Most of these approved products are for feed-­or water-­based formulations. Furthermore, sheep and goats are classed as separate species by the regulatory authorities in most countries. This means that approved drugs are specifically licensed and withdrawal times and tolerances/maximum residue limits (MRLs) have been set for each species. This situation creates a great deal of confusion for veterinarians as well as sheep and goat producers. Journal articles, textbooks, and the internet provide accessible information from clinical trials and dose regimens for a wide variety of antimicrobials that are not licensed in small ruminants. In many cases, the safety and efficacy of the drug are well documented and if the antimicrobial product is commercially available, it may not be licensed for sheep or goats.

G ­ eneral Recommendations Despite their similarities as ruminant species, the pharmacokinetics of antimicrobial drugs in sheep and goats can vary when compared to cattle, and can also vary when compared from sheep to goats. With this in mind, sheep and goats are not simply “small cows”; they react differently to certain medications and suffer from different diseases. Prudent antimicrobial use first requires a tentative diagnosis ideally followed by confirmation of the etiological agent by microbiological culture and antimicrobial susceptibility testing before commencing therapy. However, collection of samples from sheep and goats is not always feasible and even if samples are obtained, results usually take at least 2–­3 days to process, and currently there are no species-­ specific breakpoints for sheep and goats. Thus, empirical therapy is common and should be determined by a thorough physical examination and a presumptive diagnosis, knowledge of the most common pathogens, the expected antimicrobial susceptibility of those organisms, and the pharmacokinetics/pharmacodynamics of the antimicrobial in the species being treated. Table 31.1 contains information to help make these decisions.

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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Condition

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Table 31.1  Antimicrobial drug selection for common conditions of sheep and goats. Species affected

Etiological agent(s)

Recommended treatment

Comments

Enzootic abortion of ewes (EAE)/ovine enzootic abortion (OEA)

Sheep and goats

Chlamydophila abortus

Tetracycline Oxytetracycline Tylosin

Prophylaxis in high-­risk flocks: tetracycline in feed for 6–­8 weeks prior to breeding at a dose of 200–­400 mg/head/ day until lambed. Outbreak: 400–­500 mg/head/day tetracycline in feed until lambing finished. Poor efficacy if placental damage already present. Not recommended for dairy goats because of milk withdrawal. Vaccination or biosecurity should be considered. Outbreak or previous diagnosis: long-­acting oxytetracycline at label dosage starting 6–­8 weeks before start of lambing every 10–­14 days until finished.

Campylobacter spp. abortion (vibrionic)

Sheep

C. jejuni C. fetus subsp. fetus

Penicillin G-­streptomycin; tetracycline Oxytetracycline (resistance commonly reported) Tylosin Sulfamethazine

Prophylaxis: injections of penicillin-­streptomycin for 2–­5 days. Antimicrobial susceptibility patterns should be established from any isolates. Vaccination in the face of an outbreak also very successful. A tetracycline-­resistant clone of C. jejuni exists in the United States

Listeria abortion

Sheep and goats

L. monocytogenes

Oxytetracycline

Injectable long-­acting tetracycline to all animals at risk in the face of an outbreak.

Toxoplasma abortion

Sheep and goats

T. gondii

Monensin Decoquinate

Mixed in feed at a dose of 15 mg/head/day from breeding to lambing. Mixed in feed or premix to feed at a dose of 2 mg/kg/day for last 14 weeks of gestation. Use of feed-­grade drugs for this purpose may be prohibited in some regions.

Salmonella abortion

Sheep and goats

S. typhimurium, S. abortus ovis, S. montevideo, S. dublin

IM or SC broad-­spectrum antimicrobials

Often widespread by the time diagnosis is made. Requires culture and susceptibility testing. Antimicrobials may not eliminate organism; consider culling and environmental management.

Infectious abortion

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Species affected

Etiological agent(s)

Recommended treatment

Comments

Leptospira abortion

Sheep and goats

L. hardjo, L. pomona

Penicillin G-­streptomycin; tetracyclines

Treat all pregnant animals at risk with injections.

Other infectious reproductive disorders Metritis

Sheep and goats

Trueperella pyogenes, E. coli, mixed anaerobes including Clostridium spp.

Penicillin G; ceftiofur; broad-­spectrum antimicrobials

Treat for 3–­4 days after clinically normal. Uterine evacuation with prostaglandins and tetanus vaccination should also be considered.

Lamb epididymitis

Sheep

H. somni, A. seminis, Corynebacterium pseudotuberculosis

Oxytetracycline

Prophylaxis: low levels in feed in situations where rams intensively managed, or injectable long-­acting oxytetracycline (IM or SC). Responds poorly to treatment.

Enzootic posthitis

Sheep and goats

C. renale group

Penicillin G; oxytetracycline

Remove from high-­protein diet and treat locally with antimicrobial ointments. May treat systemically for severe cases.

Brucella ovis ram epididymitis

Sheep

Brucella ovis

Oxytetracycline with dihydrostreptmycin

20 mg/kg oxytetracycline at 3-­day intervals for 5 treatments and 12.5 mg/kg dihydrostreptomycin 2×/day for 7 days decreases shedding of bacteria and improves semen quality but may not cure. Should consider culling.

Infectious diseases of lambs and kids, systemic Enterotoxemia/pulpy kidney

Sheep and goats

C. perfringens type C and D

Oral virginiamycin, penicillin G, or bacitracin

Vaccinate all animals at risk. Withdraw carbohydrate source in diet, give C&D antitoxin and a balanced electrolyte solution (BES) parenterally.

Omphalophlebitis

Sheep and goats

T. pyogenes, E. coli, mixed anaerobes

Penicillin G; broad-­ spectrum antimicrobials

Antimicrobial therapy alone not often effective. Local drainage, treatment with chlorhexidine or iodine, and possibly surgical removal should be considered.

Watery mouth (lambs)

Sheep

Probable E. coli endotoxin

Oral amoxicillin; apramycin

Prevention by ensuring clean environment and good colostrum ingestion. Early prophylactic treatment with oral antimicrobials.

Tick-­borne fever (tick pyemia)

Sheep

Anaplasma phagocytophilum and/or S. aureus

Long-­acting oxytetracycline

At 1–­3 weeks of age and repeated at 5–­7 weeks, in addition to dipping with an acaricide at those times.

Erysipelothrix polyarthritis

Sheep

E. rhusiopathiae

Penicillin G

Treat minimum of 3 days.

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Condition

(Continued )

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Condition

Species affected

Etiological agent(s)

Recommended treatment

Comments

Infectious diseases of lambs and kids, digestive

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Table 31.1  (Continued)

Colibacillosis

Sheep and goats

Enterotoxigenic E. coli

Broad-­spectrum antimicrobials parenterally

Appropriate diagnosis is necessary (culture and susceptibility testing). Clean environment and adequate colostrum are important. Consider vaccination. Resistance to antimicrobials is common.

Salmonella dysentery

Sheep and goats

S. typhimurium and others

Broad-­spectrum antimicrobials

Often poor efficacy due to unpredictable susceptibility patterns. May not eliminate carriers if host-­adapted species. Risk of resistance development.

Abomasitis/abomasal hemorrhage

Sheep and goats

Clostridium spp.

Oral and systemic penicillins

Rarely effective. Should treat symptomatically with antitoxins, nonsteroidal antiinflammatory drugs, and BES. Use polyvalent clostridial vaccine.

Coccidiosis

Sheep and goats

Eimeria spp.

Monensin; lasalocid; decoquinate; salinomycin; amprolium; toltrazuril or sulfonamides

Mixing should be done at a feed mill and all feeds pelleted. Some products can be mixed with salt. Dose varies with feed management. Artificially raised lambs/kids can be medicated via milk replacer. Feed from 2 weeks of age until market age. Ionophores toxic to horses and dogs. Additional regulations may be present in some countries with feed-­grade drugs.

Infectious conditions of lambs and kids, respiratory Pneumonic pasteurellosis

Sheep and goats

M. haemolytica, P. multocida

Tilmicosin; oxytetracycline; ceftiofur; florfenicol; tulathromycin

Long-­acting oxytetracycline, tilmicosin, or florfenicol can be used as prophylaxis and during an outbreak therapeutically. Tilmicosin should not be used in goats (therapeutic dose very close to toxic dose). Ceftiofur for daily treatment of affected animals when meat or milk withdrawal is an issue (e.g., market lambs close to slaughter, lactating dairy sheep). Tulathromycin can be used but will have longer meat or milk withdrawal recommendations.

Pasteurella septicemia

Sheep

Bibersteinia trehalosi

As with M. haemolytica

B. trehalosi will show artifactual resistance, due to no established breakpoints, and because the disease is peracute, vaccination is recommended for susceptible animals.

Necrotic laryngitis

Sheep and goats

Fusobacterium necrophorum

Penicillin G; oxytetracycline

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Species affected

Etiological agent(s)

Recommended treatment

Comments

Mycoplasma pneumonia

Sheep and goats

M. ovipneumoniae, M. arginini

Oxytetracycline; tylosin

Often seen in conjunction with pasteurellosis (atypical pneumonia) or alone.

Mycoplasma mycoides

Goats

M. mycoides subsp. mycoides large colony type

Oxytetracycline; lincomycin; tylosin

Treatment of peracute septicemia often ineffective. If goat survives, it will probably be a carrier.

Infectious conditions of the integument Infectious keratoconjunctivitis (Mycoplasma)

Sheep and goats

M. conjunctivae

Spiramycin; oxytetracycline; tylosin, chlortetracycline, streptomycin; tulathromycin; tiamulin

Spiramycin or oxytetracycline repeated days 1, 5, and 10; tiamulin repeated days 1, 3, 6, and 9. Oxytetracycline eye ointment, either alone or with polymyxin B. NSAIDs typically required to decrease inflammation.

Chlamydophila keratoconjunctivitis

Sheep and goats

C. abortus and C. percorum

Spiramycin; oxytetracycline; tylosin, chlortetracycline, streptomycin; tulathromycin; tiamulin

Parenteral antimicrobials should be considered for lambs, due to concerns of polyarthritis. Treatment principles are similar to those for Mycoplasma keratoconjunctivitis.

Moraxella ovis keratoconjunctivitis

Sheep and goats

Moraxella ovis (formerly Branhamella ovis) may be opportunistic to M. conjunctivae or C. pecorum infections

Florfenicol; oxytetracycline; ampicillin; ceftiofur

Some isolates are resistant to oxytetracycline and penicillin. Topical therapies could be considered.

Secondary infection of contagious ecthyma (orf)

Sheep and goats

S. aureus

Tilmicosin; oxytetracycline; ampicillin

May also try local antimicrobials but wear gloves, as zoonosis is a concern.

Dermatomycosis (streptothricosis, lumpy wool, rain scald, rain rot)

Sheep

Dermatophilus congolensis

Long-­acting oxytetracycline; procaine pencillins G; ceftiofur

Decrease humidity (ventilation) if possible, and protect from rain. Topical treatment includes lime sulfur, iodophors, copper sulfate, zinc sulfate, and potassium aluminum sulfate. Causative organism is thought to be susceptible to sulfonamides, bacitracin, and polymyxin B.

Caseous lymphadenitis

Sheep and goats

Corynebacterium pseudotuberculosis

No generally effective treatment. Tulathromycin and penicillin have been used

Although susceptible to penicillin and tulathromycin, treatment may not be effective because of the thick abscess wall. Techniques for abscess flushing and lavage have been described but risk introducing contaminated material into the environment. Recommend cull infected animals and avoid opening abscesses as it spreads the pathogen.

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Condition

(Continued )

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Condition

Species affected

Etiological agent(s)

Recommended treatment

Comments

Long-­acting oxytetracycline; penicillin; lincomycin; florfenicol

10–­20% zinc sulfate with 2% w/v sodium lauryl sulfate, as a foot bath with or without foot trimming. Must remain in bath 20 minutes. Repeat in 5–­7 days. Topical antimicrobial administration (tetracycline) to the hoof can also be considered. Can use in conjunction with systemic antimicrobials and/or vaccination. Selenium supplementation may improve outcome. Culling chronic nonresponders should be considered.

Infectious conditions of the foot and musculoskeletal system

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Table 31.1  (Continued)

Infectious/contagious pododermatitis (foot rot)

Sheep and goats

D. nodosus, F. necrophorum

Foot scald

Sheep and goats

F. necrophorum

Digital dermatitis (hairy heel wart)

Sheep

Treponema sp.

Oxytetracycline; tetracycline

Oxytetracycline can be administered systemically. Tetracycline can be used via topical application or foot bath. Avoid aggressive hoof trimming.

Contagious ecthyma (strawberry foot rot)

Sheep and goats

D. congolensis

As with lumpy wool

Verify that condition is not chorioptic mange.

Polyarthritis

Sheep and goats

Chlamydophila pecorum

Oxytetracycline

Poor response to treatment, may relapse, treatment not recommended.

Polyarthritis

Goats

Mycoplasma mycoides subsp. mycoides, LC other Mycoplasma spp.

Oxytetracycline; florfenicol; tulathromysin; tylosin

Poor response, may relapse. Treatment may require repeated joint lavage.

S. aureus, M. haemolytica, Clostridium spp., coliforms

Tilmicosin (sheep); broad-­spectrum antimicrobials

Gland will be lost if animal survives, so should consider culling. May require aggressive therapy with NSAIDs and fluid support for endotoxemic shock. Mastectomy may be required. Tilmicosin should not be used in goats.

Zinc sulfate foot bath as above.

Infectious conditions of the mammary gland Gangrenous mastitis

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Species affected

Etiological agent(s)

Mycoplasma mastitis

Sheep and goats

Multiple species

Subclinical and clinical mastitis

Sheep and goats

M. haemolytica, environmental streptococci, coagulase-­negative Staphylococcus spp.

Tilmicosin; cloxacillin; cephapirin benzathine; oxytetracycline

Dry treatment to be used at the end of lactation in dairy goats or at weaning for prevention of new infections in high-­risk sheep flocks. Do not split IMM tubes. Tilmicosin should not be used in goats (therapeutic dose very close to toxic dose).

Staphylococcus aureus mastitis

S. aureus

IMM therapy

Unlikely to guarantee clearance of S. aureus species. May need to consider mastectomy or culling.

Mastitis

Coliform

IMM therapy combined with systemic antimicrobials exhibiting Gram-­negative activity.

If severely ill, fluid therapy and nonsteroidal antiinflammatories may be necessary.

Ampicillin; ceftiofur; tetracycline; florfenicol; gentamicin; tulathromycin. IMM: Penicillin/ novobiocin, pirlimycin

Multiple therapies described but potential for variation exists. Consider milk culture and susceptibility.

Bacillus spp. mastitis

Recommended treatment

Comments

Typically ineffective; slaughter or culling should be considered. Potential for a carrier state.

Infectious conditions of the oral cavity Tooth root abscess

Sheep and goats

Many species

Oxytetracycline; florfenicol; ceftiofur

4–­6 weeks of therapy. Consider surgical intervention if antimicrobials fail. Long-­term florfenicol administration can pose risk of bone marrow suppression.

Actinobacillosis

Sheep

Actinobacillus lignieresii

Sodium iodide; oxytetracycline

Sodium iodide intravenously every 1-­2 weeks for 2–­3 doses or until clinical signs of iodinism.

Actinomycosis

Sheep

Actinomyces bovis

Sodium iodide; sulfadimethoxine; oxytetracycline

As for actinobacillosis. Treat for weeks to months. Prognosis poor.

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Condition

(Continued )

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Condition

Species affected

Etiological agent(s)

Recommended treatment

Comments

Infectious conditions of the urinary tract Leptospirosis

Sheep and goats

Leptospira interrogans

Oxytetracycline; tilmicosin (sheep); ceftiofur

Fluid therapy or blood transfusion may be required. Drugs are potentially nephrotoxic (oxytetracycline), questionable efficacy.

Cystitis

Sheep and goats

Corynebacterium renale, other species

Beta-­lactams

Therapy should be based on culture and sensitivity and should be given for 10–­14 days. Beta-­lactams should be considered for increased concentrations due to urinary excretion.

Infectious conditions of the nervous system Bacterial meningitis

Sheep and goats

Many species

Multiple options, should be focused on culture results

Gram-­negative organisms are likely; diagnostics should be considered to guide coverage. Antiinflammatory drugs important.

Listeriosis

Sheep and goats

L. monocytogenes

Oxytetracycline; penicillin G; florfenicol

Oxytetracycline: 10 mg/kg IV q24h administered slowly. Procaine penicillin G: 22,000–­44,000 IU/kg IM twice per day; intravenous formulation (potassium penicillin G) can be considered at the same dose q6h. Florfenicol: 20 mg/kg IM q24–­48h.

0005858926.INDD 662

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Table 31.1  (Continued)

10-28-2024 13:56:04

Once an antimicrobial drug is selected, proper administration is important. The label claim (when available) should generally be followed closely for dose, frequency, route of administration, and dose volume. However, clinicians should be aware of the fact that label claims may be outdated and not currently practiced. An example of this is the procaine penicillin G label in the United States for sheep, which recommends a 6600  IU/kg dosage. Due to resistance concerns, most clinical cases are not treated with this label dose but rather a much higher one (22,000–­44,000 IU/kg) due to the length of time that has passed since the label claim was approved. Any deviation from the label constitutes extra-­label drug use. Extra-­label usage is permitted in most regions but typically carries with it a zero tolerance for any residue, as opposed to a defined tolerance or MRL for drugs used on label. For quality assurance, it is also important to administer parenteral antimicrobial drugs in a way that minimizes damage to muscle tissues. Clean syringes and fresh needles should be used. The volume of drug per injection site should generally be limited to 5 ml or less. The route of administration should be selected based on known pharmacokinetic or pharmacodynamic data. Intramuscular injections should be given only in the neck. Subcutaneous injections should be given in the neck also. Small volumes (1 μg/ml for up to 12 hours; its reported volume of distribution is smaller than many other species (Christensen et al., 1996). Thus, trimethoprim appears to offer little advantage in the alpaca, but may be useful in llamas against susceptible microorganisms. Sulfamethoxazole acts more similarly in llamas and alpacas (Christensen et  al.,  1996; Chakwenya et  al.,  2002), but with a smaller

volume of distribution and faster clearance in alpacas. Camelids are fairly similar to sheep and cattle in these regards, with considerably faster clearance than horses or people. Active secretion into the renal tubules and trapping of the excreted agent in alkaline urine may contribute to this rapid clearance. It may also reflect a significant difference between NWCs and camels, which are reported to have acidic urine and slow sulfa clearance (Kumar et al., 1998). Injectable sulfamethoxazole may have some value in treating susceptible infections, especially in llamas. At metabolically scaled doses of 55–­62 mg/kg, sulfadimethoxine in llamas has a higher volume of distribution at steady state and a shorter half-­life than in cattle, and also may reach only subtherapeutic blood concentrations (Junkins et  al.,  2003; Boxenbaum et  al.,  1977). Similar volumes of distribution and peak concentrations and even faster clearance have been described in dromedary camels (Chatfield et al., 2000). No evaluation of higher doses has been described, but a metabolically scaled dose of 69  mg/kg was suggested for camels, and a higher, and possibly unsafe, dose might be necessary to reach desirable concentrations in New World camelids. The role of protein binding of sulfadimethoxine on clearance has been investigated in other species, and shown to be essential to the long half-­life (Bevill et al., 1982). Clinicians and clients should be aware that sulfonamide usage in crias has been associated  with Cryptosporidium spp. infections (Wyrosdick et  al.,  2023; Starkey et  al.,  2007), and this zoonosis should be considered for nonantimicrobial diarrhea in the NWCs.

Florfenicol Florfenicol has become more popular in the treatment of tooth root abscesses and other conditions where a long dosing interval is desirable. Recent clinical and experimental results suggest its value may be for treatment of highly susceptible pathogens in focal

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infections rather than as a broad-­spectrum treatment. This relates to its pharmacokinetic properties and the potential adverse effects. Intravenous florfenicol had a lower volume of distribution in NWCs than in sheep, goats, or camels, and a slightly longer half-­life than in sheep or goats; a 20 mg/kg dose yields plasma concentrations >1 μg/ml in both llamas and alpacas for about 12 hours (Ali et  al.,  2003; Christensen et  al.,  2001). When given IM at 20  mg/kg, peak plasma concentrations are achieved quickly, and are comparable between llamas and beef cattle, but higher in alpacas (Holmes et al., 2011). Elimination also appears to be similar to cattle or prolonged, but plasma concentrations generally drop below 1 μg/ml within 14–­24 hours. As with other medications, the variation between individual adult camelids is considerable: the volume of distribution after intravenous administration ranged from 0.25 to 2.54 l/kg in one study, and peak plasma concentrations after 20 mg/kg IM injection were 4.3±3 μg/ml in another. Single-­dose SC administration over the dorsal thorax takes slightly longer (2–­3 hours) to reach a lower peak, followed by an extensive elimination phase (Holmes et al., 2011). Regardless of whether 20 or 40 mg/kg is given SC, plasma concentrations are generally below 1 μg/ml within 18–­24 hours. The long elimination half-­lives (31–­100 hours) following SC injection may reflect important considerations related to camelid skin. Subcutaneous injections of more than a few ml are usually given to llamas and alpacas over the dorsal thorax. In contrast, SC florfenicol in cattle is specifically administered in the neck. Camelids’ dorsal thoracic skin is seasonally covered with fiber and plays little role in thermoregulation. It is poorly vascularized compared to axillary skin, potentially slowing the uptake of agents injected there, so that the long elimination half-­life is actually a ­reflection of slow absorption. Serial injections may increase blood flow and eventually lessen the differences between the IM and SC routes (Holmes et  al.,  2011). Administering florfenicol, or any other medication, in the axillary

region may result in faster peaks and a shorter elimination half-­life, but this has not been tested. The available studies suggest that daily IM florfenicol (20  mg/kg) may be effective against very to moderately susceptible microorganisms, but not Staphylococcus aureus, Pseudomonas aeruginosa, and many Gram-­ negative enteric bacteria. An ideal SC regimen has not been described. Lower doses (20  mg/kg, SC, q 24 h) may become adequate against sensitive pathogens, if greater absorption with serial dosing may be inferred. Higher doses (40 mg/kg, SC, q 24 h) maintain therapeutic steady-­state concentrations but are associated with evidence of toxicity, including reductions in blood proteins and cell counts, abnormal feces, and clinical ­disease. Because it interferes with protein synthesis, bone marrow suppression has been reported in other ruminants with high-­dose or chronic administration of florfenicol. While this has not been reported in NWCs, clinicians should be aware of this risk.

Fluoroquinolones Intravenous and SC enrofloxacin has been studied in llamas and alpacas. Marbofloxacin has been evaluated in llama crias ranging in age from three to 80  days (Rubio-­Langre et  al.,  2018). Enrofloxacin reaches high concentrations after IV or SC dosing, but conflicting information regarding its elimination half-­life is available (Christensen et  al.,  2001; Gandolf et al., 2005). There is a single report of retinopathy after enrofloxacin administration to a guanaco (Harrison et  al.,  2006). Marbofloxacin after IV administration in crias achieved high concentrations and displayed a long elimination half-­life, but these values decreased in older crias, suggesting dose adjustment may be necessary for older animals (Rubio-­Langre et  al.,  2018). Marbofloxacin dosed at 5 mg/kg in these crias achieved concentration of AUC24/MIC that would be consistent with effective concentrations for isolates of E. coli and Staphylococcus aureus.

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­Antimicrobials Used in New World Camelid  679

Antimicrobial Therapy in New World Camelids

Tetracyclines Intravenous and IM oxytetracycline has also been studied in llamas and alpacas. Intravenous oxytetracycline in llamas had a similar volume of distribution to camels, but a much longer elimination half-­life (Oukessou et  al.,  1992). Alpacas had a larger volume of distribution but a similar elimination half-­life to camels. Subcutaneous administration is common in clinical practice, particularly for the treatment of Mycoplasma haemolama or Anaplasma phagocytophilum infection, but has not been evaluated scientifically, and is likely to have the same problems as SC florfenicol. Preparations using a propylene glycol carrier are anecdotally associated with more local muscle irritation, shaking, and collapse than those using polyvinylpyrrolidone (povidone); reactions with either carrier are rare in New World camelids.

Other Antimicrobials Agents A variety of other parenteral antimicrobials have been used in individual camelids without full knowledge of safety or efficacy. For the most part, reasonable extrapolation can be made from similar species, with caution remaining the overarching principle. An example of this is the macrolide tilmicosin, which is labeled in the US for use in cattle and sheep, but reported to have cardiotoxic effects in horses and goats, and is also reported to have toxic effects in NWCs (FDA, 2024). The pharmacokinetics of gamithromycin have been described for alpacas and are characterized by a long elimination half-­life, as well as concentration in leukocytes (Gordon et  al.,  2022). That study noted one animal with colic signs after administration. There are several reports of tulathromycin being used in llamas and alpacas, but no rigorous safety evaluations have been performed. Oral tetracycline, amoxicillin-­clavulanic acid, isoniazid, and chloramphenicol have also been used in camelids, but no pharmacokinetic studies have been

performed. Oral enroflaxacin has a 29.3% ­bioavailability and reaches therapeutic concentrations after dosing at 10 mg/ kg (Gandolf et al., 2005).

Regional Antimicrobial Administration Regional limb perfusions have become increasingly common in NWCs for the treatment of infected bone and septic synovial structures. With this technique, a tourniquet is applied to the proximal limb and antimicrobials are intravenously infused, and the tourniquet is left applied for 20–­30 minutes afterwards. In ruminating species, florfenicol, ceftiofur, ampicillin sodium, and ampicillin-­sulbactam have all been used for this purpose but a paucity of information currently exists for this practice  with NWCs. Antimicrobial-­impregnated beads, with matrices of calcium sulfate or polymethylmethacrylate, have also been described for large animals, but no NWC-­specific studies evaluate the prospective use of this technique.

­Antifungals A variety of fungal diseases are reported in camelids but few systemic antifungal drug studies have been conducted in NWCs. Systemic mycoses include aspergillosis, cocciodiomycosis, cryptococcosis, histoplasmosis, and mucormycosis. Dermal or superficial mycoses include candidiasis, dermatophytosis (ringworm), and entomophthoramycosis. Voriconazole given IV (4  mg/kg) maintains plasma concentrations >0.1 μg/ml for at least 24 hours (Chan et al., 2009). Clearance and volume of distribution are comparable to horses but lower than in humans. Absorption after oral dosing is noted within five minutes, but bioavailability is less than 23%. Absorption is generally slow and unpredictable, and the need for higher doses has been postulated. Fluconazole administered orally in alpacas at 10–­15 mg/kg achieved AUCs similar to humans, but limited pharmacodynamic data exist (Butkiewicz

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et al., 2022). Other systemic antifungal use has been largely extrapolated from ruminants. Agents used empirically include fluconazole (14 mg/kg, IV or PO as a loading doses, followed by 5 mg/kg PO, q 24 h), itraconazole (5 mg/kg, IV or PO, q 24 h), and amphotericin B (0.5 mg/kg diluted in 0.5–­1 l of 5% dextrose IV over 1 h, q 24 h; after pretreatment with flunixin meglumine [0.25 mg/kg, IV]). Miconazole and clotrimazole

have been used topically for ­fungal dermatitis. Keratomycosis caused by Scopulariopsis brevicaulis and Fusarium verticilliodes has been treated with topical application of caspofunginin solution and terbinafine ointment, after initial treatment with voriconazole was unsuccessful (Foote et al., 2021). As with antibacterial use, all antifungal use in camelids is extra-­label.

­References and Bibliography Ali BH, et al. 2003. Comparative plasma pharmacokinetics and tolerance of florfenicol following intramuscular and intravenous administration to camels, sheep and goats. Vet Res Commun 27:475. Anderson DE. 2009. Analysis of antimicrobial cultures from llamas and alpacas: a review of 1821 cultures (2001–­2005). Proceedings of the International Camelid Health Conference. Bedenice D, et al. 2012. Florfenicol pharmacokinetics in healthy adult alpacas evaluating two commercially available drug formulations. J Vet Pharmacol Therapeut 35:382. Bevill RF, et al. 1982. Disposition of sulfadimethoxine in swine: inclusion of protein binding factors in a pharmacokinetic model. J Pharmacokinet Biopharm 10(5):539. Boxenbaum HG, et al. 1977. Pharmacokinetics of sulphadimethoxine in cattle. Res Vet Sci 23:24. Buchheit TM, et al. 2010. Use of a constant rate infusion of insulin for the treatment of hyperglycemic, hypernatremic, hyperosmolar syndrome in an alpaca cria. J Am Vet Med Assoc 236:562. Butkiewicz CD, et al. 2022. A preliminary study of the plasma concentrations of orally administered fluconazole in alpacas (Vicugna pacos). J Vet Pharmacol Therapeut 45:99. Cebra CK, et al. 2005. Glucose tolerance and insulin sensitivity in crias. Am J Vet Res 66:1013. Cebra CK, et al. 2006a. Meta-­analysis of glucose tolerance in llamas and alpacas. In: Gerken M,

Renieri C (eds) South American Camelids Research, vol. 1. Proceedings of the 4th European Symposium on South American Camelids and DECAMA European Seminar. The Netherlands: Wageningen Academic Publishers, p. 161. Cebra CK, et al. 2006b. Determination of organ weights in llamas and alpacas. In: Gerken M, Renieri C (eds). South American Camelids Research, vol. 1. Proceedings of the 4th European Symposium on South American Camelids and DECAMA European Seminar. The Netherlands: Wageningen Academic Publishers, p. 233. Chakwenya J, et al. 2002. Pharmacokinetics and bioavailability of trimethoprim-­ sulfamethoxazole in alpacas. J Vet Pharmacol Therapeut 25:321. Chan HM, et al. 2009. Pharmacokinetics of voriconazole after single dose intravenous and oral administration to alpacas. J Vet Pharmacol Therapeut 32:235. Chatfield J, et al. 2001. Disposition of sulfadimethoxine in camels (Camelus dromedarius) following single intravenous and oral doses. J Zoo Wildl Med 32:430. Christensen JM, et al. 1996. The disposition of five therapeutically important antimicrobial agents in llamas. J Vet Pharmacol Therapeut 19:431. Christensen JM, et al. 2001. Comparative metabolism of oxytetracycline and florfenicol in the llama and alpaca. Unpublished data.

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  ­References and Bibliograph

Antimicrobial Therapy in New World Camelids

Cox S, et al. 2015 Pharmacokinetics of intravenous and subcutaneous cefovecin in alpacas. J Vet Pharmacol Therapeut 38:344. Dechant J, et al. 2012. Pharmacokinetics of ceftiofur crystalline free acid after single and multiple subcutaneous administrations in healthy alpacas (Vicugna pacos). J Vet Pharmacol Therapeut 36:122. Dowling PM, et al. 1996. Pharmacokinetics of gentamicin in llamas. J Vet Pharmacol Therapeut 19:161. Drew ML, et al. 2004. Pharmacokinetics of ceftiofur in llamas and alpacas. J Vet Pharmacol Therapeut 27:13. Fajt VR. 2014. Drug therapy in llamas and alpacas. In: Cebra C et al. (eds) Llama and Alpaca Care: Medicine, Surgery, Reproduction, Nutrition, and Herd Health. Elsevier, St Louis, p.365. Featherstone CA, et al. 2011. Verocytotoxigenic Escherichia coli O157 in camelids. Vet Rec 16:194. Food and Drug Administration (FDA). 2024. Animal drug safety-­related labeling changes. www.fda.gov/animal-­veterinary/drug-­labels/ animal-­drug-­safety-­related-­labeling-­changes Foote BC, et al. 2021 Case Report: successful management of refractory keratomycosis in an alpaca using penetrating keratoplasty and combination antifungal therapy (caspofungin 0.5% and terbinafine 1%). Front Vet Sci 8:644074. Gandolf AR, et al. 2005. Pharmacokinetics after intravenous, subcutaneous, and oral administration of enrofloxacin to alpacas. Am J Vet Res 66:767. Gestrich A, et al. 2018. Pharmacokinetics of intravenous gentamicin in healthy young-­ adult compared to aged alpacas. J Vet Pharmacol Therapeut 41:581. Gordon E, et al. 2022. Plasma pharmacokinetics, pulmonary disposition, and safety of subcutaneous gamithromycin in alpacas. J Vet Pharmacol Therapeut 45:283. Hadi AA, et al. 1994. Pharmacokinetics of tobramycin in the camel. J Vet Pharmacol Therapeut 17:48.

Harrison TM, et al. 2006. Enrofloxacin-­induced retinopathy in a guanaco (Lama guanicoe). J Zoo Wildl Med 37:545. Hewson J, et al. 2001. Peritonitis in a llama caused by Streptococcus equi subsp. zooepidemicus. Can Vet J 42:465. Holmes K, et al. 2011. Florfenicol pharmacokinetics in healthy adult alpacas after subcutaneous and intramuscular injection. J Vet Pharmacol Therapeut 35:382. Hutchison JM, et al. 1993. Acute renal failure in the llama (Lama glama). Cornell Vet 83:39. Jones M, et al. 2009. Outbreak of Streptococcus equi ssp. zooepidemicus polyserositis in an alpaca herd. J Vet Intern Med 23:220. Junkins K, et al. 2003. Disposition of sulfadimethoxine in male llamas (Llama glama) after single intravenous and oral administrations. J Zoo Wildl Med 34:9. Kumar R, et al. 1998. Pharmacokinetics, bioavailability and dosage regimen of sulphadiazine (SDZ) in camels (Camelus dromedarius). J Vet Pharmacol Therapeut 21:393. Lackey MN, et al. 1995. Urinary indices in llamas fed different diets. Am J Vet Res 56:859. Lackey MN, et al. 1996. Single intravenous and multiple dose pharmacokinetics of gentamicin in healthy llamas. Am J Vet Res 57:1193. Lewis CA, et al. 2009. Colonic impaction due to dysautonomia in an alpaca. J Vet Intern Med 23:1117. Middleton JR, et al. 2006. Dysautonomia and salmonellosis in an 11-­year-­old female llama (Lama glama). J Vet Intern Med 20:213. Oukessou M, et al. 1992. Pharmacokinetics and local tolerance of a long-­acting oxytetracycline formulation in camels. Am J Vet Res 53:1658. Rubio-­Langre S, et al. 2018. Pharmacokinetic evaluation of marbofloxacin after intravenous administration at different ages in llama crias, and pharmacokinetic/pharmacodynamic analysis by Monte Carlo simulation. J Vet Pharmacol Therapeut 41(6):861. Saulez MN, et al. 2004. Necrotizing hepatitis associated with enteric salmonellosis in an alpaca. Can Vet J 45:321.

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Simpson KM, et al. 2011. Acute respiratory distress syndrome in an alpaca cria. Can Vet J 52:784. Snook CS, et al. 2002. Plasma concentrations of trimethoprim and sulfamethoxazole in llamas after orogastric administration. J Vet Pharmacol Therapeut 25:383. Starkey SR, et al. 2007. An outbreak of cryptosporidiosis among alpaca crias and their human caregivers. J Am Vet Med Assoc 231:1562. Sting R, et al. 2022. Fatal infection in an alpaca (Vicugna pacos) caused by pathogenic Rhodococcus equi. Animals 12:1303.

Tillotson K, et al. 1997. Outbreak of Salmonella infantis infection in a large animal veterinary teaching hospital. J Am Vet Med Assoc 211:1554. Wyrosdick HM, et al. 2023. Cryptosporidiosis in an alpaca cria secondary to prolonged antimicrobial administration. Vet Rec Case Rep 2023:e685. Zarrin M, et al. 2020. Camelids: new players in the international animal production context. Trop Anim Health Prod 52:903.

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  ­References and Bibliograph

33 Antimicrobial Therapy in Swine Ken Steen Pedersen

G ­ lobal Perspective on Antimicrobial Use in Swine Improved management and husbandry ­systems in swine farming have helped to prevent ­infectious diseases, including improvement of ­biosecurity, sourcing of high-­health breeding animals, utilization of “all-­in and all-­out” and multisite production. Further, increasing knowledge of feeding, development of feed additives, and efficient vaccines have added to the prevention and management of infectious diseases, potentially reducing the need for and consumption of antimicrobials in swine ­production. In spite of this, not all infectious diseases are controlled sufficiently. Problems in recruiting qualified staff and economic incentives to reduce production costs and increase productivity may lead to use of lower quality feed, increased stocking density, and lower management quality. In addition, preventive medication and antimicrobial growth promoters are used in some parts of the world. Therefore, there is and will be an ongoing need for treatment of pigs with antimicrobials. Antimicrobial use in swine has always been substantial and in some cases, the swine industry has been considered overreliant on their use. As farms have evolved from small ­backyard operations to today’s substantially larger units, it is not surprising that

antimicrobials have been used to help farmers maintain production under these major management, housing, and disease pattern changes. Globally, antimicrobial use in swine is significant and accounts for a large part of the total antimicrobial use in animals. The amount of antimicrobials consumed by food animals worldwide is almost double that used in humans (Aarestrup, 2012) and the global antimicrobial consumption by food animals will increase substantially from 2010 to 2030 based on increasing demand for animal protein in low-­ and middle-­income countries. The production of pigs will be one of the main drivers of this increase (Van Boeckel et al., 2015). Significant differences exist between ­countries, but consumption data suggest that managing antimicrobials in modern and intensive pig production is important in reducing the use of antimicrobials globally. This is in agreement with an OIE published report that highlights important animal diseases for which the development of a vaccine would potentially reduce antimicrobial use in swine. In total, eight porcine bacterial pathogens and three viral agents were identified: Streptococcus suis, Pasteurella multocida, Mycoplasma ­hyopneumoniae, Actinobacillus pleuropneumoniae, Escherichia coli, Clostridium perfringens, Lawsonia intracellularis, Brachyspira spp.,

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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porcine reproductive and respiratory ­syndrome virus, swine influenza virus, and rotaviruses (OIE, 2015). Overall, this makes antimicrobial use in swine highly relevant for the swine industry, the individual farmer, and global One Health. It must be remembered that antimicrobials have a cost and that farmers are unlikely to pay for them unless they can see a benefit from their use. Responsible use calls for veterinarians and farmers to use antimicrobials “as little as possible but as much as needed.” There are a number of guidelines on how to use antimicrobials properly. At the same time, efforts are being made to reduce unnecessary use and forbid the use of critical human antimicrobials where suitable alternatives are available. It is the aim of this chapter to provide insights into the use of antimicrobials in swine and to help with decision making regarding when to use antimicrobials, selection of active substance, route of administration, and dose.

­ atterns of Antimicrobial Use P in Swine Globally, national information about administration, age groups, and indications for antimicrobial consumption is sparse, but more and more countries are setting up surveillance ­systems for antimicrobial consumption. Several studies have reported on the consumption patterns in swine, though primarily from Europe. Variation in antimicrobial use in ­relation to amount, age groups, and indications exists between herds, countries, and studies (Sjölund et al., 2016). The postweaning production stage is exposed to the highest antimicrobial use across several European countries, though in countries like Germany and Sweden, the highest usage was in suckling pigs. Similar differences exist among countries in relation to the indications for which the antimicrobials are used. In Denmark and France, the majority of antimicrobials were prescribed for gastrointestinal

diseases. In Germany, respiratory infections were the main indication for antimicrobial use in piglets, weaners, and fattening pigs, ­followed by intestinal diseases. In Austria, nonspecified metaphylactic or prophylactic treatment ­constituted the major indication, followed by respiratory and intestinal diseases. In Belgium, diarrhea was the main indication from weaning up to 70 days of age, while respiratory disease was the main indication in grower and finishing pigs. The route of administration used also varies between age groups and countries. Oral administration at group level dominates, and numbers in the region of 86–­98% of total consumption have been reported. In Denmark, antimicrobial use in pigs was administered orally for 24% of sows and piglets, 98% of weaners, and 74% of finishers. In contrast, Swedish herds received only 13% of antimicrobials orally. Antimicrobial use can be therapeutic, prophylactic, metaphylactic or for growth promotion. Therapy is the treatment of infections in clinically sick pigs. Prophylactic use of antimicrobials has been defined as the treatment of healthy pigs to prevent disease from occurring, whereas metaphylactic use has been defined as concurrent treatment of both clinically healthy and diseased pigs belonging to the same group of animals. Finally, growth promotion is defined as the continual inclusion of antimicrobial agents in animal feed to improve growth (Aarestrup, 2005). This use has been banned in Europe, Canada, and the United States, but antimicrobial growth promoters are still used in many countries. Most prophylactic and metaphylactic treatments are administered orally in pigs. The relative occurrence of these two types of medications varies among countries.

