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Copyright © 2011. Nova Science Publishers, Incorporated. All rights reserved. Social Insects: Structure, Function, and Behavior : Structure, Function, and Behavior, edited by Emily M. Stewart, Nova Science Publishers,

Copyright © 2011. Nova Science Publishers, Incorporated. All rights reserved. Social Insects: Structure, Function, and Behavior : Structure, Function, and Behavior, edited by Emily M. Stewart, Nova Science Publishers,

INSECTS AND OTHER TERRESTRIAL ARTHROPODS: BIOLOGY, CHEMISTRY AND BEHAVIOR

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SOCIAL INSECTS: STRUCTURE, FUNCTION AND BEHAVIOR

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Social Insects: Structure, Function, and Behavior : Structure, Function, and Behavior, edited by Emily M. Stewart, Nova Science Publishers,

INSECTS AND OTHER TERRESTRAIL ARTHROPODS: BIOLOGY, CHEMISTRY AND BEHAVIOR

SOCIAL INSECTS: STRUCTURE, FUNCTION AND BEHAVIOR

EMILY M. STEWART

Copyright © 2011. Nova Science Publishers, Incorporated. All rights reserved.

EDITOR

Nova Science Publishers, Inc. New York Social Insects: Structure, Function, and Behavior : Structure, Function, and Behavior, edited by Emily M. Stewart, Nova Science Publishers,

Copyright ©2011 by Nova Science Publishers, Inc. All rights reserved. No part of this book may be reproduced, stored in a retrieval system or transmitted in any form or by any means: electronic, electrostatic, magnetic, tape, mechanical photocopying, recording or otherwise without the written permission of the Publisher. For permission to use material from this book please contact us: Telephone 631-231-7269; Fax 631-231-8175 Web Site: http://www.novapublishers.com

NOTICE TO THE READER The Publisher has taken reasonable care in the preparation of this book, but makes no expressed or implied warranty of any kind and assumes no responsibility for any errors or omissions. No liability is assumed for incidental or consequential damages in connection with or arising out of information contained in this book. The Publisher shall not be liable for any special, consequential, or exemplary damages resulting, in whole or in part, from the readers‘ use of, or reliance upon, this material. Any parts of this book based on government reports are so indicated and copyright is claimed for those parts to the extent applicable to compilations of such works.

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Independent verification should be sought for any data, advice or recommendations contained in this book. In addition, no responsibility is assumed by the publisher for any injury and/or damage to persons or property arising from any methods, products, instructions, ideas or otherwise contained in this publication. This publication is designed to provide accurate and authoritative information with regard to the subject matter covered herein. It is sold with the clear understanding that the Publisher is not engaged in rendering legal or any other professional services. If legal or any other expert assistance is required, the services of a competent person should be sought. FROM A DECLARATION OF PARTICIPANTS JOINTLY ADOPTED BY A COMMITTEE OF THE AMERICAN BAR ASSOCIATION AND A COMMITTEE OF PUBLISHERS. Additional color graphics may be available in the e-book version of this book. LIBRARY OF CONGRESS CATALOGING-IN-PUBLICATION DATA Social insects : structure, function, and behavior / editor, Emily M. Stewart. p. cm. Includes index. ISBN 978-1-61761-620-4 (E-Book) 1. Insect societies. I. Stewart, Emily M. QL496.S625 2010 595.7'1782--dc22 2010031752

 New York Social Insects: Structure, Function, and Behavior : Structure, Function, and Behavior, edited by Emily M. Stewart, Nova Science Publishers,

CONTENTS Preface

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Chapter 1

vii The Role of Food Storage and Communication in the Evolution of Perennial Social Hymenopteran Colonies Timothy M. Judd

Chapter 2

Issues in the Study of Proboscis Conditioning Charles I. Abramson, Michel B. C. Sokolowski and Harrington Wells

Chapter 3

Regulation of Reproductive States and Control of Sex of Eggs by Reproductive Females in Eusocial Hymenoptera Ken Sasaki

Chapter 4

The Global Empire of an Invasive Ant Eiriki Sunamura, Hironori Sakamoto, Shun Suzuki, Koji Nishisue, Mamoru Terayama and Sadahiro Tatsuki