­ iagnostic Aspects D of Antimicrobial Use in Swine Several guidelines relating to the responsible use of antimicrobials in veterinary practice have been published, including standards for

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the prudent use of antimicrobial agents in ­veterinary medicine in the OIE Terrestrial Animal Health Code 2017 (OIE,  2018) and guidelines issued by the European Commission (European Commission, 2015). According to the OIE, the first key step is to perform a proper clinical examination of the animal and decide whether antimicrobial treatment is necessary or if the disease should be managed another way. The second step is to select the most appropriate antimicrobial drugs based on clinical experience and diagnostic laboratory information, including pathogenic agent isolation, identification, and antimicrobial resistance assessment where possible. The third step is to provide a detailed treatment protocol. The initial steps in the timely and accurate diagnosis of infectious swine diseases are often challenging. Diagnostic samples must be transported from farms to laboratories and then tested. The time lag between sampling and obtaining the result has been reported as one of the important reasons why antimicrobial susceptibility testing is not commonly performed before antimicrobial treatment (Carmo et  al.,  2018). Therefore, veterinarians need to treat pigs empirically in order to prevent disease progression. Knowledge of the prevalence of a disease in a given country can help when making an empirical diagnosis and guide the initial selection of an antimicrobial substance. Rapid and affordable point-­of-­care tests (pig-­ side or pen-­side tests) for selected diseases for which antimicrobials are most commonly used will improve antimicrobial usage. Development of diagnostics has been reported as one of the areas that must be prioritized in animal research in order to reduce antimicrobial use (WHO, 2017). For diseases in pigs, clinical decision making in relation to antimicrobial treatment should be based on evidence, using diagnostic test performance measures, knowledge of disease prevalence, post-­test probabilities, and scientific knowledge of the efficacy of different antimicrobial treatments or alternative (i.e.,

nonantimicrobial) interventions. Clinical decision making is a complex process potentially influenced by many factors and clinical decisions on antimicrobial treatment and preventive strategies in relation to diseases in pigs are likely to be a combination of clinical experience, judgment, and scientific knowledge. For a diagnostic method to be useful, it must be accurate, simple, and affordable and it should also be demonstrated that a diagnostic test has an impact, resulting in changes related to prevention or antimicrobial treatment strategies. Lack of impact is an important reason why antimicrobial susceptibility testing is not commonly performed before antimicrobial treatment, as the results are not considered to help in the clinical decision-­making process and/or selection of the antimicrobial to be used (Carmo et al., 2018). Aspects including selection of pigs to test, diagnostic sample, and diagnostic technique must be considered. These considerations are in no way trivial and require professional effort in each case. Interpretation of laboratory examinations must be made in relation to the clinical signs, pathology, and history from the farm. Modern techniques like PCR testing of swabs, oral fluids, fecal materials or pathological material are in general sensitive. Also ­techniques like MALDI-­TOF identify many bacteria after an unspecific culture. Many infectious diseases in pigs are caused by opportunistic bacterial organisms so the simple detection of such bacteria in a sample does not necessarily demonstrate the cause of disease or  a need for antimicrobial treatment. Such ­bacteria include Mycoplasma hyorhinis, Streptococcus spp., Staphylococcus spp., Escherichia coli, and Glaesserella parasuis. Quantitative or semiquantitative techniques should be preferred when possible, including quantitative PCR or culture. Additional characterization by virulence determination in enterotoxigenic E. coli should be performed. Service limitations in local laboratories may make such diagnostic tests difficult to obtain

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and increase the need for empirical ­antimicrobial treatments. Laboratory investigation should include antimicrobial resistance assessment of bacteria. This may not be possible for some important bacteria like Lawsonia intracellularis or difficult for some slow-­growing bacteria like Brachyspira spp. Further, some bacteria like Glaesserella parasuis or Mycoplasma spp. may be difficult to culture and can only be detected by molecular techniques, providing no possibility for antimicrobial resistance assessment. Molecular techniques may help with this in the future but currently, it is not possible to predict the clinical effect of an antimicrobial based on demonstration of resistance genes. Additional elements concerning susceptibility testing are outlined in the section on susceptibility testing below, and see also Chapter 2. Mixed infections pose a special situation and are very common for both enteric and respiratory diseases in swine. This may result in use of more broad-­spectrum antimicrobials or even using two different substances in some situations. For enteric infections, it is also now well documented that the infections involved can change from one batch to the next within the same herd. This has implications for both selection of antimicrobials and diagnostic strategy, since pigs representing more than one batch ideally should be included in the diagnostic workup. The concept of primary and secondary ­infections is also important and must be taken into consideration when selecting antimi­ crobials for treatment of mixed infections. Antimicrobial therapy should always be ­targeted against the primary infections while secondary infections like some Streptococcus spp. or Trueperella pyogenes do not need to be taken into consideration if efficient treatment of the primary infection is performed at the beginning of a disease episode in a pig. In other situations, the primary infection is a virus infection such as PRRS virus or PCV2 virus, in which case treatment may by targeted at the secondary bacteria. Secondary bacteria may

also cause more severe disease, for example Mycoplasma hyorhinis or Pasteurella multocida, and in those situations the treatment must also be targeted at these secondary infections.

­ dministration of Antimicrobials A in Swine Administration of antimicrobials is done by topical, intramuscular injection or oral (water/ feed/directly in mouth) routes. Intramammary and intrauterine routes are normally not used in pigs. Antimicrobials can be administrated to a single pig or as herd (batch) medication where all pigs in a “herd” are treated simultaneously. Herd medication can be done via water or feed (e.g., outbreak of pneumonia), directly in the mouth (e.g., outbreak of diarrhea), by injection (e.g., outbreak of pleuropneumonia) or by ­topical treatment (e.g., outbreak of exudative dermatitis). A herd can be anything from a litter by one sow, a small collection of pigs in a pen, multiple pens in a section/room to an entire herd, i.e., herd medication can be at pen level, section level, farm section level (e.g., all nursery pigs) or farm level. Topical administration is mainly done in cases of skin diseases like exudative dermatitis with Staphylococcus hyicus, ear necrosis or other wounds like shoulder ulcers. Also, preventive treatment of umbilical infections by spraying the umbilicus with antimicrobials like tetracyclines at birth is commonly performed. Injection of pigs is performed in many settings, but primarily to treat clinically ill pigs. Examples include pigs with acute respiratory infections, such as Actinobacillus pleuropneumoniae, enteric infections such as swine ­dysentery caused by Brachyspira hyodysenteriae, pigs with lameness or sows with mastitis. Animals which are too sick to eat or drink are best treated by injection. Injection is a

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highly effective route of administration to an individual animal and is the best way to make sure the pig receives the full dose of the antimicrobial. Some antimicrobials require repeated injections on a daily basis which reduces their practical application, because repeated injections make it more difficult to treat the same pig. Increasing use of loose housing systems for breeding animals has increased the relevance of this challenging aspect of administration. The development of antimicrobial drugs in long-­acting formulations has improved the ease of practical application and made it possible to use injections for more preventive or metaphylactic reasons. Examples of such use of injections with or without long-­acting formulations include preventive or metaphylactic treatment of navel or respiratory infections in piglets. Since each animal caretaker or veterinarian has to manage many pigs and injections are laborious, oral administration of antimicrobials is the most common route of application in swine medicine globally. Oral administration can be performed by oral dosers (drench/ individual pump), mixing in to water or feed and by top dressing on feed. The administration of antimicrobials in water or feed has often been the subject of discussion in the political debate regarding antimicrobial resistance as this has become synonymous with herd medication. However, in practice, all medicinal products mixed in either water or feed can easily be administered at single-­ animal level as well. The advantages of administering antimicrobials in water or feed are that many pigs can be treated in a short time, it is labor saving, less stressful for the pig, and treatment of all infected pigs including the subclinical cases is ensured. However, there are also a number of disadvantages, including uncertainty regarding dosage as many diseased pigs drink and particularly eat less and the intake of the antimicrobial can be uncertain at the individual level. Other disadvantages are potential problems with

antimicrobial residue in feed pipes and water pipes plus an unnecessarily high antimicrobial consumption due to treatment of potentially healthy pigs. Piglets can easily be treated individually by drenches or oral dosers, which have proven useful for the control of neonatal intestinal infections. Increasing focus of optimized antimicrobial use may lead to more individual administration by oral dosers in slightly older pigs, such as treatment of E. coli postweaning diarrhea in newly weaned pigs. Administering antimicrobial products to single animals via feed or water is done if this is easier or if the antimicrobial selected is not available for injection. Water or feed is also used when several animals with the same ­disease/treatment are collected in one or more pens. An example would be hospital pens with pigs that have diarrhea due to infection with Lawsonia intracellularis. Finally, medicated water and feed are used when many animals need treatment at the same time (herd treatment). Situations where herd medication (batch medication) is used include cases of high disease occurrence (many clinically sick animals at the same time), very acute disease courses in a group of pigs and cases of many subclinically infected pigs that will develop clinical disease if left untreated (incubation-­phase treatment). It is a requirement that the majority of the animals are clinically sick, subclinically sick or in an incubation phase before it is reasonable to carry out herd treatment. Those cases where seemingly healthy animals are treated routinely at a fixed time in order to avoid an infection must be considered as preventive treatment and therefore be reserved for special situations such as eradicating an infection from the herd. Administration of antimicrobials in water can be performed directly in troughs to single animals or a few animals in one or a few pens. If a larger number of animals need treatment, a medicine mixer (e.g., Dosatron®) is used where entire pens, entire sections, entire farm

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sections or the entire herd can be treated at the same time. Such a medicine mixer can be placed in front of a pen, a section or centrally in the production system with a medicine pipe connected to all the different parts of the farm. When using a medicine mixer, the antimicrobial is dissolved in a concentrate, which flows into the main water system at a set target rate (approximately 1–­2%, depending on drug solubility). However, the antimicrobial needs to be sufficiently water soluble. Calculations of the dosing rate can be made by determining the pigs’ water consumption, but it is important to monitor that the total daily dose for the group of pigs has been consumed. Making new concentrate every day is recommended. Administration of antimicrobials via the feed can be done with top dressing for single animals or a few animals in one or few pens. On farms with home-­mixed feed, the antimicrobial can be mixed into the feed during or after manufacturing. On farms using purchased feed, the feed can be ordered mixed with an antimicrobial product, i.e., a medicated feed. Feed medication is still the main route of antimicrobial administration in the swine industry in many countries. However,

for treatment of disease it can be argued that it is the least efficient route as it may take some days for the feed to be manufactured, delivered, and work through the storage bin system to get to the pigs. There are some pharmacokinetic disadvantages to administering antimicrobials via the feed, as sometimes the feed interferes with the absorption of a drug and reduces its bioavailability (Nielsen, 1997) and thereby plasma concentrations. This can have an impact particularly when treating systemic or respiratory infections (Figure 33.1). Drug dose and inclusion rate are also critical and the former depends on feed intake and the body weight of the pig. The daily feed intake depends on many things, including the age of the pig, production phase for breeding animals, feeding system (wet-­feed, ad libitum, restrictive), housing temperature, genetics, and gender. Therefore, it is essential to adjust the inclusion rate to achieve an effective dose rate. Both water and in-­feed antimicrobials, administered over the day, usually give a lower but flatter pharmacokinetic plasma concentration curve than following a single injection or oral dose (Figure 33.2).

100 90 Bioavailability (%)

80 70 60 50 40

Fasted Fed

30 20 10

llin V te tra cy cl C in hl e or te tra cy cl in e Sp ira m yc in Li nc om yc in En ro flo xa ci n

ci

O xy

Pe ni

et h Tr im

Su

lfa

di a

zi

op rim

ne

0

Figure 33.1  Bioavailability (%) of antimicrobials when given fasted or after feeding to pigs (Nielsen, 1997).

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690

Antimicrobial conc. (μg/ml)

7

Maximum concentration (C max)

6 5

Area under the curve (AUC)

4 Steady state (Css)

3

MIC for bacterium

2 1 0 0

4

8

Injection

12 Hours Water/feed

16

20

24

MIC

Figure 33.2  Comparative antimicrobial pharmacokinetic curves following injection or in-­feed or in-­water medication.

­ ntimicrobial Growth Promoters A in Swine Antimicrobial growth promotion is defined as the continual inclusion of antimicrobial agents in animal feed to improve growth (Aarestrup,  2005). In the 1990s, antimicrobial growth promoters were used in most countries. This has changed and antimicrobial growth promoters are now banned in some countries (EU, Canada, US) but are still used in other countries. The benefit of antimicrobial growth promoters is increased productivity from weaning to slaughter. Further, inclusion of antimicrobial growth promoters in sow lactation feed may increase growth in piglets. Many successful growth promoters actually have a prevention of disease effect/claim, such as virginiamycin, which prevents Clostridium perfringens infections, carbadox which prevents swine dysentery (Brachyspira hyodysenteriae), and tylosin which prevents porcine proliferative enteropathy “ileitis” (Lawsonia intracellularis). Therefore, the use of growth promoters also contributes to controlling enteric infections in pigs. This explains some of the mechanism by which antimicrobial growth promoters increase productivity. However, this indicates that the

application of therapeutic ­antimicrobials to control intestinal disease may increase following a global ban on antimicrobial growth promoters. This effect was observed in Denmark after the voluntary ban in 1999, but the total consumption of antimicrobials was still significantly reduced. An antimicrobial growth promotion effect can be obtained by continuous inclusion of low-­dose antimicrobials in water or feed for 3–­4  weeks during the nursery or grower production phase as preventive medication of infections. Such usage of antimicrobials for promotion of growth cannot be considered prudent antimicrobial use.

­ ypes of Antimicrobials Used T in Swine A wide variety of antimicrobials are available for use in swine, as summarized in Table 33.1. The active substances used vary across age groups and countries. The most commonly used active substances include tetracyclines, TMP/ sulfonamides, aminoglycosides, macrolides, amoxicillin, colistin, and penicillin. Fluoroquinolones and cephalosporins are also used in some countries but are banned in others.

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­Types of Antimicrobials Used in Swin  691

Antimicrobial Therapy in Swine

Table 33.1  Antimicrobials used in swine.

Table 33.1  (Continued)

Tetracyclines

Macrolides

Oxytetracycline

Tylosin

Chlortetracycline

Tylvalosin

Tetracycline

Tilmicosin

Doxycycline

Tildipirosin

Diaminopyrimidine/sulfonamide

Triamilide

Trimethoprim/sulfadiazine

Tulathromycin

Trimethoprim/sulfadoxine

Gamithromycin

Trimethoprim/sulfamethoxasole

Lincosamides

Penicillins

Lincomycin

Benzylpenicillin procaine (penicillin G)

Pleuromutilins

Phenoxymethylpenicillin (penicillin V)

Valnemulin

Synthetic penicillins

Tiamulin

Amoxicillin

Orthosomycin

Ampicillin

Avilamycin

Also in combination with clavulanic acid

Ionophores

Cephalosporins

Salinomycin

Cephalexin

Growth promoters

Ceftiofur

Virginiamycin

Cefquinome

Bacitracin methylenedisalicylate

Fluoroquinolones

Zinc bacitracin

Enrofloxacin

Flavophospholipol

Danofloxacin

Bambermycin

Marbofloxacin

Carbadox

Thiamphenicols Thiamphenicol Florfenicol Aminoglycosides Dihydrostreptomycin Neomycin Apramycin Gentamicin Amikacin Paromomycin Aminocyclitols Spectinomycin Polymixin Colistin

­ ntimicrobial Susceptibility A of Pathogenic Bacteria in Swine Surveillance of antimicrobial resistance is performed and reported in a number of countries around the world, but is for the most part restricted to indicator bacteria like E. coli or food-­borne zoonoses like Salmonella spp. For porcine pathogenic bacteria, less systematic ­surveillance results are reported, though a number of peer-­reviewed publications and reports have been available for different regions, countries or laboratories over the

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years. In relation to selection of antimicrobial substance for treatment of diseases in pigs, the focus must be on antimicrobial resistance in porcine pathogenic bacteria. Antimicrobial resistance in porcine bacterial pathogens poses treatment problems in some situations, including the occurrence of multiresistant bacteria. Antimicrobial resistance has been reported in many different porcine pathogenic species including, though not exclusively, Actinobacillus pleuropneumoniae, Escherichia coli, Mycoplasma hyopneumoniae, Brachyspira hyodysenteriae, and Pasteurella multocida. It is well documented that antimicrobial use on the individual farm selects for resistance over time and can result in lack of efficiency for such antimicrobials. From clinical practice, it is also well known that after termination of use of a specific antimicrobial substance, the resistant bacteria can disappear, making it possible to use the antimicrobial again after some time. The process is dynamic within a farm and not well understood. The antimicrobial susceptibility patterns are different between farms, pigs, and countries. As an example, a comparative study looking at 30 UK and 30 Spanish isolates of G. parasuis (Martin de la Fuente et al., 2007) highlights the difference in susceptibility patterns in different countries, showing that it is important to develop local farm and national data. In those countries or regions where resistance data are available, such data should be consulted before selecting antimicrobial substances for empirical treatment. A better alternative is to use historical data from the same farm from earlier demonstration of the same bacteria pathogen or alternatively other bacteria pathogens demonstrated in the same farm. However, ­caution must be exercised as different bacterial species on the same farm may have different susceptibility patterns. In the optimum situation, assessment of antimicrobial susceptibility should be

performed regularly for those diseases that are treated on the individual farm. However, in clinical practice, it sometimes can be difficult to predict the clinical effect of an antimicrobial based on susceptibility assessment in the laboratory. Additionally, assessment is not possible in routine laboratories for many significant porcine bacteria pathogens and there is also a lack of internationally recognized species-­ or disease-­specific breakpoints for some significant porcine bacteria pathogens. Finally, resistance is not static and has been shown to vary between different groups of pigs with the same clinical disease in the same herd over time.

­ mpirical Treatment of Common E Bacterial and Mycoplasma Infections in Swine Table  33.2 includes suggestions for prudent empirical selection of antimicrobial substances for common swine diseases. The table is based on clinical signs since these are the basis for decision making on empirical treatment in clinical practice. The substances and dose suggested are based on clinical experience from Danish farms with low use of antimicrobials. Duration of treatment must be for as long as the pig or group of pigs has clinical signs. After clinical signs have disappeared, treatments can be terminated. Special situations apply to batch medication which in many cases must be a bit longer to make sure all animals are treated since it may take some days before all animals have been infected. Early termination of ­treatment in such situations may result in repeated disease episodes within the same group of pigs. For individual treatments, three days are normally sufficient while batch medications should have a duration of 4–­5  days, although for E. coli postweaning diarrhea this may only be three days.

693

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Empirical Treatment of Common Bacterial and Mycoplasma Infections in Swine

Antimicrobial Therapy in Swine

Table 33.2  Suggestion for prudent empirical selection of antimicrobial substances for common swine diseases. The substances suggested are from clinical experiences in Danish farms with low use of antimicrobials. In other countries, drugs such as avilamycin and tilmicosin are approved for treatment use in swine. Disease/clinical signs

Age group

Pathogen

Antimicrobial

Diarrhea

Piglets

Escherichia coli

Trimethoprim/sulfadiazine

Clostridium perfringens type A and C

Benzylpenicillin procaine

Weaning to finishing Escherichia coli

Trimethoprim/sulfadiazine

Lawsonia intracellularis

Tiamulin

Brachyspira pilosicoli

Tiamulin

Salmonella enterica spp.

Trimethoprim/sulfadiazine

Brachyspira hyodysenteriae

Tiamulin

Upper respiratory tract Piglets, weaning to finishing

Bordetella bronchoseptica

Trimethoprim/sulfadiazine

(sneezing)

Toxigenic Pasteurella multocida

Trimethoprim/sulfadiazine

Mycoplasma hyorhinis

Tiamulin

Bordetella bronchiseptica

Trimethoprim/sulfadiazine

Toxigenic Pasteurella multocida

Trimethoprim/sulfadiazine

Mycoplasma hyorhinis

Tiamulin

Lower respiratory tract Piglets, weaning to finishing

Nontoxigenic Pasteurella multocida

Trimethoprim/sulfadiazine

(cough, dyspnea)

Bordetella bronchiseptica

Trimethoprim/sulfadiazine

Mycoplasma hyopneumoniae

Tiamulin

Actinobacillus pleuropneumoniae

Benzylpenicillin procaine

Trueperella pyogenes

Benzylpenicillin procaine

Streptococcus spp.

Benzylpenicillin procaine

Staphylococcus spp.

Benzylpenicillin procaine

Streptococcus suis

Benzylpenicillin procaine

Haemophilus parasuis

Benzylpenicillin procaine

Mycoplasma hyosynoviae

Tiamulin

Mycoplasma hyorhinis

Tiamulin

Escherichia coli

Trimethoprim/sulfadiazine

Streptococcus spp.

Benzylpenicillin procaine

Staphylococcus spp.

Benzylpenicillin procaine

Escherichia coli

Trimethoprim/sulfadiazine

Lameness

Mastitis, metritis, agalactia (MMA) syndrome

Septicemia

Piglets, weaning to finishing, breeding animals

Sows

Piglets, weaning to finishing

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Table 33.2  (Continued) Disease/clinical signs

Meningitis

Age group

Piglets, weaning to finishing

Pathogen

Antimicrobial

Streptococcus spp.

Benzylpenicillin procaine

Escherichia coli

Trimethoprim/sulfadiazine

Streptococcus spp.

Benzylpenicillin procaine

Haemophilus parasuis

Benzylpenicillin procaine

Erysipelas dermatitis

Piglets, weaning to finishing, breeding animals

Erysipelothrix rhusiopathiae

Benzylpenicillin procaine

Exudative epidermitis, “greasy pig disease”

Piglets, weaning-­ growing pigs

Staphylococcus hyicus

Trimethoprim/sulfadiazine

­References and Bibliography Aarestrup FM. 2005. Veterinary drug usage and antimicrobial resistance in bacteria of animal origin. Basic Clin Pharmacol Toxicol 96:271. Aarestrup F. 2012. Get pigs off antibiotics. Nature 486(7404):465. Carmo LP, et al. 2018. Veterinary expert opinion on potential drivers and opportunities for changing antimicrobial usage practices in livestock in Denmark, Portugal, and Switzerland. Front Vet Sci 5(29):1. European Commission. 2015. Guidelines for the prudent use of antimicrobials in veterinary medicine. Official Journal of the European Union. C299. Martin de la Fuente AJ, et al. 2007. Antimicrobial susceptibility patterns of Haemophilus parasuis from pigs in the United Kingdom and Spain. Vet Microbiol 120:184. Nielsen P. 1997. The influence of feed on the oral bioavailability of antibiotics/chemotherapeutics

in pigs. J Vet Pharmacol Therapeut 20 Suppl 1:30. OIE. 2015. Report of the Meeting of the OIE Scientific Commission for Animal Diseases. World Organisation for Animal Health, Paris. OIE. 2018. Terrestrial Animal Health Code. www.oie.int/en/standard-­setting/terrestrial-­ code/access-­online/ Sjölund M, et al. 2016. Quantitative and qualitative antimicrobial usage patterns in farrow-­to-­finish pig herds in Belgium, France, Germany and Sweden. Prev Vet Med 130:41. Van Boeckel TP, et al. 2015. Global trends in antimicrobial use in food animals. Proc Natl Acad Sci USA 112(18):5649. World Health Organization (WHO). 2017. Global Framework for Development & Stewardship to Combat Antimicrobial Resistance –­Draft Roadmap. World Health Organization, Geneva.

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  ­References and Bibliograph

34 Antimicrobial Therapy in Poultry Jenny A. Nicholds, David French, Daniel Parker, and Peter O’Kane

The poultry industry is different from many other livestock sectors in that it is unusual for the veterinarian to be presented with single animals to examine and treat. Due to the scale of farms, it is not unusual for a poultry barn to house up to 200,000 commercial layers or 80,000 broilers. Furthermore, poultry farms can have multiple houses so the number of birds on these epidemiological units can easily exceed 1,500,000 in the case of commercial layers and 400,000 in the case of broilers. At the hatchery level, depending on the type of equipment in place, one incubator can contain more than 120,000 eggs or developing embryos. Thus the poultry veterinarian’s main focus needs to be on disease prevention Historically, antimicrobials were widely used in both the prevention and treatment of diseases, but with more focus on responsible use of antimicrobials, poultry producers and veterinarians should consider biosecurity, hygiene, and management as their primary tools for disease prevention and target antimicrobial use for treatment of clinical infections. When husbandry, hygiene, and biosecurity procedures fail to prevent the introduction of a disease agent, appropriate antimicrobial therapy may become necessary to prevent pain and suffering in affected birds as well as economic losses to the producer. The poultry veterinarian will then be required to investigate, make a

diagnosis, and if antimicrobial therapy is considered necessary, they must then determine the appropriate drug formulation, duration of treatment, and route of administration.

­ ategories of Antimicrobial Drug C Use in the Poultry Industry Antimicrobial drug use in poultry can be divided into three categories: therapeutic, ­preventive/prophylactic, and growth promotion. Antimicrobial drugs in the therapeutic category are used to treat or cure a clinically detectable disease. Because sick birds often have reduced feed consumption or feed bins at the farm may be full, therapeutic antimicrobials are typically administered via the drinking water to ensure rapid intake and adequate uptake of the medication. Certain circumstances or disease conditions, however, may dictate administration in feed instead or concomitantly with water. Prophylactic or preventive antimicrobials are administered prior to the appearance of clinical signs of disease in a flock. The route of antimicrobial administration will depend on the timing or age of bird when the treatment is applied. Due to the structure and scale of the poultry industry, the hatchery is a critical control point in the management of health in

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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Antimicrobial Therapy in Poultry

poultry flocks. Hatching eggs from multiple flocks are often co-­mingled in the incubators and hatchers, so the microbiological status of the eggs and chicks from these eggs will be shared. If increased bacterial contamination has been identified in association with hatching eggs coming from a particular breeder flock, progeny from that flock and progeny of other flocks that were in the same hatchery may be treated preventively using in ovo (eggs) or subcutaneous (day-­old chicks or poults) injection of an antimicrobial until the underlying cause for such contamination is identified and corrected. Other routes for administration of prophylactic antimicrobials include oral administration via drinking water or feed. The last category of antimicrobial use, growth promotion, is the most controversial. Many antimicrobials were first approved for use in poultry based on their observed growth-­ promoting effects: improved feed efficiency and growth rates. The improved production resulted in an economic benefit that was greater than the cost of the antimicrobial drug. Due to increasing concerns that the growth-­ promoting use of antimicrobials had the potential for a negative impact on human health due to the development of antimicrobial resistance, there has been voluntary and regulatory removal of antimicrobial growth promoters from poultry production in many jurisdictions. With discontinued use, it became evident that much of the growth promotion effect was due to the control and prevention of subclinical enteric disease. In the US and Canada, some antimicrobials were authorized for both therapeutic use and growth promotion; however, the dose level per bird for growth promotion was generally lower than the therapeutic dose. Regulatory changes definitively preventing growth promotion use became effective in early 2017 and late 2018 in the US and Canada respectively. In the United Kingdom (UK) and European Union (EU), the use of antimicrobials for growth promotion was phased out through the 1990s and finally became illegal in the UK and EU on 1 January

2006. Since January 2022, in the EU antimicrobial medicinal products are not permitted for prophylaxis other than in exceptional circumstances. In considering these broad categories of antimicrobial use in poultry, the distinction between therapeutic and preventive categories is not always clear. The poultry veterinarian is faced with making a decision regarding treatment of a population; individual treatment is often not possible or practical. When disease is identified in a flock, not all birds will be clinically ill. The antimicrobial treatment will be therapeutic for those clinically affected birds and preventive for the remaining birds. This introduces the concept of metaphylaxis  –­ treating part of the population that are not clinically ill but are in contact with sick animals. To further complicate the matter, growth-­promoting antimicrobials are known to kill or inhibit the growth of disease-­ causing agents, including bacteria or coccidia. These products are particularly effective at prevention of necrotic enteritis, a condition triggered by enteric overgrowth of Clostridium perfringens (Grave, 2006; Smith, 2011). While the exact mode of action for growth promotion associated with the use of antimicrobials is debatable (Dibner and Richards,  2005; Neiwold, 2007), growth promotion is clearly a “side-­effect” of disease prevention.

­ ntimicrobial Drug Use in the A Poultry Industries of Canada, United States, and UK Antimicrobial use, in animals or humans and in any of the previously described categories, has the potential to select for bacterial strains that are resistant to the antimicrobial used (Roth et al., 2019). For this reason, antimicrobial use in food-­producing animals, particularly for growth promotion, has been and still is a focus of scientific, political, and ­consumer debate (Casewell et  al.,  2003; Phillips et  al.,  2004; Kelly et  al.,  2004; Cox and

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698

Popken,  2004; Cox,  2005; Phillips,  2007). The association of antimicrobial use in food animals with antimicrobial resistance impacting human health resulted in different approaches to the control of antimicrobial use in a number of countries. Over the past 50 years, use of antimicrobials in these categories has changed substantially, including since the last edition of this text. Steps have been taken in many countries to voluntarily and/or regulatorily reduce the use of Medically Important Antimicrobials (MIAs). The World Health Organization’s Critically Important Antimicrobials for Human Medicine (WHO CIA) list (WHO, 2017) defines MIAs that are used in human medicine and are divided into categories according to specific criteria (Table  34.1). The WHO List has recently been updated, WHO 2024; Table 23.2. Different

countries have referred to this list in defining their own lists of MIAs and definitions vary by country (Scott et al., 2019; Table 23.2). The significant variation in approach between different countries across the globe is beyond the scope of this chapter. We will attempt to highlight some major differences between Canada, the US, and the UK.

Growth Promotion Products previously used for growth promotion have included penicillins, tetracyclines, avoparcin, virginiamycin, bacitracin, flavomycin, and avilamycin. In the UK in 1969, the Swann Report concluded that “the administration of antibiotics to farm livestock, particularly at sub-­therapeutic levels, poses

Table 34.1  Medically Important Antimicrobials –­WHO categories in relation to Canada, US, and UK categories and the relevant poultry drugs implicated. WHOa

Canadab

Category I, Very High Critically Important and Importance –­ceftiofur, highest priority –­ceftiofur, enrofloxacin macrolides (erythromycin, tilmicosin, tylosin) polymyxins, quinolones, and fluoroquinolones (enrofloxacin)   Critically Important and high priority –­ aminoglycosides (gentamicin, apramycin, neomycin, streptomycin), penicillins (aminopenicillins; amoxicillin, ampicillin) Highly Important –­ lincosamides (lincomycin), antistaphylococcal penicillins –­ narrow spectrum (penicillin G), streptogramins (virginiamycin), sulfonamides and dihydrofolate reductase inhibitors and combos (sulfas, pyrimethamine, trimethoprim), tetracyclines (chlortetracycline, oxytetracycline, tetracycline), amphenicols (florfenicol)

Category II, High Importance –­ virginiamycin, penicillins, tylosin, aminoglycosides (except topical agents) –­ gentamicin, lincosamides and extra-­label use of trimethoprim-­ sulfadiazole

USc,e

UKd

Critically Important –­ ceftiofur, fluoroquinolones, erythromycin, trimethoprim sulfa

Category A: Avoid: not authorized for use in veterinary medicine Category B: Restricted: ceftiofur, fluoroquinolones

Highly Important –­ penicillin G, gentamicin, neomycin, tetracycline, chlortetracycline, oxytetracycline, virginiamycin

Category C: Caution: aminoglycosides (except spectinomycin), aminopenicillins in combo with beta-­ lactamase inhibitors (amoxicillin-­clavulanic acid, ampicillin-­ sublactam), lincosamides, pleuromutilins, marcolides (Continued )

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­Antimicrobial Drug Use in the Poultry Industries of Canada, United States, and U  699

Antimicrobial Therapy in Poultry

Table 34.1  (Continued) WHOa

Canadab

USc,e

Important –­aminocyclitols (spectinomycin), polypeptides (bacitracin), pleuromutilins (tiamulin)

Important –­ Category III, Medium Importance –­ no poultry drugs bacitracin, sulfonamides, listed aminocyclitols (spectinomycin) and topical aminoglycosides (neomycin), aminoglycosides not used in human medicine: apramycin, tetracyclines

Antimicrobial classes currently not used in humans –­aminocoumarins (noboviocin), arsenicals (roxarsone, nitarsone), orthosomycins (avilamycin), phosphoglycolipids (bambermycin, flavomycin), ionophores (lasalocid, monensin, narasin, salinomycin)

Category IV, Low Importance –­ flavophospholipols (bambermycin), ionophores   Uncategorized –­ avilamycin

Not listed as medically important –­ bacitracin, avilamycin, ionophores

UKd

Category D: Prudence: aminopenicillins without beta-­lactamase inhibitors (amoxicillin, ampicillin), tetracyclines, narrow-­ spectrum penicillins, spectinomycin, antistaphylococcal penicillins, potentiated sulfas (trimethoprim sulfa) Not categorized as medicines: ionophores

a

 WHO (2018) Critically Important Antimicrobials for Human Medicine, 6th Revision, 2018. https://apps.who.int/ iris/bitstream/handle/10665/312266/9789241515528-­eng.pdf?ua=1 b  Category III reduction –­everything you need to know, Chicken Farmers of Canada (no date). www. chickenfarmers.ca/category-­3-­reduction/ c  CVM GFI #152 Evaluating the Safety of Antimicrobial New Animal Drugs with Regard to Their Microbiological Effects on Bacteria of Human Health Concern. FDA (no date). www.fda.gov/regulatory-­information/search-­fda-­ guidance-­documents/ cvm-­gfi-­152-­evaluating-­safety-­antimicrobial-­new-­animal-­drugs-­regard-­their-­microbiological-­effects d  British Veterinary Poultry Association. Antimicrobials Guidelines. 2021. www.ema.europa.eu/docs/en_GB/ document_library/Other/2014/07/WC500170253.pdf e  Scott HM, et al. 2019. Critically important antibiotics: criteria and approaches for measuring and reducing their use in food animal agriculture. Ann NY Acad Sci 1441:8.

certain hazards to human and animal health.” In particular, it led to resistance in enteric bacteria of animal origin (Swann et al., 1969). The outcome of this report was a recommendation that some antimicrobials that were important for therapeutic use (e.g., penicillins and tetracyclines) should no longer be used in feed for growth promotion. Following the ban on all growth-­promoting antimicrobials in Sweden in 1986, and the ban on avoparcin and virginiamycin in Denmark in 1995 and 1998, the EU banned the use of avoparcin in 1997 and four other antimicrobials used for growth promotion in 1999, on the basis of

the “Precautionary Principle.” These four antibiotics were bacitracin (a polypeptide), spiramycin and tylosin (macrolides), and virginiamycin (a streptogramin combination).  Two remaining antimicrobial growth promoters licensed in the EU, flavomycin and avilamycin, were banned from use in 2006 (Dibner and Richards, 2005; Castanon, 2007). In the US, Food and Drug Administration (FDA) guidelines came into effect in 2017, preventing use of antimicrobials considered important for human medicine for growth ­promotion. This has effectively eliminated use  of virginiamycin. According to the US

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700

classification of MIAs, bacitracin is the only remaining product with a growth promotion claim. Despite this claim, to the authors’ knowledge bacitracin is now seldom used for growth promotion and its main use is for disease prevention (e.g., necrotic enteritis). In Canada, a regulatory foundation for changes to the use of MIAs began in 2014 with the publication of the Federal Framework for Action. In early 2015 the Federal Action Plan on AMR and AMU in Canada was released, building on the Framework for Action (Government of Canada,  2015). Later that year, Health Canada and the pharmaceutical industry began work to phase out growth promotion claims on antimicrobial labels. Currently, all growth promotion doses and label claims have been removed but preventive and therapeutic label claims for virginiamycin and bacitracin remain. This initial wave of changes had minimal impact on feed medication for poultry as producers were not using the implicated products for growth promotion. Additional restriction on use of products formerly labeled for growth promotion has largely targeted preventive use and has been voluntary and/or in response to customer and/or consumer pressure and the need to fill a market 2019

3%

25%

8%

2018

24%

11%

2017

15%

Preventive Use Since January 2022, the EU banned antimicrobials for prophylactic treatment for all animals, other than in exceptional cases limited only to individual animals or restricted numbers of animals. This effectively prohibits prophylactic use in commercial poultry as the numbers of birds in a commercial barn are seldom restricted. The terms prophylaxis and metaphylaxis are often poorly defined and easily confused. A short-­ term targeted treatment in response to epidemiological evidence could appear similar to widespread, routine application of the same treatment to compensate for poor husbandry, but the former may be considered justifiable whereas the latter would not. The use of certain antimicrobials for disease prevention in poultry in the US and Canada is currently still permitted. It should be noted, however, that the market for poultry products raised without the use of antimicrobials for prevention or treatment has moved the respective industries away from antimicrobial use (Figure 34.1). Full spectrum (program includes virginiamycin and/or other medications deemed medically important to humans by FDA)

58%

18%

32%

30%

2016

demand for meat “raised without the use of antibiotics.”

51%

16%

24%

Reduced use (program includes avilamycin, bacitracin and/or bambermycin)

40%

25%

lonophores only (WHO guidelines)

21%

No antibiotics ever 33%

2015

19%

35%

13%

†

† 2015 numbers extrapolated from feed tonnage.

0

10

20

30

40 50 60 PERCENT

70

80

90

100

*Data by year may not total 100% due to rounding. Source: Rennier Associates, Inc.

Figure 34.1  Graphic representation of the percentage of broiler chickens raised by antimicrobial use marketing programs (full spectrum, reduced use, ionophores only, or no antibiotics ever) in the USA.

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­Antimicrobial Drug Use in the Poultry Industries of Canada, United States, and U  701

Antimicrobial Therapy in Poultry

A prohibition on preventive use in the United States was implemented in January 2012  with a ban on the extra-­label use of cephalosporins. This ban particularly targeted the extra-­label use of ceftiofur when administered in ovo for metaphylaxis in cases of known or anticipated E. coli challenge (FDA, 2012a,b). Such use was associated with E. coli isolations from poultry carcasses containing genes that rendered them resistant to third-­generation cephalosporins used in humans. While extra-­label use is prohibited, ceftiofur remains approved for subcutaneous administration to day-­old chicks and turkey poults in the USA. Since broiler chicks in the USA are generally vaccinated against Marek disease in ovo, and not handled for vaccination individually as day-­old chicks, the use of ceftiofur in broiler chicks has effectively been discontinued. In Canada, voluntary industry-­led changes in antimicrobial use (AMU) unfolded alongside the regulatory changes that impacted growth-­promoting use. The Chicken Farmers of Canada (CFC), a national farm animal group overseeing national programs of ­on-­farm food safety and quality assurance, voluntarily made changes to permitted use of MIAs. In 2014, the preventive use of Category I antimicrobials was eliminated. In 2018 the preventive use of Category II antimicrobials was also prohibited. Together, these measures effectively eliminated the use of ceftiofur, gentamicin or lincomycin-­spectinomycin as in ovo preventives for hatchery use and the use of virginiamycin in feed for prevention of necrotic enteritis. The CFC also plans to eliminate the preventive use of Category III antimicrobials, though the initial 2020 timeline for this has lapsed and an updated timeline has yet to be announced. Elimination of preventive use in this category will effectively eliminate in-­feed use of bacitracin for the prevention of necrotic enteritis. Thus far, use of category IV antimicrobials (bambermycin and ionophores) and uncategorized products (avilamycin) will ­continue to be permitted. The CFC AMU

reduction strategy will also continue to permit use of antimicrobials for treatment of disease, the use of ionophores to prevent coccidiosis, and the use of chemical coccidiostats. At the turn of the century in Europe, there was growing concern regarding the development of antimicrobial resistance in human medicines and the use of antimicrobials in livestock industries was blamed for this increase. In some EU countries, legislation was introduced to reduce antimicrobial use in the livestock sectors. In the UK, the poultry industry recognized these consumer and legislator concerns and took the initiative in responsible use of antimicrobials, following WHO guidelines on highest priority critically important antimicrobials by banning the use of some antimicrobial classes (e.g., third-­ and fourth-­ generation cephalosporins, glycopeptides, and ketolides) and restricting the use of other classes (e.g., fluroquinolones and macrolides), thus preempting the need for legislation. Furthermore, when the O’Neill report on antimicrobial resistance (O’Neill,  2016) recommended restrictions on the amount of antimicrobials used in agriculture to 50 mg/kg, the UK poultry industry was already well below this level. Following the 2016 finding of the Mcr-­1 resistance gene to colistin in China (Liu et  al.,  2016), the UK poultry meat and layer industries voluntarily restricted colistin use. Ionophores, which are used to control coccidiosis in poultry, are not considered to be MIAs but because of their activity against some Gram-­positive bacteria, they are considered antimicrobials in the US and Canada; for this reason, they cannot be used in “antimicrobial-­free” production. This is unfortunate because the ionophores are very effective at controlling coccidiosis and preventing subsequent disease in poultry. There has been debate about whether the use of ionophores will drive antimicrobial resistance in bacteria important for human health (Nilsson et  al.,  2012,  2016,  2019). However, there is scant evidence that this is

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the case (Naemi et  al.,  2020). In EU and UK, ionophore coccidiostats are approved as feed additives and not as veterinary medicines and therefore can be added to feed without veterinary prescription. The UK has not to date developed “antimicrobial-­free” or “no antimicrobial ever” labels and so ionophores are still widely used in poultry diets for coccidiosis control. There is good evidence that the removal of ionophore coccidiostats from poultry diets has both financial and environmental impact because of the reduction in feed efficiency (Parker et al., 2021). It is worth noting that based on this key difference in classifying the ionophores, the approach that each country has been able to take in terms of reducing antimicrobial use, specifically for the prevention and treatment of enteric disease, namely necrotic enteritis, has been impacted.

Therapeutic Use Therapeutic antimicrobial uses have not as yet been banned in Europe but have been targeted in the US. Two fluoroquinolones that were approved as therapeutic antimicrobials for the control of colibacillosis in the US were banned in 2005 from use in poultry but not in cattle. The primary reason for this ban was to allay concerns regarding rising fluoroquinolone resistance rates in human cases of campylobacteriosis (FDA, 2005). In Canada, there have been no bans on antimicrobials for therapeutic use. Extra-­label drug use (ELDU) in Canada is not codified like the US; Canadian veterinarians have the privilege of prescribing ELDU as long as the drug is not banned by federal law for use in food animals. Health Canada defines ELDU as “the use or intended use of a drug approved by Health Canada in an animal in a manner not in accordance with the label or package insert” (Health Canada,  2014). While not recommended by Health Canada, it is possible for Canadian veterinarians to use the cattle injectable enrofloxacin formulation in poultry.

In the United States, veterinarians may have some flexibility with how antimicrobials are administered via the water. ELDU is permitted with the exception of antimicrobials that have been banned from use for poultry (such as the fluoroquinolones) or in food production animals in general (such as chloramphenicol) so long as a prescription is utilized. Deviations from the label for dose and duration may be allowed, but the veterinarian is expected to take adequate precautions to ensure that those deviations do not cause violative residues. In most cases, antimicrobials that are added to the feed must be accompanied by a veterinary feed directive (VFD). There are a few exceptions to this rule where the antimicrobials are not considered to have any human significance (e.g., ionophores, bacitracin methylene disalicylate, bambermycin, anthelmintics). While US veterinarians have some degree of flexibility with water additive antimicrobials, they have no flexibility regarding dose, duration, species, or indications for use with antimicrobials that are added to feed. Such antimicrobials, whether a VFD is required or not, have to be used with no deviations from the label. In the UK, veterinarians can prescribe antimicrobials extra-­label, but a standard minimum withdrawal period of seven days for eggs and 28 days for meat is generally applied. As these restrictions are often impractical or financially prohibitive, veterinarians rarely elect to use these products extra-­label.

­ onsequences of Antimicrobial C Bans The consequences of the ban on enrofloxacin and sarafloxacin use in poultry in the United States has been studied but not widely reported. The bans have removed effective treatments of bacterial disease from the poultry veterinarian’s arsenal and treatment options for Gram-­ negative organisms are lacking. Veterinarians have valid concerns that antimicrobial bans cause serious animal welfare issues in the face

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­Consequences of Antimicrobial Ban  703

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of an untreatable disease outbreak. In a somewhat extreme example, the potentially difficult decision regarding early slaughter of entire flocks may need to be considered should no approved therapy exist. The outcomes or concerns relating to bans on antimicrobial use for growth promotion and/or prevention are better documented and understood. The EU bans on growth-­promoting antimicrobials initially led to significant animal and human health concerns. Clostridial related diseases, including necrotic enteritis and cholangiohepatitis, initially increased and required more therapeutic antimicrobial treatments. However, poultry diets were refined and these clostridial diseases are now much less common in the EU and UK poultry industries. However, the dietary changes and interventions required to mitigate the removal of antibiotic growth promoters (AGPs) have impacted the feed costs for poultry (Wierup, 2001; Casewell et al., 2003; Dibner and Richards, 2005; Grave et al., 2006). In the EU, ionophores are still used for the control of coccidiosis and these ionophores have some moderating effect on clostridial disease. Similar observations were made in the US when poultry-­producing companies voluntarily removed in-­feed antimicrobials in order to produce an “antibiotic-­free” product for specific markets (Smith, 2011). Of concern for human health in the EU was the increased use of therapeutic antimicrobials in poultry to treat clinical disease, primarily necrotic enteritis but also other forms of infectious enteritis (Casewell et  al.,  2003; Grave et al., 2006). Unlike the majority of in-­feed antimicrobials approved for growth promotion, many of the antimicrobials used for therapy were related to or were the same as those used in human medicine (Casewell et  al.,  2003; Phillips et  al.,  2004; Phillips,  2007). Another potential unintended consequence for human health was the importance of the AGPs in maintaining intestinal integrity. This is especially important during slaughter and processing of birds as the normal poultry intestinal tract can contain zoonotic pathogens. Inflammation and

disease of the intestinal tract weaken the gut wall and increase the risk of intestinal breakage and the potential for greater contamination of the final product (Russell,  2003). While raw meat is not sterile, good intestinal health is vital in reducing the bacterial load on poultry products provided to the consumer; alternative strategies for gut health management are therefore important in the absence of AGPs. The use of antimicrobials considered critically important in human medicine for therapy of food animals will continue to be scrutinized. The benefits of these products for health, both human and animal, also need to be considered. Consumer and retailer pressure in some regions has resulted in removal of these antimicrobials from broiler diets and in some cases resulted in a ban on their use through other routes (e.g., in water or parenterally). Producers supplying export markets with poultry products may be required to discontinue use of antimicrobials if they wish to continue to supply certain markets where bans are in place, or where consumers demand that antimicrobial use is discontinued (Dibner and Richards, 2005). The general trend for the future is reduced and more responsible antimicrobial use. This ultimately means that when the question of whether or not to treat a flock is raised for the poultry veterinarian, there are more factors than ever to consider in the decision-­making process: effectiveness against the disease agent, pharmacokinetics and pharmacodynamics of the medication, withdrawal times, pathology and physiology, economics, animal welfare, impact on food-­borne pathogens, and impact on the ability to market the final product.

­ actors Influencing Antimicrobial F Administration in the Poultry Industry Husbandry and Economics Under current husbandry conditions in the poultry industry, segregation and medication of individual sick birds are not feasible. The

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low economic value of the individual bird makes it cost-­prohibitive to individually dose each bird in a house, which eliminates parenteral administration of antimicrobials. An additional argument against parenteral administration is that the stress on birds when individually handled can result in more rapid progression of the disease. Since sick birds continue to drink, therapeutic antimicrobials labeled for use in drinking water are most often used. Antimicrobial interventions must be administered early in the course of disease. Bacterial infections in birds tend to progress rapidly, and there is frequently a very short time from initial infection to death. In addition, birds are adept at producing inflammatory responses, but poor at resolving the products of such responses. As prey species, poultry tend to hide clinical signs of disease. Spotting the prodromal, subtle signs of infection in individuals in a flock of 10,000–­100,000 birds is both important and challenging. Metaphylactic treatment of all individuals in contact and at high risk of exposure (i.e., the entire flock) is the only practical approach to disease outbreaks in large flocks. Thus, the decision to treat a “sick flock” of birds means veterinarians will be administering antimicrobials not only to the sick birds but also to all birds in that flock that have been or will be exposed to the disease agent. In making this decision to treat the “sick flock,” the poultry veterinarian must also decide, based on clinical judgment, whether the “flock” to be treated includes the entire farm or only the house containing the most clinically affected birds. A rapidly spreading disease may necessitate targeted preventive treatment of all houses on the farm. When considering treatment in the drinking water or feed, the poultry veterinarian must also take into account lighting schedules and feed programs, which can strongly influence both feed and water consumption. Laying hens begin to eat when the lights are turned on and then consume water after eating. Broiler chickens and turkeys are reared on a variety of

different lighting programs which impact feeding and drinking patterns. The majority of water intake in replacement breeders under feed restriction occurs for only a few hours after feeding.

Production Type/Bird Type Within the poultry industry, integrated or not, there is a continuum or flow of birds and bird types. For example, in the chain of production of a commercial broiler (meat-­type chicken), the parents of that bird are hatched at a hatchery, reared and brought into egg production. Eggs collected from that flock will return to a hatchery for incubation, to hatch into broiler chickens that will be raised and ultimately slaughtered for meat. Prevention of disease at all levels within this continuum is extremely important; otherwise, there can be serious downstream consequences. The consequences are also affected by the type of bird and the point in this chain of production at which the disease occurs. For example, disease in a flock producing hatching eggs can not only have a severe impact on overall health and productivity of that flock, but some bacterial diseases such as Mycoplasma can be vertically transmitted to offspring and, if not treated, the spread of disease is amplified (Bradbury, 2005). Conversely, treatment of flocks in egg production can impact the production or quality of the eggs depending on the antimicrobial used. The use of tetracyclines in flocks in egg production can adversely affect the amount of calcium available to the hen for eggshell formation as these medications are known to chelate with divalent cations. Poor shell quality in the hatching eggs in turn increases the risk of bacterial contamination of the egg, which when placed in an incubator with hatching eggs from another flock increases the risk of bacterial disease for all embryos in the incubator. Disease in a broiler flock, where there is only a short time between hatching and slaughter,

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­Factors Influencing Antimicrobial Administration in the Poultry Industr  705

Antimicrobial Therapy in Poultry

can be challenging from the perspective of medication withdrawal times. When flocks approach market age, the number of antimicrobial treatment options available for ­different diseases diminishes because their administration would result in violative antimicrobial residues present at the time of slaughter. Antimicrobial residues would result in condemnation of slaughtered birds. The other option to comply with drug withdrawal times would be to postpone slaughter but this results in other challenges such as oversized birds that do not meet slaughterhouse and customer specifications, or overstocking in the house/ barn with welfare issues as the birds will continue to grow. Oversize birds are a further challenge for the processor as the slaughterhouse equipment may not be able to cope with larger birds, creating potential welfare concerns for humane slaughter and carcass damage and increased risk of microbial contamination of the final poultry product.

Feed and Water Consumption Flock treatment is the method of choice, with drinking water and feed as the primary means of delivering antimicrobials to commercial poultry (Vermeulen et  al.,  2002). When birds become sick, there is a significant reduction in consumption of both feed and drinking water. The decline in drinking water consumption is usually less than that of feed. Additionally, there may be a large amount of nonmedicated feed already on the farm which delays the onset of treatment if medicating via the feed. Therefore, the route of choice for administering antimicrobials in the early stages of a disease is usually by the drinking water. If therapy lasts more than 5–­7 days, the veterinarian may choose to have the antimicrobial drug added to the birds’ feed if an approved feed-­grade product is available. This change to feed can be based upon the flock beginning to recover and eating more. In general, feed-­grade antimicrobials are less expensive than water-­soluble ones and are preferred when a suitable

drug with a clinically effective inclusion rate is available. Another consideration when selecting the appropriate antimicrobial is the ambient temperature since poultry have limited means of eliminating body heat. In large part, they cool themselves by drinking water; therefore, water consumption increases significantly as the ambient temperature increases. This affects dosage calculation and makes it possible for birds to be overdosed when a drug is administered in drinking water. This is especially important with the use of sulfonamides, as the therapeutic dose is close to the concentration that can cause toxicity (Goren et al., 1984).

Pathology and Disease Etiology Escherichia coli is the leading cause of bacterial disease-­related economic loss for the poultry industry throughout the world (Barnes et al., 2008). In most instances, E. coli infections are secondary infections following a primary viral or environmental insult (Glisson,  1998). Therefore, therapeutic antimicrobials in commercial poultry are almost always used to relieve the suffering of sick birds, control morbidity and mortality, and minimize the financial impact of the disease on bird performance until the primary insult can be identified and controlled or eliminated. The use of therapeutic antimicrobials also decreases the public health risk associated with slaughtering birds from sick flocks. Poultry that are sick eat greater amounts of bedding material (litter), resulting in higher rates of Salmonella and Campylobacter spp. in their intestinal tracts (Corrier et al., 1999). Birds from flocks having higher air sacculitis condemnation had higher levels of E. coli and Campylobacter contamination (Bull et al., 2008; Russell, 2003). The choice of therapeutic antimicrobials available to treat infections caused by E. coli is limited (Glisson et  al.,  2004). Tetracyclines, aminoglycosides, sulfonamides, and penicillins are registered for use in poultry in many countries. The average resistance rates in

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E. coli to representatives of these antimicrobials are >40% in all countries, with the exception of ampicillin in the US. The resistance rates to fluoroquinolones in the US, where fluoroquinolones are not used in poultry, are 40% in Brazil, China, and the EU, where use of fluoroquinolones is approved (Roth et al., 2019).

Pharmacology The success of antimicrobial therapy depends upon many interacting factors, including pharmacodynamics (drug interaction with the organism/bird), pharmacokinetics (drug absorption, distribution, excretion), and the components of the host immune system (see Chapters  4 and  5). The activity of an antimicrobial agent against a particular microbe is often expressed as the minimal inhibitory concentration (MIC). When interpreting antimicrobial susceptibility information, the poultry veterinarian must keep in mind that this is an in vitro test that does not take into consideration whether the drug can reach the site of infection or whether the drug is bacteriostatic or bactericidal. It should also be remembered that the MIC is usually performed by the laboratory on only one isolate and, as was previously stated, many infections of poultry are secondary, so a “sick flock” is often affected by multiple isolates that can have a wide range of MICs. Also, MIC breakpoint criteria in veterinary medicine are not uniform worldwide and are often based on standards for human medicine (see Chapter  2). Additionally, pharmacokinetic data determined in mammals are not always applicable to poultry because birds have higher body temperatures, higher metabolic rates, and shorter alimentary tracts, which often results in shorter elimination half-­ life times for medications. This frequently leaves the poultry veterinarian with an antimicrobial therapy decision in part based on clinical judgment from previous cases as well as antimicrobial susceptibility

data from the case investigation. The primary criterion for measuring success of treatment under poultry industry conditions is reduction of morbidity and mortality. Other important parameters include return to regular water and feed consumption, normal growth rate, and normal egg production.

­ ractical Antimicrobial Drug P Application under Commercial Poultry Conditions Since commercial poultry are food animals, the choice of antimicrobials to treat the most common bacterial diseases is limited. The decision to treat may need to be made prior to the results of culture and susceptibility testing. Oral treatment of poultry requires that the drug be stable and be uniformly distributed in either feed or water. When a feed-­ based antimicrobial is prescribed, the time required for the medicated feed to be manufactured, transported, and delivered through the feeding system at the farm must be considered. Administering the antimicrobial in the drinking water allows for more rapid treatment. The most accurate method is to calculate the dose based upon the total body weight of birds in the house, and then include that dose in the volume of water or feed the birds are expected to consume during each dosing interval. The drinking water medication can either be administered by adding the medication to the bulk tank or through a water proportioner. If giving via the bulk tank then the amount of water in the bulk tank may need to be adjusted to match the water consumption for the duration of the dosage period of the birds being treated. A water proportioner is a device that meters the antimicrobial from a highly concentrated stock solution into the drinking water to achieve the appropriate concentration. If delivery of the antimicrobial through a stock solution and proportioner is expected to occur over a 24-­hour period, care must be

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­Practical Antimicrobial Drug Application under Commercial Poultry Condition  707

Antimicrobial Therapy in Poultry

taken to ensure that delivery continues through the night and does not run out. Some veterinarians administer antimicrobials to poultry based solely on concentration of the active ingredient in the drinking water, but this does not always take account of physiological, pathological, and husbandry conditions and can lead to highly inaccurate dosing. Dosing based on water consumption can result in a toxic overdose if the ambient temperature increases, or the amount of drug consumed may result in subtherapeutic concentrations if the ambient temperature declines. Additionally, younger birds consume more water daily per unit of body weight than older birds, so dosing at a constant rate per volume of drinking water can result in overdosing of young chicks or underdosing of older birds. In addition, hens producing eggs will drink more per unit of weight than nonlaying hens or roosters. In situations where the birds’ water consumption is limited, a short, intensive treatment with certain antimicrobials may be administered as a pulse dose (Charleston et al., 1998). This method should only be used with bactericidal antimicrobials and those with a wide margin of safety. Pulse dosing requires that all of the medication to be administered for a 24-­hour period is mixed into the water the birds will consume in, for example, six hours.

­ harmacological Characteristics P of Poultry Antimicrobials Avilamycin Avilamycin is an oligosaccharide antimicrobial of the orthosomycin group that is produced by fermentation of Streptomyces viridochromogenes. It is primarily active against Gram-­positive bacteria through inhibition of protein synthesis. Avilamycin has been marketed in various locations around the world since the late 1980s with a claim as an AGP. Risk assessment concluded that this product presents no risk to

human health (Shryock and Belanger,  2004). As antimicrobial use for growth promotion was falling from favor and discontinued around the world, a label claim in the US and Canada for this antimicrobial for the prevention and/or reduction of mortality caused by necrotic enteritis associated with Clostridium perfringens was approved in 2013 and 2014 respectively. Being one of the first new antimicrobials approved for use in poultry in the US and Canada in several years, it was approved for use with a VFD or a veterinary prescription respectively, even though it is not considered an antimicrobial of human health significance. Along with the approval for use in poultry in the US and Canada came several restrictions for dose, consecutive days or duration of treatment, and ages of birds to which this product can be fed. In many locations outside the US and Canada, registration as a therapeutic option was not pursued, and product use was discontinued as antimicrobial use for growth promotion ceased. Avilamycin was only licensed as an AGP in the EU and was removed from the market in 2006.

Beta-­lactams (Cephalosporins and Penicillins) Phenoxymethylpenicillin (penicillin V) is available in the UK for treatment of necrotic enteritis. In other countries, only human formulations are available. Despite years of use, penicillin G (benzylpenicillin) is still an effective antimicrobial for Gram-­positive bacterial infections in poultry. This drug is particularly important for the therapy of clostridial infections causing necrotic enteritis (Gadbois et  al.,  2008). One Gram-­negative bacterium routinely treated with penicillin is Pasteurella multocida. Historical publications indicated the susceptibility of this pathogen to penicillin and supported the selection of this medication for the treatment of pasteurellosis or fowl cholera (Huang et  al.,  2009; Sellyei et  al.,  2009). However, Jones (2013) reported mixed penicillin susceptibility in P. multocida, with only

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54% of chicken isolates fully susceptible. In a recent publication by the American Veterinary Medical Association Committee on Antimicrobials (AVMA, 2020) P. multocida is identified as an animal pathogen of heightened resistance concern in chickens, turkeys, and cattle. Penicillin G is formulated for both drinking water and feed administration, with water administration being the preferred initial route. The broader spectrum beta-­lactams, such as ampicillin and amoxicillin, are more effective for Gram-­negative infections such as E. coli air sacculitis; however, there are limited data published on the use and clinical efficacy of these medications in poultry species. The reportedly short elimination half-­lives of both amoxicillin and ampicillin when administered to poultry species are a desirable characteristic from the perspective of managing withdrawal times in broiler flocks, as this can be a factor limiting the treatment options (El-­Sooud et  al.,  2004; Fernandez-­Varon et  al.,  2006). One potential factor that may limit the use of these products is the reportedly poor stability of amoxicillin in aqueous solution (Jerzsele and Nagy, 2009). No products are currently available for use in poultry in the US, but water-­soluble amoxicillin trihydrate powder(s) are available in Canada and the UK. The UK 2021 VARRS report states that amoxicillin accounted for 71% of the antimicrobials administered to meat poultry in the UK in 2020. Concerns regarding increasing bacterial resistance to amoxicillin and ampicillin have prompted some EU researchers to investigate the pharmacokinetics of these antimicrobials in combination with beta-­lactamase inhibitors clavulanic acid and sulbactam in poultry (Fernandez-­Varon et al., 2006; Jerzsele et al., 2010). The only cephalosporin used in poultry production is the third-­generation ceftiofur. Since it has poor oral absorption, ceftiofur is only approved for subcutaneous injection in day-­ old chicks (US) and poults (US and Canada). It was historically administered along with Marek disease vaccine SC to day-­old chicks

(Kinney and Robles, 1994), or in an extra-­label fashion by in ovo injection at approximately 18  days of incubation. The extra-­label in ovo administration of ceftiofur in the US has been banned (FDA, 2012b) and therefore the use of this antimicrobial in broilers in the US has effectively been discontinued. Ceftiofur was never licensed for use in poultry in the UK, but was used extra-­label along with Marek disease vaccine in breeders and layers. Marek vaccines are not routinely given to standard broilers in the UK. However, following increased identification of extended-­spectrum beta-­lactamase (ESBL) E. coli and Salmonella in poultry and poultry products across the EU, this extra-­label use was voluntarily withdrawn. Following the voluntary withdrawal from use in UK poultry, levels of ESBL E. coli isolated from poultry and poultry products dropped significantly (Parker and Elvidge,  2020; Randall et  al.,  2021). The need to use a third-­generation cephalosporin should be assessed against the risk of selecting for resistance to this important group of drugs, including the danger of selection of multidrug-­ resistant Salmonella carrying the blaCMY2 resistance gene, since such isolates would also be resistant to ceftriaxone, a drug used to treat salmonellosis in people (Carson et al., 2019).

Streptogramins The only poultry-­labeled streptogramin antimicrobial in the US and Canada is virginiamycin. Virginiamycin has a Gram-­positive spectrum of action and prevents bacterial protein synthesis by binding to bacterial ribosomes. This product has been used for growth promotion as well as prevention of necrotic enteritis in poultry flocks in the US but was only ever licensed for growth promotion in Europe. Additionally, virginiamycin is useful for the prevention of ulcerative enteritis due to Clostridium colinum in quail. The streptogramin combination quinipristin-­dalfoprisin (QD) is used in human medicine for the treatment of vancomycin-­resistant Enterococcus faecium infections. Avoparcin is a glycopeptide

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­Pharmacological Characteristics of Poultry Antimicrobial  709

Antimicrobial Therapy in Poultry

antibiotic which was used as an AGP in poultry feed in Europe. It has a chemical structure similar to the glycopeptide drug vancomycin used in human medicine. Despite risk assessments indicating low to no risk to human health (Cox and Popken,  2004), the isolation of QD-­ resistant E. faecium from poultry in the US, categorization as a MIA, and market growth for “raised without antibiotics” poultry products all fueled the EU ban on avoparcin in 1997.

Polypeptides Bacitracin is the only poultry-­approved polypeptide antimicrobial. Its effect is local, as it is essentially not absorbed when administered orally. Bacitracin is a very effective antimicrobial for prevention and treatment of Gram-­ positive enteric infections such as necrotic enteritis caused by Clostridium perfringens (Hofacre et  al.,  1998). It is available in both drinking water and feed additive formulations in the US, with the feed-­grade form commonly used as a preventive for necrotic enteritis. Since it is not considered a medically important antimicrobial in humans, it can currently be used in feed in the US without a VFD or in Canada with a veterinary prescription. Consumer pressure for “raised without antibiotics” products, however, has greatly reduced use of this antimicrobial in the poultry industry. Bacitracin was only licensed as an AGP in Europe and was withdrawn from the market in 1999.

Aminoglycosides and  Aminocyclitols Three aminoglycosides are used in poultry: gentamicin, streptomycin, and neomycin. Because aminoglycosides are poorly absorbed from the gastrointestinal tract when administered orally, their primary usage in poultry has been by subcutaneous injection. Gentamicin was the most widely used aminoglycoside in the US, and it was used primarily as a subcutaneous injection in day-­old

birds or in ovo injection in chickens or turkeys (McCapes et  al.,  1976; Vernimb et al., 1977). With increased consumer pressure for “raised without antibiotics” products, far less gentamicin is used by the poultry industry today. A dose of 5  mg/kg body weight in broiler chickens has been reported to be a suitable therapeutic dose when administered intravenously, intramuscularly or subcutaneously. Subcutaneous administration has 100% bioavailability while oral administration has an absolute bioavailability of zero unless enteritis is present (Abu-­Basha et al., 2007a). Because gentamicin is a highly basic compound, it can damage cell-­associated Marek disease vaccine if used in ovo at too high a dose (greater than 0.2 mg/chick) or improperly mixed with the vaccine (Kinney and Robles, 1994). Streptomycin is partially absorbed from the intestines and therefore can be considered for use to treat systemic E. coli infections. Neomycin is commonly used to treat enteric infections, administered in either the feed or water. Interestingly, despite poor absorption from the gastrointestinal tract, there are reports that administration of neomycin has resulted in clinical efficacy in the treatment of colibacillosis in poultry, likely due to a local effect (Marrett et al., 2000). Spectinomycin is a poultry-­approved aminocyclitol. It is a relatively safe antimicrobial in poultry that when administered once orally, at doses of 50–­100 mg/kg body weight, has limited absorption from the gastrointestinal tract with bioavailability reported as 11.8% and 26.4%, respectively (Abu-­Basha et  al.,  2007b). Similar to neomycin, spectinomycin has been reported to be highly efficacious for E. coli infections when administered in the drinking water (Goren et al., 1984). This antimicrobial is available commercially alone or in combination with lincomycin. This combination has been reported as efficacious in controlling early chick mortality associated with E. coli and Staphylococcus aureus when administered subcutaneously (Hamdy et  al.,  1979) and has

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been used as an alternative to gentamicin or ceftiofur for prophylaxis in some hatcheries. However, rapid development of resistance and higher cost limit the use of spectinomycin. The combination product with lincomycin was historically extensively used prophylactically in the UK for chick quality issues but use has reduced significantly with the reduction in prophylactic administration of antimicrobials. Apramycin is another aminocyclitol approved for use in poultry in some European countries and can be used in an extra-­label fashion where permitted, such as in the US and Canada. Consistent with the observations of other antimicrobials in this class, oral absorption is poor (Afifi and Ramadan, 1997). There are, however, reports that oral administration of apramycin for treatment of E. coli infections has been associated with a clinical response (reduced mortality, improved final body weight, and feed conversion) and reduced intestinal colonization by E. coli (Cracknell et al., 1986; Leitner et al., 2001).

Macrolides, Pleuromutilins and Lincosamides The macrolides occasionally used in poultry include erythromycin, tylosin, tiamulin, tilmicosin, and tylvalosin. While use of these antimicrobials may not be permitted in all countries, all have been available in formulations for administration in either drinking water or feed. Erythromycin was most frequently used in poultry to treat Staphylococcus aureus arthritis. This product is not currently manufactured or available in Canada or the US. Tylosin has been one of the most effective antimicrobials to treat mycoplasma infections in laying hens to restore egg production, reduce transovarial transmission, and reduce clinical signs (Bradbury et al., 1994; Kleven, 2008). The macrolides are only bacteriostatic, which may be one reason why their use will not entirely eliminate Mycoplasma spp. infections from a flock and thus treatment is not considered a long-­term solution. Clinical and subclinical

necrotic enteritis in poultry flocks can also be successfully treated with tylosin (Brennan et  al.,  2001a; Collier et  al.,  2003; Lanckriet et al., 2010). Tilmicosin, like the other antimicrobials in this family, has proven effective for control mycoplasma infections and has also been used to treat Pasteurella multocida and Ornithobacterium ­rhinotracheale bacterial infections (Jordan and Horrocks,  1996; Kempf et  al.,  1997; Jordan et  al.,  1999, Abu-­Basha et  al.,  2007c; Warner et  al.,  2009). Tylvalosin is  licensed in the UK  to  treat Mycoplasma ­gallisepticum and Ornithobacterium rhinotracheale. Tiamulin, a pleuromutilin, is available in water-­soluble formulations in the US, Canada, and UK. Of these countries, only in the UK does a poultry label indication exist; use in the US or Canada would be ELDU. This  drug has excellent efficacy against Mycoplasma spp. infections (Laber and Schutze,  1977) and has proven efficacious in the treatment of avian intestinal spirochetosis (Stephens and Hampson, 2002; Burch et al., 2006; Islam et al., 2009). With the exception of lasalocid, tiamulin is incompatible with the ionophore anticoccidials: monensin, salinomycin, narasin, maduramicin, and semduramicin. Administration of tiamulin with these ionophores results in clinical signs consistent with ionophore toxicity, and it seems to interfere with metabolism and excretion of these compounds (Islam et al., 2009). The only poultry-­approved lincosamide is lincomycin. Although it is absorbed following oral administration in feed or water, lincomycin is primarily used to treat enteric infections in poultry such as Clostridium perfringens-­ induced necrotic enteritis or intestinal spirochaetosis (Lanckriet et al., 2010; Stephens and Hampson, 2002). As previously described, this antimicrobial is also available in combination with spectinomycin and has been used effectively to control clinical signs and lesions associated with infections due to Mycoplasma species in poultry (Hamdy et al., 1976, 1982).

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­Pharmacological Characteristics of Poultry Antimicrobial  711

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Florfenicol The potential for fatal aplastic anemia in humans resulted in the prohibition of chloramphenicol in food-­producing animals throughout most of the world. However, the closely related antimicrobial florfenicol lacks the para-­nitro group associated with aplastic anemia in humans and is available for use in food-­producing animals for the treatment of susceptible Gram-­positive and/or Gram-­ negative infections. A water formulation of florfenicol was approved but is no longer marketed in Canada. Currently, there are no poultry-­labeled florfenicol products available in Canada, the US or the UK. A water-­soluble product for swine is available in the US and UK and there are several injectable products available for food animals in all 3 countries. Of publications on pharmacokinetics of florfenicol in poultry species, the oral bioavailability of this antimicrobial is relatively high; reports vary from 55.3% to 94% (Afifi and El-­ Sooud,  1997; Shen et  al.,  2002,  2003; Switala et  al.,  2007). Shen et  al. (2003) suggest that some of the discrepancy in the numbers reported may relate to timing of oral administration in relation to feeding as there have been reports of variable bioavailability between fasted and fed animals. Clinical response to treatment with florfenicol appears to be somewhat inconsistent. The multiplication of E. coli and Ornithobacterium rhinotracheale in a dual bacterial infection model, as well as the associated clinical signs, were significantly reduced in turkeys treated with 20  mg/kg body weight of florfenicol for five days (Marien et al., 2007). Clinical experience suggests that the use of florfenicol to treat E. coli infections in broiler chickens was not successful. There may be several reasons for this observation, including incompatibility with water administration via a proportioner when water hardness is >275  ppm (label ­directions of former approved product). Additionally, suitable therapeutic plasma concentrations for the targeted pathogen may not

be achieved as there is scant information published on the florfenicol MIC values for poultry pathogens such as E. coli. Several publications on the pharmacokinetics of florfenicol in poultry concur that plasma concentrations above 2 μg/ml for 11 hours can be achieved after a single dose of 30  mg/kg body weight florfenicol (Shen et al., 2003; Switala et al., 2007). As the activity of florfenicol is time dependent, it is important that plasma concentrations can be maintained above the MIC during treatment. In the absence of MIC data for poultry pathogens, many have looked to the MIC data for bacteria isolated from other species and have extrapolated these values to conclude that florfenicol should also be effective in poultry (Anadon et al., 2008). This may be inappropriate, as there are several publications documenting florfenicol MIC90 data against E. coli to be 8 μg/ml and 16 μg/ml or higher in turkeys and chickens respectively (Salmon and Watts,  2000; Dai et  al.,  2008). There has been one report of severe muscle degeneration in broiler chickens treated concurrently with both lasalocid and chloramphenicol (Perelman et  al.,  1986); there is no information as to whether or not this may occur with concurrent use of lasalocid and florfenicol. Finally, it is important to note that ELDU of injectable formulations of florfenicol for the treatment of disease in birds producing fertile eggs has been associated with embryo toxicity (Al-­Shahrani and Naidoo,  2015; Hu et al., 2020).

Tetracyclines The tetracyclines are the most widely used antimicrobials in poultry. This is largely due to their broad spectrum of activity, including Mycoplasma, Gram-­positive and Gram-­ negative bacteria, and wide margin of safety. This class of antimicrobials is also one of few with label claims permitting use in egg-laying chickens, at the specified dosage, with a zero-­ day egg withdrawal.

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The tetracyclines are available in formulations that can be administered in feed and/or water. This group of drugs are only slightly soluble in water above pH 7.0, so concurrent use of citric acid greatly enhances their absorption from the gastrointestinal tract (Clary et al., 1981). Tetracyclines are readily chelated in the intestine by divalent cations such as calcium or magnesium, resulting in reduced absorption. Therefore, the dosage of tetracyclines to laying hens on a high-­calcium diet should be increased. After administration is complete, it is recommended to include additional calcium in the diet to improve eggshell thickness and make up for calcium lost to tetracycline binding and intestinal excretion during therapy. For the same reason, tetracyclines are incompatible with concurrently administered oral electrolytes. The tetracyclines most commonly used in poultry are chlortetracycline, oxytetracycline, tetracycline, and doxycycline. It appears that any differences in clinical efficacy of these tetracyclines are primarily due to differences in solubility, absorption, drug distribution, or rate of excretion. Doxycycline is available for in-­ water medication in the UK. Although licensed for treatment of Pasteurella multocida and Ornithobacterium rhinotracheale, it is frequently used for E. coli infection.

Sulfonamides The sulfonamides are broad-­spectrum antimicrobials widely used to treat or prevent coccidiosis in poultry. There are a wide variety of sulfonamides available for feed and/or water administration. Sulfonamides are more soluble in an alkaline pH. Therefore, when administering sulfonamides in acidic water, it may be necessary to raise the pH of the water with unscented household ammonia to prevent drug precipitation in the bulk tank or stock solution. If the poultry water supply is being acidified for any reason, this process should be discontinued prior to and during treatment.

The use of sulfonamides has been limited in poultry in the US and Canada because of their narrow margin of safety, concerns regarding risk of tissue residues at slaughter, and/or increased use of coccidia vaccination programs. Conversely, sulfonamides are used more commonly in younger broilers in Europe, likely because they are one of few licensed products and toxic effects are not commonly observed in younger birds. Toxic effects of sulfonamides include bone marrow suppression, thrombocytopenia, and depression of the lymphoid and immune function of birds. This is frequently manifested as pale, almost yellow bone marrow and petechial or ecchymotic hemorrhages on the breast, thigh, and leg muscles (Daft et al., 1989). The most frequent toxic adverse effect of sulfonamide therapy in laying hens is a decline in egg production and eggshell quality (loss of brown pigment). The combination of sulfonamides with ionophores may predispose birds to toxic effects. The mechanism for this toxicity has not yet been elucidated; however, the effect that the drug combination has on the cytochrome P450 enzyme system has been hypothesized as one possible explanation and is being investigated (Ershov et al., 2001). There is one potentiated sulfonamide in  the US (sulfadimethoxine/ormetoprim) approved for use in feed and labeled for the prevention of coccidiosis, or bacterial infections such as Avibacterium paragallinarum, E. coli, and Pasteurella multocida. In Canada, there are several sulfadiazine/trimethoprim products that are approved for use in salmon or horses, and are used in an extra-­label manner in feed to treat bacterial infections caused by E. coli and/or P. multocida. The combination of these drugs allows for a therapeutic dose at a much lower level of each product, lessening the risk of sulfonamide toxicity. In the UK, there are several potentiated sulfonamide products licensed for administration through drinking water and feed to poultry.

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­Pharmacological Characteristics of Poultry Antimicrobial  713

Antimicrobial Therapy in Poultry

Quinolones and Fluoroquinolones Many of the quinolones, such as naladixic acid or oxolinic acid, have been used in poultry to treat primarily Gram-­negative bacterial infections. However, when these compounds are used, resistance in the bacterial population in these flocks develops quickly and can eventually result in more rapid resistance developing to the fluoroquinolones (Glisson,  1997). Therefore, poultry veterinarians should not recommend the use of these older quinolones in commercial poultry. While now banned for use in poultry in the US, fluoroquinolones are approved for therapeutic use in many countries and permitted for ELDU in others. Fluoroquinolones are still licensed for use in poultry in the UK, but voluntary antimicrobial stewardship programs limit use to treatments of last resort. According to the British Poultry Council Antimicrobial stewardship report, in 2022  no fluoroquinolones were used in the production of chicken meat (British Poultry Council, 2023). The fluoroquinolones are some of the most effective antimicrobial compounds developed for use in poultry and are highly effective against Gram-­positive, Gram-­negative, and Mycoplasma infections. It was shown that one of the fluoroquinolones, enrofloxacin, eliminated Mycoplasma gallisepticum infection in laying hens (Stanley et al., 2001). However, the fluoroquinolones are ineffective against anaerobic bacteria, such as Clostridium perfringens. The fluoroquinolones have a wide margin of safety in poultry. They are rapidly absorbed from the gastrointestinal tract, reaching peak blood concentrations within 1–­2 hours after ingestion. Their long elimination half-­life gives the poultry veterinarian the opportunity to administer the fluoroquinolones by a “pulsed dose” method in the drinking water (Charleston et  al.,  1998), which takes advantage of concentration-­dependent killing to help prevent the emergence of resistance. Rapid development of resistance to fluoroquinolones is a significant problem, and has resulted in resistance increasing in Campylobacter jejuni.

In ovo injection of fluoroquinolones causes complex adverse effects on avian embryonic development, considerably reducing the performance of incubated eggs and hatching chicks (Hruba et  al.,  2019). The presence of multivalent cations in the intestine or in the drinking water (water hardness ≥1300  ppm) reduces the absorption of oral fluoroquinolones (Sumano et al., 2004). Therefore, it is not recommended to concurrently administer electrolytes with a fluoroquinolone.

Ionophores The primary use of ionophore antimicrobials in poultry is to control coccidiosis. However, they also have activity against Gram-­positive bacteria, especially anaerobes such as Clostridium perfringens (Brennan et al., 2001b; Lanckriet et al., 2010). Since the ionophores function by altering cell permeability of both prokaryotic and eukaryotic cells, the toxic effects in poultry are reluctance to move and paralysis. This is caused by muscle weakness resulting from passive transport of potassium out of the cells, with calcium entering. Ionophore toxicity is more severe in adult birds and especially turkeys, even at a safe therapeutic dose for young chickens (Van Assen, 2006).

Novobiocin Novobiocin is rarely used in commercial poultry. It is primarily used to treat juvenile pullets or hens early in the laying house for Staphylococcus aureus arthritis. Novobiocin is poorly water soluble and so must be administered in the feed. High cost is a major reason for its limited use.

­ esponsible Use of Antimicrobials R in Poultry The responsible use of antimicrobial drugs in poultry producing meat and eggs for human consumption is based upon good professional

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judgment, laboratory results, medical knowledge, and information about the flock to be treated. When a flock of commercial poultry begins to exhibit signs of illness, the birds should be physically examined (antemortem and postmortem). If possible, bacterial cultures should be performed to confirm the clinical diagnosis and determine the susceptibility of the isolate to the chosen antimicrobial. The potential for rapid spread of disease on a poultry farm often necessitates empirical treatment prior to the results of bacterial culture and susceptibility testing. When laboratory results are available, the poultry veterinarian must use clinical judgment to decide between continuing or changing therapy. Also, a flock will usually have birds in three stages of disease development

when symptoms are first noted: clinically ill, incubating with no outward signs of illness, and unaffected but susceptible. Therefore, the entire flock is treated instead of just the clinically ill birds. Such strategic medication in anticipation of major disease spread can be justified, but antimicrobials should not be used as a substitute for good management, hygiene, and biosecurity. Clinical outcomes following each antimicrobial treatment should be critically reviewed by the veterinarian and the ­outcomes used to help formulate future treatment plans. Finally, responsible use should also ensure sufficient withdrawal time is applied following antimicrobial therapy to avoid residues in meat or eggs destined for human consumption.

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American Veterinary Medical Association (AVMA). 2020. Antimicrobial resistant pathogens affecting animal health in the United States. www.avma.org/sites/default/ files/2020-­10/ AntimicrobialResistanceFullReport.pdf Anadon A, et al. 2008. Plasma and tissue depletion of florfenicol and florfenicol-­ amine in chickens. J Agric Food Chem 56:11049. Barnes HJ, et al. 2008. Colibacillosis. In: Saif YM (ed.) Diseases of Poultry, 12th edn. Blackwell, Ames, pp.691–­732. Blom L. 1975. Residues of drugs in eggs after medication of laying hens for eight days. Acta Vet Scand 16:396. Bradbury JM. 2005. Poultry mycoplasmas: sophisticated pathogens in simple guise. Br Poultry Sci 46:125. Bradbury JM, et al. 1994. In vitro evaluation of various antimicrobials against Mycoplasma gallisepticum and Mycoplasma synoviae by the microbroth method and comparison with a commercially prepared test system. Avian Pathol 23:105.

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Food and Drug Administration (FDA). 2012b. Cephalosporin order of prohibition goes into effect. www.gpo.gov/fdsys/pkg/FR-­2012-­ 01-­06/pdf/2012-­35.pdf Fernandez-­Varon E, et al. 2006. Pharmacokinetics of an ampicillin-­sulbactam (2:1) combination after intravenous and intramuscular administration to chickens. Vet Res Commun 30:285. Gadbois P, et al. 2008. The role of penicillin G potassium in managing Clostridium perfringens in broiler chickens. Avian Dis 52:407. Glisson JR. 1997. Correct use of fluroquinolones in the poultry industry. Turkey World Mar–­Apr:24. Glisson JR. 1998. Bacterial respiratory diseases of poultry. Poult Sci 77:1139. Glisson JR, et al. 2004. Comparative efficacy of enrofloxacin, oxytetracycline, and sulfadimethoxine for the control of morbidity and mortality caused by Escherichia coli in broiler chickens. Avian Dis 48:658. Goren E, et al. 1984. Some pharmacokinetic aspects of four sulphonamides and trimethoprim, and their therapeutic efficacy in experimental Escherichia coli infection in poultry. Vet Quart 6:134. Government of Canada. 2015. Federal Action Plan on Antimicrobial Resistance and Use in Canada: Building on the Federal Framework for Action. Government of Canada, Ottawa. Grave K, et al. 2006. Usage of veterinary therapeutic antimicrobials in Demark, Norway and Sweden following termination of antimicrobial growth promoter use. Prev Vet Med 75:123. Gupta RC, Sud SC. 1978. Sulphaquinoxaline in the poultry. Ind J Anim Res 12:91. Hamdy AH, et al. 1976. Efficacy of linco-­ spectin water medication on Mycopasma synoviae airsacculitis in broilers. Avian Dis 20:118. Hamdy AH, et al. 1979. Effect of a single injection of lincomycin, spectinomycin, and linco-­spectin on early chick mortality caused

by Escherchia coli and Staphylococcus aureus. Avian Dis 23:164. Hamdy AH, et al. 1982. Efficacy of lincomycin-­ spectinomycin water medication on Mycoplasma meleagridis airsacculitis in commercially reared turkey poults. Avian Dis 26:227. Health Canada. 2014. Extra-­label drug use in animals. www.canada.ca/en/health-­canada/ services/drugs-­health-­products/veterinary-­ drugs/extra-­label-­drug-­use.html Hofacre CL, et al. 1998. Use of Aviguard, virginiamycin or bacitracin MD in experimental Clostridium perfringens associated necrotizing enteritis. J Appl Poult Res 7:412. Hruba H, et al. 2019. Reproductive toxicity of fluoroquinolones in birds. BMC Vet Res 15(1):209. Hu D, et al. 2020. Growth and cardiovascular development are repressed by florfenicol exposure in early chicken embryos. Poultry Sci 99:2736. Huang TM, et al. 2009. Antimicrobial susceptibility and resistance of chicken Escherchia coli, Salmonella spp., and Pasteurella multocida isolates. Avian Dis 53:89. Islam KMS, et al. 2009. The activity and compatibility of the antibiotic tiamulin with other drugs in poultry medicine –­a review. Poult Sci 88:2353. Jerzsele A, Nagy G. 2009. The stability of amoxicillin trihydrate and potassium clavulanate combination in aqueous solutions. Acta Vet Hung 57:485. Jerzsele A, et al. 2010. Oral bioavailability and pharmacokinetic profile of the amoxicillin-­clavulanic acid combination after intravenous and oral administration in turkeys. J Vet Pharmacol Therapeut 34:202. Jones KH, et al. 2013. A 5-­year retrospective report of Gallibacterium anatis and Pasteurella multocida isolates from chickens in Mississippi. Poult Sci 92(12):3166. Jordan FT, Horrocks BK. 1996. The minimum inhibitory concentration of tilmicosin and

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tylosin for Mycoplasma gallisepticum and Mycoplasma synoviae and a comparison of their efficacy in the control of Mycoplasma gallispeticum infection in broiler chicks. Avian Dis 40:326. Jordan FT, et al. 1999. The comparison of an aqueous preparation of tilmicosin with tylosin in the treatment of Mycoplasma gallisepticum infection of turkey poults. Avian Dis 43:521. Kelly L, et al. 2004. Animal growth promoters: to ban or not to ban? A risk assessment approach. Int J Antimicrob Ag 24:7. Kempf I, et al. 1997. Efficacy of tilmicosin in the control of experimental Mycoplasma gallisepticum infection in chickens. Avian Dis 41:802. Kinney N, Robles A. 1994. The effect of mixing antibiotics with Marek’s disease vaccine. Proceedings of the 43rd Western Poultry Disease Conference, pp. 96–­97. Kleven SH. 2008. Control of avian mycoplasma infections in commercial poultry. Avian Dis 52:367. Laber G, Schutze E. 1977. Blood level studies in chickens, turkey poults, and swine with tiamulin, a new antibiotic. J Antibiot 30:1112. Lanckriet A, et al. 2010. The effect of commonly used anticoccidials and antibiotics in a subclinical necrotic enteritis model. Avian Pathol 39:63. Leitner G, et al. 2001. The effect of apramycin on colonization of pathogenic Escherichia coli in the intestinal tract of chickens. Vet Quart 23:62. Liu YY, et al. 2016. Emergence of plasmid-­ mediated colistin resistance mechanism MCR-­1 in animals and human beings in China: a microbiological and molecular biological study. Lancet Infect. Dis 16:161. Marien M, et al. 2007. Efficacy of enrofloxacin, florfenicol and amoxicillin against Ornithobacterium rhinotracheale and Escherichia coli O2:K1 dual infection in turkeys following APV priming. Vet Microbiol 121:94. Marrett LE, et al. 2000. Efficacy of neomycin sulfate water medication on the control of mortality associated with colibacillosis in growing turkeys. Poult Sci 79:12.