Chapter 5

Modification of Morphological Characteristics by Endoparasites in Workers of the Swarm-Founding Wasp Polybia Occidentalis Kazuyuki Kudô, Hitomi Ohka and Ronaldo Zucchi

Chapter 6

Chapter 7

Asymmetric Trophallaxis between Workers of the Stingless Bee Melipona Quadrifasciata (Apidae, Meliponini) Felipe Andrés León Contrera, Vera Lúcia Imperatriz-Fonseca and Dirk Koedam Social Insects: Morphophysiology of the Nervous System Roberta C. F. Nocelli, Thaisa C. Roat, Elaine C. M. Silva Zacarin, Mario Sérgio Palma and Osmar Malaspina

Social Insects: Structure, Function, and Behavior : Structure, Function, and Behavior, edited by Emily M. Stewart, Nova Science Publishers,

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51 73

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vi Chapter 8

Contents Structure and Function of the Intestine and Malpighian Tubules: From Bee Biology to Cell Marker Development for Toxicological Analysis Elaine C. M. Silva-Zacarin, Rafael A. Costa Ferreira, Roberta C. F. Nocelli, Thaisa C. Roat, Mário Sérgio Palma and Osmar Malaspina

Chapter 9

Hydrocarbons and Insects‘ Social Physiology E. Provost, O. Blight, A. Tirard and M. Renucci

Chapter 10

Biogenic Amines and Division of Reproduction in Social Insects Ken Sasaki

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Index

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143

199 225

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PREFACE Social insects are among the most diverse and ecologically important organisms on Earth. This book presents current research in the study of social insects, including food storage behavior in social Hymenoptera; the global empire of an invasive ant; asymmetric trophallaxis between workers of the stingless bee; termite breeding strategies; as well asbiogenic amines and division of reproduction in social insects. Chapter 1 - Social insect colonies have been traditionally divided in to primitive (less social) and highly eusocial colonies. A trait used to distinguish these two categories in Hymenoptera is the perennial or annual existence of the colony. One aspect that differs between an annual and perennial colony is the flow of nutrients into a colony and the level of food storage. This chapter reviews food storage behavior in social Hymenoptera. It then discusses how this behavior ties in with nutrient flow, recruitment behavior and the evolution of perennial life histories. Four traits energy storage, nitrogen (protein) storage, the presence of a food-receiving caste, and directed recruitment (the ability to relay location information to recruits) are examined in their relative importance to the transition from an annual to a perennial life history in both fluctuating climates (climates that alternate between favorable and harsh conditions) and non-fluctuating environments. Of the four traits, only energy storage and the food-receiving caste are universal among all perennial colonies thus are probably essential for the maintenance of a perennial life history. Nitrogen storage and directed recruitment appear to only be essential for perennial colonies in fluctuating environments. Chapter 2 - This paper discusses methodological and theoretical issues associated with proboscis conditioning in neglected pollinators. The case is made that the search for learning curves similar to those found in Apis is hindered by several factors. Among the most important are the lack of consensus on what is classical conditioning and the allure of cognitive explanations of insect learning. However, the problem is fueled by few examples of individual data and no generally accepted learning taxonomies. On the other hand, the possibility exists that the proboscis learning found in Apis is overestimated as a research methodology. The few cases of proboscis conditioning in pollinators not generally used as model systems (not Apis or Bombus) are compared to results found with honey bees. Although generally conditioning is less complete, what is also noteworthy is the spotty occurrence of cases across the pollinator taxa. Finally, suggestions are provided on the type of