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Randall L, et al. 2021. A decline in the occurrence of extended-­spectrum β-­ lactamase-­producing Escherichia coli in retail chicken meat in the UK between 2013 and 2018. J Appl Microbiol 130:247. Righter HF, et al. 1970. Tissue-­residue depletion of sulfaquinoxaline in poultry. Am J Vet Res 31:1051. Righter HF, et al. 1973. Tissue residue depletion of sulfaquinoxaline in turkey poults. J Agr Food Chem 21:412. Roth N, et al. 2019. The application of antibiotics in broiler production and the resulting antibiotic resistance in Escherichia coli: a global overview. Poult Sci 98(4):1791. Russell SM. 2003. The effect of airsacculitis on bird weights, uniformity, fecal contamination, processing errors and population of Campylobacter spp. and Escherichia coli. Poult Sci 82:1326. Salmon SA, Watts JL. 2000. Minimum inhibitory concentration determinations for various antimicrobial agents against 1570 bacterial isolates from turkey poults. Avian Dis 44:85. Scott HM, et al. 2019. Critically important antibiotics: criteria and approaches for measuring and reducing their use in food animal agriculture. Ann N Y Acad Sci 1441(1):8. Sellyei B, et al. 2009. Antimicrobial susceptibility of Pasteurella multocida isolated from swine and poultry. Acta Vet Hung 57:357. Shen J, et al. 2002. Pharmacokinetics of florfenicol in healthy and Escherichia coli-­ infected broiler chickens. Res Vet Sci 73:137. Shen J, et al. 2003. Bioavailability and pharmacokinetics of florfenicol in broiler chickens. J Vet Pharmacol Ther 26:337. Shryock TR, Belanger AE. 2004. Qualitative risk assessment for the use of antibiotics in poultry production –­human health implications: avilamycin risk assessment. Proceedings of the 16th Australian Poultry Science Symposium, p.194. Smith JA. 2011. Experiences with drug-­free broiler production. Poult Sci 90:2670. Stanley WA, et al. 2001. Case report –­monitoring Mycoplasma gallisepticum and Mycoplasma synoviae infection in an experimental line of

broiler chickens after treatment with enrofloxacin. Avian Dis 45:534. Stephens CP, Hampson DJ. 2002. Evaluation of tiamulin and lincomycin for the treatment of broiler breeders experimentally infected with the intestinal spirochaete Brachyspira pilosicoli. Avian Pathol 31:299. Sumano LH, et al. 2004. Influence of hard water on the bioavailability of enrofloxacin in broilers. Poult Sci 83:726. Swann MM, et al. 1969. Report of the Joint Committee on the use of Antibiotics in Animal Husbandry and Veterinary Medicine. HMSO, London. Switala M, et al. 2007. Pharmacokinetics of florfenicol, thiamphenicol and chloramphenicol in turkeys. J Vet Pharmacol Therapeut 30:145. Van Assen EJ. 2006. A case of salinomycin intoxication in turkeys. Can Vet J 47(3):256. Vermeulen B, et al. 2002. Drug administration to poultry. Adv Drug Deliver Rev 54:795. Vernimb GD, et al. 1977. Effect of gentamicin on early morality and later performance of broiler and leghorn chickens. Avian Dis 20:706. Warner K, et al. 2009. Control of Ornithobacterium rhinotracheale in poultry. Vet Rec 165:668. Wierup M. 2001. The Swedish experience of the 1986 year ban of antimicrobial growth promoters, with special reference to animal health, disease prevention, productivity, and usage of antimicrobials. Microb Drug Resist 7:183. World Health Organization (WHO). 2017. WHO guidelines on use of medically important antimicrobials in food-­producing animals. World Health Organization, Geneva. World Health Organization (WHO). 2024. WHO’s List of Medically Important Antimicrobials: A risk management tool for mitigating antimicrobial resistance due to non-human use. World Health Organization, Geneva. Zamon Q, et al. 1995. Experimental furazolidone toxicosis in broiler chicks: effect of dosage, duration and age upon clinical signs and some blood parameters. Acta Vet Hung 43:359. Zhao S, et al. 2005. Antimicrobial susceptibility and molecular characterization of avian pathogenic Escherichia coli isolates. Vet Microbiol 107:215.

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  ­References and Bibliograph

35 Antimicrobial Therapy in Companion Birds Marike Visser

Companion birds include members of the orders Psittaciformes (e.g., parakeets, parrots, lories, cockatoos, and macaws), Passeriformes (e.g., canaries and finches), and Columbiformes (e.g., pigeons and doves). Psittacine birds are the most common pet birds; over 50 species are  commonly seen in veterinary practice. Microbial diseases are common and use of antimicrobials is an important part of avian practice. Optimal treatment regimens can be developed if the principles of rational antimicrobial therapy are integrated with the unique behavioral and physiological characteristics of birds. The general approach to selecting an avian antimicrobial treatment regimen is similar to other species. However, the husbandry must be especially scrutinized to insure a healthy plane of nutrition and that any habitat corrections are implemented. The site and cause of infection should be identified and the minimal inhibitory concentrations (MIC) of potentially effective antimicrobial drugs determined. However, there are no CLSI breakpoints for pathogens of birds. Selection of the most appropriate drug will thus depend on the severity of illness, site of infection, pharmacokinetic and pharmacodynamic properties of the selected drugs (if available), and the routes of administration that can be accomplished by the owner or veterinary staff. Additional

considerations are adverse drug effects, t­ oxicity, and cost. The Federation of European Companion Animal Veterinary Associates (FECAVA) provides a flow diagram in multiple languages that can aid the clinician in assessing whether antimicrobial therapy is warranted (www. fecava.org/policies-­actions/guidelines/).

­ stablishing the Cause and Site E of Infection A wide variety of primary and secondary ­bacterial pathogens have been identified in companion birds; however, some are more common than others. Often, poor husbandry and nutrition are contributing factors. In ­psittacine birds, Gram-­negative bacterial ­infections are most common, especially those caused by Escherichia coli, Klebsiella spp., and  Pseudomonas aeruginosa. Other Gram-­ negative bacteria include Enterobacter spp., Proteus, Citrobacter, and Serratia marcescens. Gram-­positive bacterial pathogens include Staphylococcus aureus, S. pseudintermedius, Clostridium spp., Streptococcus spp., and Enterococcus spp. Chlamydophila psittaci is the most important intracellular pathogen; Mycobacterium avium and M. genavense ­infections are occasionally seen. Anaerobes are

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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721

Antimicrobial Therapy in Companion Birds

relatively uncommon, although clostridial infections of the alimentary tract do occur. Similar pathogens are found in canaries and pigeons; Enterococcus faecalis is an important cause of respiratory disease in canaries and there is a higher incidence of Salmonella spp. and Streptococcus gallolyticus infections in pigeons. Mycotic infections are also important. Yeasts most commonly affect the alimentary tract and common pathogens include Candida albicans and Macrorhabdus ornithogaster. Hyphal fungi are important pathogens of the respiratory tract and, occasionally, the eye and skin. Aspergillus fumigatus and A. niger are the most common isolates; Mucor spp., Penicillium spp., Rhizopus spp., and Scedosporium spp. and other opportunist molds may rarely infect immunocompromised birds. Usually, there are predisposing factors that result in infection, particularly in parrot species, ranging from poor husbandry, aspiration of food or medicine, to underlying disease. In companion birds, septicemia and infections of the alimentary tract, respiratory tract, and liver are the most common sites of microbial infection. It is important to note that simply culturing a potential pathogen is not an indication for antimicrobial treatment. It is not unusual to culture small numbers of Gram-­ negative bacteria or yeasts from the cloaca and choana of apparently healthy birds. Treatment may be indicated if the organism is present in large numbers and there are accompanying clinical signs. Physical exam findings, results of clinical laboratory tests, and a Gram stain of material from the suspected site of infection can help determine if a microbial infection is the cause of illness.

­ hoosing an Antimicrobial C Regimen To be effective, the pathogen must be susceptible to the drug at concentrations that are achievable in birds. Some pathogens have

known susceptibility (e.g., Chlamydophila ­psittaci is invariably susceptible to doxycycline), but most will require susceptibility ­testing to guide therapy. Susceptibility tests reporting MIC values are quantitative and provide the most useful information to guide drug selection. Disk diffusion tests can be used, but it is important to recognize that the designations of susceptible, intermediate, and resistant may not correlate with treatment success in birds as there are no validated breakpoints. These designations are based on the achievable drug concentrations in humans (or in a limited number of animal species) and it may be difficult to achieve similar concentrations in birds. Chapter 2 discusses susceptibility testing. Companion birds often hide signs of disease and may present at an advanced stage of ­illness. If a bacterial infection is strongly suspected, it may be necessary to start empirical treatment before the results of culture and susceptibility tests are available. In companion birds, Gram-­negative bacterial infections are most common, especially those caused by E. coli, Klebsiella spp., and P. aeruginosa. Chlamydiosis most commonly occurs in birds recently obtained from commercial sources (e.g., pet stores, flea markets, and breeders). Salmonella is common in pigeons. If these organisms are suspected, a broad-­spectrum antimicrobial with excellent Gram-­negative activity is most appropriate for initiating empirical treatment; doxycycline is preferred if chlamydiosis is likely. The treatment plan can be modified once the bird is stable and results of laboratory testing are available. The frequency and route of administration are important considerations when choosing a dosage regimen. Most birds will need to be captured and restrained to deliver medication, so that treatment regimens with longer dosage intervals are preferred. In sick birds, a ­parenteral route of administration should be used to rapidly establish effective drug concentrations. Once a bird is clinically stable, it may be relinquished to the owner’s care to complete antimicrobial therapy. Birds can be difficult to

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722

medicate, and the procedure is often stressful for both the bird and owner. If oral medication is used, low-­volume, palatable drug formulations can aid treatment success. Some avian veterinarians favor use of IM injection because bird restraint and drug delivery may be easier with this route. Additional pros and cons of different routes of administration are discussed below. Regardless of the treatment regimen, it is useful to check compliance and offer assistance after a few days of treatment. Choosing the dose can be challenging because drug formularies often list a wide range of recommended dosages. This is partly because there are sparse data on the pharmacokinetics of antibiotics in many species of psittacine birds. Many dosage regimens are empirically derived or extrapolated from other species. Table  35.1 provides suggested doses for selected commonly used antimicrobial drugs. However, even doses based on pharmacokinetic studies often represent only a single-­ dose study in a limited number of individuals of a single species. Therefore, all treated birds should be monitored carefully since safety and efficacy have not been investigated for widespread use of many of the drug dosages listed. Basic pharmacodynamic principles should be considered when evaluating which dose to use. Drugs showing time-­dependent efficacy (e.g., beta-­lactams, macrolides, tetracyclines, and trimethoprim-­sulfonamides) must be dosed frequently enough to maintain plasma concentrations above the target MIC for most of the dosing interval. Birds rapidly excrete most beta-­lactam drugs, so penicillins and cephalosporins should be dosed at least 3–4 times daily unless pharmacokinetic data demonstrate that less frequent administration is adequate. Cephalosporins that show prolonged activity in other species (e.g., cefovecin in dogs) may have short activity in birds, especially if high protein binding is necessary for the prolonged half-­life (Thuesen et  al.,  2009). Concentration-­dependent antimicrobials (e.g., fluoroquinolones and aminoglycosides) can probably be dosed once daily if high peak concentrations and large area under

the curve values are achieved. Since these values may depend on the route of administration, ­parenteral routes may be required to achieve the desired concentration for resistant organisms. Controlled studies involving large numbers of different avian species are lacking, so veterinarians should carefully monitor treatment efficacy and potential toxicity. This is ­especially important when using drugs with a narrow therapeutic range or treating an unfamiliar species. Relevant chapters in this book should be consulted on specific antimicrobials and their potential adverse effects and contraindications. Using broad-­spectrum antimicrobials may impact normal intestinal microflora. With the advent of advanced genomic sequencing, the microbiome has been found to be significantly different between species, even within the same class. Fecal samples from lovebirds and cockatiels have high numbers of Mycoplasma in presumed healthy birds, suggesting this may be a commensal organism. Pooled fecal ­samples from budgerigars, cockatiels, and domestic canaries report that Lactobacillaceae and Firmicutes (Clostridia and Bacilli class) are abundant (Garcia-­Mazcorro et  al.,  2017). Studies in chickens have reported abundant Lactobacillus in the crop, gizzard and small intestine and large intestine (Proszkowiec-­ Weglarz, 2022). The use of a quality probiotic containing Lactobaccilus may help mitigate the impact of orally administered antibiotics.

­ natomical and Physiological A Considerations Differences in anatomy and physiology may alter drug pharmacology in birds as compared to mammals. For example, granuloma formation is a common avian response to infection by many microbial agents. Granuloma formation can inhibit drug penetration so that ­surgical debridement, use of lipophilic drugs, and prolonged treatment may be needed for treatment success.

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­Anatomical and Physiological Consideration  723

Drugs

Dose (mg/kg)

Interval (h)

Route

Studyb/species

Referencec

Comments

Penicillins Ampicillin sodium Ampicillin trihydrate

150

12–24

IM

PK/pigeons

1

50 –100

6–8

IM

PK/Amazon parrot

2

125–175

4–8

PO

PK/pigeons

1

Gram-­positive bacteria only.

Gram-­positives only. IM bioavailability 57% so double the IV dose.

150–200

4–­8

PO

PK/Amazon parrots

2

Amoxicillin sodium

250

12–24

IM

PK/pigeons

1

Gram-­positives only.

Amoxicillin trihydrate

100

12–24

PO

PK/pigeons

1

Gram-­positives only.

150–175

4–8

PO

Empirical/psittacines

1

125–250

8

PO

PK/collared doves

3

100/25

8–12

PO

PK/collared doves

3

60–120

8–12

IM

PK/collared doves

3

125

8

PO

PK/blue-­fronted Amazon parrots

4

Amoxicillin + clavulanic acid

Piperacillin/tazobactam

100

8–12

IM, IV

PK/bsittacines

5

Ticarcillin

200

2–4

IM

PK/blue-­fronted Amazon parrots

6

Cephalothin

100

6

IM

PK/pigeon

7

Cephalexin

35–50

6

PO

PK/pigeon

7

Cefadroxil

100

12

PO

Psittacines, pigeons

8

Ceftiofur sodium free

10

4

IM

PK/cockatiels

9

10

8

IM

PK/orange-­winged Amazon parrots

9

50

96

IM

PK/ring-­necked doves

Cephalosporins

Ceftiofur crystalline-­free acid

0005858930.indd 724

10

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Table 35.1  Conventional dosage regimens for antimicrobial drugs in companion birds.a

Gram-­positives only.

10/25/2024 10:40:22 AM

75–100

4–8

IM

PK/blue-­fronted Amazon parrots

11

Ceftazidime

50–100

4–8

IM

Ceftriaxone

75–100

4–8

IM

PK/blue-­fronted Amazon parrots

11 11

Aminoglycosides Amikacin

15–40

24

IM, IV

PK/cockatiels, blue-­fronted Amazon parrots

12, 13

Preferred aminoglycoside; potentially nephrotoxic.

Gentamicin

2.5–10

24

IM

PK/cockatiels, scarlet macaws, rose-­breasted cockatoos

12

Nephrotoxic.

Tobramycin

2.5–10

24

IM

Empirical

Fluoroquinolones Enrofloxacin

7.5–15

12–24

IM

PK/African gray parrots

14

IM injection causes muscle irritation.

7.5–15

12–24

SC

PK/African gray parrots

14

Inject into SC fluid pocket containing lactated Ringer’s solution. Double the dose when using q 24 h administration.

15–30

24

PO

PK/African gray parrots

14

High oral doses result in plasma concentrations that may be effective with once-­daily dosing.

Marbofloxacin

2.5–5

24

PO

PK/blue and gold macaw

15

Tetracyclines Oxytetracycline, long acting (LA-­200®, Zoetis)

50–100

48–72

IM, SC

PK/Goffin’s cockatoo

11

Chlortetracycline

40–50

8 (w/ grit) 12 (w/o grit)

PO

pigeons

16

Doxycycline

25

12

PO

PK/pigeon

1

Chlamydophila psittaci; dose in birds with access to grit.

7.5

12

PO

PK/pigeon

1

Chlamydophila psittaci; dose in birds with no access to grit.

Empirical, based on gentamicin studies; used for Pseudomonas aeruginosa.

Chlamydophila psittaci; causes irritation at the site of injection.

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Cefotaxime

(Continued)

0005858930.indd 725

10/25/2024 10:40:22 AM

Dose (mg/kg)

Interval (h)

Route

Studyb/species

Referencec

Comments

35

24

PO

Cockatiel/ PD

17

Chlamydophila psittaci; at least 21-­day therapy

300 mg/kg food

24

Food

Plasma concentration/ budgerigars

18

Chlamydophila psittaci. Diet = 1:4 mixture of hulled oat groats and hulled millet; coat seed with sunflower oil (~6 ml/kg seed).

300–500 mg/ kg food

24

Food

Plasma concentration/ cockatiels

19

Diet = 60:40 mixture of hulled millet and hulled sunflower seeds; coat seed with sunflower oil (~6 ml/kg seed).

Doxycycline

300 mg/l

24

Water

Plasma concentration/ cockatiels

19

May be effective for treating chlamydiosis.

Doxycycline

400 mg/l

24

Water

Plasma concentration/ cockatiels

20

May be effective for treating spiral bacteria.

400–800 mg/l

24

Water

Plasma concentration/ orange-­winged Amazon parrot, African gray parrot, Goffin’s cockatoo

21

75–100

5–7 days

IM

PK/pigeons

10

24

PO

Blue and gold macaw

17

40

24

PO

Cockatiel

17

Azithromycin, sustained release granules

25–35

24

PO

Pigeons/PK, PD; MIC ≤0.03 mg/ml

22

Tylosin

25

6

IM

PK/pigeons

23

Clarithromycin

60–85

24

PO

Psittacines

24

Clindamycin

25–50

8–12

PO

Empirical

Drugs

Doxycycline, injectable

1

Use lower doses in macaws and cockatoos.

Macrolides Azithromycin

0005858930.indd 726

In combination with doxycycline

Gram-­positives and anaerobes.

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Table 35.1  (Continued)

10/25/2024 10:40:22 AM

Trimethoprim

15–20

8

PO

PK/pigeons

1

Trimethoprim-­sulfadiazine

20–30

12

SC, IM

Psittacines

24 25

60

12

PO

Pigeons

Trimethoprim-­ sulfamethoxazole

10/50

12

PO

PK/pigeons

1

Trimethoprim-­ sulfatroxazole

10/50

24

PO

PK/pigeons

1

Trimethoprim-­ sulfamethoxazole

20/100

12

PO

Empirical

Metronidazole

20–50

12

PO

Empirical

Isoniazid

15–30

24

PO

Empirical

1.5

8

IV

Empirical

Aspergillus and hyphal fungi.

1.0

8–12

IT

Empirical

Aspergillus and hyphal fungi.

1.0 mg/ml

8–12

Neb

Empirical

Aspergillus and hyphal fungi.

100 mg/kg

12

PO

Empirical

Ketoconazole

20–30

12

PO

PK/Amazon parrots and cockatoos

27

Yeast ± Aspergillus.

Fluconazole

10–20

24

PO

PK/African gray parrots, blue-­fronted Amazon parrots, Goffin’s cockatoos

28

Yeast. Higher dose may be toxic in African gray parrots.

75–100 mg/l

24

Water

PK/cockatiels

29

Candida.

5–10

24

PO

PK/blue-­fronted Amazon parrots

30

Aspergillus and hyphal fungi.

6

12

PO

PK/pigeon

31

Aspergillus and hyphal fungi.

2.5–5

24

PO

Empirical/African gray parrot

May cause regurgitation, especially in macaws.

Other Anaerobes. 26

Antimycobacterial agent; should be used in combination with other drugs.

Antifungals Amphotericin B

Itraconazole

Avian gastric yeast.

Itraconazole may be toxic in some African gray parrots, even at the low dose indicated here.

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Trimethoprim and sulfonamides

(Continued)

0005858930.indd 727

10/25/2024 10:40:23 AM

Drugs

Dose (mg/kg)

Interval (h)

Route

Studyb/species

Referencec

Voriconazole

18

12

PO

PK/African gray parrot

32

Voriconazole

18

8

PO

PK/Hispaniolan Amazon parrot

33

Voriconazole

10 20

12 24

PO

PK/Pigeon

34

Nystatin

200,000– 300,000 IU/kg

8–12

PO

Empirical

a

Comments

May cause hepatic toxicity. Yeast; not absorbed from the GI tract; must come in contact with the yeast.

 Adapted from Dorrestein (2000).  PK means dose recommendations based on pharmacokinetic studies in the listed species. Empirical means studies based on anecdotal reports; no published kinetic data available for pigeons or psittacine birds. c  References: 1, (Dorrestein et al., 1987); 2, (Ensley and Janssen, 1981); 3, (Dorrestein et al., 1998); 4, (Orosz et al., 2000); 5, (Carpenter et al., 2017); 6, (Schroeder et al., 1997); 7, (Bush et al., 1981); 8 (Harlin, 2006); 9, (Tell et al., 1998); 10, (Sanchez-­Migallon Guzman, 2014); 11, (Flammer, 1990); 12, (Ramsay and Vulliet, 1993); 13, (Schroeder et al., 1997); 14, (Flammer et al., 1991); 15, (Carpenter et al., 2006); 16, (Frazier et al., 1995); 17, (Guzman et al., 2010) ; 18, (Flammer et al., 2003); 19, (Powers et al., 2000); 20, (Evans et al., 2008); 21, (Flammer et al., 2001); 22, (Zań et al., 2020); 23, (Locke et al., 1982); 24, (Lennox, 2007); 25, (Harlin and Wade, 2009); 26, (Dorrestine, 2000); 27, (Kollias et al., 1986); 28, (Flammer and Papich, 2006); 29, (Ratzlaff et al., 2011); 30, (Orosz et al., 1996); 31, (Lumeij et al., 1995); 32, (Flammer et al., 2008); 33, (Sanchez-­Migallon Guzman et al., 2010); 34, (Beernaert et al., 2009). Disclaimer: As noted in the text, safety and efficacy data for widespread use of drugs in birds are lacking; none of these drug doses are warranted to be either safe or effective. b

0005858930.indd 728

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Table 35.1  (Continued)

10/25/2024 10:40:23 AM

In mammals, gastric emptying and drug ­dissolution are often the rate limiting steps for oral drug absorption. Companion birds have a crop, and passage of ingesta from the crop may delay oral drug absorption. For example, a lag phase of 20–40 minutes was observed in studies investigating the pharmacology of oral suspensions of doxycycline in fasted birds (Flammer, unpublished observation, 2005). There is little absorption from the crop, and its neutral pH may precipitate some drugs that are solubilized in acid or base (e.g., chlortetracycline), further delaying absorption. Alimentary tract motility in birds also differs from mammals. Birds have a two-­part stomach composed of the proventriculus and ventriculus. Grit is retained in the ventriculus and may expose orally administered drugs to high concentrations of calcium and magnesium. This can reduce the absorption of tetracyclines and fluoroquinolones. There is also both normograde and retrograde movement of ingesta through the proventriculus, ventriculus, and small intestine, which might expose acid-­ sensitive drugs to greater degradation by gastric acids (Proszkowiec-­Weglarz, 2022). Companion birds also have a short intestinal tract that may limit drug absorption, especially when food is present and competes for absorption. The lower respiratory system of birds consists of the lungs and air sacs. The air sacs are poorly vascularized and local drug delivery via nebulization may augment systemic drug administration. At rest, birds may ventilate only a small portion of their total air sac volume, so that nebulization may be enhanced by gently stimulating the bird to increase respiration and promote greater drug penetration (Powell, 2022). The renal system of birds differs considerably from mammals. Avian kidneys contain both mammalian and reptilian nephrons and may excrete drugs differently than expected from mammalian physiology. Uric acid is the major end-­product of avian nitrogen metabolism and is produced in the liver. Birds lack a bladder, and waste from the kidney is transported directly to

the cloaca. Cloacal ­contents can be refluxed into the colon to promote additional water absorption. Consequently, avian water balance may be independent of the glomerular filtration rate and renally excreted drugs may face reabsorption in the colon. As a final consideration, birds have a renal portal system (Goldstein,  2022). Theoretically, renally excreted drugs could face a first-­pass effect before reaching systemic circulation if injected into the leg muscles.

­Routes of Administration The route of administration will depend on the drug, available drug formulation, condition of the bird, and ability of the owner and/or veterinary staff to deliver the drug. Severely ill birds should be treated using parenteral routes to quickly establish effective drug concentrations. Achievable plasma concentrations are often route dependent due to differences in bioavailability.

Intravenous (IV) Route It is difficult to deliver IV drugs in birds so this method is usually reserved for one-­time administration of antimicrobials or emergency drugs. Birds can be catheterized, but it is more difficult to maintain IV catheters in birds than in other small animals. The right jugular and right and left brachial veins are the most accessible in psittacines. The medial metatarsal vein is accessible in pigeons.

Intraosseous (IO) Route Fluids given via the intraosseus route quickly reach the systemic circulation (Aguilar et  al.,  1993). Intraosseous catheters can be installed in the distal ulna or tibiotarsus. This route is most often used to administer fluids but it is an acceptable route for IV antimicrobial drug formulations. Care should be taken to flush fluid through the IO catheter and bone to avoid leaving concentrated drug in the IO site.

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Intramuscular (IM) Route The pectoral muscles are the most accessible sites for IM administration in parrots and passerines; the leg muscles are sometimes used in racing pigeons. Small needle size (25–30 gauge) and small volume of injection are necessary. The author prefers to use injection volumes that are less than 1 ml/kg. Irritating drugs (e.g., enrofloxacin and tetracyclines) should be avoided unless there is a compelling reason to use this route.

Subcutaneous (SC) Route Medications can be given subcutaneously in the groin, axilla, and dorsal region between the shoulders. Nonirritating drugs are preferred. Injectable tetracyclines (e.g., oxytetracycline) have been used but can cause skin sloughs (Flammer et  al.,  1990). Enrofloxacin can be injected into a SC pocket of lactated Ringer’s solution and achieves plasma concentrations comparable to IM injection without causing severe irritation.

Oral (PO) Route Liquid solutions and suspensions are often used. Capsules can be given to pigeons but are difficult to administer to parrots and small passerines. Drugs that are unpalatable or require large volumes are more difficult to administer. Only nonirritating drugs should be used, as birds may aspirate drug into the trachea or pass it rostrally into the choanal slit. It can be surprisingly difficult to medicate psittacines via the oral route, so owner compliance should be verified if this route is chosen. As an alternative, drugs can be administered via a crop tube; however, this method is technically difficult and is usually performed in a veterinary hospital setting.

Medicated Food Medications can be added to palatable food vehicles such as mash diets and treat foods. It is difficult to monitor food (and therefore drug)

consumption, so this route should be reserved for treatment of clinically stable birds with proven dosage regimens. Lower plasma drug concentrations are usually achieved than with other routes and can vary significantly between patients, so this method is used only to treat  highly susceptible bacteria (Flammer et al., 2013). It is important to use the same diet as is used in published methods, since food consumption is largely based on the energy content of the diet. Medicated food recipes for treating chlamydiosis are available for some species.

Medicated Water Delivering medication via this route usually provides only low plasma drug concentrations. This route should be avoided unless there are data proving therapeutic plasma drug concentrations can be achieved. For example, water medicated with enrofloxacin at 200  mg/l achieves low, sustained plasma concentrations of 0.05–0.2 μg/ml (Flammer and Whitt-­ Smith,  2002). Doxycycline medicated water has been shown to achieve plasma drug concentrations that are greater than 1 μg/ml and should be effective for treating chlamydiosis and spiral bacteria in cockatiels treated with 300–400 mg/l (Evans et al., 2008; Powers et al., 2000) and cockatoos and gray parrots treated with 400–800  mg/l (Flammer et  al.,  2001). Water medicated with fluconazole at 100 mg/l achieved plasma drug concentrations that should be effective for treating candidasis in cockatiels (Ratzlaff et al., 2011).

Topical Topical drugs can be applied to the skin or eye. A minimal amount of topical cream or ointment should be used, as birds may ingest or spread medications into their feathers when preening. Where possible, water-­soluble formulations are preferred as they are easier to wash off if the bird spreads them into the feathers. Silver sulfadiazine cream is a popular choice

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for treating avian skin infections because it has broad-­spectrum activity and is easy to clean up. Topical products containing corticosteroids should be avoided since birds may be more ­susceptible to the immunosuppressive effects.

Injection Antimicrobials are occasionally injected directly into the site of infection. Intratracheal injection can be used to deliver topical amphotericin B (~1 ml/kg) to treat fungal infections of the trachea. Amphotericin B and clotrimazole have been used topically to treat fungal lesions on the air sacs. Topical antimicrobials are sometimes used to treat upper respiratory infections via injection into the nares (nasal flush) or periorbital sinus (sinus flush).

Nebulization Nebulization can be used to deliver topical medication to portions of the air sacs and lungs. It is most often used when treating respiratory fungal infections. A nebulizer that produces particles less than 1 μm in diameter should be used and the birds should be exposed for 2–4 hours to ensure deposition of drug into the air sacs (Tell et al., 2012).

­Compounding Considerations The prescribing and/or dispensing of compounded products is vital to treating and preventing disease in avian patients. Reasons for compounding include, but are not limited to, the need for smaller concentrations and volumes, variable patient size, the need for easily administrable formulations, palatability, and minimizing patient stress. One of the reasons why compounded ­products present a greater risk than approved products is quality. Prior to using a compounded product, the veterinarian must assess the quality of the product and identify how the therapeutic response (efficacy and/or safety)

will be monitored. Product quality is based on stability, strength, and purity, each of which can vary based on a variety of factors. Notable among them is the vehicle used to deliver the active pharmaceutical ingredient (API). Stability reflects the physical, chemical, and microbiological properties of a compound. Instability can alter product potency (strength), leading to therapeutic failure, an increased risk of toxicity (for example, due to degradation products) or infection due to product contamination (Shrewsbury,  2015). Any product with visible changes suggesting instability should be discarded, even if the product is still within its use date. However, the potency of a product can be affected with no visible indicators. Stability testing is not simply determination of strength but involves rigorous methodology development and testing over a set period of time to determine whether a product ­continues to have the same pH, potency, and solubility in a variety of light and temperature conditions. Two compounded antimicrobials frequently utilized in avian medicine have significantly different reported stabilities. Whereas enrofloxacin remained stable for at least 56  days when prepared in corn syrup-­distilled water, cherry syrup-­distilled water or liquid sweetener and stored in amber bottles at room ­temperature, compounded doxycycline hyclate prepared in 50:50 syrup-­suspension vehicle remained stable for only seven days (Papich et al., 2013; Petritz et  al.,  2013). Furthermore, the stability can be significantly affected depending on the vehicle used. Fluoroquinolones combined with metal-­ containing products such as antacids or Lixotinic® can cause chelation and instability (Papich,  2005). Beyond use dates should be utilized for any compounded product, whether for in-­clinic use or if dispensed to a client. Compounded drugs can also have different bioavailability compared to manufactured products. Bioavailability refers to the percentage of a drug that reaches systemic distribution, with absolute bioavailability comparing intravenous administration to an alternative

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route and relative bioavailability comparing two extravenous routes. Bioavailability can impact both safety, if more API is absorbed than anticipated, and therapeutic failure (efficacy) if the API is not delivered. In general, the more sophisticated the delivery system, the greater the need for evidence of bioavailability. However, evidence of bioavailability of novel compounded preparations is limited. These pharmacokinetic studies can be time-­ consuming, expensive, and not generally of interest to funding agencies, yet these reports increase confidence that the recommended dose will reach therapeutic concentrations. A cross-­over study comparing compounded itraconazole to the approved product (Sporanox®) in black-­footed penguins reported significantly decreased absorption and subtherapeutic concentrations from the compounded product (Smith et  al.,  2010). If a commercial product cannot be used, then it is important

that some measure of therapeutic response is  instituted, such as therapeutic drug monitoring. Challenging cases sometimes require alternative drug delivery systems like antimicrobial impregnated beads for local implantation, ophthalmic solutions or transdermal gel or patchs. Among the novel products being promoted are transdermal pluronic-­lecithin (PLO) gels into which the API is dissolved. Proper mixing presumably results in the formation of micelles which, when topically applied, render the stratum corneum more permeable because of the lecithin. Few studies have demonstrated effective delivery and several demonstrated limited or no drug absorption. Impregnated antimicrobial beads have been reported for treating pododermatitis in raptors and a polymethyl methacrylate-­based synthetic grit containing nicarbizin has been reported to achieve plasma therapeutic concentrations.

­References and Bibliography Aguilar RF, et al. 1993. Osseous-­venous and central circulatory transit times of technetium-­99m pertechnetate in anesthetized raptors following intraosseous administration. J Zoo Wildlife Med 24:488. Beernaert LA, et al. 2009. Designing a treatment protocol with voriconazole to eliminate Aspergillus fumigatus from experimentally inoculated pigeons. Vet Microbiol 139:393. Bush M, et al. 1981. Pharmacokinetics of cephalothin and cephalexin in selected avian species. Am J Vet Res 42:1014. Carpenter JW, et al. 2006. Pharmacokinetics of marbofloxacin in blue and gold macaws (Ara ararauna). Am J Vet Res 67:947. Carpenter JW, et al. 2017. Single-­dose pharmacokinetics of piperacillin/tazobactam in Hispaniolan Amazon parrots (Amazona ventralis). J Avian Med Surg 31:95. Dorrestein G, et al. 1987. Comparative study of ampicillin and amoxycillin after intravenous,

intramuscular and oral administration in homing pigeons (Columba livia). Res Vet Sci 42:343. Dorrestein G, et al. 1998. Comparative study of Synulox® and Augmentin® after intravenous, intramuscular and oral administration in collared doves (Streptopelia decaocto). Proceedings of the 11th Symposium on Avian Diseases, Munich, pp. 42–54. Dorrestein G. 2000. Antimicrobial drug use in companion birds. In: Prescott J, Baggot J, Walker R (eds) Antimicrobial Therapy in Veterinary Medicine. Iowa State University Press, Ames. Ensley P, Janssen D. 1981. A preliminary study comparing the pharmacokinetics of ampicillin given orally and intramuscularly to psittacines: Amazon parrots (Amazona spp.) and blue-­naped parrots (Tanygnathus lucionensis). J Zoo Anim Med 12:42. Evans EE, et al. 2008. Administration of doxycycline in drinking water for treatment of

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spiral bacterial infection in cockatiels. J Am Vet Med Assoc 232:389. Flammer K. 1990. An update on psittacine antimicrobial pharmacokinetics. Proc Assoc Avian Vet 218. Flammer K, Papich M. 2006. Pharmacokinetics of fluconazole after oral administration of single and multiple doses in African grey parrots. Am J Vet Res 67:417. Flammer K, Whitt-­Smith D. 2002. Plasma concentrations of enrofloxacin in psittacine birds offered water medicated with 200 mg/l of the injectable formulation of enrofloxacin. J Avian Med Surg 16:286. Flammer K, et al. 1990. Potential use of long-­ acting injectable oxytetracycline for treatment of chlamydiosis in Goffin’s cockatoos. Avian Dis 34:228. Flammer K, et al. 1991. Intramuscular and oral disposition of enrofloxacin in African grey parrots following single and multiple doses. J Vet Pharmacol Ther 14:359. Flammer K, et al. 2001. Plasma concentrations of doxycycline in selected psittacine birds when administered in water for potential treatment of Chlamydophila psittaci infection. J Avian Med Surg 15:276. Flammer K, et al. 2003. Assessment of plasma concentrations of doxycycline in budgerigars fed medicated seed or water. J Am Vet Med Assoc 223:993. Flammer K, et al. 2008. Pharmacokinetics of voriconazole after oral administration of single and multiple doses in African grey parrots (Psittacus erithacus timneh). Am J Vet Res 69:114. Flammer K, et al. 2013. Assessment of plasma concentrations and potential adverse effects of doxycycline in cockatiels (Nymphicus hollandicus) fed a medicated pelleted diet. J Avian Med Surg 27:187. Frazier D, et al. 1995. Pharmacokinetic considerations of the renal system in birds: Part II. review of drugs excreted by renal pathways. J Avian Med Surg, 9:104. Garcia-­Mazcorro JF, et al. 2017. Comprehensive molecular characterization

of bacterial communities in feces of pet birds using 16s marker sequencing. Microb Ecol 73:224. Goldstein DL. 2022. Renal and extrarenal regulation of body fluid composition. In: Scanes CG, Dridi S (eds) Sturkie’s Avian Physiology, 7th edn. Academic Press, San Diego. Guzman DS, et al. 2010. Evaluating 21-­day doxycycline and azithromycin treatments for experimental Chlamydophila psittaci infection in cockatiels (Nymphicus hollandicus). J Avian Med Surg 24:35. Harlin R. 2006. Practical pigeon medicine. Proceedings of the Annual Conference of the Association of Avian Veterinarians, pp. 249–262. Harlin R, Wade L. 2009. Bacterial and parasitic diseases of Columbiformes. Vet Clin North Am Exot Anim Pract 12:453. Kollias G, et al. 1986. The use of ketoconazole in birds: preliminary pharmacokinetics and clinical applications. Proceedings of the Annual Conference of the Association of Avian Veterinarians, pp. 103–104. Lennox AM. 2007. Mycobacteriosis in companion psittacine birds: a review. J Avian Med Surg 21:181. Locke D, Bush M, Carpenter JW. 1982. Pharmacokinetics and tissue concentrations of tylosin in selected avian species. Am J Vet Res 43:1807. Lumeij J, Gorgevska D, Woestenborghs RJ. 1995. Plasma and tissue concentrations of itraconazole in racing pigeons (Columba livia domestica). J Avian Med Surg 9:32. Orosz SE, et al. 1996. Pharmacokinetic properties of itraconazole in blue-­fronted Amazon parrots (Amazona aestiva aestiva). J Avian Med Surg 10:168. Orosz SE, et al. 2000. Pharmacokinetics of amoxicillin plus clavulanic acid in blue-­ fronted Amazon parrots (Amazona aestiva aestiva). J Avian Med Surg 14:107. Papich MG. 2005. Drug compounding for veterinary patients. AAPS J 7:E281. Papich MG, et al. 2013. Doxycycline concentration over time after storage in a

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Antimicrobial Therapy in Companion Birds

compounded veterinary preparation. J Am Vet Med Assoc 242:1674. Petritz OA, et al. 2013. Stability of three commonly compounded extemporaneous enrofloxacin suspensions for oral administration to exotic animals. J Am Vet Med Assoc 243:85. Powell FL. 2022. Respiration. In: Scanes CG, Dridi S (eds) Sturkie’s Avian Physiology, 7th edn. Academic Press, San Diego. Powers LV, et al. 2000. Preliminary investigation of doxycycline plasma concentrations in cockatiels (Nymphicus hollandicus) after administration by injection or in water or feed. J Avian Med Surg 14:23. Proszkowiec-­Weglarz M. 2022. Gastrointestinal anatomy and physiology. In: Scanes CG, Dridi S (eds) Sturkie’s Avian Physiology, 7th edn. Academic Press, San Diego. Ramsay EC, Vulliet R. 1993. Pharmacokinetic properties of gentamicin and amikacin in the cockatiel. Avian Dis 37:628. Ratzlaff K, et al. 2011. Plasma concentrations of fluconazole after a single oral dose and administration in drinking water in cockatiels (Nymphicus hollandicus). J Avian Med Surg 25:23. Sanchez-­Migallon Guzman D. 2014. Advances in avian clinical therapeutics. J Exotic Pet Med 23:6.