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Emily M. Stewart

training variables that should be manipulated in an effort to find learning curves similar to those found in Apis. Chapter 3 - In Hymenoptera, sex of an egg is determined by egg fertilizations on a haplodiploidy sex determination system. Female reproductive organs in eusocial species have unique functions to regulate ovarian development and control the sex of an egg. Egg fertilizations could be controlled by muscle activities of the spermathecal pump and/or secretion of the spermathecal glands. Activities of the spermathecal pump are regulated by neuronal systems and may inhibit release of sperm for unfertilized male eggs. Secretions of the spermathecal glands could activate spermatozoa from the spermathecal receptacle and therefore, lack of spermathecal gland secretions may cause unfertilization of eggs. In highly eusocial species, especially honeybees, queens can control the egg fertilization in response to comb cell types for oviposition. They can make a choice of the cell types and behaviorally control the egg sex ratios in the colonies. Workers in honeybees can develop reproductive organs in the absence of a queen and lay only unfertilized eggs. They select preferentially male comb cells for the oviposition to produce larger males for mating competition between males. I introduce general structure and function of female reproductive organs for controlling egg fertilizations, physiological processes for regulating reproductive states and behavioral controls of the egg sex ratio in colonies in Hynemoptera. Chapter 4 - Social insects have obtained their prosperity by cooperation among individuals. This can be applied particularly to the success of invasive ants, which form unusual social structure called supercolonies, within which individuals can move freely among physically separated nests, and thereby gain high population densities to dominate indigenous ants. Native to South America, the Argentine ant Linepithema humile has been unintentionally introduced into many parts of the world during the last 150 years. Although it is well known that the introduced Argentine ant populations form much larger and fewer supercolonies than the native populations, the relationship among beyond-ocean populations has been poorly understood. Recent studies, however, are uncovering the behavioral, chemical and genetic relationships among introduced Argentine ant populations worldwide. Individuals from the dominant supercolonies around the world have very similar cuticular hydrocarbon profiles (nestmate recognition cue), and do not show aggressive behavior toward each other, when artificially put into contact. The supercolonies constitute the largest cooperative unit ever known. Their genetic closeness suggests a common introduction pathway. Considering historical records, descendants of the most ancient introduced population have spread to many parts of the world, without losing memory of their roots. In this chapter, we introduce the nestmate recognition system and mechanism of supercolony expansion in invasive ants, with the global empire of Argentine ants as an example. Chapter 5 - Two kinds of parasites, i.e., a strepsipteron, possibly Xenos myrapetrus (Trois) and an undescribed gregarine were recognized in the neotropical swarm-founding wasp, Polybia occidentalis (Olivier). Although the proportions of workers that were infected by these parasites varied greatly among colonies, a prevalence of infected workers was recognized. Five external characteristics and the number of hamuli were measured and compared among uninfected workers, stylopized (i.e., infected by Strepsiptera) workers and workers infected by gregarines. Stylopized workers were significantly smaller than uninfected workers, while workers infected by gregarines were significantly larger than uninfected workers. Moreover, number of hamuli in workers infected by gregarines tended to be greater than that in uninfected workers. In one of our colonies, in which a large number of workers

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Preface

ix

had been stylopized, the number of strepsipteran larvae per host tended to be negatively correlated with mean body length of strepsipteran larvae, indicating that the greater the number of strepsipteran larvae, the smaller the mean body length was. These findings were consistent with those of our previous study conducted in a consubgeneric wasp, P. paulista. Since the nutrients of stylopized workers may be plundered during their growth period, their body size may be consequently reduced. On the contrary, gregarines may manipulate host larvae to solicit more food from adults or increase the development time of larvae longer, and therefore produce more parasites from a larger host. Chapter 6 - In several species of social insects, there is a positive relationship between the food exchange that occurs during trophallaxis and the ovarian development of workers, indicating that food transfers are important for the establishment of reproductive hierarchies between the colony members. In this chapter, we tested if such correlations also occur between workers of the stingless bee Melipona quadrifasciata anthidioides. Due to the difficulty to follow the totality of worker behavior inside the nest, we housed six newlyemerged workers in confined conditions, away from the mother colony and older workers, with pollen and honey ad libitum and under stable conditions of temperature and illumination. We measured their trophallactic behavior until the moment the first worker died; after that all workers from the group were dissected for the measure of their ovarian development. We found significant differences in the individual trophallactic behavior exhibited by workers, since some workers were mainly food donors whereas others were mainly receivers, and all contacts were initiated by the soliciting bee. Moreover, only 15% of the trophallactic contacts were longer than one second, and thus, possibly resulted in an effective food exchange. However, there was no correlation between the ovarian development and the trophallactic behavior of workers, which might be caused by the experimental conditions the workers were kept, and the small age they died, which was five days after the start of the experiment. However, considering that Melipona workers from different species highly vary in their ovarian development and egg-laying capacities, and that individual workers exhibit significant differences in their trophallactic behavior, it is reasonable to presume that under natural conditions trophallaxis might have a relevant role in their ovarian development, but this possibility yet remains to be tested in regular colonies. Chapter 7 - Social insects are an interesting model to neurobiological studies due to the simplicity of its brain commanding the complex behaviors demanded by eusocial relationships and its capacity of learning and memorizing. In their social structure, usually have differentiated sex and caste, where individuals have different morphology, physiology and behavioral patterns correlated with their functions in the society and represented by some brain polymorphism. They have a rich behavioral repertoire and perform various tasks throughout their lives that involve a complex and diverse system of learning. The brain is the main center of association of the insect. Receives sensory impulses coming from the sense organs of the head and from the ventral ganglia nerve chain, through ascending interneurons. Orders emanating from the brain to the antennae muscles and the posterior part of the body, passing through the descending pathways, pre-motor, going to the ganglia of the ventral nerve chain. In addition, the brain is the center of the integration of activities, which produce organized patterns of long-term behavior driving their changes by learning. In general, the brain of insects is divided into three regions: the protocerebrum, deuterocerebrum and tritocerebrum. The protocerebrum is the largest fraction of the brain, including the optical lobes, a pair of lateral-dorsal mushroom bodies, the cerebral bridge and the central body; the