Sanchez-­Migallon Guzman D, et al. 2010. Pharmacokinetics of voriconazole after oral administration of single and multiple doses in Hispaniolan Amazon parrots (Amazona ventralis). Am J Vet Res 71:460. Schroeder EC, et al. 1997. Pharmacokinetics of ticarcillin and amikacin in blue-­fronted Amazon parrots (Amazona aestiva aestiva). J Avian Med Surg 11:260. Shrewsbury RP. 2015. Applied Pharmaceutics in Contemporary Compounding. Morton Publishing Company, Englewood. Smith JA, et al. 2010. Effects of compounding on pharmacokinetics of itraconazole in black-­ footed penguins (Spheniscus demersus). J Zoo Wildlife Med 41:487. Tell L, et al. 1998. Pharmacokinetics of ceftiofur sodium in exotic and domestic avian species. J Vet Pharmacol Therapeut 21:85. Tell L, et al. 2012. Study of nebulization delivery of aerosolized fluorescent microspheres to the avian respiratory tract. Avian Dis 56:381. Thuesen LR, et al. 2009. Selected pharmacokinetic parameters for cefovecin in hens and green iguanas. J Vet Pharmacol Therapeut 32:613. Zań R, et al. 2020. Pharmacokinetics and pharmacodynamics of a single dose of sustained-­release azithromycin formulation in pigeons. Pol J Vet Sci 23:43.

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36 Antimicrobial Therapy in Rabbits, Rodents, and Ferrets Colette L. Wheler and Patricia M. Dowling

I­ ntroduction Veterinarians may treat small mammals as laboratory animals (e.g., rabbits, rodents), as household pets (e.g., rabbits, rodents and ­ferrets), and as food-­producing animals (e.g.,  rabbits, guinea pigs). There are many challenges when using antimicrobials in these species. Some antimicrobials are known to be toxic to rabbits and some rodents, so careful selection of the most appropriate drugs is critical. Globally, there are very few antimicrobials specifically approved for treatment of these patients, necessitating extra-­label drug use (ELDU). An alternative source or formulation of drug may be needed, which may involve compounding or importing medications from other countries (following strict federal regulations) or the use of human drug formulations. Many antimicrobials must be reconstituted prior to administration, and have a fairly short shelf-­life, even if refrigerated. Often, very little of the drug formulation is needed to treat an individual patient, so the remainder is often frozen in aliquots for economic reasons and to avoid wastage. However, information on the stability of these frozen, reconstituted products is often unavailable or difficult to find.

While many antimicrobial stewardship ­ rograms have been developed for other areas p of  food and companion animals, attention to small mammals has lagged. Antimicrobials are used extensively for the clinical treatment of infectious disease in laboratory animals, meat  animals and pets, and spontaneous and research-­related conditions in research animals. This is an area where stewardship calls for ­adding two newer “Rs” of review and responsibility to the original “3Rs” of reduce, replace, and refine promoted in experimental use of animals (Narver, 2017). ●●

Reduce: systemic administration of antimicrobials to small mammals should be infrequent and reserved for systemic or severe disease, preferably with documented susceptible bacteria. Replace: improve hygiene and sanitation practices to prevent infections and use immunizations or other nonantimicrobial therapies to control diseases. Many conditions requiring antimicrobial therapy are actually secondary to inadequate nutrition or husbandry, so these issues must also be addressed for a positive therapeutic outcome. Nail trims may lessen skin and soft tissue damage from scratching. Environmental manipulations, such as cage enrichment to

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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Antimicrobial Therapy in Rabbits, Rodents, and Ferrets

●●

●●

distract from scratching, may be helpful. With rodents, transferring a small amount of  bedding from the dirty cage to the clean cage at cage change may decrease aggression within a cage by carrying over the hierarchy established by scent marking. Modifying cage enrichment also may decrease fighting in mice. Refine: use pharmacokinetic/pharmacodynamic information to determine the best dose and route of administration for the ­species treated and decrease the duration of antimicrobial therapy. Drug dosages are ­generally based on extrapolation from other species and/or clinical experience, and many small mammal pharmacokinetic and pharmacodynamic studies are actually models for human drug development. In addition, most drugs are not manufactured in a form that is convenient for administration to small, easily stressed patients, so unique treatment methods must be developed to ensure patient acceptance and owner compliance. The number of animals being treated and their intended use must also be taken into consideration, since the treatment of one patient kept as a companion animal will differ significantly from that of hundreds being purpose bred for the pet trade, used as ­laboratory animals, being farmed for fur, or raised for meat. Review: antimicrobial use should be reviewed in each area of small mammal care. The ­laboratory animal community and meat rabbit industry should follow other veterinary organizations in developing antimicrobial stewardship guidelines that would benefit veterinarians serving the other owners and producers of small mammals.

The following sections discuss these many challenges in more detail, and conclude with a series of tables listing some reported dosages of antimicrobials and common conditions in small mammal pets. Some information is also included for hedgehogs and sugar gliders, since their popularity as pets is increasing in

North America, and this information can be difficult to find.

­Antimicrobial Use and Resistance Much of the antimicrobial use in small ­mammals is empirical; that is, without identification of a pathogen and subsequent susceptibility testing. Even if veterinarians are able to collect and submit samples to a diagnostic ­laboratory, there are no validated breakpoints for bacteria affecting small mammals and the quality of the testing may be below the standards acceptable in human diagnostic microbiology (Boot, 2012). Additionally, the minimum inhibitory concentration (MIC) breakpoint values for most antimicrobials are based on studies in humans or larger animals; the high metabolic rates of small lab animals such as rodents make it challenging to achieve drug concentrations above the pathogen MIC for many drugs. Infections should be treated as soon as possible to maximize the efficacy for the initial bacterial kill, either to eradicate the bacteria at the site of infection or to decrease the bacteria to numbers that can be eradicated by the ­animal’s immune system. Some antimicrobials (e.g., sulfonamides) are time dependent, with their efficacy dependent on the duration for which the plasma concentrations of the drug exceed the MIC for the pathogen. Concentration-­dependent antimicrobials (e.g., fluoroquinolones) rely on the peak plasma concentration for efficacy. Enteral dosing in the feed and water may be effective for time-­ dependent antimicrobials, while parenteral injectable (bolus) dosing is more appropriate for concentration-­dependent antimicrobials. If systemic antimicrobials are used, then they should be administered for the shortest time that is clinically appropriate; there is no ­minimum duration of treatment necessary for antimicrobials. Ideally, the animal’s total exposure to an antimicrobial should be limited,

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both to minimize perturbations to the normal microbiome and physiology and to decrease selecting for resistant bacteria. Antimicrobial resistance in bacteria from small mammals has not been well studied. The close interaction between pet small mammals and their owners facilitates the transmission of  pathogenic bacteria between humans and ­animals. While gerbils are first, rabbits are the second most common specialty/exotic pet mammals among US households, and are the most well studied for AMR (AVMA, 2022). According to a Spanish study of bacterial diseases of pet rabbits, the most prevalent ­genera were Staphylococcus spp. (15.8%), Pseudomonas spp. (12.7%), Pasteurella spp. (10%), Bordetella spp. (9.6%), and Streptococcus spp. (6.8%). Enterobacterales represented around 18% of the isolates, with Enterobacter spp., E. coli, and Klebsiella spp. being the most frequent isolates (Fernández et  al., 2023). Infections of the respiratory tract (53%) were most common, followed by otitis (18%), abscesses, principally located in the head (16%), conjunctivitis (5%), reproductive tract (3%), skin disease/dermatitis (2%), urinary tract (2%), and  dental disease (1%). Gram-­ positive cocci (Staphylococcus, Streptococcus and Corynebacterium [Trueperella] pyogenes) and Gram-­negative bacteria, such as Pasteurella multocida, were susceptible to a  wide panel of  conventional antimicrobials. While fluoroquinolones are the most common therapeutic option in small mammals, for respiratory infections caused by P. multocida or B. bronchiseptica, results of this study suggest that trimethoprim/sulfonamides would be a good candidate for first-­line treatment in pet rabbits.

A ­ ntimicrobial Toxicity Most veterinary practitioners are aware that some antimicrobials are toxic to rabbits and some rodents, especially when given orally. Disruption of the normal population of

i­ ntestinal flora occurs, and this dysbiosis allows proliferation of clostridial or coliform bacteria, and subsequent release of toxins. Hindgut ­fermenters, such as rabbits, guinea pigs, chinchillas, and hamsters, are particularly susceptible to this condition, and narrow-­spectrum antimicrobials such as beta-­lactams, macrolides, and lincosamides are most responsible. Diarrhea usually appears within 24–­48 hours following administration of the drug, and most cases of dysbiosis are fatal. Pathogenic conditions and sudden alterations in diet may also predispose the animal to dysbiosis, and even antimicrobials that are considered safe can sometimes cause problems. Rats, mice, gerbils, and ferrets are less vulnerable to this condition. As in other species, aminoglycosides are  potentially nephrotoxic and ototoxic to small mammal species. Streptomycin has been reported to be toxic in gerbils. Although normally safe in rabbits, rodents, and ferrets, fluoroquinolone antimicrobials (e.g., enrofloxacin) may cause arthropathies in young animals. Chloramphenicol is generally safe to use in small mammals, and many bacteria infecting these animals are highly susceptible to this drug. However, chloramphenicol has been associated with a dose-­independent irreversible aplastic anemia in humans, so appropriate directions for prevention of exposure, such as wearing gloves and hand washing, must be given when this antimicrobial is prescribed. In addition, chloramphenicol is prohibited for use in food-­producing animals, such as meat rabbits. Potential toxicities must always be kept in mind when selecting an antimicrobial based on culture and susceptibility results, as the most appropriate choice may result in dysbiosis or  other problems in a particular species. Supportive ancillary therapies, such as administration of fluids along with aminoglycosides and good nursing care, as well as provision of adequate nutrition and a comfortable, stress-­ free environment, will also aid in successful treatment.

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­Antimicrobial Toxicit 

Antimicrobial Therapy in Rabbits, Rodents, and Ferrets

­ xtra-­label Use, Compounding, E and Importation In Canada and the US, there are a limited number of drugs labeled for use in rabbits, rodents, and ferrets, and very few of these are  antimicrobials. Some antimicrobials are approved for use in mink, and dosages for mink would likely be valid in ferrets since they are closely related species. In Canada, chlortetracycline feed premix is approved for the treatment of mink. In Canada, antimicrobials labeled for use in rabbits include procaine ­penicillin G (IM use only) and tilmicosin (SC and in feed), and salinomycin and robenidine for the control of coccidiosis. In the US, only sulfaquinoxaline as an antimicrobial and lasalocid for coccidiosis are approved for use in rabbits. In the EU, most meat rabbit production occurs in France, Spain, and Italy, and there are a number of approved antimicrobials including trimethoprim/sulfas, sulfadimethoxine, chlortetracycline, apramycin, and colistin. Therefore, in order to provide appropriate care for small mammal patients, veterinarians are required to use many drugs in an extra-­ label manner. In Canada and the US, ELDU refers to the use of a federally approved drug in a manner that is not in accordance with the label or package insert. It is the responsibility of the veterinarian to be aware of, and follow, the rules and regulations in their particular jurisdiction. In the US, further clarification of ELDU was provided in 1994  with the introduction of the Animal Medicinal Drug Use Clarification Act (AMDUCA). This act clearly explains legitimate extra-­label drug use by veterinarians, and outlines the specific conditions that must be followed for acceptable extra-­ label drug use (see Chapter 26). Extra-­label use of human antimicrobial formulations is fairly common for treatment of  small mammals. Many of these products are  single-­dose vials that have a fairly short ­shelf-­life once reconstituted (e.g., cefazolin). Treatment of the small mammal patient may occur for a longer period than the shelf-­life of

the drug, or the total amount needed may be very small. Rather than discard the remainder, veterinarians often freeze small aliquots of the product for future use. Details on the stability of these reconstituted products after freezing are often not easy to find; however, some information can be found in the Handbook of Injectable Drugs by Lawrence Trissel, which is available in hardcopy and electronic format, and Plumb’s Veterinary Drug Handbook by Donald Plumb, as well as in some package inserts. An alternative source of drug sometimes needs to be explored by veterinarians for the treatment of small mammal patients, such as compounding or importing medications from other countries. Compounding is a type of ELDU whereby the original drug dosage form is manipulated by a veterinarian or pharmacist, or an entirely new product is manufactured by a compounding pharmacy, to create a customized medication to meet a specific need. This could involve anything from altering the concentration of a drug by diluting it other than according to the package instructions, or mixing a crushed tablet into a liquid, to the custom creation of a medicated tablet or liquid that is particularly palatable to the intended species. Importation of a more suitable drug or drug formulation from another country is another option for veterinarians. For example, a ­suspension of metronidazole is available in some countries that is much more accurate for ­dosing small patients than the tablet form available in the US. Regulations for such importations must be followed.

­Drug Dosages Although there are many antimicrobial dosages published for rabbits, rodents, and ferrets, very few pharmacokinetic studies or clinical trials have been performed specifically for these animals; rather, they are carried out primarily to establish information for human trials. Because of this, antimicrobial dosages for these patients are generally based on extrapolations from other species and/or clinical experience.

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738

Lack of scientifically derived dosages, combined with the extra-­label use of most antimicrobials, are daily challenges of veterinarians who care for small mammal patients. Clients should be informed of this, and give written consent for treatment of their animals. Extrapolation of drug dosages from one ­species to another can be done in several ways. Straightforward linear extrapolations based on body weight alone tend to result in overdosing of larger animals and underdosing of smaller ones. This method is only appropriate with drugs that have large margins of safety and wide therapeutic margins, or if the two animals are similar in taxonomy, body size, and physiology. Metabolic scaling is a method popular in zoological medicine, and uses a formula based on body weight, a constant based on the energy group of the animal, and the known pharmacokinetic data of the drug in one species to calculate the dosage of the drug in other species. Allometric scaling uses mathematical ­equations to analyze differences in anatomy, physiology, biochemistry, and pharmacokinetics in animals of different sizes. Known pharmacokinetic parameters in several species are used in the equations to estimate the pharmacokinetic parameter in an unknown species, and thus predict drug dosage. Allometric scaling is commonly used in the pharmaceutical industry to determine the first dosage in human trials. There are several reports in the  literature validating the use of allometric ­scaling to predict pharamacokinetic parameters in  small mammal species for several drugs, including some fluoroquinolone antimicrobials. Tables 36.1–­36.3 present drug dosages for the treatment of common microbial diseases. Tables  36.4–­36.11 present clinical signs and suggested drugs for common bacterial diseases.

­Drug Administration Rabbits and rodents are prey species, and are generally less tolerant of handling and other manipulations than predator species such as  ferrets, dogs, and cats, especially when

debilitated. Administration of antimicrobials in these prey species must be performed in a way that allows for the entire dose to be given without unduly stressing the patient. The method of administration must also be achievable for the client, otherwise frustration and noncompliance may result. Available antimicrobial formulations are often too large and/or too concentrated for small mammals and need to be split up or diluted for accurate dosing. Routes of antimicrobial administration in rabbits, rodents, and ferrets include oral (liquid, pill, or capsule), subcutaneous (usually in the loose skin over the shoulders), intraperitoneal (generally reserved for very small rodents), intramuscular (generally avoided in very small animals), topical, and less commonly, via intravenous or intraosseous catheter, nebulization, gavage, nasoesophageal or esophagostomy tube (rabbits, ferrets), or antimicrobial-­impregnated implants. Injections are more commonly used in clinic than at home; however, some clients are willing to master the procedure, particularly if the pet objects excessively to being medicated orally, or if it has a sore mouth, or tends to nip. Self-­administration, where the animal willingly takes the entire dose on its own, preferably with minimal or no restraint, is the best and least stressful method of medication (for  both the animal and the administrator). Flavored antimicrobial preparations, such as trimethoprim/sulfa or chloramphenicol palmitate suspensions, are willingly consumed by some of these patients. Crushed pills, liquids, or capsule contents can be mixed with small amounts of palatable liquids, gels, or food to encourage consumption. Small rodents, such as rats and mice, willingly take vanilla-­flavored human nutritional supplements, such as Boost® or Ensure®, directly from a syringe or small dish. Hamsters favor rice-­based baby cereal, rabbits like bananas, chinchillas are partial to  raisins, and ferrets enjoy malt-­flavored cat ­laxatives or pet nutritional supplements such as Nutri-­Cal® . The internet abounds with suggestions from clients and veterinarians alike, and these include Cool Whip®, maple syrup,

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­Drug Administratio  739

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Table 36.1  Reported antimicrobial drug dosages in rabbits, guinea pigs, and chinchillas. Caution: most uses and dosages are extra-­label. Drug

Rabbita

Guinea pig

Chinchilla

Amikacin

2–­5 mg/kg q 8–­12 h; SC, IM

2–­5 mg/kg q 8–­12 h; SC, IM

2–­5 mg/kg q 8–­12 h; SC, IM, IV

Azithromycin

5 mg/kg q 48 h; IM OR 15–­30 mg/kg q 24 h; PO

15–­30 mg/kg q 12–­24 h; PO

15–­30 mg/kg q 24 h; PO

Captan powder

–­

–­

5 ml/475 ml bathing dust

Cephalexin

11–­22 mg/kg q 8–­12 h; SC

50 mg/kg q 24 h; IM

–­

Chloramphenicol

30 mg/kg q 8–­12 h; PO, SC, IM, IVb

20–­50 mg/kg q 6–­12 h; PO, SC, IM, IV

30–­50 mg/kg q 12 h; PO,SC,IM, IV

Chlortetracycline

50 mg/kg q 24 h; PO

–­

50 mg/kg q 12 h; PO

Ciprofloxacin

5–­20 mg/kg q 12 h; PO

5–­20 mg/kg q 12 h; PO

5–­20 mg/kg q 12 h; PO

Clindamycin

Do not use

7.5 mg/kg q 12 h; SC; Do not use PO

7.5 mg/kg q 12 h; SC; Do not use PO

Doxycycline

2.5 mg/kg q 12 h; PO

2.5 mg/kg q 12 h; PO

2.5 mg/kg q 12 h; PO

Enrofloxacin

5–­10 mg/kg q 12 h; PO, SC, IM OR 200 mg/L dw q 24 h

0.05–­0.2 mg/mL dw q 24 h OR 5–­15 mg/kg q 12 h; PO, SC, IM

5–­15 mg/kg q 12 h; PO, SC, IM

Fluconazole

38 mg/kg q 12 h; PO

16–­20 mg/kg q 24 h × 14 d; PO

16 mg/kg q 24 h × 14 d; PO

Gentamicin

1.5–­2.5 mg/kg q 8 h; SC, IM, IV

2–­4 mg/kg q 8–­12 h; SC, IM

2 mg/kg q 12 h; SC, IM, IV

Griseofulvin (avoid in pregnant animals)

25 mg/kg q 24 h × 30–­45d; PO

25–­50 mg/kg q 12 h × 14–­60d; PO OR 1.5% in DMSO for 5–­7 d; topically

25 mg/kg q 24 h × 30–­60d; PO

Itraconazole

20–­40 mg/kg q 24 h; PO

5–­10 mg/kg q 24 h; PO

5 mg/kg q 24 h; PO

Ketoconazole

10–­40 mg/kg q 24 h; PO

10–­40 mg/kg q 24 h; PO

10–­40 mg/kg q 24 h; PO

Lime sulfur dip

Dilute 1:40 with water, dip q 7d for 4–­6 wk

Dilute 1:40 with water, dip q 7d for 4–­6 wk

Dilute 1:40 with water, dip q 7d for 4–­6 wk

Marbofloxacin

2 mg/kg q 24 h; IM, IV OR 5 mg/kg q 24 h; PO

4 mg/kg q 24 h; PO, SC

4 mg/kg q 2 h4; PO, SC

Metronidazole

20 mg/kg q 12 h; PO

25 mg/kg q 12 h; PO

10–­20 mg/kg q 12 h; PO; use with caution

20–­50 mg/kg q 24 h; PO

Fenbendazole

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50 mg/kg q 12 h; PO OR 1 mg/mL dw

–­

50 mg/kg q 12 h; PO

Penicillin G, benzathine

42,000–­60,000 IU/kg q 48 h; SC, IM

Toxic

Avoid

Penicillin G, procaine

42,000–­84,000 IU/kg q 24 h; SC, IM

Toxic

Avoid

Sulfadimethoxine

10–­15 mg/kg q 12 h × 10d; PO

10–­15 mg/kg q 12 h; PO

25–­50 mg/kg q 24 h × 10–­14d; PO

Sulfamethazine

1 mg/mL dw

1 mg/mL dw

1 mg/mL dw

Sulfaquinoxaline

1 mg/mL dw

1 mg/mL dw

–­

Terbinafine

100 mg/kg q 12–­24 h; PO

10–­40 mg/kg q 24 h × 4–­6 wk; PO

10–­30 mg/kg q 24 h × 4–­6 wk; PO

Tetracycline

50 mg/kg q 8–­12 h; PO OR 250–­1000 mg/L dw q 24 h

10–­40 mg/kg q 24 h; PO

0.3–­2 mg/mL dw q 24 h OR 10–­20 mg/kg q 8–­12 h; PO

Trimethoprim-­sulfa

30 mg/kg q 12–­24 h; PO, SC, IM

15–­30 mg/kg q 12 h; PO, SC

15–­30 mg/kg q 12 h; PO, SC

Tylosin

10 mg/kg q 12 h; PO, SC

10 mg/kg q 24 h; PO, SC; use with caution

10 mg/kg q 24 h; PO, SC

a

 Observe correct withdrawal time in meat rabbits.  Do not use in meat rabbits. PO, per os; SC, subcutaneous; IM, intramuscular; IV, intravenous; dw, drinking water.

b

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Oxytetracycline

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Drug

Hamster

Gerbil

Amikacin

2–­5 mg/kg q 8–­12 h; SC

Ampicillin

Toxic

Cephalexin

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Table 36.2  Reported antimicrobial dosages in hamsters, gerbils, rats, and mice. Caution: most uses and dosages are extra-­label. Rat

Mouse

2–­5 mg/kg q 8–­12 h; SC

10 mg/kg q 12 h; SC

10 mg/kg q 8–­12 h; SC

6–­30 mg/kg q 8 h; PO

20–­100 mg/kg q 12 h; PO, SC

20–­100 mg/kg q 12 h; PO, SC OR 500 mg/L dw

–­

25 mg/kg q 24 h; SC

15 mg/kg q 12 h; SC

60 mg/kg q 12 h; PO

Cephaloridine

10–­25 mg/kg q 24 h; SC

30 mg/kg q 12 h; IM

10–­25 mg/kg q 24 h; SC

10–­25 mg/kg q 24 h; SC

Cephalosporin

–­

–­

–­

30 mg/kg q 12 h; SC

Chloramphenicol palmitate

50–­200 mg/kg q 8 h; PO

50–­200 mg/kg q 8 h; PO

50–­200 mg/kg q 8 h; PO

0.5 mg/mL dw OR 50–­200 mg/kg q 8 h; PO

Chloramphenicol succinate

20–­50 mg/kg q 12 h; SC

20–­50 mg/kg q 12 h; SC

30–­50 mg/kg q 12 h; SC

30–­50 mg/kg q 12 h; SC

Azithromycin

30 mg/kg q 24 h; PO

Chlortetracycline

20 mg/kg q 12 h; PO, SC

–­

–­

25 mg/kg q 12 h; PO, SC

Ciprofloxacin

7–­20 mg/kg q 12 h; PO

7–­20 mg/kg q 12 h; PO

7–­20 mg/kg q 12 h; PO

7–­20 mg/kg q 12 h; PO

Doxycycline

2.5–­5 mg/kg q 12 h; PO; do not use in young or pregnant animals

2.5–­5 mg/kg q 12 h; PO; do not use in young or pregnant animals

5 mg/kg q 12 h; PO

2.5 –­5 mg/kg q 12 h; PO

Enrofloxacin

0.05–­0.2 mg/mL dw × 14d OR 5–­10 mg/kg q 12 h; PO, SC

0.05–­0.2 mg/mL dw × 14d OR 5–­10 mg/kg q 12 h; PO, SC

0.05–­0.2 mg/mL dw × 14d OR 5–­10 mg/kg q 12 h; PO, SC

0.05–­0.2 mg/mL dw × 14d OR 5–­10 mg/ kg q 12 h; PO, SC 20 mg/kg q 12 h; PO

Erythromycin

–­

–­

20 mg/kg q 12 h; PO

Gentamicin

5 mg/kg q 24 h; SC

2–­4 mg/kg q 8 h; SC

5–­10 mg/kg divided q 8–­12 h; SC

2–­4 mg/kg q 8–­12 h; SC

Griseofulvin (avoid in pregnant animals)

25–­50 mg/kg q 12 h × 14–­60d; PO OR 1.5% in DMSO for 5–­7 d; topically

25–­50 mg/kg q 12 h × 14–­60d; PO OR 1.5% in DMSO for 5–­7 d; topically

25–­50 mg/kg q 12 h × 14–­60d; PO OR 1.5% in DMSO for 5–­7 d; topically

25–­50 mg/kg q 12 h × 14–­60d; PO OR 1.5% in DMSO for 5–­7 d; topically

Ketoconazole

10–­40 mg/kg q 24 h × 14d; PO

10–­40 mg/kg q 24 h × 14d; PO

10–­40 mg/kg q 24 h × 14d; PO

10–­40 mg/kg q 24 h × 14d; PO

Metronidazole

7.5 mg/70–­90gm animal q 8 h

7.5 mg/70–­90gm animal q 8 h

10–­40 mg/kg q 24 h; PO

2.5 mg/mL dw × 5 d OR 20–­60 mg/kg q 8–­12 h; PO

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0.5 mg/mL dw OR 100 mg/kg q 24 h; PO

2 g/L dw OR 100 mg/kg q 24 h; PO

2 g/L dw OR 25 mg/kg q 12 h; PO

2 g/L dw

Oxytetracycline

0.25–­1 mg/mL dw or 16 mg/kg q 24 h; SC

0.8 mg/mL dw or 10 mg/kg q 8 h; PO or 20 mg/kg q 24 h; SC

500 mg/L dw or 10–­20 mg/kg q 8 h; PO

500 mg/L dw OR 10–­20 mg/kg q 8 h; PO

Sulfadimethoxine

10–­15 mg/kg q 12 h; PO

10–­15 mg/kg q 12 h; PO

10–­15 mg/kg q 12 h; PO

10–­15 mg/kg q 12 h; PO

Sulfamerazine

1 mg/mL dw q 24 h

0.8 mg/mL dw q 24 h

1 mg/mL dw

1 mg/mL dw OR 500 mg/L dw

Sulfamethazine

1 mg/mL dw q 24 h

0.8 mg/mL dw q 24 h

1 mg/mL dw

1 mg/mL dw

Sulfaquinoxaline

1 mg/ml dw q 24 h

1 mg/ml dw q 24 h

Tetracycline

0.4 mg/mL dw q 24 h OR 10–­20 mg/kg q 8–­12 h; PO

2–­5 mg/mL dw q 24 h OR 10–­20 mg/ kg q 8–­12 h; PO

2–­5 mg/mL dw OR 10–­20 mg/kg q 8 h; PO

2–­5 mg/mL dw OR 10–­20 mg/kg q 8 h; PO

Trimethoprim-­sulfa

15–­30 mg/kg q 12–­24 h; PO, SC

30 mg/kg q 12–­24 h; PO, SC

15–­30 mg/kg q 12 h; PO, SC

30 mg/kg q 12 h; PO, SC

Tylosin

2–­8 mg/kg q 12 h; SC, PO OR 500 mg/mL dw

0.5 mg/mL dw q 24 h OR 10 mg/kg q 24 h; PO, SC

0.5 mg/mL dw OR 10 mg/kg q 24 h; PO, SC

0.5 mg/mL dw OR 10 mg/kg q 24 h; PO, SC

PO, per os; SC, subcutaneous; IM, intramuscular; dw, drinking water.

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Neomycin

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Drug

Ferrets

Hedgehogs

Sugar gliders

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Table 36.3  Reported antimicrobial dosages in ferrets, hedgehogs, and sugar gliders. Caution: most uses and dosages are extra-­label.

Amikacin

10–­15 mg/kg q 12 h; SC, IM

2–­5 mg/kg q 8–­12 h; SC, IM

10 mg/kg q 12 h; IM

Amoxicillin

20–­30 mg/kg q 8–­12 h; PO

15 mg/kg q 12 h; PO, SC

30 mg/kg divided q 12–­24 h; PO, SC

Amoxicillin/ clavulanate

12.5–­25 mg/kg q 8–­12 h; PO

12.5 mg/kg q 12 h; PO

12.5 mg/kg divided q 12–­24 h; PO, SC

Ampicillin

5–­30 mg/kg q 8–­12 h; SC, IM, IV

–­

–­

Azithromycin

5 mg/kg q 24 h; PO

–­

–­

Ceftiofur

–­

20 mg/kg q 12–­24 h; SC

–­

Cephalexin

15–­30 mg/kg q 8–­12 h; PO

25 mg/kg q 12 h; PO

30 mg/kg divided q 12–­24 h; PO, SC

Chloramphenicol

25–­50 mg/kg q 12 h; PO, SC, IM

30–­50 mg/kg q 6–­12 h; PO, SC, IV

–­

Chlortetracycline

–­

5–­20 mg/kg q 12 h; PO

–­

Ciprofloxacin

5–­15 mg/kg q 12 h; PO

5–­20 mg/kg q 12 h; PO

10 mg/kg q 12 h; PO

Clarithromycin

12.5–­25 mg/kg q 12 h; PO

5.5 mg/kg q 12 h; PO

–­

Clindamycin

5–­10 mg/kg q 12 h; PO

5.5–­10 mg/kg q 12 h; PO

–­

Doxycycline

–­

2.5–­10 mg/kg q 12 h; PO, SC

–­

Enrofloxacina

10–­20 mg/kg q 12–­24 h; PO, SC, IM

5 mg/kg q 12 h; PO, SC

2.5–­5 mg/kg q 12–­24 h; PO, SC, IM

Erythromycin

10 mg/kg q 6 h; PO

10 mg/kg q 12 h; PO

20 q 12 h;

Fluconazole

50 mg/kg q 12 h; PO

–­

–­

Griseofulvin (avoid in pregnant animals)

25 mg/kg q 12–­24 h; PO

50 mg/kg q 24 h; PO

20 mg/kg q 24 h; PO

PO

Itraconazole

15 mg/kg q 24 h; PO

5–­10 mg/kg q 12–­24 h; PO

5–­10 mg/kg q 12 h; PO

Ketoconazole

10–­30 mg/kg q 12–­24 h; PO

10 mg/kg q 12–­24 h; PO

–­

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Lincomycin

–­

–­

30 mg/kg divided q 12–­24 h; PO, IM

Metronidazole

20 mg/kg q 12 h; PO

20 mg/kg q 12 h; PO

25 mg/kg q 24 h; PO

Neomycin

10–­20 mg/kg q 6 h; PO

–­

–­

Nystatin

–­

30,000 IU/kg q 8–­24 h; PO, topical

5000 IU/kg q 8 h × 3d; PO

Oxytetracycline

–­

25–­50 mg/kg q 24 h; PO, in food

–­

Penicillin G procaine

40,000 IU/kg q 24 h; SC

40,000 IU/kg q 24 h; SC, IM

22,000–­25,000 IU/kg q 12–­24 h; SC, IM

Piperacillin

–­

10 mg/kg q 8–­12 h; SC

–­

Sulfadimethoxine

–­

2–­20 mg/kg q 24 h; PO, SC

5–­10 mg/kg q 12–­24 h; PO, SC

Tetracycline

25 mg/kg q 12 h; PO

–­

–­

Trimethoprim-­sulfa

15–­30 mg/kg q 12 h; PO, SC, IM

30 mg/kg q 12 h; PO, SC

15 mg/kg q 12 h; PO

Tylosin

10 mg/kg q 8–­12 h; PO, SC

10 mg/kg q 12 h; PO, SC

–­

a

 Dilute if giving by subcutaneous or intramuscular route to avoid tissue necrosis at injection site. PO, per os; SC, subcutaneous; IM, intramuscular; dw, drinking water.

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Site

Clinical signs/diagnosis

Common infecting organisms

Comments

Suggested drugs

Integument

Scabbing over dorsum and perineum; dermatitis; abscesses

Staphylococcus aureus, Proteus spp., Streptococcus spp.

Secondary to fighting/bite wounds or self-­trauma due to acariasis. Trim toenails.

Ampicillin, chloramphenicol, tetracyclines

Mastitis

S. aureus

Lance and drain in addition to antimicrobials.

Ampicillin, chloramphenicol, fluoroquinolones

Pruritus, weight loss, hyperkeratosis, alopecia

Corynebacterium bovis

Affects immunocompromised mice. Low mortality. Treatment not curative.

Ampicillin, penicillin

Alopecia, erythema, crusting on face, head, neck, tail

Trichophyton mentagrophytes, Microsporum gypseum (less frequently)

Uncommon, zoonotic.

Griseofulvin (avoid in pregnant animals)

Rhinitis, dyspnea, otitis media, Mycoplasma pulmonis, upper respiratory tract disease, Staphylococcus aureus, Streptococcus spp. pneumonia

Often concurrent with Sendai virus or CAR bacillus; decrease intracage ammonia levels.

Tylosin, fluoroquinolones, tetracyclines; enrofloxacin PLUS doxycycline for its immunomodulating effect

Dacryoadenitis, sneezing, dyspnea, pneumonia

Pasteurella pneumotropica, Klebsiella pneumoniae, Bordetella bronchiseptica

Often concurrent with Sendai virus or CAR bacillus; decrease intracage ammonia levels.

Chloramphenicol, fluoroquinolones, tylosin, aminoglycosides

Pneumonia

Filobacterium rodentium

Primary or opportunist with other Sulfamerazine, ampicillin, respiratory pathogens. trimethoprim-­sulfa

Stunted growth, diarrhea, rectal prolapse, death; transmissible murine colonic hyperplasia

Citrobacter rodentium, Clostridium piliforme

Genotype, age, and diet influence course and severity of disease.

Respiratory

Gastrointestinal

Liver disease, death, chronic Helicobacter hepaticus active hepatitis, rectal prolapse

0005858931.INDD 746

Tetracyclines, neomycin, metronidazole

Amoxicillin (1.5–­3 mg/30 g/d) PLUS metronidazole (0.69 mg/30 g/d) PLUS bismuth subsalicylate (0.185 mg/30 g/d), combined PO

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Table 36.4  Antimicrobial treatment in mice. Caution: most uses and dosages are extra-­label.

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CNS

General

0005858931.INDD 747

Clostridium piliforme

Concurrent fluid therapy essential.

Tetracyclines

Anorexia, weight loss, lethargy, dull coat

Salmonella enteriditis, S. typhimurium

Zoonotic; recommend culling infected animals.

Treatment not recommended

Oophoritis, salpingitis, metritis, infertility, abortions

Mycoplasma pulmonis, Pasteurella pneumotropica, Klebsiella oxytoca

Tylosin, fluoroquinolones, tetracyclines

Urethral gland obstruction, preputial gland abscesses

Pasteurella pneumotropica, Staphylococcus aureus

Chloramphenicol, fluoroquinolones, aminoglycosides

Head tilt, torticollis

Mycoplasma pulmonis, Streptococcus spp.

Chloramphenicol, tylosin, fluoroquinolones

Eye abscesses, conjunctivitis, panophthalmitis

Pasteurella pneumotropica

Tetracyclines, aminoglycosides

Septicemia, death; mice that survive acute infection may have chronic arthritis, limb deformity, limb amputation; streptobacillosis

Streptobacillus moniliformis

Zoonotic potential.

Rough hair coat, hunched posture, inappetence, nasal and ocular discharge, arthritis

Corynebacterium kutscheri

Antimicrobial treatment not curative.

Ampicillin, tetracycline

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Urogenital

Anorexia, dehydration, diarrhea, death (Tyzzer disease)

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Table 36.5  Antimicrobial treatment in hamsters. Caution: most uses and dosages are extra-­label. Site

Clinical signs/diagnosis

Common infecting organisms

Comments

Suggested drugs

Integument

Cheek pouch abscesses, bite wound abscesses

Staphylococcus aureus, Streptococcus spp., Pasteurella pneumotropica, Actinomyces spp.

Drain and flush; complete excision of abscess beneficial.

Chloramphenicol, tetracyclines, fluoroquinolones

Swollen lymph nodes, lymphadenitis

Staphylococcus aureus, Streptococcus spp.

Mastitis

Beta-­hemolytic streptococci

Glands warm and swollen. Supportive treatment; self-­limiting infection.

Alopecia, dry skin, yellow flaky seborrhea

Trichophyton mentagrophytes

Zoonotic. Sometimes pruritic; improve cage ventilation.

Griseofulvin (avoid in pregnant animals)

Sneezing, dyspnea, upper respiratory tract disease, pneumonia

Pasteurella pneumotropica, Streptococcus pneumoniae

Secondary to poor nutrition and husbandry.

Chloramphenicol, tetracyclines, fluoroquinolones

Filobacterium rodentium

Opportunist with other respiratory pathogens.

Sulfamerazine, sulfonamides

Diarrhea, stained perineum, lethargy, anorexia, rectal prolapsed, proliferative ileitis (“wet tail”)

Lawsonia intracellularis

Especially in 3–­10-­week-­olds; difficult to treat successfully; concurrent fluid therapy, analgesics, supportive care.

Chloramphenicol, tetracyclines, fluoroquinolones, trimethoprim-­ sulfa. Start with double recommended dose for 1 day

Enteritis

E. coli, Clostridioides difficile

Concurrent fluid therapy, analgesics, supportive care essential.

Fluoroquinolones, metronidazole, tetracyclines

Anorexia, dehydration, diarrhea, death, Tyzzer disease

Clostridium piliforme

Concurrent fluid therapy, analgesics, supportive care essential.

Tetracyclines

Catarrhal enteritis in weanlings

Giardia muris

Squinting, rubbing eye, corneal ulceration

Pasteurella spp., Streptococcus spp.

Respiratory

Chloramphenicol, tetracyclines, fluoroquinolones

Streptococcus spp.

Gastrointestinal

Central nervous system

0005858931.INDD 748

Metronidazole Topical treatment. Prone to proptosis.

Chloramphenicol, tetracyclines

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Table 36.6  Antimicrobial treatment in gerbils. Caution: most uses and dosages are extra-­label. Site

Clinical signs/diagnosis

Common infecting organisms

Comments

Suggested drugs

Integument

Red, crusty nares, staining on forepaws, nasal dermatitis (“sore nose” or “red nose”)

Staphylococcus aureus, Staphylococcus spp., Streptococcus spp.

Secondary to irritation due to Harderian gland secretions.

Chloramphenicol, trimethoprim-­sulfa, fluoroquinolones

Midventral marking gland infection, dermatitis

Staphylococcus aureus, Streptococcus spp.

Alopecia, hyperkeratosis

Trichophyton mentagrophytes, Microsporum gypseum

Zoonotic.

Griseofulvin (avoid in pregnant animals)

Respiratory

Sneezing, dyspnea, weight loss

Beta-­hemolytic streptococci, Pasteurella pnuemotropica

Rare; concurrent therapy with oxygen, mucolytics, bronchodilators may help.

Fluoroquinolones, oxytetracycline, sulfonamides

Gastrointestinal

Lethargy, anorexia, diarrhea, death, Tyzzer disease

Clostridium piliforme

Highly susceptible.

Tetracyclines

Diarrhea, salmonellosis symptoms, death

Salmonella enteriditis, S. typhimurium

Zoonotic; recommend culling affected animals. Fluid therapy is essential if treatment attempted.

Chloramphenicol, fluoroquinolones

Enteritis, diarrhea, dehydration

E. coli

0005858931.INDD 749

Chloramphenicol, trimethoprim-­sulfa, fluoroquinolones

Chloramphenicol, fluoroquinolones

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Site

Clinical signs/diagnosis

Integument

Abrasions/ulcerations over shoulders and back; ulcerative dermatitis

Respiratory

0005858931.INDD 750

Common infecting organisms

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Table 36.7  Antimicrobial treatment in rats. Caution: most uses and dosages are extra-­label.

Comments

Suggested drugs

Staphylococcus aureus

Secondary to primary wounds caused by self-­trauma. Trim toenails.

Ampicillin, chloramphenicol,

Abscesses, furunculosis

P. pneumotropica, K. pneumoniae

Secondary opportunist; drain and flush abscesses.

Chloramphenicol, fluoroquinolones, aminoglycosides

Mastitis

P. pneumotropica, Staphylococcus aureus

Hot compress, drainage.

Chloramphenicol, fluoroquinolones, aminoglycosides

Alopecia, pruritus

Microsporum spp.

Zoonotic.

Griseofulvin (avoid in pregnant animals)

Snuffling, sneezing, dyspnea, vestibular disease, depression chromodacryorrhea, upper respiratory tract disease and/or pneumonia; murine respiratory mycoplasmosis (MRM)

Mycoplasma pulmonis

Common; improve nutrition, husbandry; decrease intracage ammonia levels.

Combination enrofloxacin 10 mg/kg and doxycyline 5 mg/kg beneficial; tetracycline, tylosin

Filobacterium rodentium

Often concurrent with mycoplasmas or viruses.

Sulfamerazine, ampicillin, chloramphenicol, enrofloxacin

Serosanguinous to mucopurulent nasal discharge, rhinitis, conjunctivitis, otitis media

Streptococcus pneumoniae

Immunocompromised animals at greatest risk.

Oxytetracycline

Rough coat, hunched, oculonasal discharge, dyspnea, granulomatous pneumonia; pseudotuberculosis

Corynebacterium kutscheri

Immunocompromised animals at greatest risk; antimicrobials will not eliminate infection.

Ampicillin, chloramphenicol, tetracyclines

Conjunctivitis, panophthalmitis, oculonasal discharge, dyspnea, head tilt

Pasteurella pneumotropica

Immunocompromised animals at greatest risk.