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deuterocerebrum is part of the brain containing the olfactory antennal centers, and the cell bodies of motor nerves of the antennae muscles and usually form a pair of lateral lobes distinct. The tritocerebrum is very small in insects, consisting of two small lobes after the deuterocerebrum connected by circum-esophageal commissures to the subesofagean ganglia. Forward, this region connects through nerves, with the oral region and the ganglia of estomogastric nervous system. Despite this apparent simplicity, have systems of spatial orientation and communication very complex and studies have shown that neurotransmitters and receptors are common to those found in vertebrates. According to the developmental stage and the activity performed, the pattern of protein expression is different, reflecting differential gene expression in neurons according to environmental and / or physiological stimuli received, many of these genes similar to those expressed in neurons of vertebrates. This ability to respond physiologically to the varied stimulus, turn on or turn off some genes, stimulating or blocking specific neuronal receptors, makes these insects can be used as models in studies of new drugs possibly neuroactive and also to analyze the effects of neurotoxic substances, as some studies have shown. Chapter 8 - The purpose of this chapter is to exhibit research developed about morphological studies in honeybee organs and others social insects, specifically in midgut and Malpighian tubules. The chapter will include a review of relevant articles about structure and function of these organs to provide knowledge of social insect biology with emphasis in larvae and adults of bees, as in Apis mellifera as in Meliponini. This section of the chapter will discuss the basic tissue architecture of midgut and Malpighian tubules and some differences according species and groups. Since the histology and ultra-morphology of midgut and Malpighian tubules are well characterized, any modification in their structure and function or in the physiological process of programmed cell death, induced by xenobiotics, can be diagnosed. The midgut and Malpighin tubules of larvae and adults of bees are involved in the absorption an excretion of chemical compounds, respectively, an assessment of their morphology can reveal ultrastructural alterations induced by environmental stressors such as pesticides. Then, in a second section of this article, the subject is to report on the advances of the morphological studies of honeybee organs in response to pesticide exposure, which will contribute to data acquisition for building shared knowledge on bee biology and toxicology, providing arguments for effective policy marking in this field of research. Chapter 9 - Since the middle of the 20th century, improvements in analytical technologies have permitted the identification of cuticular hydrocarbons present on the cuticle of almost all insects. Concomitantly, a great deal of attention has been paid to the determination of their role at the individual, colonial, and populational level. Many studies pointed to the importance of the surface lipids which cover the insects' cuticle. They are important for the survival of individuals and at a higher level, to maintain colony integrity and species survival. The lipid layer is responsible for the water-repellent character of the cuticle which is its primary function. It reduces surface transpiration and protects terrestrial insects from desiccation. Furthermore, it creates a barrier against micro-organisms and inorganic chemical components. More recently, bioassays have shown that surface hydrocarbons are important tools for recognition systems for solitary insects but above all, for social insects where the chemical communications between individuals are complex. Cuticular hydrocarbons serve as contact pheromones when insects encounter each other. They permit insects to identify « friends » from « foes », nestmates from non-nestmates. They constitute a true chemical signature. They