Enrofloxacin

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Urogenital

Central nervous system

0005858931.INDD 751

Tetracyclines

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Gastrointestinal

Diarrhea, dehydration, anorexia, death; Tyzzer disease

Clostridium piliforme

Diarrhea, dry coat, unthriftiness, death

Salmonella enteriditis

Infertility, oophoritis, salpingitis, metritis, pyometra

Mycoplasma pulmonis

Tylosin, fluoroquinolones, tetracyclines

Preputial gland abscess

P. pneumotropica, Staphylococcus aureus

Chloramphenicol, fluoroquinolones

Head tilt, circling, torticollis, otitis interna

Mycoplasma pulmonis ± secondary bacterial invaders

Fluoroquinolones, chlormaphenicol, tylosin

Zoonotic; recommend culling affected animals.

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Table 36.8  Antimicrobial treatment in guinea pigs. Caution: most uses and dosages are extra-­label. Site

Disease/clinical signs

Common infecting organisms

Comments

Suggested drugs

Integument

Enlarged lymph nodes; cervical lymphadenitis

Streptococcus zooepidemicus; also Streptococcus moniliformis and Yersinia pseudotuberculosis

May cause septicemia; complete surgical excision of infected lymph nodes beneficial.

Chloramphenicol, fluoroquinolones

Abscesses

Staphylococcus aureus, Streptococcus spp., Pseudomonas aeruginosa, Pasteurella multocida, Corynebacterium pyogenes

Secondary to bite wounds (especially males), trauma, dental disease.

Fluoroquinolones, trimethoprim-­ sulfa, chloramphenicol, azithromycin, metronidazole

Mastitis

Klebsiella spp., Staphylococcus spp., Streptococcus spp., Pasteurella spp., E. coli, Proteus spp.

Hot compresses; milk out infected glands.

Chloramphenicol, trimethoprim-­sulfa

Swollen, ulcerated foot; ulcerative pododermatitis; osteomyelitis

Staphylococcus aureus, Actinomyces spp.

Secondary to inappropriate bedding, hypovitaminosis C, trauma.

Chloramphenicol, fluoroquinolones, trimethoprim-­ sulfa, azithromycin, metronidazole

Circular areas of alopecia, crusts; pruritus

Trichophyton mentagrophytes, Microsporum canis

Zoonotic.

Fluconazole, itraconazole, ketoconazole, terbinafine, griseofulvin (avoid in pregnant animals)

Rhinitis, tracheitis, otitis media, oculonasal discharge, upper respiratory tract disease and/or pneumonia

Bordetella bronchiseptica, Streptococcus zooepidemicus, Streptococcus pneumoniae, Streptococcus zooepidemicus

Bordetella bronchiseptica commonly carried by dogs and rabbits; some success with Bordetella bacterins.

Amikacin, fluoroquinolones, chloramphenicol

Respiratory

S. pneumoniae, S. zooepidemicus, K. pneumoniae Gastrointestinal

0005858931.INDD 752

Anorexia, diarrhea, enteritis, death

Clostridiodes difficlie, Salmonella typhimurium, S. enteriditis, E. coli, Yersinia pseudotuberculosis, Pseudomonas aeruginosa, Listeria monocytogenes

Amikacin, fluoroquinolones, chloramphenicol Concurrent fluid therapy essential; amikacin best for P. aeruginosa.

Chloramphenicol, systemic amikacin or gentamicin

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Diarrhea, dehydration, bloating, death

Clostridiodes difficile, E. coli

Spontaneous, or following administration of antimicrobials.

Metronidazole, chloramphenicol, control pain

Anorexia, ascites, diarrhea, death; Tyzzer disease

Clostridium piliforme

Recent weanlings, predisposed by crowding, poor sanitation.

Tetracyclines

Diarrhea; coccidiosis

Eimeria caviae

Most common in juveniles.

Sulfonamides

Failure to gain, weight loss, diarrhea, death; cryptosporidiosis

Cryptosporidium wrairi

In humans, newer macrolides such as roxithromycin and azithromycin have shown some efficacy.

Sulfonamides

Metritis, pyometra, abortions, stillbirths

Bordetella bronchiseptica, Streptococcus spp., Corynebacterium pyogenes, Staphyloccus spp., E. coli

Ovariohysterectomy recommended in nonbreeding sows.

Fluoroquinolones, trimethoprim-­ sulfa, chloramphenicol

Orchitis, epididymitis

Bordetella bronchiseptica, Streptococcus spp.

Cystitis

Staphylococcus pyogenes, Staphylococcus spp., fecal coliforms

Urinary calculi often present.

Trimethoprim-­sulfa, fluoroquinolones

Eye

Ocular discharge; conjunctivitis

Chlamydophila caviae, Bordetella bronchiseptica, Streptococcus pneumoniae

Topical treatment; often secondary to hypovitaminosis C.

Tetracyclines, fluoroquinolones, chloramphenicol

Ear

Head tilt, otitis media/interna

Streptococcus pneumoniae, Streptococcus zooepidemicus, Bordetella bronchiseptica, Staphylococcus aureus

General

Anorexia, soft stools, dyspnea, hepatitis, lymphadenitis, septicemia, death

Salmonella typhimurium, S. enteriditis

Urogenital

0005858931.INDD 753

Chloramphenicol, systemic amikacin or gentamicin

Fluoroquinolones, trimethoprim-­ sulfa, chloramphenicol, metronidazole Zoonotic; recommend culling infected animals.

Treatment not recommended

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Site

Disease/clinical signs

Common infecting organisms

Comments

Suggested drugs

Integument

Dermatitis, abscesses

Staphylococcus spp., Streptococcus spp., Corynebacterium spp., Pasteurella spp., Actinomyces spp., E. coli

Secondary to bite wounds; debride and flush.

Ampicillin, chloramphenicol, fluoroquinolones

Cervical masses with sinus tracts containing thick yellow-­green pus, actinomycoses

Actinomyces spp.

Debride and flush.

Clavulanic acid-­amoxicillin, chloramphenicol

Skin black, dam ill, dehydrated; acute gangrenous mastitis

Staphylococcus spp., coliforms

Immediate surgical excision of infected gland; contagious between dams.

Clavulanic acid-­amoxicillin, chloramphenicol

Glands firm, scarred, not painful or discolored; chronic mastitis

Staphylococcus spp., E. coli

Contagious between dams; appears insidiously when kits 3 weeks old.

Treatment generally ineffective

Alopecia, crusts, hyperkeratosis, broken hair shafts

Trichophyton mentagrophytes, Microsporum canis

Zoonotic.

Itraconazole, griseofulvin (avoid in pregnant animals)

Dyspnea, cyanosis, upper respiratory tract disease and/or pneumonia

Streptococcus zooepidemicus, Streptococcus pneumoniae, E. coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Bordetella bronchiseptica, Listeria monocytogenes

Secondary to influenza virus, respiratory syncytial virus, canine distemper virus.

Ampicillin, tetracyclines, fluoquinolones

Respiratory

0005858931.INDD 754

Pneumocystis jiroveci

Trimethoprim-­ sulfamethoxazole

S. pneumoniae, S. zooepidemicus, K. pneumoniae

Amikacin, fluoroquinolones, chloramphenicol

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Table 36.9  Antimicrobial treatment in ferrets. Caution: most uses and dosages are extra-­label.

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Urogenital

0005858931.INDD 755

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Gastrointestinal

Dental tartar, gingivitis, periodontal disease

Multiple etiologies

Improve diet; dentistry.

Metronidazole

Inappetence, vomiting, bruxism, diarrhea, melena, hypersalivation, anemia, gastritis, gastric/duodenal ulceration; Helicobacter mustelae gastritis

Helicobacter mustelae

Rule out foreign body, lymphoma, Aleutian disease, coronavirus.

Amoxicillin 10 mg/kg PLUS metronidazole 20 mg/kg PLUS bismuth subsalicylate 17 mg/kg (1 ml/kg) combined and given PO, q 12 h for 14–­21 days OR clarithromycin 25 mg/kg PLUS omeprazole 1 mg/kg PO, q 24 h OR enrofloxacin 4 mg/kg PLUS colloidal bismuth subcitrate 6 mg/kg PO, q 12 h

Diarrhea, wasting, tenesmus, prolapsed rectum; proliferative bowel disease

Lawsonia intracellularis

Acute gastric distension, dyspnea, cyanosis, sudden death; gastric bloat

Clostridium perfringens

Treat as for bloat in canine patients.

Metronidazole

Fever, bloody diarrhea, lethargy

Salmonella newport, S. typhimurium, S. choleraesuis

Zoonotic; recommend culling infected animals.

Treatment not recommended

Weight loss, diarrhea, vomiting, granulomatous inflammation; mycobacteriosis

Mycobacterium spp.

Zoonotic potential –­consider culling.

Chloramphenicol, tylosin

Diarrhea; coccidiosis

Coccidia spp.

Sulfonamides

Diarrhea; giardiasis

Giardia spp.

Metronidazole

Straining, hematuria, cystitis

Staphylococcus spp., Proteus spp.

Urolithiasis often present.

Fluoroquinolones, ampicillin, sulfonamides

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Table 36.10  Antimicrobial treatment in chinchillas. Caution: most uses and dosages are extra-­label. Site

Disease/clinical signs

Common infecting organisms

Comments

Suggested drugs

Integument

Abscesses

Staphylococcus aureus, Streptococcus spp., Pseudomonas spp.

Secondary to wounds; complete surgical excision beneficial.

Chloramphenicol, tetracyclines, fluoroquinolones

Patches of alopecia, scales on nose, ears, and feet

Trichophyton mentagrophytes

Zoonotic.

Griseofulvin (avoid in pregnant animals), itraconazole, fluconazole

Respiratory

Anorexia, upper respiratory tract disease, dyspnea and/or pneumonia

Pasteurella multocida, Bordetella spp., Streptococcus pneumoniae, Pseudomonas aeruginosa

Overcrowding, high humidity, poor ventilation are predisposing factors. Amikacin best for Pseudomonas aeruginosa.

Fluoroquinolones, trimethoprim-­ sulfa, chloramphenicol, amikacin

Gastrointestinal

Anorexia, decreased fecal output, diarrhea, enteritis, sudden death

Yersinia enterocolitica, Clostridium perfringens, E. coli, Proteus spp., Salmonella typhimurium, S. enteriditis, Pseudomonas aeruginosa, Listeria monocytogenes, Corynebacterium spp.

Concurrent fluid therapy essential; sulfonamides best for Listeria monocytogenes.

Chloramphenicol, trimethoprim-­ sulfa, fluoroquinolones, metronidazole (use with caution)

Diarrhea, dehydration, bloating, death

Clostridium spp., E. coli

Spontaneous, or following administration of antimicrobials.

Metronidazole (use with caution), chloramphenicol, trimethoprim-­sulfa

Diarrhea ± rectal prolapse; giardiasis

Giardia spp.

Depression, abortions

Listeria monocytogenes

Metritis, fever, purulent vaginal discharge

E. coli, Pseudomonas spp., Staphylococcus spp., Streptococcus spp.

Aminoglycosides, fluoroquinolones

Otic

Vestibular signs, head tilt, anorexia; otitis media and/or interna

Pseudomonas aeruginosa, Listeria monocytogenes, anaerobes

Fluoroquinolones, trimethoprim-­ sulfa, chloramphenicol

Central nervous system

Depression, ataxia, convulsions, sudden death

Listeria monocytogenes

Highly susceptible.

Trimethoprim-­sulfa, tetracyclines

General

Septicemia, death

Streptococcus spp., Enterococcus spp., Pasteurella multocida, Klebsiella pneumoniae, Actinomyces spp., Fusobacterium necrophorum

Zoonotic; recommend culling infected animals.

Chloramphenicol, fluoroquinolones

Urogenital

0005858931.INDD 756

Fenbendazole, metronidazole (use with caution) Highly susceptible.

Sulfonamides, tetracyclines

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Site

Disease/clinical signs

Integument

Abscesses

Common infecting organisms

Comments

Suggested drugs

Pasteurella multocida, Staphylococcus aureus, Pseudomonas spp., Streptococcus spp., Bacteroides spp.

Can be located anywhere on body; complete surgical excision beneficial.

Chloramphenicol, tetracyclines, fluoroquinolones

Ulcerative podocermatitis; “sorehock”

Staphylococcus aureus, Pasteurella multocida

Often secondary to inappropriate substrate.

Chloramphenicol, tetracyclines, fluoroquinolones

Dermatitis

Staphylococcus aureus

Usually secondary to poor husbandry.

Chloramphenicol, tetracyclines, fluoroquinolones

Ulceration/necrosis of face, feet; dental and internal abscesses (Schmorl disease)

Fusobacterium necrophorum

Associated with poor hygiene and husbandry.

Cephalosporins, chloramphenicol, tetracyclines, metronidazole

Wet chin, dewlap (“slobbers”) or urine scald (“hutch burn”); exudative dermatitis

Pseudomonas aeruginosa, Streptococcus spp., Staphylococcus spp.

Secondary to moist skin. P. aeruginosa may turn fur green. Correct underlying cause (dental disease, obesity, inappropriate waterers).

Fluoroquinolones, amikacin, gentamicin

Mastitis

Staphylococcus aureus, Pasteurella spp., Streptococcus spp.

Hot compresses; milk out affected glands often.

Amikacin, fluoroquinolones, chloramphenicol, tetracyclines

Alopecia, scaling, crusting on eyelids, at base of ears, and muzzle

Trichopyton spp., Microsporum spp.

Zoonotic.

Griseofulvin (avoid in pregnant animals), itraconazole, ketoconazole, terbinafine

Crusty lesions on nose and lips ± concurrent genital lesions

Treponema cuniculi

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Table 36.11  Antimicrobial treatment in rabbits. Caution: most uses and dosages are extra-­label.

Parenteral penicillin, cephalexin, tetracyclines, chloramphenicol

(Continued)

0005858931.INDD 757

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Site

Disease/clinical signs

Respiratory

Snuffling, oculonasal discharge, conjunctivitis, upper respiratory tract disease and/or pneumonia

Gastrointestinala

Urogenital

0005858931.INDD 758

Common infecting organisms

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Table 36.11  (Continued)

Comments

Suggested drugs

Pasteurella multocida

Very common; treatment rarely curative.

Parenteral penicillin, fluoroquinolones, tetracyclines, amikacin, gentamicin

Bordetella bronchiseptica, Staphylococcus aureus, Pseudomonas aeruginosa

Usually secondary to Pasteurella multocida.

Amikacin, fluoroquinolones, tetracyclines

Diarrhea, death; iota-­enterotoxemia

Clostridium spiroforme

Spontaneous, or following administration of antimicrobials.

Metronidazole, chloramphenicol

Diarrhea; coccidiosis

Eimeria spp.

Hepatic or intestinal; improve sanitation.

Sulfonamides

Diarrhea, death; colibacillosis

E. coli

Especially neonates 1–­14 days old and weanlings.

Sulfonamides, fluoroquinolones, amikacin

Diarrhea, death

Salmonella spp., Pseudomonas spp.

Concurrent fluid therapy essential.

Chloramphenicol, fluoroquinolones

Diarrhea, death; Tyzzer disease

Clostridium piliforme

Tetracyclines

Reddening, edema to dry, scaly, slightly raised areas of external genitalia; venereal spirochetosis (“rabbit syphilis”)

Treponema paraluiscuniculi

Parenteral penicillin, tetracyclines, chloramphenicol

Abortion

Listeria monocytogenes, Pasteurella multocida

Trimethoprim-­sulfa, chloramphenicol, tetracyclines

Cystitis

E. coli, Pseudomonas spp.

Trimethoprim-­sulfa, fluoroquinolones

Orchitis, metritis, uterine abscesses

Pasteurella multocida, Staphylococcus aureus

Chloramphenicol, tetracycline, gentamicin

Polydypsia, polyuria, depression, anorexia, renal failure

Leptospira spp.

Contact with wild rodents; diagnosis by serology.

Parenteral penicillin

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Clear to white discharge from one or both eyes, conjunctivitis

Pasteurella multocida, Staphylococcus aureus

Treat topically; flush tear ducts.

Chloramphenicol, tetracycline, aminoglycosides

Central nervous system

Head tilt, nystagmus, torticollis; “wry neck”

Pasteurella multocida

Usually due to otitis media.

Chloramphenicol, fluoroquinolones

General

Ataxia, torticollis, tremors, convulsions, lethargy, anorexia, pyrexia, septicemia

Encephalitozoon cuniculi, Pasteurella multocida, Listeria monocytogenes

Diagnosis by clinical signs and serology.

Fenbendazole, albendazole, tetracyclines, fluoroquinolones, aminoglycosides, tetracyclines, chloramphenicol

a  Where applicable, provide aggressive supportive care, including fluids (SC, IV, intraosseous), analgesics, high-­fiber diet (via syringe or nasogastric tube if necessary), cisapride or metaclopramide, excellent nursing care; cholestyramine at 2 g/20 ml water q 24 h by gavage may bind bacterial toxins.

0005858931.INDD 759

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Ocular

11-05-2024 12:34:58

Antimicrobial Therapy in Rabbits, Rodents, and Ferrets

V.A.L. syrup, canned pumpkin, cooked sweet potato, coconut milk, raspberry-­flavored gelatin, etc. The availability of a suitable vehicle, compatible with both the antimicrobial and the patient, is limited only by the imagination of the veterinarian. Manual administration of pills, capsules, and liquids to rabbits and rodents is made ­challenging by their relatively long, narrow oral cavity, large tongue, and small gape. Ferrets have the wider gape characteristic of most carnivores, but generally dislike having their mouths pried open, and may bite. In ­ferrets, rabbits, and some larger rodents, administration of pills and capsules can be accomplished with careful use of a pilling device designed for cats. Guinea pigs and ­chinchillas have fleshy cheek invaginations just behind the incisors that act as “one-­way valves.” Small pills or pill pieces can be pushed past these invaginations with the fingers (through the diastema). The presence of the substance in the mouth stimulates a chewing response that aids intake, especially if the medication is somewhat palatable. In rabbits and rodents, liquids can be given using a small syringe inserted part way into the  mouth to avoid dribbling and stimulate a  ­swallowing response. Gentle restraint and careful cleaning of the face and chin will help minimize stress and prevent skin irritation. Several companies specialize in incorporating test compounds or medical therapies into  palatable diets, treats, or feed for a wide variety of animals used in biomedical research. Administration in the food or water is generally reserved for treatment of large numbers of ­animals, such as in research facilities, rabbitries, chinchilla farms, and pet-­breeding operations, where individual dosing would be time-­consuming and impractical. Problems inherent with mass medication include variable intake by sick animals, reduced palatability of the food or water, uneven distribution of the drug, and possible water quality effects on the chemical composition of the compound. Injectable antimicrobials are most often administered to small mammals subcutaneously

in the loose skin over the shoulders. The procedure is quick and minimally stressful when performed correctly. Concurrent fluid therapy can also be given in this large space, provided the two compounds are compatible. Small rodents can be restrained with one hand and injected with the other. The rodent is firmly grasped by the scruff, and either left standing or partially lifted off the exam table, while the injection is administered. Positive reinforcement following the procedure facilitates repeat treatments. Larger rodents and rabbits can be wrapped in a towel or restrained by an assistant to facilitate injection. Ferrets should be securely grasped by the scruff or around the neck in a “turtle-­neck” hold to prevent excessive wriggling. Careful restraint is particularly necessary for rabbits to prevent thrashing and spinal fractures, for chinchillas to prevent damage to the fur (“furslip”), for gerbils to prevent degloving of the tail, and for ferrets and hamsters to minimize the risk of bites. Intraperitoneal injection is suitable for smaller rodents, and is a common route of drug administration in laboratory animals. The procedure is quick and easy to perform, which minimizes patient discomfort. The rodent is firmly scruffed and turned upside down to expose the abdomen. Injections are given in the mid to lower right quadrant, to avoid puncturing the cecum. Intraperitoneal injection in animals with voluminous intestines, such as guinea pigs and rabbits, is not recommended. Small rodents generally lack sufficient muscle mass to accommodate intramuscular injections. Soft tissue trauma and irritation, leading to self-­mutilation, may occur, and drug uptake can be unreliable. In larger rodents, rabbits, and ferrets, intramuscular injections can be given in the lumbar, gluteal, or quadriceps muscles, taking care not to penetrate the bone or sciatic nerve. Alternative routes of drug administration are generally easier and safer, and therefore preferable. Topical antimicrobial preparations, especially those containing corticosteroids, should be used sparingly and cautiously in rabbits and

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rodents. Due to their fastidious grooming ­habits, ingestion of amounts of drug sufficient to cause undesirable systemic effects may result. In addition, use of oil-­based products should be avoided if possible, especially in chinchillas and gerbils, which require dust-­baths to keep their fur healthy. Ophthalmic preparations are less concentrated than other topical medications, and are useful not only for the eyes but for other parts of the body too. These preparations can also be injected into the nasolacrimal ducts of rabbits following flushing, or instilled into the nares. The intravenous route of antimicrobial administration is not used routinely and is usually reserved for initial treatment of ­critically ill patients. In ferrets and rabbits, a catheter can be placed in the cephalic vein for administration of fluids and other drugs, and the ear vein is sometimes catheterized in ­rabbits but may result in subsequent ear necrosis. Injections can be attempted directly into the ear, cephalic, lateral saphenous, medial saphenous, femoral, or tail vein of some small mammals, but this requires a large amount of skill and often anesthesia of the patient. In severely debilitated patients in which venous access is  not possible, placement of an intraosseous catheter in the tibia or femur may be indicated. Nebulization of antimicrobial drugs is ­sometimes used to treat upper and lower airway disease in small mammal pets. A mask may be tolerated by some animals, or a small chamber, such as an anesthesia induction chamber, can be used. Patients should be supervised at all times during confinement to detect undue stress, overheating, or other problems. Gavage is used primarily in experimental studies, where accurate administration of the drug is critical. In rabbits and ferrets, soft plastic feeding tubes or catheters can be introduced into the esophagus through a speculum. The barrel of a 3  ml syringe with the end cut off works well in rabbits. In rodents, curved, ­ball-­ended metal or plastic feeding needles are commercially available for oral dosing. Correct

restraint and gentle insertion, allowing gravity and the swallowing reflex to pass the tube into the esophagus, are critical for success but once the technique is mastered, it is quick and ­relatively stress free for the animal. Administration of oral antimicrobials in very debilitated patients can also be accomplished through placement of a nasogastric tube (in rabbits) or esophagostomy tube (in rabbits, larger rodents, and ferrets), especially if repeated administrations are necessary and the  animal becomes unduly stressed by oral manipulations, or in animals that also require nutritional supplementation. Nasogastric tube placement in rabbits is not technically difficult to perform and several descriptions of the  procedure are available in the literature. Esophagostomy tube placement requires general anesthesia; however, postoperative animal comfort is greater than with nasogastric tubes, breathing is not compromised, and tubes are rarely pulled out. Antimicrobial-­impregnated implants are used primarily for treatment of facial and tooth-­root abscesses in rabbits. In biomedical research, small mammal models have been used to study the elution kinetics and other effects on bone formation and implant integration of various antimicrobial-­coated orthopedic implants for use in people.

­Animal Numbers and Use The number of animals requiring treatment and their intended use must always be kept in mind when prescribing antimicrobial medications in small mammals. The method of treating a single small mammal patient will often differ significantly from that of treating hundreds of animals being bred for the pet trade, used as research subjects, being farmed for fur, or, in the case of rabbits or guinea pigs, being raised for meat. The cost and feasibility of treatment, the effect of the drug on the animal, the deleterious effects that handling may have on the animal, and the possibility of the

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­Animal Numbers and Us  761

Antimicrobial Therapy in Rabbits, Rodents, and Ferrets

animal being consumed by humans are some of the factors that will affect the choice of antimicrobial, its formulation, and the method of administration. Veterinary treatment of animal colonies used for biomedical research must be compatible with the intended scientific use of the animals, so as not to render them useless. Unlike physicians and veterinarians treating pet small ­animals, laboratory animal veterinarians are unlikely to face pressure from their clients (i.e., research scientists) to use antimicrobials in their animals. A colony treatment approach, rather than an individual animal approach, is  commonly implemented and medications are generally incorporated into feed or water. Particular attention must be paid to treatment of genetically modified animals; not only are they usually very valuable and often irreplaceable, they sometimes do not metabolize drugs in predictable ways due to their altered genome. With particularly valuable animals, treatment of a small number and observation for adverse effects may be indicated prior to treating the larger population. Because of the interactions between bacterial populations, changes in the microbiome of antimicrobial-­treated animals are not limited to those bacteria that are susceptible to the administered drug. Antimicrobial treatment leads to transient and/or sustained shifts in host immunity and physiology. As researchers become more sensitized to the effects of antimicrobials, the clinical use of antimicrobials in laboratory animals may be seen as an unacceptable variable in research (Narver, 2017). A number of antimicrobials are prohibited for use in food-­producing animals. One of these is chloramphenicol, a drug frequently used in pet rabbits and guinea pigs due to its effectiveness and relative safety but which has been associated with irreversible aplastic anemia in  humans. Metronidazole is also banned in food-­producing animals in many countries. Strategic and tactful questioning must sometimes take place to determine whether the

patient is strictly a pet or might eventually be used as a food source for humans. Industrial production of rabbits raised for meat, despite being limited to a few countries, has some of the highest antimicrobial administration levels of any food-­producing animals. In an Italian study, antimicrobial classes used in meat rabbit production included tetracyclines (28.2%), polymyxins (17.5%), pleuromutilins (14.0%), sulfonamides (12.6%), bacitracin (12.4%), quinolones (6.1%), diaminopyrimidines (5.1%), aminoglycosides (2.9%), and macrolides (1.2%) (Agnoletti et  al., 2018). Other antimicrobials, such as beta-­lactams and phenicols, were consumed in negligible amounts. Extra-­label medications are occasionally used in meat rabbits; for example, antimicrobial mixtures labeled for prevention of necrotic enteritis in chickens are sometimes added to commercial rabbit feed to manage enteritis problems. Both the producer and the veterinarian are responsible for ensuring no drug residues are present in meat produced for human consumption; however, withdrawal times are not readily available. In these situations, specific withdrawal recommendations can be obtained from a food animal residue avoidance databank (in Canada: www.cgfarad. usask.ca; in the United States: www.farad.org).

­Enhancing Therapeutic Success Treatment of small mammal patients usually involves more than just choosing the correct antimicrobial agent. Many infections are ­secondary to a compromised immune system caused by stress or inadequate nutrition and/ or husbandry. A thorough history is necessary to detect preexisting problems that may be unknown to the client. For example, guinea pigs are unable to synthesize vitamin C and require supplementation of 10–­25  mg daily. Undersupplementation is very common in these animals, leading to subclinical hypovitaminosis C, altered immune function, and ­secondary bacterial infections. Although

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ascorbic acid is present in guinea pig food, the stability varies and the actual amount available depends on the milling date and storage ­conditions. Owners may not be aware that most chows should be used within 90–­180  days of being milled. Other options for provisions of vitamin C include water supplementation, flavored tablets, or daily feeding of small amounts of vitamin C-­rich fruit or vegetables, such as kale, parsley, beet greens, kiwi fruit, broccoli, orange, or cabbage. Oversupplementation of this water-­soluble vitamin is generally not a concern since excess amounts are eliminated from the body. Altered immune function can also occur due to stress. Prey species, such as rabbits and rodents, are particularly susceptible to stressors and the effects can be long-­lasting. Studies in laboratory animals have shown that transport, separation from cage mates, and confinement in an unfamiliar container can affect the cardiovascular, endocrine, immune, central nervous, and reproductive systems, and it can take at least two days for rabbits to acclimate to a new environment after travel. Noise and odors are also stressful to rabbits and rodents, and their heart and respiratory rates can increase rapidly in response to catecholamine release. Prey species are particularly sensitive to predator odors so an attempt should be made to minimize exposure of rabbits and rodents to dogs, cats, and ferrets, and their vocalizations and smells while in the clinic. Prey species instinctively mask any signs of weakness or illness. In addition, their quiet nature, secretive habits, and confinement to a cage can allow them to become debilitated before the average owner notices a problem. Being handled by unfamiliar people with unfamiliar voices and scents, having samples collected, and the unfamiliar hospital environment will add to the overall stress load. Rodents, especially guinea pigs, seem to lack a strong will to live, so good nursing and ­supportive care are necessary for both physical  and psychological health. Due to their small  size, sick rodents can quickly become

hypothermic and debilitated from not eating or drinking. Administration of warmed fluids subcutaneously, provision of supplemental heat (taking care not to overheat the animal), and hospitalization in a safe, quiet environment are almost always indicated. Ample amounts of soft, comfortable bedding and placement of food and water stations within easy reach are also important. If the animal is  not eating, ­gentle handling and syringe feeding a palatable diet are indicated, taking care to administer the food slowly to prevent aspiration. Pain behaviors can be difficult to detect in  rodents but include anorexia, unkempt fur,  piloerection, immobility or restlessness, lethargy, pressing of the abdomen to the floor or table, bruxism, hunched posture, half-­ closed eyes, isolation from a group, unusual aggression, and guarding of specific areas of the body. Analgesics should be used as necessary. Ensuring the animal stays clean and well groomed is also important. During hospitalization, assessment of the animal’s condition, especially body weight, should be performed 2–­3 times a day to monitor progress. Facilities housing large groups of animals, such as rabbitries, fur farms, and pet-­breeding farms, must have sufficient environmental controls in place to maintain the animals at the appropriate temperature and ensure adequate ventilation. Heat or cold stress can compromise the animals, and excessive build-­up of ammonia can cause irritation to the mucous membranes, creating a portal for entrance of bacteria. It is important to recognize that, in addition to prescribing antimicrobial therapy in small mammal patients, and demonstrating the correct method of administration to ensure compliance, veterinarians must also advise clients on correct management, nutrition, and husbandry practices. A good understanding of the normal anatomy, physiology, and behavior of these vulnerable patients will help both the veterinarian and client provide them with the best care possible.

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­Enhancing Therapeutic Succes  763

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­References and Bibliography Agnoletti F, et al. 2018. Longitudinal study on antimicrobial consumption and resistance in rabbit farming. Int J Antimicrob Agents 51(2):197. Alt V, et al. 2011. Effects of gentamicin and gentamicin-­RGD coatings on bone ingrowth and biocompatibility of cementless joint prostheses: an experimental study in rabbits. Acta Biomater 7:1274. American Veterinary Medical Association (AVMA). 2022. AVMA 2022 Pet Ownership and Demographic Sourcebook. www.avma. org/resources-­tools/reports-­statistics/ us-­pet-­ownership-­statistics Animal Medicinal Drug Use Clarification Act (AMDUCA) of 1994. www.fda.gov/animal-­ veterinary/guidance-­regulations/animal-­ medicinal-­drug-­use-­clarification-­act-­1994-­ amduca#:~:text=The%20Animal%20 Medicinal%20Drug%20Use,for%20animals% 20under%20certain%20conditions Bennett RA. 2004. Advances in the treatment of rabbit abscesses. North American Veterinary Conference, Orlando, FL. Boot R. 2012. Frequent major errors in antimicrobial susceptibility testing of bacterial strains distributed under the Deutsches Krebsforschungszentrum Quality Assurance Program. Lab Anim 46(3):253. Booth R. 2000. General husbandry and medicinal care of sugar gliders. In: Bonagura J (ed.) Kirk’s Current Veterinary Therapy XIII. WB Saunders, Philadelphia, pp. 1157–­1163. Capdevila S, et al. 2007. Acclimatization of rats after ground transportation to a new animal facility. Lab Anim 41:255. Compendium of Veterinary Products. www.vetalytix.com/en/products/cvp-­ compendium-­of-­veterinary-­products Cox SK. 2007. Allometric scaling of marbofloxacin, moxifloxacin, danofloxacin and difloxacin pharmacokinetics: a retrospective analysis. J Vet Pharmacol Therapeut 30:381.

Cox SK, et al. 2004. Allometric analysis of ciprofloxacin and enrofloxacin pharmacokinetics across species. J Vet Pharmacol Therapeut 27:139. Donnelly TM, Brown CJ. 2004. Guinea pig and chinchilla care and husbandry. Vet Clin Exot Anim 7:351. Fendt M. 2006. Exposure to urine of canids and felids, but not herbivores, induces defensive behaviour in laboratory rats. J Chem Ecol 32:2617. Fernández M, et al. 2023. Current situation of bacterial infections and antimicrobial resistance profiles in pet rabbits in Spain. Vet Sci 10(5):352. Gebru E, et al. 2011. Allometric scaling of orbifloxacin disposition in nine mammalian species: a retrospective analysis. J Vet Med Sci 73(6):817. Graham JE, Schoeb TR. 2011. Mycoplasma pulmonis in rats. J Exotic Pet Med 20:270. Harkness JE, et al. 2010. Harkness and Wagner’s Biology and Medicine of Rabbits and Rodents, 5th edn. Wiley-­Blackwell, Ames. Hirakawa Y, et al. 2010. Prevalence and analysis of Pseudomonas aeruginosa in chinchillas. BMC Vet Res 6(52):1. Hunter RP, Isaza R. 2008. Concepts and issues with interspecies scaling in zoological pharmacology. J Zoo Wild Med 39:517. Lipman NS, Perkins SC. 2002. Factors that may influence animal research. In: Fox JG, et al. (eds) Laboratory Animal Medicine, 2nd edn. Academic Press, San Diego, pp. 1156–­1159. Makidon P. 2005. Esophagostomy tube placement in the anorectic rabbit. Lab Anim 34(8):33. Marin P, et al. 2007. Pharmacokinetic-­ pharmacodynamic integration of orbifloxacin in rabbits after intravenous, subcutaneous and intramuscular administration. J Vet Pharmacol Therapeut 31:77. Masini CV, et al. 2005. Non-­associative defensive responses of rats to ferret odor. Physiol Behav 87:72.

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Mitchell MA, Tully Jr TN. 2009. Manual of Exotic Pet Practice. Saunders Elsevier, St Louis. Morris TH. 1995. Antibiotic therapeutics in laboratory animals. Lab Anim 29:16. Narver HL. 2017. Antimicrobial stewardship in laboratory animal facilities. J Am Assoc Lab Anim Sci 56(1):6. Obernier JA, Baldwin RL. 2006. Establishing an appropriate period of acclimatization following transportation of laboratory animals. Inst Lab Anim Res J 47:364. Oglesbee BL. 2011. Blackwell’s Five Minute Veterinary Consult: Small Mammal, 2nd edn. Wiley-­Blackwell, Ames. Papich MG. 2005. Drug compounding for veterinary patients. AAPS J 7:E281. Pollock C. 2003. Fungal diseases of laboratory rodents. Vet Clin Exot Anim 6:401. Porter WP, et al. 1985. Absence of therapeutic blood concentrations of tetracycline in rats after administration in drinking water. Lab Anim Sci 35:71. Powers LV. 2006. Techniques for drug delivery in small mammals. J Exotic Pet Med 15:201. Quesenberry KE, Carpenter JW. 2012. Ferrets, Rabbits, and Rodents: Clinical Medicine and Surgery, 3rd edn. Saunders Elsevier, St Louis. Quesenberry KE, Hillyer EV (eds). 1994. The Veterinary Clinics of North America Small Animal Practice Exotic Pet Medicine II. WB Saunders, Philadelphia. Ramirez HE. 2006. Antimicrobial drug use in rodents, rabbits, and ferrets. In: Giguère SS, et al. (eds) Antimicrobial Therapy in Veterinary Medicine, 4th edn. Blackwell, Ames, pp. 565–­580.

Rosenthal KL. 2004. Therapeutic contraindications in exotic pets. Sem Avian Exot Pet Med 13:44. Rosenthal KL, et al. 2008. Rapid Review of Exotic Animal Medicine and Husbandry. Manson Publishing, London. Sedgwick CJ. 1993. Allometric scaling and emergency care: the importance of body size. In: Fowler ME (ed.) Zoo and Wild Animal Medicine, 3rd edn. WB Saunders, Philadelphia, pp. 235–­241. Smith AJ. 1992. Husbandry and medicine of African hedge-­hogs (Atelerix albiventris). J Small Exotic Anim Med 2:21. Smith AJ. 1999. Husbandry and nutrition of hedgehogs. Vet Clin North Am Exotic Anim Pract 2:1. Spenser EL. 2004. Compounding, extralabel drug use, and other pharmaceutical quagmires in avian and exotics practice. Sem Avian Exot Pet Med 13:16. Staples LG, McGregor IS. 2006. Defensive responses of Wistar and Sprague-­Dawley rats to cat odour and TMT. Behav Brain Res 172:351. Streppa HK, et al. 2001. Applications of local antibiotic delivery systems in veterinary medicine. J Am Vet Med Assoc 219:40. Swallow J, et al. 2005. Guidance on the transport of laboratory animals. Lab Anim 39:1. Turner PV, et al. 2012. Oral gavage in rats: animal welfare evaluation. J Am Assoc Lab Anim Sci 51:25. Van Praag E, et al. 2010. Skin Diseases of Rabbits. MediRabbit.com. Wightman SR, et al. 1980. Dihydrostreptomycin toxicity in the Mongolian gerbil, Meriones unguiculatus. Lab Anim Sci 40:71.

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 ­References and Bibliograph

37 Antimicrobial Therapy in Reptiles J. Scott Weese

Antimicrobial therapy is an important ­component in clinical management of reptiles affected with various infectious diseases but is complicated by limited species-­specific ­pharmacokinetic and safety data, limitations in extrapolation of in vitro susceptibility testing designed for endothermic species, and little objective efficacy data. Selecting the appropriate antimicrobial agent for reptiles is based on similar principles and considerations common to antimicrobial selection in domestic species. However, this process is more complicated in reptiles because of the number and diversity of species, their unique anatomical and physiological features, the diversity of infectious agents, and even behavioral characteristics that make safety an important factor in drug and route considerations. This chapter will focus on the process of antimicrobial selection in reptiles while highlighting the unique differences and challenges associated with selecting antimicrobial agents for these species.

I­ nfectious Agents A diverse range of bacterial, viral, and fungal infections are important causes of ­morbidity  and mortality in captive reptiles (Austwick  and  Keymer,  1981; Cooper,  1981;

Jacobson,  1999,  2007). While some of these also affect humans and domestic animals, a large proportion are mainly or solely pathogens of reptiles and amphibians, limiting the amount of available data. Infections caused by Gram-­negative ­bacterial pathogens are common in captive reptiles (Paré et al., 2006), often with bacteria from environmental reservoirs and water. Although not as well documented, Gram-­negative ­bacterial infectious diseases are also reported in wild populations of reptiles. For instance, die-­offs of American alligators (Alligator mississippiensis) have been associated with Aeromonas hydrophila infections (Shotts et al., 1972). Gram-­positive aerobic bacteria, anaerobes, and other organisms such as mycobacteria, Mycoplasma, and Chlamydophila can also cause disease (Jacobson,  2007; Stewart,  1990; Homer et  al.,  1994; Jacobson and Telford,  1990; Jacobson et al., 1989, 2002; Soldati et al., 2004). Fungal infections are also common in captive reptiles (Paré et  al.,  2006; Austwick and Keymer, 1981; Migaki et al., 1984), including a wide range of fungi of environmental origin. Fungal infections are often difficult to treat because disease can be advanced by the time the diagnosis is made and/or there may be ­serious co-­morbidities that have predisposed to fungal infection. While knowledge about antibacterials is limited, there are even less

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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Antimicrobial Therapy in Reptiles

clincal and pharmacological data on antifungals in reptiles. Other pathogens, including protozoal, helminth, and viral agents, have been described in reptiles (Jacobson,  2007). However, given the relative importance of bacterial and fungal infections and the greater body of evidence regarding treatment, this chapter will focus on  bacterial and fungal pathogens. Tables  37.1–­37.3 provide examples of more common diseases and potential treatment options. Selected dosage regimens for reptiles are shown in Table 37.4.

D ­ iagnostic Testing Principles of diagnostic testing for reptiles are no different than for other species and both lack of testing and suboptimal diagnostics can compromise treatment decisions. However, there are various challenges that must be considered in reptiles. Once a bacterial or mycotic infection is ­suspected in a reptile patient, the accurate ­identification of the primary pathogen is an essential step in choosing the most appropriate antimicrobial. Proper sampling is at the core of

Table 37.1  Antimicrobial drug selection in chelonian infections.

Site or type

Diagnosis

Skin, shell, and subcutis

Epidermitis/ dermatitis

Abscesses

Common infecting organisms

Potential empirical antimicrobial choices

Various

Topical antimicrobials or biocides should be considered for superficial infections in terrestrial species

Citrobacter freundii

Amikacin, gentamicin, enrofloxacin, marbofloxacin, trimethoprim-­sulfonamide

Serratia

Ceftazidime

Proteus morganii

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Providencia rettgeri

Trimethoprim-­sulfa, enrofloxacin, marbofloxacin, amikacin, gentamicin, chloramphenicol

Pseudomonas aeruginosa

Amikacin, gentamicin, ceftazidime, enrofloxacin, marbofloxacin

Dermatophilus chelonae

Penicillin G, ampicillin, tetracycline

Mycobacterium chelonei

Amikacin, clarithromycin

Mucor

Immersions in malachite green solution

Aspergillus

Fluconazole

Various

Incision and drainage is the cornerstone of treatment. Antimicrobials are only indicated if there is significant cellulitis or systemic disease.

Pasteurella testudinis

Amikacin, ceftiofur, ceftazidime

Escherichia coli

Enrofloxacin, marbofloxacin, gentamicin, amikacin

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768

Table 37.1  (Continued)

Site or type

Oral cavity

Respiratory tract

Diagnosis

Stomatitis

Pneumonia

Rhinitis

Gastrointestinal tract

Enteritis

Liver abscesses

Common infecting organisms

Potential empirical antimicrobial choices

Providencia

Trimethoprim-­sulfa, enrofloxacin, marbofloxacin, amikacin, gentamicin, chloramphenicol, piperacillin-­tazobactam

Bacteroides

Metronidazole

Fusobacterium

Penicillin G, metronidazole

Aeromonas hydrophila

Enrofloxacin, marbofloxacin, gentamicin, amikacin, tetracyclines, ceftiofur, ceftazidime

Pseudomonas aeruginosa

Amikacin, gentamicin ceftazidime, enrofloxacin, marbofloxacin

Vibrio

Ticarcillin, enrofloxacin, marbofloxacin

Pseudomonas aeruginosa

Amikacin, gentamicin ceftazidime, enrofloxacin, marbofloxacin

Morganella morganii

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Serratia marcescens

Ticarcillin

Acinetobacter calcoaceticus

Enrofloxacin, marbofloxacin, amikacin

Bacteroides

Metronidazole

Fusobacterium

Metronidazole

Aspergillus

Ketoconazole, fluconazole, itraconazole

Geotrichum candidum

Itraconazole

Beauvaria

Fluconazole

Mycoplasma spp.

Doxycycline, clarithromycin, enrofloxacin, marbofloxacin

Pasteurella testudinis

Enrofloxacin, marbofloxacin, ceftiofur, ceftazidime

Mycoplasma agassizii

Clarithromycin, enrofloxacin, doxycycline

Salmonella

Antimicrobials are not normally indicated for enteric salmonellosis. Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, chloramphenicol

Aeromonas hydrophila

Enrofloxacin, marbofloxacin, gentamicin, amikacin, tetracyclines, ceftiofur, ceftazidime

Salmonella

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Bacteroides

Metronidazole (Continued)

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­Diagnostic Testin 

Antimicrobial Therapy in Reptiles

Table 37.1  (Continued)

Site or type

Diagnosis

Septicemia

Skeletal

Osteomyelitis/ arthritis

Common infecting organisms

Potential empirical antimicrobial choices

Clostridium

Metronidazole, penicillin

Fusobacterium

Metronidazole

Salmonella

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Aeromonas hydrophila

Enrofloxacin, marbofloxacin, gentamicin, amikacin, ceftiofur, ceftazidime

Pseudomonas aeruginosa

Amikacin, gentamicin, ceftazidime, enrofloxacin, marbofloxacin

Pseudomonas

Amikacin, gentamicin, ceftazidime, enrofloxacin, marbofloxacin

Klebsiella

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime. chloramphenicol

Mycobacterium chelonae

Clarithromycin, amikacin, gentamicin

Nocardia

Azithromycin, trimethoprim-­sulfonamide

Eye and adnexa

Conjunctivitis

Mycoplasma agassizii

Topical aminoglycoside, fluoroquinolone, or chloramphenicol

Ear

Otitis interna

Pseudomonas

Amikacin, gentamicin ceftazidime, enrofloxacin, marbofloxacin

Escherichia coli

Amikacin, gentamicin, ceftiofur, ceftazidime, enrofloxacin, marbofloxacin

Proteus

Enrofloxacin, marbofloxacin, amikacin, gentamicin, trimethoprim-­sulfa

Pasteurella testudinis

Enrofloxacin, marbofloxacin, ceftiofur, ceftazidime

Bacteroides Fusobacterium

Metronidazole

Table 37.2  Antimicrobial drug selection in crocodilian infections.

Site or type

Diagnosis

Common infecting organisms

Oral cavity

Stomatitis

Aeromonas hydrophila

Enrofloxacin, marbofloxacin, gentamicin, amikacin, tetracyclines, ceftiofur, ceftazidime

Candida

Nystatin

Various

Topical antimicrobials or biocides should be used when possible for superficial lesions. Systemic antimicrobials may not be required.

Skin

Epidermitis/ dermatitis

Suggested drugs

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770

Table 37.2  (Continued)

Site or type

Respiratory tract

Diagnosis

Pneumonia

Common infecting organisms

Suggested drugs

Dermatophilus

Procaine penicillin G, tetracycline

Morganella morganii

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Pseudomonas aeruginosa

Amikacin, gentamicin, ceftazidime, enrofloxacin, marbofloxacin

Serratia

Enrofloxacin, gentamicin, amikacin, marbofloxacin, chloramphenicol

Klebsiella

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime. chloramphenicol

Aspergillus

Itraconazole

Trichophyton

Fluconazole

Aeromonas hydrophila

Enrofloxacin, marbofloxacin, gentamicin, amikacin, tetracyclines, ceftiofur, ceftazidime

Citrobacter freundii

Amikacin, gentamicin, enrofloxacin, marbofloxacin, trimethoprim-­sulfonamide

Morganella morganii

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Providencia rettgeri

Trimethoprim-­sulfa, enrofloxacin, marbofloxacin, amikacin, gentamicin, chloramphenicol

Escherichia coli

Ampicillin, doxycycline, enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Salmonella

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Beauvaria

Ketoconazole

Fusarium

Itraconazole

Mucor

Fluconazole

Mycoplasma alligatoris

Enrofloxacin, marbofloxacin, oxytetracycline

Yolk infection

Omphalitis

Aeromonas hydrophila

Enrofloxacin, marbofloxacin, gentamicin, amikacin, tetracyclines, ceftiofur, ceftazidime

Liver

Hepatitis

Escherichia coli

Ampicillin, doxycycline, enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Salmonella

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol (Continued)

771

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­Diagnostic Testin 

Antimicrobial Therapy in Reptiles

Table 37.2  (Continued)

Site or type

Diagnosis

Common infecting organisms

Suggested drugs

Aeromonas hydrophila

Enrofloxacin, ofloxacin, marbofloxacin, gentamicin, amikacin, tetracyclines, ceftiofur, ceftazidime

Eye

Uveitis

Aeromonas hydrophila

Enrofloxacin, ofloxacin, marbofloxacin, gentamicin, amikacin, tetracyclines, ceftiofur, ceftazidime

Cardiovascular

Septicemia

Salmonella

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Aeromonas hydrophila

Enrofloxacin, marbofloxacin, gentamicin, amikacin, tetracyclines, ceftiofur, ceftazidime

Mycoplasma alligatoris

Enrofloxacin, oxytetracycline, doxycycline, marbofloxacin

Serosa/joints

Polyserositis/ arthritis

Table 37.3  Antimicrobial drug selection for infections in snakes and lizards.

Site or type

Diagnosis

Oral cavity

Stomatitis

Skin and subcutis

Abscesses

Common infecting organisms

Suggested drugs

Pseudomonas aeruginosa

Amikacin, gentamicin ceftazidime, enrofloxacin, marbofloxacin

Aeromonas hydrophila

Enrofloxacin, marbofloxacin, gentamicin, amikacin, tetracyclines, ceftiofur, ceftazidime

Various

Incision and drainage is the cornerstone of treatment. Antimicrobials are only indicated if there is significant cellulitis or systemic disease.

Proteus

Enrofloxacin, marbofloxacin, amikacin, gentamicin, trimethoprim-­sulfa

Providencia

Trimethoprim-­sulfa, enrofloxacin, marbofloxacin, amikacin, gentamicin, chloramphenicol

Pseudomonas

Amikacin, gentamicin ceftazidime, enrofloxacin, marbofloxacin

Salmonella

Amikacin, gentamicin, ceftiofur, ceftazidime, enrofloxacin, marbofloxacin

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772

Table 37.3  (Continued)

Site or type

Diagnosis

Bacterial dermatitis

Mycotic dermatitis

Respiratory tract

Pneumonia

Common infecting organisms

Suggested drugs

Serratia

Enrofloxacin, gentamicin, amikacin, marbofloxacin, chloramphenicol

Clostridium

Penicillin, metronidazole

Pseudomonas aeruginosa

Amikacin, gentamicin ceftazidime, enrofloxacin, marbofloxacin

Fusobacterium

Metronidazole

Bacteriodes

Metronidazole

Various

Topical antimicrobials or biocides should be used when possible for superficial lesions. Systemic antimicrobials may not be required.

Citrobacter

Amikacin, gentamicin, enrofloxacin, marbofloxacin, trimethoprim-­sulfonamide

Devriesea agamarum

Ceftazidime

Klebsiella

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime. chloramphenicol

Pseudomonas

Enrofloxacin, amikacin, gentamicin, ceftazidime

Geotrichum

Ketoconazole

Fusarium

Itraconazole

Chrysosporium

Fluconazole

Pseudomonas

Amikacin, gentamicin ceftazidime, enrofloxacin, marbofloxacin

Providencia rettgeri

Trimethoprim-­sulfa, enrofloxacin, marbofloxacin, amikacin, gentamicin, chloramphenicol, piperacillin-­tazobactam

Aeromonas hydrophila

Enrofloxacin, marbofloxacin, gentamicin, amikacin, tetracyclines, ceftiofur, ceftazidime

Morganella morganii

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Mycoplasma

Clarithromycin, oxytetracycline (Continued)

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­Diagnostic Testin 

Antimicrobial Therapy in Reptiles

Table 37.3  (Continued)

Site or type

Diagnosis

Gastrointestinal tract

Enteritis

Hepatitis

Skeletal

Osteomyelitis

Common infecting organisms

Suggested drugs

Pseudomonas aeruginosa

Amikacin, gentamicin, ceftazidime, enrofloxacin, marbofloxacin

Aeromonas hydrophila

Enrofloxacin, marbofloxacin, gentamicin, amikacin, tetracyclines, ceftiofur, ceftazidime

Escherichia coli

Ampicillin, doxycycline, enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Salmonella

Antimicrobials are not normally indicated for enteric salmonellosis. Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, chloramphenicol

Pseudomonas aeruginosa

Amikacin, gentamicin, ceftazidime, enrofloxacin, marbofloxacin

Morganella morganii

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Providencia rettgeri

Trimethoprim-­sulfa, enrofloxacin, marbofloxacin, amikacin, gentamicin, chloramphenicol

Aeromonas hydrophila

Enrofloxacin, marbofloxacin, gentamicin, amikacin, tetracyclines, ceftiofur, ceftazidime

Escherichia coli

Ampicillin, doxycycline, enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Salmonella

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Clostridium

Metronidazole

Salmonella

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

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774

Table 37.3  (Continued)

Site or type

Eye

Diagnosis

Subspectacle infections

Uveitis

Conjunctivitis

Common infecting organisms

Suggested drugs

Escherichia coli

Ampicillin, doxycycline, enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime, chloramphenicol

Pseudomonas aeruginosa

Amikacin, gentamicin, ceftazidime, enrofloxacin, marbofloxacin

Pseudomonas aeruginosa

Ophthalmic aminoglycoside or fluoroquinolone

Providencia rettgeri

Trimethoprim-­sulfa, enrofloxacin, marbofloxacin, amikacin, gentamicin, chloramphenicol

Proteus

Enrofloxacin, marbofloxacin, amikacin, gentamicin, trimethoprim-­sulfa

Pseudomonas aeruginosa

Amikacin, gentamicin, ceftazidime, enrofloxacin, marbofloxacin

Serratia

Enrofloxacin, gentamicin, amikacin, marbofloxacin, chloramphenicol

Klebsiella

Enrofloxacin, gentamicin, amikacin, marbofloxacin, ceftiofur, ceftazidime. chloramphenicol

Pseudomonas aeruginosa

Amikacin, gentamicin, ceftazidime, enrofloxacin, marbofloxacin

testing, and this can be complicated in some reptiles based on their housing (e.g., water contamination in aquatic species), infection site (difficulty in safely obtaining a sample), animal size (e.g., too small to collect an optimal volume of blood for culture), and the potential impacts of physical or chemical restraint. Diagnostic specimens should be collected, whenever possible, using approaches to minimize contamination with commensal, surface, and environmental bacteria. If a discrete lesion is present, a biopsy specimen is ideally obtained for both cytological and histological ­examination. Concurrent with

the morphological assessment, a specimen of the lesion is also submitted for culture. Concurrent use of other diagnostic methods such as PCR or immunohistochemistry should be considered, when available. Laboratory issues must also be considered. Reptile specimens would typically comprise a very small percentage of specimens tested by commercial veterinary diagnostic laboratories which can lead to interpretation challenges. For example, laboratories are supposed to only report growth of organisms that are considered potentially clinically relevant and not report

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­Diagnostic Testin 

Antimicrobial Therapy in Reptiles

Table 37.4  Selected dosage regimens for antimicrobial drugs in reptiles. Route of administration Dose

Dose interval

References

American alligator

IM

2.25 mg/kg

96 h

Jacobson, 1988

Gopher tortoise

IM

5 mg/kg

48 h

Caligiuri, 1990

Snakes

IM

5 mg/kg; 2.5 mg/kg

Mader, 1985 1st loading dose; thereafter 72 h

Ball python

IM

3.5 mg/kg

Not given

Various

IM, SC

10–­20 mg/kg 12h

a

Tortoises

IM, SC

50 mg/kg

12h

a

2–­7 days

Drugs

Species

Amikacin

Ampicillin Azithromycin

Carbenicillin

Ceftazidime

Johnson, 1997

Ball python

PO

10 mg/kg

Freshwater crocodile

IM

10 mg/kg

Coke, 2003

Snakes

IM

400 mg/kg

24 h

Lawrence, 1984a

Tortoises

IM

400 mg/kg

48 h

Lawrence, 1986

Sukkheewan, 2022

Snakes

IM

20 mg/kg

72 h

Lawrence, 1984b

Loggerhead sea turtle

IM; IV

20 mg/kg

72 h

Stamper, 1999

Snakes

IM

15 mg/kg

120 h

Adkesson, 2011

Bearded dragons

IM, SC

30 mg/kg

10-­12 days

Churgin, 2014

Chloramphenicol

Snakes

SC

50 mg/kg

12–­72 h depending on species

Clark, 1985

Clarithromycin

Desert tortoise

Oral

15 mg/kg

48–­72 h

Wimsatt, 1999

Oral gavage

15 mg/kg

84 h

Freshwater crocodile

IM, IV

2.5 mg/kg

Poapolathep, 2022

Green sea turtle Hawksbill sea turtle

  IM, IV

6 mg/kg

Wanmad, 2022

Freshwater crocodiles

IM

6 mg/kg

Poapolathep, 2022

Gopher tortoise

IM

5 mg/kg

Ceftiofur crystalline free acid

Danofloxacin

Enrofloxacin

24–­48 h

Wimsatt, 2008

Prezant, 1994

Star tortoise

IM

5 mg/kg

12–­24 h

Raphael, 1994

Loggerhead sea turtle

PO

20 mg/kg

Not given

Jacobson, 2005

5 mg/kg

Not given

James, 2003

PO

10 mg/kg

Not given

IV

5 mg/kg

36 h

Red-­eared slider IM American alligator

Helmick, 2004a

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776

Table 37.4  (Continued)

Drugs

Species

Estuarine crocodile

Route of administration Dose

Dose interval

References

PO, IM, IV

Not given

Martelli, 2009

Green iguana

IM

5 mg/kg

24 h

Maxwell, 2007

Burmese python

IM

10 mg/kg

48 h

Young, 1997

Eastern box turtles Yellow-­bellied sliders River cooters

SC

10 mg/kg

Griffioen, 2020

Green sea turtles

IV, IM

5 mg/kg

Poapolathep, 2021

Fluconazole

Loggerhead sea turtle

SC

21 mg/kg; 10 mg/kg

1st dose; thereafter 5 days

Mallo, 2002

Itraconazole

Kemp’s Ridley sea turtle

PO

15 mg/kg

72 h

Manire, 2003

5 mg/kg

24 h Daily

Spiny lizard

PO

23.5 mg/kg

Ketoconazole

Tortoise

PO

15–­30 mg/kg 24 h

Page, 1991

Marbofloxacin

Loggerhead sea turtle

IM, IV

2 mg/kg

Lai, 2009

Green sea turtle

IM, IV

4 mg/kg

Ball python Metronidazole

Green iguana

10 mg/kg

24 h

Gamble, 1997

Poapolathep, 2020 48 h

Coke, 2006

PO

20 mg/kg

48 h

Kolmstetter, 1998

Yellow rat snake PO

20 mg/kg

48 h

Kolmstetter, 2001

Red rat snake

PO

50 mg/kg

48 h

Bodri, 2006

Red-­eared slider IC turtle

20 mg/kg

48 h

Innis, 2007

American alligator

IV

10 mg/kg

5 days

Helmick, 2004b

Loggerhead sea turtle

IM

41 mg/kg, followed by 21 mg/kg q72h. 82 mg/kg, followed by 42 mg/kg q72h

3 days

Harms, 2004

Piperacillin

Snakes

IM

100 mg/kg

24 h

Hilf, 1991

Ticarcillin

Loggerhead sea turtle

IM

50 mg/kg

24 h

Manire, 2005

100 mg/kg

48 h

Oxytetracycline

a

5 mg/kg

 www.merckvetmanual.com/multimedia/table/antimicrobial-­drugs-­used-­in-­reptiles

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­Diagnostic Testin 

Antimicrobial Therapy in Reptiles

commensal bacteria or those that are likely contaminants. This can create problems with specimens from species where uncommon (for the laboratory) organisms are more likely and where environmental opportunists may be much more relevant than they would be in domestic mammals. This can result in lack of reporting of potentially relevant organisms. Testing conditions also need to be considered, as culture plates are usually incubated at mammalian body temperature. Organisms that grow suboptimally at those higher temperatures may be missed. Some reptile pathogens such as Chlamydophila, Mycoplasma, and mycobacteria are relatively difficult to isolate from routine cultures and also often difficult to see in standard histopathological preparations. Special histological stains, immunohistochemical stains, and molecular techniques are sometimes necessary to detect their presence (Bodetti et al., 2002; Jacobson et  al.,  2004; Johnson et  al.,  2007). Similarly, antimicrobial susceptibility testing is usually performed at 35 °C, and it is unclear how well results would reflect susceptibility in  vivo and at lower temperatures. Another issue is the lack of clinical breakpoints for any antimicrobials for reptiles. Thus, susceptible/ intermediate/resistant determination (if provided) would be based on the susceptibility of the organism to drug levels achieved in serum of mammalian species, something that may not well (or at all) reflect the situation in the target species. It is important to inform the laboratory that the culture specimen is from a reptile and may need special laboratory handling to isolate the pathogen (Origgi and Paré, 2007).

­ usbandry and Immunological H Considerations The next consideration in the process of antimicrobial selection should be an understanding that captive husbandry and the immunological status of the reptile are important. Bacterial and fungal infections tend to become more invasive and clinically apparent in captive

reptiles when husbandry conditions are suboptimal (Cooper, 1981). For example, maintaining reptiles below their optimal temperature range may induce an immunocompromised ­condition in the patient. Furthermore, Vaughn et  al. (1974) demonstrated that some lizards experimentally infected with Gram-­negative bacteria voluntarily selected higher ambient temperatures. This behavior was interpreted as an induced fever, and is thought to help the lizards fight bacterial infections. Given that reptile body temperature affects immune system function, it is imperative to maintain the ill reptile under optimum environmental conditions as an important part of the therapeutic plan. Our understanding of reptile bacterial and mycotic infections has advanced to recognize that reptiles become more susceptible to bacterial diseases when exposed to other pathogens. For example, primary viral infections, such as ophidian paramyxovirus pneumonia and herpes virus stomatitis of tortoises, are associated with severe secondary bacterial infections (Jacobson, 1992; Origgi et al., 2004). Exposure to contaminated environments and a lack of proper quarantine program are important and potentially modifiable risk factors for infection with multiple pathogens.

­ natomical and Physiological A Considerations Reptile anatomy and physiology differ significantly from domestic mammals and can also differ greatly between different reptile species. Reptiles have several unique features that can potentially influence the pharmacokinetics of antimicrobials and the subsequent response to treatment. The carapace and plastron form the ­characteristic shell of chelonians. This unique anatomical feature is composed of an outer keratinized epidermis overlying a base of ­dermal cartilage and bone. The dermal bone is highly vascularized and considered a ­metabolically active tissue (Jacobson,  2007).

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778

The relative metabolic activity and blood ­perfusion of the chelonian shell have led to the recommendation that antimicrobials should be dosed based on their entire body weight and not adjusted to subtract the weight of the ­turtle shell. An anatomical feature of all snakes with eyes and some lizards is the transparent palpebral spectacle (Millichamp et  al.,  1983). This spectacle embryologically represents a fusion of the upper and lower eyelids that permanently covers the cornea, leaving a potential subspectacular space. Infections of this subspectacular space have been reported and are difficult to treat with topically applied antimicrobial agents that do not appear to penetrate this barrier (Millichamp et al., 1983). In treating reptiles with subspectacular infections, a wedge can be carefully excised from the lower half of the spectacle and the appropriate ­antimicrobial drug applied directly through the wedge-­shaped hole onto the surface of the cornea. Most species of reptile have a renal portal system that can shunt blood from the caudal half of the body through the kidneys before reaching the systemic circulation. This blood flow pattern can potentially alter the pharmacokinetics of drugs and is the basis for ­recommendations that intramuscular and subcutaneous injections be given in the cranial half of the reptile body. However, few studies have tested this hypothesis and the theoretical impacts would vary between different antimicrobials. Holz et  al. (1997a) reported that in red-­eared sliders (Trachemys scripta elegans), the blood from the caudal region of the body did not necessarily flow through the kidney via the renal portal system. Instead, the blood draining the caudal portion of the body perfused both the liver and the kidneys, indicating that the renal portal shunt was only partially functional. In a related study, Holz et  al. (1997b) also found that red-­eared sliders receiving gentamicin in either a forelimb or hindlimb had no significant differences in pharmacokinetic parameters, indicating a

minimal pharmacokinetic effect from the renal portal system. In contrast, the same study noted that red-­eared sliders receiving carbenicillin in the hindlimb had significantly lower blood concentrations for the first 12 hours post injection than those that received the same dose in a forelimb. Despite this finding for ­carbenicillin, the authors concluded that this difference was not clinically important and questioned the necessity of forelimb injections (Holz et  al.,  1997b). Because the renal portal system varies in development, anatomy, and function between various groups of reptiles and the pharmacokinetic evidence is conflicting, many clinicians still recommend injecting potentially nephrotoxic drugs and drugs eliminated primarily through the renal system in the cranial half of the body. In contrast to mammalian pus, reptiles infected with bacterial and fungal pathogens tend to develop solid exudates within discrete granulomatous lesions (Montali,  1988; Jacobson,  2007). These pathogens are located within the necrotic center of heterophilic granulomas, within histiocytes (macrophages) in histiocytic granulomas, or near the capsule of chronic granulomas. Granulomas can limit the penetration of many antimicrobial agents into the sites of infection. When possible, surgical removal of the granulomatous mass prior to antimicrobial therapy can improve the chances of a positive therapeutic outcome. Physiological and husbandry factors can also influence drug pharmacokinetics and therefore drug selection in reptiles. The ambient temperature of the reptile enclosure directly affects the pharmacokinetics of antimicrobials. Mader et al. (1985) studied gopher snakes (Pituophis melanoleucus catenifer) given amikacin and housed at ambient temperatures of either 25 °C or 37 °C. When housed at 37 °C, the apparent volume of distribution was larger and body clearance of amikacin was faster. In another study of gopher tortoises (Gopherus polyphemus), the mean residence time of amikacin was significantly shorter in tortoises acclimated to 30 °C than those kept at 20 °C,

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­Anatomical and Physiological Consideration  779

Antimicrobial Therapy in Reptiles

and clearance at 30 °C was approximately twice that in the tortoises kept at 20 °C (Caligiuri et  al.,  1990). In contrast, Johnson et al. (1997) found no significant pharmacokinetic differences among snakes given amikacin and housed at 25 °C and 37 °C. No explanation for this discrepancy was offered, suggesting that the effect of temperature on drug pharmacokinetics is either species specific or requires further evaluation. It is challenging to determine how to apply results such as these to clinical situations. However, it is important to consider the impact of species and environment when determiningg antimicrobial dosing regimens. Therefore, while it can be useful to base dosing decisions on published pharmacokinetic data, it is important to remember that those data may not reflect results at different temperatures or in different reptile species.

­ ehavioral and Safety B Considerations The size and temperament of a reptile can influence antimicrobial drug selection and the route of administration. Some reptiles are extremely timid and nervous, and may not be suitable for repeated handling and intramuscular injections. In such cases, the antimicrobial must be administered orally, preferably in food if the animal is still eating. Most species of reptiles weigh less than 100 g and many lizards are under 30 g as adults. This may limit antimicrobials that can easily be diluted to a concentration that can be precisely and safely injected. At the other end of the spectrum, some reptiles are quite large in size and dangerous to approach. In such cases, a choice may need to be made between a drug that can be administered in a relatively small volume via remote injection dart or orally in food. Oral administration in food can also be challenging when sick animals are hyporexic or anorexic, or in species that go long periods of time between feedings. Venomous snakes

present a similar treatment challenge, since they are dangerous to handle and manipulate for administration of drugs. For these dangerous species, drugs that can be administered every few days are preferred over drugs that must be administered each day.

­ outes of Antimicrobial R Administration Oral antimicrobials are generally used in species not tolerant of injections, when the optimal antimicrobial is available in an oral formulation, or when safety considerations make injections dangerous. Oral medication may also be indicated in rare situations when large numbers of reptiles are infected and must be treated simultaneously. In these situations, the individual administration of drugs may not be practical, and the usage of medicated food may be warranted. However, several problems exist with oral medication of reptiles. Very few pharmacokinetic studies have been performed on drugs administered orally to reptiles. Thus, for the vast majority of antimicrobials, the dose selected will not be based on existing literature. Gastrointestinal transit time impacts drug absorption and varies greatly among the various reptile species. Transit time is usually slowest in the large herbivorous reptiles. For example, the transit time in large tortoises may be as much as 21 days. Even in some carnivorous reptiles, the transit time may be quite prolonged. Carnivorous reptiles, such as pythons, are adapted to infrequent meals and increase their gastric and intestinal mucosa in response to feeding (Secor, 2008). This massive change in gastrointestinal metabolism is likely to influence antimicrobial absorption and treatment frequency. Thus, in reptiles it may be difficult to achieve optimum and consistent therapeutic concentrations of antimicrobials in blood following oral administration. Martelli et  al. (2009) published a pharmacokinetic study of enrofloxacin in estuarine crocodiles

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(Crocodylus porosus) where delayed absorption and subtherapeutic drug concentrations were measured with the oral route. In contrast, repeated twice-­weekly oral doses of clarithromycin in desert tortoises (Gopherus agassizii) attained target drug concentrations (Wimsatt et al., 2008). Clearly, oral absorption in reptiles is species and drug specific and requires further investigation. While clinicians can often administer oral antimicrobials in the food of reptiles actively feeding, orally medicating reptiles that are anorexic or that feed infrequently is often difficult. In giant tortoises, extracting the head beyond the shell margins and then forcing the  mouth open is usually impossible. Furthermore, these overzealous efforts to force the mouth open can injure the keratinized epidermal hard parts over the mandibles and dentary bones. In general, giant tortoises must be anesthetized and a pharyngostomy tube inserted for oral medication. Pharyngostomy tubes are easy to insert and routinely used in tortoises and other chelonians (Norton et al., 1989). As a generalization, nonvenomous snakes are the easiest group of reptiles to medicate orally. The mouth of most snakes is simple to open and the glottis is easy to see and avoid. In these snakes, a lubricated French catheter or nasogastric tube is passed down the esophagus with minimal resistance. Since the cranial esophagus is extremely thin in most snake species, the end of the catheter should be round and smooth. The use of excessively rigid catheters should be avoided as they may penetrate the esophageal mucosa. While the stomach of most snakes is located from one-­third to half the distance from the head to the cloaca, it is not necessary to pass a catheter this far. In most situations, passing the catheter halfway between the stomach and oral cavity is satisfactory. Most of the injectable antimicrobials commonly used in reptiles are injected intramuscularly, subcutaneously and occasionally intracoelomically. Intravenous administration

is challenging because peripheral vessels are difficult to catheterize (Jacobson et al., 1992). While blood can be collected from several vascular sites in different species of reptiles, most of this sampling is “blind” and is not suitable for repetitive intravenous infusions (Olson et al., 1975; Samour et al., 1984). Intramuscular and subcutaneous injections are practical and provide the most predicable drug absorption. Snakes and lizards are the easiest reptiles to inject intramuscularly because of the large epaxial dorsal muscles of the body associated with the ribs and vertebrae. In lizards, the forelimb muscle masses are usually small, which limits injection volumes. The best site for intramuscular injections in chelonians is the pectoralis musculature located medial and caudal to the base of the forelimbs just within the cranial margins of the shell. Despite the ease of intramuscular and ­subcutaneous drug administration, placing large volumes of irritating drugs into reptile muscles can result in significant irritation and tissue damage, including development of necrotizing skin and soft tissue disease from a single injection. Anecdotally, numerous severe reactions have been reported with enrofloxacin injection. This may be in part because of frequent use of the drug but it is not unreasonable to suspect that this drug truly poses a higher risk for tissue damage, and enrofloxacin administered by injection is best avoided in reptiles whenever possible. Injectable drugs with a prolonged elimination are potentially useful in reptiles that are difficult or dangerous to handle. Adkesson et al. (2011) reported that a long-­acting formulation of ceftiofur maintained adequate plasma concentrations for five days in ball pythons (Python regius). However, care must be taken when extrapolating across species. For ­example, cefovecin has a long elimination half-­ life in dogs (133 h) (Stegemann, 2006a) and cats (166 h) (Stegemann, 2006b) and is typically dosed at 14-­days interval, but much shorter or more variable half-­lives have been

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­Routes of Antimicrobial Administratio  781

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reported in some other species, including green iguanas (3.9 h) (Thuesen et  al.,  2009), red-­eared sliders (6.8h) (Sypniewski et al., 2017), and Hermann’s tortoises (20.8 h) (Nardini et al. 2014). Intracoelomic injections are a potential route of delivery, but this is infrequently used because of limited pharmacokinetic and safety data. The potential injury from an inappropriately placed or irritating drug in the coelomic space requires further investigation and intracoelomic injection should be avoided unless adequate safety and dosing data are available for the particular animal species and drug. Local and topical antimicrobials may be useful in some situations. Topical administration may be viable for superficial infections in nonaquatic species that are amenable to frequent handling. Local antimicrobial approaches, through antimicrobial impregnated beads or gels, can provide sustained therapeutic levels at deep sites, and can be particularly useful in situations where frequent administration of systemic antimicrobials is not possible or the optimal antimicrobial cannot be safety administered at the doses required for systemic use. Local antimicrobials can also be used alongside parenteral antimicrobials. Absorbable materials (e.g., gels, calcium sulfate) are preferred over PMMA beads as they do not require a secondary surgical procedure for removal.

Testing decisions are ideally made based on clinical trial data, alongside species-­and drug-­ specific pharmacokinetic data that are relevant to the disease (e.g. location of infection) and management conditions (e.g. temperature). These are also uncommonly available. Of the 7,500 species of reptiles, pharmacokinetic studies have been reported for a few drugs in a small number of species commonly kept in captivity. As in most scientific literature there is a publication bias toward reporting pharmacokinetic studies that lead to dosage recommendations, versus those that fail to produce useful recommendations (Stamper et al., 2003; Thuesen et  al.,  2009). Studies focused on the metabolism, tissue concentrations, and potential toxicity of antimicrobials in reptiles are rare in the literature (Hunter et al., 2003). Understanding of likely pathogens, typical susceptibility patterns and clinical experience with response to treatment are often the maintay of antimicrobial decision-­making, albeit with numerous potential limitations. Various formularies can be found, and some can provide excellent guidance. However, limitations of these must be recognized as dosing recommendations are often made with little or no supporting data, and some recommendations are not consistent with current understanding of pharmacology or antimicrobial therapy.

­Antimicrobial Drug Selection

­ llometric Scaling to Estimate A Drug Dosages

Ideally, antimicrobial selection is based on ­culture and susceptibility results from a proper specimen, using validated methods, and ­evaluated with breakpoints that are ­relevant to the treated animal species. This unfortunately is rarely (if ever) available. Despite the limitations described above, culture and susceptibility testing can be an important guide. However, empirical treatment is often needed in lieu of, or in advance of, laboratory testing results. Suggestions are made in Tables 37.1–37.3.

The lack of pharmacokinetic studies and even the paucity of empirical dosage recommendations may require extrapolation of drug dosages from domestic mammals. It should be recognized that this is a highly tenuous approach, and it is presumably best to extrapolate dosing from other reptile species, whenever available. However, data gaps may require highly empirical decision making. In practice, there are three main methods to estimate proper therapeutic drug dosages (Hunter and Isaza, 2008).

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The first method is to use an established drug dose derived from pharmacokinetic studies in other species. By this method, a 20 mg/kg dose of amoxicillin in dogs is applied across all reptile species regardless of size. Using a set dose results in a linear increase in the amount of drug administered as body weight increases. Although common, this method tends to overdose larger animals and underdose smaller animals. The second method is similar except that it takes the established dosage in a specific species and makes an additional assumption that links the dosage to the metabolic rates of both species. Using this method, the established drug dosage from one species is adjusted based on the ratio of the calculated metabolic rate of the patient over the calculated metabolic rate of the target species: Patient dose set dose species X 

Pmet

patient

/ Pmet

species X

This method, termed metabolic scaling, is popular in zoological medicine and has been described for use in reptiles (Pokras et al., 1992; Mayer et al., 2006). Unfortunately, this method of allometric scaling is controversial because formulas to estimate reptile metabolic rates are inconsistent between reptile species. For many mammals, the following allometric equation is considered the best estimate of the basal metabolic rate: Pmet

70 Kg0.75

where Pmet is the minimum energy costs and Kg is the body weight in kilograms (Kleiber, 1961). In contrast, a similar allometric equation: Pmet 10 Kg0.75 is suggested for use in all reptile species (Pokras et al., 1992; Mayer et al., 2006). However, when Jacobson (1996) reviewed the subject, this ­single reptile equation was not considered

appropriate for all reptiles, since the constant varied from 1 to 5 for snakes and 6 to 10 for lizards, with no values for chelonians or crocodilians available. Additionally, significant data variability in those groups was noted where scientific studies have been performed. For instance, Bartholomew and Tucker (1964) measured the metabolic rate in lizards ranging in size from 0.002 to 4.4 kg and calculated the allometric equation to be Pmet = 6.84(Kg0.62). This is different from findings by Bennet and Dawson (1976) for 24 species of lizards, ranging from 0.01 to 7 kg, for which the equation: Pmet

7.81 Kg0.83

was determined. Further, when one looks at studies with snakes, still more equations can be calculated (Galvao et  al.,  1965). In determining resting metabolic rates of 34 species of boas and pythons, the mass exponents of different species showed considerable variation (Chappell and Ellis, 1987). The problem with metabolic scaling is that reptiles represent a very heterogeneous group of vertebrates and because of this, no single equation relating metabolic rate to body mass can be developed for calculating antimicrobial dosages. Differences in body temperature, season, reproductive status, nutrition, and overall physiology are just a few of the variables that may ultimately influence metabolic rates, making application of a single equation impossible. While at first glance, metabolic scaling may appear better than extrapolation, using a single equation for all reptile species may not be valid. In the third method, the allometric scaling of measured pharmacokinetic parameters is used for extrapolation of drug doses between species. This method is commonly used in the pharmaceutical industry to extrapolate pharmacokinetic parameters between laboratory mammal species to humans (Hunter and Isaza,  2008). Using known pharmacokinetic parameters as the basis for extrapolation has  theoretical advantages over calculated metabolic rates. However, when Maxwell and Jacobson (2007)

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­Allometric Scaling to Estimate Drug Dosage  783

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compared the pharmacokinetics of enrofloxacin over a wide range of green iguana sizes, they found that clearance and other pharmacokinetic parameters did not scale adequately allometrically. Thus, this method of allometric scaling using pharmacokinetic parameters also needs further investigation in reptiles.

A ­ ntimicrobial Stewardship The most widely recommended and used antimicrobials in reptiles have tended to be from higher tier drug classes, such as those categorized as “highest priority critically important antimicrobial” by the World Health Organization. This includes fluoroquinolones and third-­generation cephalosporins, which likely account for most of the antimicrobial use in reptiles in some situations. There are various reasons for this, including limited or no alternatives being authorized for use in reptiles and the availability of pharmacokinetic data for some species/drug combinations. Much also likely relates to habit and comfort with a limited range of antimicrobials, leading to very common empirical use of drugs such as enrofloxacin and ceftazidime. While antimicrobial use in reptiles accounts for a miniscule amount of antimicrobial use in animals, there are some issues that must be considered, for both animal and public health. Indeed, public health considerations may be greater in pet reptiles compared to other common pet species because of the frequency of reptile-­associated salmonellosis and the public health consequences of fluoroquinolone and third-­generation cephalosporin resistance. While fluoroquinolones or third-­generation cephalosporins may be indicated as primary treatment options for some pathogens, there should be more consideration of use of lower tier drugs such as tetracyclines, potentiated ­sulfonamides, and penicillins. These may be equally effective, and in some situations safer, than common higher tier drugs, while creating fewer antimicrobial resistance concerns. When

higher tier drugs are used empirically, culture and susceptibility results should be evaluated to determine not only if the empirical treatment will likely be effective, but also to see if a lower tier drug (e.g., potentiated sulfonamide, tetracycline) might be equally effective, allowing for ­de-­scalation for continued treatment. Lower tier options should also be considered more often in animals with nonlife-­threatening disease. Topical treatments may be effective for skin and superficial soft tissue infections in terrestrial species that are amenable to frequent handling and can potentially replace systemic treatment.

C ­ onclusion Antimicrobial therapy decisions can be challenging in reptiles. The paucity of data often results in a need for empirical decisions, often with very limited relevant supporting data. Good diagnostic testing, logical consideration of likely pathogens, and understanding of typical susceptibility patterns can provide a reasonable basis for treatment decisions. Dosing is a further challenge and is not likely to be easier in the near future given limited study in the field. Consideration of available dosing data for the target or related species, general pharmacological principles and anecdotal clinical data will continue to be the basis of most decisions. However, it is important to remember that drug formularies and recommendations, including those in this chapter, often lack much foundation. Formularies also sometimes contain outdated recommendations that may have limited likelihood of being useful and that could be harmful. Therefore, it is important to continually assess available data to optimize treatment plans for this diverse range of animals.

Acknowledgment This chapter is based on that in the previous edition so the current author wishes to acknowledge Ramiro Isaza and Elliott R. Jacobson for their work.

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 ­References and Bibliograph

Antimicrobial Therapy in Reptiles

Nardini G, et al. 2014. Pharmacokinetics of cefovecin sodium after subcutaneous administration to Hermann’s tortoises (Testudo hermanni). Am J Vet Res 75 (10):918. Norton TM, et al. 1989. Medical management of a Galapagos tortoise (Geochelone elephantopus) with hypothyroidism. J Zoo Wildl Med 20:212. Olson GA, et al. 1975. Techniques for blood collection and intravascular infusions of reptiles. Lab Anim Sci 25:783. Origgi FC, Paré JA. 2007. Isolation of pathogens. In: Jacobson ER (ed.) Infectious Diseases and Pathology of Reptiles. CRC Press, Boca Raton, p. 667. Origgi FC, et al. 2004. Experimental transmission of a herpes-­virus in Greek tortoises (Testudo graeca). Vet Pathol 41:50. Page CD, et al. 1991. Multiple dose pharmacokinetics of ketoconazole administered to gopher tortoises (Gopherus polyphemus). J Zoo Wildl Med 22:191. Paré JA, et al. 1997. Cutaneous mycoses in chameleons caused by the Chrysosporium anamorph of Nannizziopsis vriesii (Apinis) Currah. J Zoo Wildl Med 28:443. Paré JA, et al. 2003. Cutaneous mycobiota of captive squamate reptiles with notes on the scarcity of Chrysosporium anamorph of Nannizziopsis vriesii. J Herp Med Surg 13:10. Paré JA, et al. 2006. Microbiology: Fungal and bacterial diseases of reptiles. In: Mader DR (ed.) Reptile Medicine and Surgery, 2nd edn. Saunders Elsevier, St Louis, p. 217. Plowman CA, et al. 1987. Septicemia and chronic abscesses in iguanas (Cyclura cornuta and Iguana iguana) associated with a Neisseria species. J Zoo Anim Med 18:86. Poapolathep S, et al. 2020. Pharmacokinetics of marbofloxacin in Green sea turtles (Chelonia mydas) following intravenous and intramuscular administration at two dosage rates. J Vet Pharmacol Therapeut 43 (2):215. Poapolathep S, et al. 2021. Enrofloxacin and its major metabolite ciprofloxacin in green sea turtles (Chelonia mydas): an explorative

pharmacokinetic study. J Vet Pharmacol Therapeut 44(4):575. Poapolathep S, et al. 2022. Pharmacokinetic profiles of clarithromycin in freshwater crocodiles (Crocodylus siamensis). J Vet Pharmacol Therapeut 45(2):147. Pokras MA, et al. 1992. Therapeutics. In: Beynon PH, et al. (eds) Manual of Reptiles. British Small Animal Veterinary Association, Gloucester. Prezant RM, et al. 1994. Plasma concentrations and disposition kinetics of enrofloxacin in gopher tortoises (Gopherus polyphemus). J Zoo Wildl Med 25:82. Raphael BL, et al. 1994. Pharmacokinetics of enrofloxacin after a single intramuscular injection in Indian star tortoises (Geochelone elegans). J Zoo Wildl Med 25:88. Samour HJ, et al. 1984. Blood sampling techniques in reptiles. Vet Rec 114:472. Secor SM. 2008. Digestive physiology of the Burmese python: broad regulation of integrated performance. J Exp Biol 211:3767. Shotts EB, et al. 1972. Aeromonas induced death among fish and reptiles in an eutrophic inland lake. J Am Vet Med Assoc 161:603. Soldati G, et al. 2004. Detection of mycobacteria and chlamydiae in granulomatous inflammation of reptiles: a retrospective study. Vet Pathol 41:388. Stamper MA, et al. 1999. Pharmacokinetics of ceftazidime in loggerhead sea turtles (Caretta caretta) after single intravenous and intramuscular injections. J Zoo Wild Med 30:32. Stamper MA, et al. 2003. Pharmacokinetics of florfenicol in loggerhead sea turtles (Caretta caretta) after single intravenous and intramuscular injections. J Zoo Wildl Med 34:3. Stegemann MR, et al. 2006a. Pharmacokinetics and pharmacodynamics of cefovecin in dogs. J Vet Pharmacol Therapeut 29(6):501. Stegemann MR, et al. 2006b. Pharmacokinetics of cefovecin in cats. J Vet Pharmacol Therapeut 29(6):513.

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Stewart JS. 1990. Anaerobic bacterial infections in reptiles. J Zoo Wildl Med 21:180. Sukkheewan R, et al. 2002. Pharmacokinetic characteristics of azithromycin in freshwater crocodiles (Crocodylus siamensis) after intramuscular administration at three different dosages. J Vet Pharmacol Therapeut 45(6):501. Sypniewski L, et al. 2017. Cefovecin pharmacokinetics in the red-­eared slider. J Exotic Pet Med 26(2):108. Thuesen LR, et al. 2009. Selected pharmacokinetic parameters for cefovecin in hens and green iguanas. J Vet Pharmacol Therapeut 32:613. Vaughn LK, et al. 1974. Fever in the lizard Dipsosaurus dorsalis. Nature 252:473.

Wanmad W, et al. 2022. Pharmacokinetic characteristics of danofloxacin in green sea (Chelonia mydas) and hawksbill sea (Eretmochelys imbricata) turtles. J Vet Pharmacol Therapeut 45(4):402. Wimsatt J, et al. 1999. Clarithromycin pharmacokinetics in the desert tortoise (Gopherus agassizii). J Zoo Wildl Med 30:36. Wimsatt J, et al. 2008. Long-­term and per rectum disposition of clarithromycin in the desert tortoises (Gopherus agassizii). J Am Assoc Lab Anim Sci 47:41. Young LA, et al. 1997. Disposition of enrofloxacin and its metabolite ciprofloxacin after IM injection in juvenile Burmese pythons (Python molurus bivittatus). J Zoo Wildl Med 28:71.

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 ­References and Bibliograph

38 Antimicrobial Therapy in Zoo and Wildlife Species Ellen Wiedner and Robert P. Hunter

I­ ntroduction Although the knowledge base of zoo and ­wildlife medicine continues to grow rapidly, basic conundrums continue to stymie the ­clinician seeking to treat nondomestic species. Often, simply determining whether an animal is actually ill, let alone affected with a treatable infectious organism, can be remarkably difficult. Nondomestic species, both wild and captive, tend to conceal disease extremely well. Frequently, sudden death is the only indication that an animal harbored significant disease, although necropsy may indicate a long-­ standing infection. Nevertheless, even if an antemortem diagnosis of infection is made in a wild animal ­species, evidence-­based decisions for antimicrobial use may be extremely challenging. Pharmacological studies relevant to zoo and wildlife species continue to be scant. This is due in part to the technical difficulty of performing drug studies in wild animals as well as the perceived risks of such research in rare or endangered species. Additionally, the unusual physiology of nonmammalian species such as reptiles, amphibians, avians, and fish can add yet another layer of complexity to drug studies. Even nondomestic mammalian taxa may metabolize drugs in surprising ways compared to domestic counterparts.

Lacking hard data on pharmacokinetics, safety, and efficacy of most antimicrobials, ­clinicians caring for zoo and wildlife species must extrapolate drugs and doses from unrelated species, a less than ideal situation. Finally, the technical aspects of providing a course of antimicrobial therapy to wild animals, most of which are uncooperative and potentially dangerous even if sick, provide a final level of complexity.

­ linical Breakpoint Interpretation C in Zoological Medicine Global events and perceptions regarding the use of antimicrobial agents in animals have placed even more importance on the essential role of antimicrobial susceptibility testing of bacteria isolates from animals. However, little information is available on microorganism–­ antimicrobial–­host interactions with zoo species. Currently, veterinary-­specific breakpoints have been compiled and published in the Clinical Laboratory Standards Institute’s (CLSI) VET01S (CLSI, 2021). This also ­indicates the criteria for several antimicrobials that are commonly used in veterinary medicine yet do not have veterinary species-­specific breakpoints. The number of drugs in the latter category gets smaller with each revision, but

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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even the veterinary-­specific criteria must be interpreted with caution when applied to zoological species for treatment considerations and therapy. This is emphasized in VET09 from CLSI (CLSI, 2019a). The first reason for this caution is how the breakpoints are determined by the Veterinary Antimicrobial Susceptibility Testing subcommittee. The path for breakpoint determination is outlined in the CLSI VET02 document “Development of Quality Control Ranges, Breakpoints, and Interpretive Categories for Antimicrobial Agents Used in Veterinary Medicine” (CLSI, 2019b). This document outlines the pathway for antimicrobial agents for the setting and recommendation of veterinary-­ specific clinical breakpoints. “S” is the susceptible interpretive test category implying that the infection, due to the isolate, may be effectively treated with the normal dosage regimen of an antimicrobial recommended for that type of infection and causative bacterial species. This is the fundamental piece of information that is often not understood by the attending zoo veterinarian because of limited information about pharmacokinetics in the species to be treated and the lack of accurate prediction of pharmacokinetics across species (Hunter and Isaza, 2008). The recorded results indicate that the isolate is S or I or R for the culture submitted when tested against the diagnostic lab’s standard array of antimicrobial agents. The S/I/R are reported using the information provided in VET01S previously mentioned. The reporting institution does not know, in many cases, the host species, route of administration, or pharmacokinetics of the antimicrobial agent in the treated species being evaluated and reported. They generally only have two pieces of the puzzle: microorganism and class representative antimicrobial agent tested. Most veterinarians then assume that if the microorganism is reported as S, they simply treat it with that agent and positive results will follow. Antimicrobial susceptibility testing of bacteria of animal origin is ultimately intended for the selection of antimicrobial agents for

better clinical outcomes. The premise of ­veterinary antimicrobial susceptibility testing (VAST) is that in vitro test results can be used to guide the veterinarian in antimicrobial drug therapeutic decision making when the testing is performed in a standardized and reproducible manner. Just as important, clinicians must understand that antimicrobial resistance is not necessarily an inherent or absolute characteristic of bacteria, but rather that resistance indicates the crossing of a threshold. Although “S” and “R” are usually considered binary characteristics, in fact resistance can only be identified if a clinical breakpoint or threshold of antimicrobial concentration is predetermined and agreed upon by regulatory agencies and/or standard-­setting organizations such as the CLSI. The threshold (“interpretive criterion”) cannot be arbitrarily determined (e.g., by saying that all bacteria with a zone of inhibition of less than “X” mm are resistant) but must be validated with the appropriate data, including knowledge of concentrations of antimicrobial drug that can be achieved in an animal (pharmacokinetics), the best presentation of the drug to the bacteria in the host (pharmacodynamics), range of concentrations of antimicrobial drug required to inhibit the growth of populations of wild-­type bacterial pathogens, and clinical outcome of treatment of the pathogen with approved or commonly accepted doses of an antimicrobial drug. It goes without saying that the determination of S is a complex process for the indication(s) on the product label. Attempting to determine this value for each and every “bug/drug/species” combination in zoological medicine would be an astronomical undertaking. To avoid misinterpretation, CLSI VET01S and VET09 recommend that diagnostic laboratories only test and report breakpoints for antimicrobials appropriate for therapeutic or control use. Antimicrobials could be added based on specific therapeutic needs (such as for specific zoological species where a specific agent and formulation are commonly used).

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Given the limited number of antimicrobial agents approved for use in some animal species, “extra-­label” use of antimicrobial agents is common. The US Congress, in the Animal Medicinal Drug Use Clarification Act (AMDUCA), has defined extra-­label use as the “actual use or intended use of a drug in an animal in a manner that is not in accordance with the approved labeling.” This includes, but is not limited to, use in species or for indications (disease or conditions) not listed in the labeling; use at a dosage level higher than that stated in the label; and use of routes of administration other than those stated in the labeling. This type of use has regulatory acceptance in many countries (e.g., extra-­label use permitted under the AMDUCA regulations). While laboratory personnel should be familiar with the extra-­label use of antimicrobial agents in animals, the laboratory client is responsible for using the compound appropriately in the animal. The laboratory client is also responsible for using the agent appropriately for the various animal types or categories (e.g., calves, lactating dairy cattle). The laboratory client assumes all responsibility for efficacy, safety, and residue avoidance with extra-­label uses of antimicrobials. The laboratory should be prepared to offer advice to the veterinarian to enable appropriate decision making. Although the laboratory may choose to modify the list of antimicrobials it tests and reports, on the basis of public health concerns, it needs to be done in consultation with appropriate experts, based on good clinical judgment, and in accordance with recognized principles of judicious use. Veterinarians working with minor or zoological species should make themselves aware of the tables provided in VET01S and VET09 so as to understand what the breakpoints are and what bug/drug/species indications they are based on. Numerous antimicrobials are approved for use in different animal species by the US FDA-­ CVM or comparable regulatory authorities in other countries. Factors such as microbiological activity, clinical efficacy, and pharmacology

should be considered for therapy, indications, and restrictions. CLSI document VET01S lists compounds in groups in which drugs are approved for use in the indicated animal ­species by the US FDA-­CVM (Groups A, B, C, and D). It is most appropriate to report those antimicrobials that have veterinary-­specific interpretive criteria over those using human interpretive ­criteria (Group A). These antimicrobials have demonstrated an acceptable correlation between in  vitro susceptibility test results and clinical criteria outlined in VET01S. While antimicrobials evaluated using human interpretive guidelines (Group B) may perform adequately in diseased animals, the interpretive relationship for veterinary applications has not been ­determined. Some antimicrobials are FDA-­CVM approved for use in a specific animal species but have neither veterinary-­specific nor human-­specific interpretive criteria (Group C) and reporting interpretive criteria from one animal species to another (extra-­label use, Group D) is not recommended due to various differences in dosages and pharmacokinetics.

I­ ntra-­and Interspecies Dose Extrapolation Species differences in drug absorption, ­distribution, metabolism, and excretion (ADME) for numerous antimicrobials have been well documented for domestic species; however, there is limited information concerning the ADME of drugs in nondomestic species (Hunter, 2017). Lack of approved drugs and/or pharmacokinetic data in the literature for zoo species is a major issue for veterinarians attempting to treat these animals. Zoological medicine practitioners take approved antimicrobials (veterinary or human) and extrapolate their use to nonapproved species. The range of animals a zoo veterinarian cares for varies from very small invertebrates (e.g., honeybees) to megavertebrates such as elephants and whales. The decision on the dose, duration,

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­Intra-­ and Interspecies Dose Extrapolatio  793

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and treatment interval is often made with ­limited species-­specific pharmacokinetic and/ or efficacy information. Because of the monetary value of these animals or their status as endangered species, the method of “trial and error” for antimicrobial dosage selection is inappropriate. In zoological medicine, various methods have been used in an attempt to extrapolate or predict safe and effective dosage regimens (Hunter and Isaza,  2008). The simplest and most typical method of extrapolating a dosage to a nondomestic species is to use a mg/kg dose established for another domestic species or humans. However, this calculation results in a linear increase in the amount of drug administered as body weight increases. Although common, this method tends to overdose large animals and underdose small animals. A second method is similar except that it takes the approved dose in a specific species and makes an additional assumption that links the dosage to a physiological function or anatomical feature. Examples are the use of basal metabolic rate or body surface area as the basis for dosage extrapolation. Allometric scaling of pharmacokinetic parameters is the final method of dosage extrapolation between species. This is commonly used in the pharmaceutical industry to establish the first dosage in human drug investigations. Adaptation of this method for zoological medicine is believed to enhance the ability to estimate therapeutic dosages for nondomestic species. This tool, when used appropriately, can provide an estimate for designing dosage regimens. The example of differences in ketoprofen inversion across species emphasizes the need to understand and be aware of the assumptions when designing treatment regimens based on allometric scaling data (Hunter et al., 2003). Just as mammals can range from a few grams to thousands of kilograms, reptiles and birds can also vary in body weight across a wide range. It has been suggested that it is impossible to derive a single equation correlating body

mass to metabolic rate for all 6000 species of reptiles (Funk,  2000). Without knowledge of the extent and route of elimination of an administered antimicrobial, extrapolation of dosage regimens from one class to another is difficult, if not impossible, with any certainty. Some reports question the practical use of this approach (Hunter et  al.,  2008; Mahmood et al., 2006; Martinez et al., 2006). Before extrapolation of any drug dose, the veterinarian should appreciate the mathematical and physiological assumptions involved and the limitations that are associated with allometry. Careful consideration of the available literature to understand the route of elimination and the extent of metabolism of antimicrobials will greatly assist in determining allometric relationships of pharmacokinetic parameters. There is a continuing need to consider and apply methods for reducing the size and risk of extrapolation error, as this can affect both target animal safety and therapeutic response. Data from at least one large animal (nonhuman and a body weight >70  kg) should be included to reduce potential error (Mahmood et al., 2006).

­ Practical Example of Allometry A and Breakpoints An example of how the above information can be interpreted and potentially misused is the case of Mycobacterium tuberculosis (TB) susceptibility testing and the treatment of this bacterial disease in elephants (Loxodonta africana and Elephas maximus). Unlike cattle and other livestock, which are more likely to be infected with M. bovis and are euthanized if positive, in the US, elephants are recognized for their rarity and value and are treated rather than culled. Mandatory testing and treatment of elephants with TB is overseen by the US Department of Agriculture (USDA), and guidelines for drug administration in pachyderms have been derived from those established for humans (USDA, 2008). This is

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different from the situation in Europe, where zoonotic transmission to humans is the greater concern and individual animals are likely to be euthanized. Susceptibility testing for this pathogen is described in detail, for human isolates, in the CLSI M24 document (CLSI, 2018). Using the human pharmacokinetics of the primary antituberculosis drugs, the results of in  vitro susceptibility testing of these drugs appear to correlate well with the clinical effectiveness of these antimicrobials in human patients. The interpretive criteria, or breakpoints, are provided in Table 38.1. In elephants, the antituberculosis drugs differ significantly regarding their pharmacodynamics and pharmacokinetics in humans. In addition, the metabolic state of Mycobacterium tuberculosis significantly affects its susceptibility to antimicrobials. Optimization of dosage of antituberculosis drugs is necessary to achieve maximum drug exposure at the site of infection in order to maximize the reduction in viable M. tuberculosis and minimize the ­emergence and selection of resistance (de Steenwinkel et al., 2010). Table 38.1  Breakpoints for selected antituberculous drugs used in elephants.

Agent

Breakpoint concentration (μg/ml)

7H10 agar

7H11 agar

Isoniazid

0.2

0.2

Rifampin

1.0

1.0

Ethambutol

5.0 10

7.5 NR

Pyrazinamide

NR

NR

Levofloxacin

1.0

ND

Moxifloxacin

0.5

0.5

Ofloxacin

2.0

2.0

Streptomycin

2.0 10

2.0 10

Where multiple values are provided, the second is when resistance has occurred and the drugs are used as “second-­line therapies” (modified from CLSI, 2018). NR, not recommended; ND, not determined.

There are published reports on the “­population” pharmacokinetics of several antituberculosis drugs in African and/or Asian elephants which were used to develop the multidrug treatment protocols for elephants published in the USDA Elephant TB Guidelines, and were modeled after human disease (Peloquin et  al.,  2006). The issues with these types of extrapolations have been previously discussed (Hunter and Isaza, 2008). Using the human breakpoints for isoniazid from the M24 and the plasma concentrations reported by Maslow et al. (2005), one could conclude that the likelihood for efficacy is high with all reported concentrations >0.2 μg/ml for the doses and routes of administration evaluated, but many concentrations were greater than 5×, which seems excessive and could be contributing to the adverse drug reactions reported by some clinicians (Isaza, personal communication). Maslow’s group (2005) suggests that the area under the curve (AUC) may be the driving pharmacodynamic parameter, which is not surprising given the slow growth of the target pathogen, but the target PK/PD relationship is currently unknown in either elephant species and is very likely to be different from that reported for humans. This idea is further supported when fluoroquinolones are evaluated. While in human medicine, an AUC:MIC ratio of ≥125 for fluoroquinolones has been shown to eradicate a particular bacterial disease, this ratio cannot be directly extrapolated across species, indication, or pathogen, nor has it been determined for antituberculosis drugs. The effective AUC:MIC ratio has been reported to be different between species (Aliabadi et  al.,  2003). Opinions also differ within the human literature, where some report that a ratio >25 is best while others state that the ratio must be greater than 350 (Barger et al., 2003). This is complicated by the fact that for the fluoroquinolone ciprofloxacin, 100% of successfully treated patients had an AUC:MIC ratio >3.6 (Barger et al., 2003). It should be remembered that the in  vivo antimicrobial effect is the result of

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­A Practical Example of Allometry and Breakpoint  795

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dynamic exposure of the pathogen to the ­antimicrobial and the host immune system (Mueller et  al.,  2004). The comments and issues raised here also apply to rifampin (Peloquin et al., 2006) and ethambutol (Maslow et al., 2005). Unfortunately, numerous serious adverse drug effects have occurred in the majority of elephants undergoing treatment. In many cases, these were severe enough that treatment needed to be discontinued, at least temporarily. Reported adverse effects include anorexia, depression, diarrhea, nephrotoxicity and hepatotoxicity, blepharospasm, and death. The high incidence of severe adverse effects suggests that the doses of drugs required to achieve therapeutic human serum concentrations may be toxic to elephants (Wiedner and Schmitt, 2007).

­Administration Techniques Administration of antimicrobials to zoo and wildlife species can be made considerably ­easier by training the animals to accept them. Such training has increasingly become part of the general animal care routine at many zoological institutions, and is a responsibility assumed by keepers. A wide variety of species has been taught to tolerate injections, swallow tablets, and accept various other forms of drug administration. Such training requires a significant time commitment both for teaching the behaviors and for ongoing practice to maintain them by using placebos when the animal is healthy.

Oral Administration Oral medications for zoo species are typically hidden in food. Generally, this requires that the patient be physically separated from its social group for feeding. For some species, such as large carnivores, this is routine. For others, separation from conspecifics can cause stress. Typically, the medicine is hidden in something the animal particularly enjoys such

as a meatball for a carnivore or a piece of apple for hoofstock. Nonhuman primates are often more compliant if medication is mixed with sweet substances such as jams or juices or occasionally with savory flavors such as salsa. Compounding pharmacies that make flavored medications for children may be helpful in developing mixtures that are appealing to ­species which require oral antimicrobials. In one study, enrofloxacin’s bitterness was found to be best concealed using cherry-­flavored syrup (Petritz et al., 2013). Pachyderms can be trained to accept oral medications using a bite block. Even using this device, elephants often learn to hide medications within their massive mouths for hours, only to spit them out hours later when they are unobserved (Isaza and Hunter,  2004). Several elephants have been taught to swallow gelatin drug capsules without tasting them first, the way that humans do. When medications are hidden in feed, antimicrobials requiring an empty stomach for absorption must be avoided, such as ampicillin and tetracycline. A frequent complication is that zoo animals often become adept at identifying “doctored” food items and will eat around concealed ­medications, leaving them untouched. Some animals will become suspicious of nonmedicated feed as well and will stop eating altogether.

Injectable Administration Under anesthesia, even dangerous animals can be given medications via parenteral routes of administration. However, anesthetizing a sick animal repeatedly for the purpose of administering a course of antimicrobials is undesirable. Thus, most injectable antimicrobials are given intramuscularly in zoo and wildlife ­species. Intramuscular injections can be given by hand to animals trained to present body parts against the pen, chute, or cage bars (gluteal or thigh muscles for large carnivores and hoofstock, limbs, back, or flat palms of hands or feet for primates). A squeeze cage can be used for uncooperative animals, although

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i­ deally the animal should have received adequate training to enter the squeeze without becoming stressed. In some species, manual restraint by experienced personnel is possible. However, repeatedly restraining the animal can be dangerous and stressful to both handler and animal and is not recommended for long courses of antimicrobial treatment (Hunter and Isaza, 2002). Intramuscular injections can also be administered using remote delivery techniques (i.e., with darting equipment). While a discussion of darting techniques is beyond the scope of this chapter, any veterinarian planning to work with wildlife and/or zoological species should understand the relevant equipment and techniques, along with their risks, which include exertional myopathy, accidental bone fractures, accidental penetration of internal organs, and equipment failure (e.g., the dart fails to inject its contents either partially or entirely into the animal). The use of long-­ lasting antimicrobial depot formulations can decrease the frequency with which the animal needs to be darted. However, the duration of these formulations may vary from species to species. In specific circumstances and with certain species, intravenous or intraosseous catheters can be placed in captive animals requiring repeated antimicrobial therapy. Reptiles, birds, and severely debilitated animals housed in a hospital environment and maintained under close observation are best suited for this. In determining whether a particular patient is an appropriate candidate for an indwelling catheter, the clinician should assess the ease of maintaining the patency and cleanliness of the catheter and the likelihood of the animal’s removing it. If any of these are concerns then an indwelling catheter is not suitable. In several taxa, anatomical and/or living situations make indwelling catheters inappropriate or extremely difficult. These include very small animals, such as songbirds, animals with extremely thick skins such as hippos, aquatic animals that cannot be dry docked and species with significant ­dexterity,

such as primates and elephants. Intraosseous catheters are used in birds and neonates but are often painful and may impair movement.

Other Routes of Administration Great apes with air sacculitis have been ­successfully trained to tolerate routine nebulization with antibiotics (Gresswell and Goodman, 2011). Rectal administration of certain antimicrobials has been used successfully in elephants. Antibiotic-­impregnated beads, placed under anesthesia, have been used in the treatment of mandibular osteomyelitis in macropods (Hartley and Sanderson 2003). The use of an osmotic pump was tested for amikacin delivery in a corn snake (Elaphe guttata) (Sykes et  al.,  2006). Although complications, such as migration of the pump, were noted with this technique, it eliminated repeated handling of an animal needing medication and its use warrants further investigation. Fish and cetaceans housed in aquaria are administered antimicrobial therapy most often via immersion (i.e., the antimicrobial is placed directly in the tank with the animal). While effective, immersion can be hard on filtration systems, and can have unforeseen effects on co-­habitating species, such as corals and other aquatic invertebrates. Evidence demonstrating that hippos are most closely related to whales (Geisler and Theodor,  2009) has led to some veterinarians attempting to treat hippos using antimicrobial immersion therapy. Hippos are a difficult species to medicate due to their lack of easily accessible external blood vessels, very thick skin, and thick subcutaneous fat, so if immersion can be demonstrated to work in this species, it will provide a welcome improvement in hippo care.

­Treating Groups of Animals Medicating herds or flocks of animals presents other considerations. If the animals are to receive medication in feed or water, a total

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­Treating Groups of Animal  797

Antimicrobial Therapy in Zoo and Wildlife Species

dosage of medication for all animals needs to be calculated, but if the individual dose is based on the size of the smallest animal in the group for safety reasons, the largest animal will receive a subtherapeutic dose. Conversely, the smallest animals may receive an overdose if the dosage is calculated for the median-­sized animal. In hierarchical species such as wild equids and some ruminants, higher ranked animals will have increased access to any feed materials and will eat considerably more than those lower in rank. The risks of overdose or underdose must be considered. Other considerations are that sick or ill animals may not be eating or drinking their normal amounts, resulting in underdosing. If the animals to be treated are kept outdoors, ambient temperature must also be considered. Mass medication of zoo megavertebrates in feed and water is not done commonly, and there are scant reports of this technique.

­ pecific Examples of Antimicrobial S Use in Zoo and Wildlife Species Prophylactic antibiotic treatment against pneumonia in groups of free-­ranging bighorn sheep (Ovis canadensis) (Weiser et al., 2009) as well as reindeer (Rangifer tarandus) (Pietsch et  al.,  1999) continues to be used to manage these animals in North America. Following capture by various methods, the animals are physically examined, then hand-­injected with florfenicol or a long-­acting formulation of oxytetracycline prior to release or translocation. The goal of these one-­time antimicrobial ­injections has been to decrease the likelihood of stress-­induced respiratory disease; however, it has been difficult to confirm the efficacy of this approach. Tetracycline baits represent another use of antimicrobials in wildlife species. When ingested, tetracycline is incorporated into bones and teeth. Under ultraviolet light, the teeth fluoresce. The drug can also be detected in histological sections from tooth and bone in

necropsy specimens. Tetracycline baits have been used for mark-­capture population studies of American black bear (Ursus americanus) (Peacock et  al.,  2011), polar bears (Ursus arctos) (Taylor and Lee,  1994), and feral swine (Reidy et al., 2011), as well as for determining the use of supplemental feed in herds of white-­ tailed deer (Odocoileus virginianus) (Bastoskewitz et al., 2003). Tetracycline is also a component of oral rabies vaccines that are scattered as baits in areas inhabited by vector species such as ­raccoons (Procyon lotor) and skunks (Mephitis mephitis). To determine with what frequency the vaccines are being ingested by target species, the animals are captured a period of time after the vaccine baits are distributed. Under anesthesia, a tooth is removed and analyzed for evidence of tetracycline deposition to provide an estimate of the performance of the baits and thus vaccine ingestion (Fehlner-­ Gardiner et  al.,  2012). However, mounting concern about antimicrobial resistance and its reliability as a biomarker is stimulating efforts to replace tetracycline in these baits with a nonantibiotic biomarker (Slate et al., 2009). An interesting area of antimicrobial research involves wildlife species that actually produce their own antimicrobial substances. The Nile hippopotamus (Hippopotamus amphibius) releases a red sweat from its skin that has been found to have antimicrobial properties. At low concentrations (lower than those actually occurring on the skin), one of the pigments inhibits the growth of Pseudomonas aeruginosa and Klebsiella pneumoniae (Saikawa et al., 2004). Antimicrobial peptides have also been found in the platypus (Ornithorhyncus anatinus) and tammar wallaby. In the wallaby, some of these compounds are expressed in maternal milk and are hypothesized to protect the pouch-­dwelling, immunologically naïve, and underdeveloped young. In the common seal (Phoca vitulina), saccharide residues are produced by apocrine skin glands and inhibit the adherence of bacteria and fungi to the epidermis (Meyer et  al.,  2000,  2003). In both

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common seals and northern fur seals (Callorhinus ursinus), lysozyme and beta-­ defensins are produced by skin glands (Lynn and Bradley, 2007).

­ nvironmental Issues E and Nontarget Wildlife Species A number of studies have documented ­antimicrobial resistant bacteria in wild and stranded marine mammals, raccoons, wild and captive nonhuman primates, seabirds, fish, rodents, and wild hoofstock, and even in zoo animals. Many of these bacteria, E. coli in ­particular, demonstrate multidrug resistance. The importance of these organisms as ­reservoirs of multidrug resistance and their ability to spread into human and domestic animal populations are unknown but under study. From the perspective of the zoo and wildlife clinician, such findings emphasize

the importance of obtaining culture and s­ usceptibility results from wild patients. Evidence of harmful antimicrobial residues in wildlife has been found to cause increased pathology in affected species. Studies ­conducted in Spain identified residues of ­antimicrobials commonly used in livestock, specifically enrofloxacin, ciprofloxacin, amoxicillin, and oxytetracycline, in nestlings of three threatened species: Griffon vultures (Gyps fulvus), cinereous vultures (Aegypius monachus), and Egyptian vultures (Neophron percnopterus), avian scavengers that feed on carcasses. Affected nestlings showed liver and kidney damage as well as compromised immune systems that could be directly correlated with the antibiotic residues (Blanco et  al.,  2009). Another study demonstrated that fluoroquinolones that could be tracked back to livestock operations were causing embryonic death in the eggs of griffon vultures and red kites (Milvus milvus) (Lemus et al., 2009).

­References and Bibliography Aliabadi FS, et al. 2003. Pharmacokinetics and PK-­PD modelling of danofloxacin in camel serum and tissue cage fluids. Vet J 165:104. Barger A, et al. 2003 Pharmacological indices in antibiotic therapy. J Antimicrob Chemother 52:893. Bastoskewitz ML, et al. 2003. Supplemental feed use by free-­ranging white-­tailed deer in southern Texas. Wildlife Soc Bull 31:1218. Blanco G, et al. 2009. Ingestion of multiple veterinary drugs and associated impact on vulture health: implications of livestock carcass elimination practices. Animal Conserv 12:571. Clinical and Laboratory Standards Institute (CLSI). 2018. Susceptibility Testing of Mycobacteria, Nocardia spp., and other Aerobic Actinomycetes, 3rd edn. CLSI standard M24. Clinical and Laboratory Standards Institute, Wayne.

Clinical and Laboratory Standards Institute (CLSI). 2019a Understanding Susceptibility Test Data as a Component of Antimicrobial Stewardship in Veterinary Settings. CLSI report VET09. Clinical and Laboratory Standards Institute, Wayne. Clinical and Laboratory Standards Institute (CLSI). 2019b. Development of Quality Control Ranges, Breakpoints, and Interpretive Categories for Antimicrobial Agents Used in Veterinary Medicine, 4th edn. CLSI guideline VET02. Clinical and Laboratory Standards Institute, Wayne. Clinical and Laboratory Standards Institute (CLSI). 2021. Performance Standards for Antimicrobial Disk and Dilution Susceptibility Tests for Bacteria Isolated From Animals, 5th edn. CLSI supplement VET01S. Clinical and Laboratory Standards Institute, Wayne. de Steenwinkle JEM, et al. 2010. Time kill kinetics of anti-­tuberculous drugs, and

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  ­References and Bibliograph

Antimicrobial Therapy in Zoo and Wildlife Species

emergence of resistance, in relation to metabolic activity of Mycobacterium tuberculosis. J Antimicrob Chemother 65:2582. Fehlner-­Gardiner C, et al. 2012. Comparing Onrab (R) and Raboral V-­RG (R) oral rabies vaccine field performance in raccoons and striped skunks, New Brunswick, Canada, and Maine, USA. J Wildlife Dis 48(1):157. Funk RS. 2000. A formulary for lizards, snakes, and crocodilians. Vet Clin North Am Exot Anim Pract 3:333. Geisler JH, Theodor JM. 2009. Hippopotamus and whale phylogeny. Nature 458(7236):E1. Gresswell C, Goodman G. 2011. Case study: Training a chimpanzee (Pan troglodytes) to use a nebulizer to aid the treatment of airsacculitis. Zoo Biol 30:570. Hartley MP, Sanderson S. 2003. Use of antibiotic impregnated polymethylmethacrylate beads for the treatment of chronic mandibular osteomyelitis in a Bennett’s wallaby (Macropus rufo-­griseus rufogriseus). Australian Vet J 81:742. Hunter RP. 2017. Zoological pharmacology. In: Riviere E, Papich MG (eds) Veterinary Pharmacology and Therapeutics, 10th edn. Iowa State University Press, Ames, pp. 1395–­1404. Hunter RP, Isaza R. 2002. Zoological pharmacology –­current status, issues, and potential. Adv Drug Deliv Rev 54:787. Hunter RP, Isaza R. 2008. Concepts and issues with interspecies scaling in zoological medicine. J Zoo Wildlife Med 39:517. Hunter RP, et al. 2003. Oral bioavailability and pharmacokinetic characteristics of ketoprofen enantiomers after oral and intravenous administration in Asian elephants (Elephas maximus). Am J Vet Res 64:109. Hunter RP, et al. 2008. Prediction of xenobiotic clearance in avian species using mammalian or avian data: how accurate is the prediction? J Vet Pharmacol Therapeut 31:281. Isaza R, Hunter RP. 2004. Drug delivery to captive Asian elephants –­treating goliath. Current Drug Deliv 1:291.

Lemus JA, et al. 2009. Fatal embryo chondral damage associated with fluoroquinolones in eggs of threatened avian scavengers. Environment Pollut 157:2421. Lynn DJ, Bradley DG. 2007. Discovery of alpha-­defensins in basal mammals. Dev Compar Immunol 31(10):963. Mahmood I, et al. 2006. Interspecies allometric scaling. Part I: prediction of clearance in large animals. J Vet Pharmacol Therapeut 29:415. Martinez M, et al. 2006 Interspecies allometric scaling: prediction of clearance in large animal species: Part II: mathematical considerations. J Vet Pharmacol Therapeut 29:425. Maslow JN, et al. 2005. Population pharmacokinetics of isoniazid in the treatment of Mycobacterium tuberculosis among Asian and African elephants (Elephas maximus and Loxodonta africana). J Vet Pharmacol Therapeut 28:21. Meyer W, et al. 2000. Aspects of general antimicrobial properties of skin secretions in the common seal (Phoca vitulina). Dis Aquat Organ 41:77. Meyer W, et al. 2003. Further aspects of the general antimicrobial properties of pinniped skin secretions. Dis Aquat Organ 53:177. Mueller M, et al. 2004. Issues in pharmacokinetics and pharmacodynamics of anti-­infective agents: kill curves versus MIC. Antimicrob Agents Chemother 48:369. Peacock E, et al. 2011. Mark-­recapture using tetracycline and genetics reveal record-­high bear density. J Wildlife Manage 75(6):1513. Peloquin CA, et al. 2006. Dose selection and pharmacokinetics of rifampin in elephants for the treatment of tuberculosis. J Vet Pharmacol Therapeut 29:581. Petritz OA, et al. 2013. Stability of three commonly compounded extemporaneous enrofloxacin suspensions for oral administration to exotic animals. J Am Vet Med Assoc 43(1):85. Pietsch GS, et al. 1999. Antibiotic treatment and post-­handling survival of reindeer calves in Alaska. J Wildlife Dis 35(4):735.

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Reidy MM, et al. 2011. A mark-­recapture technique for monitoring feral swine populations. Rangeland Ecol Manage 64(3):316. Saikawa Y, et al. 2004. Pigment chemistry: the red sweat of the hippopotamus. Nature 429(6990):363. Slate D, et al. 2009. Oral rabies vaccination in North America: opportunities, complexities, and challenges. PLoS Neglect Trop Dis 3(12):e549. Sykes JM, et al. 2006. Evaluation of an implanted osmotic pump for delivery of amikacin to corn snakes (Elaphe guttata guttata). J Zoo Wildlife Med 37(3):373. Taylor M, Lee J. 1994. Tetracycline as a biomarker for polar bears. Wildlife Soc Bull 22:83. United States Department of Agriculture (USDA). 2008. Guidelines for the control

of tuberculosis in elephants. http:// nasphv.org/Documents/ElephantTB_ NASPHV.pdf Wang J, et al. 2011. Ancient antimicrobial peptides kill antibiotic-­resistant pathogens: Australian mammals provide new options. PLoS One 6(8):1. Weiser GC, et al. 2009. Variation in Pasturella (Bibersteinia) and Mannheimia spp. following transport and antibiotic treatment in free-­ ranging and captive Rocky Mountain bighorn sheep (Ovis canadensis canadensis). J Zoo Wildlife Med 40(1):117. Wiedner E, Schmitt DL. 2007. Preliminary Report of Side Effects Associated with Drugs Used in the Treatment of Tuberculosis in Elephants. International Elephant Foundation, Orlando.

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  ­References and Bibliograph

39 Antimicrobial Therapy in Aquaculture Patrick Whittaker, Timothy S. Kniffen, and Simon Otto

I­ ntroduction As the human population and income ­continue to grow, so too does the demand for high-­ quality protein sources, including seafood. The ability to meet the increasing global seafood demand from wild-­caught resources is unsustainable and untenable. Aquaculture has rapidly grown globally to meet this additional demand for high-­quality protein to the extent that in 2018, almost 50% of all seafood consumed was provided by aquaculture (Food and Agricultural Organization, 2020). As with any other farmed food animal species, it is the moral and ethical obligation of the farmer and veterinarian to ensure the health and welfare of the animals in their care. One of the key tenets of this obligation is the prudent treatment of illness with antimicrobials when necessary and appropriate. Prudent is the key word in this statement, considering that products from aquaculture are destined for human consumption and present a potential transmission pathway for antimicrobial resistance (AMR) from the aquatic environment and products to humans. All antimicrobial use (AMU) must be evaluated via a One Health lens wherein human medicine, terrestrial animal agriculture, and aquaculture  all  contribute antimicrobials to the

environment, resulting in potential effects on nontarget organisms as well as the risk of bacterial AMR development. Salmonid aquaculture is the largest contributor as far as tonnage produced and economic relevance in North America, South America, and western Europe are concerned. Due to efforts by governments, third-­party certification groups and data reporting by publicly traded companies, salmonid aquaculture has the most robust data collected and reported on annual antimicrobial usage. Salmonid aquaculture is also an example of an industry that has shown an impressive ability to reduce AMU. The largest global salmon producer, Norway, has reduced AMU by 99% over the last  25 years while simultaneously increasing total salmon production (Norwegian Veterinary Institute (Veterinaerinstituttet), 2021). Effective vaccines, improved biosecurity, and management changes and improvements have nearly eliminated the need for antimicrobials in Norwegian salmon production.  However, viral diseases of Atlantic salmon remain a formidable challenge.  In other Atlantic salmon-­farming regions around the globe, there are gaps in successful vaccine development to address local bacterial challenges.  Ongoing research will hopefully find and develop solutions to guide the use of

Antimicrobial Therapy in Veterinary Medicine, Sixth Edition. Edited by Patricia M. Dowling, John F. Prescott, and Keith E. Baptiste. © 2025 John Wiley & Sons, Inc. Published 2025 by John Wiley & Sons, Inc.

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Antimicrobial Therapy in Aquaculture

biologics and the best management practices to mitigate these challenging diseases, as the industry recognizes the need to reduce AMU for sustainability. This chapter focuses on the prudent use of antimicrobials guided by the ethical treatment of disease while avoiding residues, as well as minimizing environmental impacts and the risk of the development of AMR. Due to a lack of research and robust data regarding AMU and AMR in many aquatic species and geographic regions, the chapter will primarily focus on salmonid production. There is a common but erroneous belief that antimicrobials are used in aquatic species for growth promotion. The cost of antimicrobials is high relative to the cost of production and market value of most aquatic species. There are no approved indications for any antimicrobial to improve production parameters in any aquatic species. For example, one study demonstrated that oral administration of oxytetracycline to channel catfish, hybrid striped bass, Nile tilapia, and rainbow trout for eight weeks did not improve survival, average daily gain, or feed conversion ratio (Trushenski et al., 2018). With no economic value to feeding antimicrobials to healthy finfish, there is no reason or incentive to utilize antimicrobials in a nontherapeutic fashion. The use of antimicrobials in all types and forms of aquaculture is and must remain limited to the treatment of disease due to susceptible pathogenic bacteria.

­Aquaculture Definition Globally, over six hundred different aquatic species are under cultivation in aquaculture compared to terrestrial agriculture, which produces approximately one dozen species, albeit with multiple breeds (Henriksson et al., 2018). Forty-­four ectothermic species make up 90% of total global aquaculture production, in comparison to five predominant terrestrial livestock species. Modern aquaculture is conducted in both fresh-­water and salt-­water (marine)

environments. Animals raised in aquaculture systems are classified as cold-­water, warm-­ water, or tropical species. Therefore, an entire book, not just one chapter, could be dedicated to pharmacology in all the aquaculture applications, not to mention aquarium species and marine mammals. However, the focus of this chapter is finfish aquaculture, with emphasis on the most economically relevant species. The authors recognize there is concern regarding both the type and quantity of antimicrobials used with crustaceans; however, this will not be addressed in the chapter as robust data for this use are sparse. The chapter focuses primarily on data from Atlantic salmon as they are generally available and robust in quality. Atlantic salmon aquaculture could be a model for many forms of aquaculture and terrestrial agriculture where biosecurity, technology (e.g, vaccines), management practices, regulation, and third-­ party certification have reduced or eliminated the need for antimicrobials in production systems. In 2017, over 80% of global aquaculture biomass produced was consumed in China (57.9%), India (11.3%), Indonesia (8.6%), and Vietnam (5%) (Schar et  al.,  2020). In these countries, accurate data regarding the types and quantities of AMU are difficult to access. For example, carp represent more than one-­ third of global aquaculture production tonnage by species, but only one survey of farms in Vietnam explicitly identified carp among the species for which AMU data were collected (Schar et  al.,  2020). Antimicrobial use also crosses socioeconomic boundaries, emphasizing the need to teach and use a One Health policy to evaluate AMU in humans, terrestrial agriculture, and aquaculture (Ayukekbong et al., 2017). Medicated feed with oral delivery is the route of administration used to treat the major aquaculture species discussed here. This mode of antimicrobial administration presents its own challenges, similar to those faced in all other species, in that the affected animal must be consuming feed in order to obtain the

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medication, and the need to treat groups of animals rather than the individual. The challenge to the aquaculture veterinarian responsible for treating aquatic species is to evaluate the population, prescribe an efficacious dose of an approved product, and ensure that a very high proportion of the population consumes the prescribed dose.

Poikilotherms versus Homeotherms Homeotherms are organisms that have the ability to maintain a relatively constant body temperature independent of their environmental temperature. Some examples of homeotherms are humans, terrestrial and marine mammals, and birds. Poikilotherms are organisms that do not have the ability to regulate their body temperature because they lack the physiological ability to generate heat. Examples of poikilotherms include amphibians, reptiles, insects, and aquatic animals such as fish and shrimp. The body temperature of aquatic species equilibrates with the temperature of the water in which they are located. Water temperature must be considered in the production and management of any aquatic species. Each species, whether cold-­water, warm-­water, or tropical, will have a preferred optimal water temperature range. Exposure to water temperatures outside the optimal range results in a decreased rate of biological functions, physiological stress, or death. The impact of water temperature on drug metabolism and elimination is crucial in ­establishing drug withdrawal recommendations. An Atlantic salmon can survive in an environment from approximately 0 °C to temperatures in the low 20 °C range. This wide range of temperatures is a predicament for the prescribing veterinarian as an equivalent amount of antimicrobial will be metabolized more slowly with low water and body temperatures. Some approved product labels include instructions that the drug is not to be used if water ­temperature is