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Preface

xi

also maintain the social structure of insect colonies by separating individuals according to their functions within the colony, according to their physiological states and their hierarchical rank. They constitute a fertility signal for the queen which indicates its presence for the workers which retain their reproduction. Furthermore, cuticular hydrocarbons can play a major role in sexual behavior as an attractant or repellent. They can also be involved in courtship behavior. These are kairomones or allomones in host-parasite and prey-predator relationships, and contribute to mutualistic relations between insects. Nowadays, the identification of all these functions allows biologists to use cuticular hydrocarbons as tools to conduct eco-ethological studies. For example, it is possible to use hydrocarbon composition as taxonomic character to identify cryptic species which are morphologically close. This could be useful in conservation planning. These components should equally allow a better understanding of ant colonies structure, for instance, the boundaries of polydomous colonies or especially in the context of an invasion when ants form super-colonies. Chapter 10 - Division of reproduction in social insects is a behavioral specialization to enhance the efficiency of individual behaviors, and the growth and reproductive success of the colony. Queens engage directly in reproduction, whereas workers perform other tasks including taking care of the queen and broods, guarding the nest and foraging. Workers can change their reproductive states and lay eggs under certain conditions such as in the absence of a queen. During the transition of reproductive states, a brain specialized to worker‘s behaviors can be reconstructed to a queen‘s type morphologically and physiologically. Physiological processes of the transition for reproduction in social insects are recently elucidated. Biogenic amines are key substances for behavioral and physiological changes during the transition. They have roles as neurotransmitters, neuromodulators and neurohormones in the peripheral and central nervous systems by acting on target cells through binding to receptors in insects. Dopamine and tyramine may play regulatory roles in the transition of the reproductive individuals from normal workers in honeybees. The differences of brain dopamine levels between reproductive and non-reproductive individuals in several species of social hymenopteran may contribute to caste–specific behavioral expression. These biogenic amines may interact with other hormones regulating the physiological states for reproduction. A part of the physiological processes is confirmed by molecular biological experiments detecting the gene.

Social Insects: Structure, Function, and Behavior : Structure, Function, and Behavior, edited by Emily M. Stewart, Nova Science Publishers,

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In: Social Insects: Structure, Function, and Behavior Editor: Emily M. Stewart

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Chapter 1

THE ROLE OF FOOD STORAGE AND COMMUNICATION IN THE EVOLUTION OF PERENNIAL SOCIAL HYMENOPTERAN COLONIES Timothy M. Judd Department of Biology, Southeast Missouri State University, Cape Girardeau, Missouri

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ABSTRACT Social insect colonies have been traditionally divided in to primitive (less social) and highly eusocial colonies. A trait used to distinguish these two categories in Hymenoptera is the perennial or annual existence of the colony. One aspect that differs between an annual and perennial colony is the flow of nutrients into a colony and the level of food storage. This chapter reviews food storage behavior in social Hymenoptera. It then discusses how this behavior ties in with nutrient flow, recruitment behavior and the evolution of perennial life histories. Four traits energy storage, nitrogen (protein) storage, the presence of a food-receiving caste, and directed recruitment (the ability to relay location information to recruits) are examined in their relative importance to the transition from an annual to a perennial life history in both fluctuating climates (climates that alternate between favorable and harsh conditions) and non-fluctuating environments. Of the four traits, only energy storage and the food-receiving caste are universal among all perennial colonies thus are probably essential for the maintenance of a perennial life history. Nitrogen storage and directed recruitment appear to only be essential for perennial colonies in fluctuating environments.

INTRODUCTION It has long been recognized that social Hymenoptera colonies have two basic organizational forms. Traditionally, these forms have been classified as primitively eusocial or highly (advanced) eusocial. These classifications were originally conceived to describe the

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level of social behavior in bees by Michener (1969). Primitively eusocial colonies are those that have an annual lifecycle and minor differences between castes. Highly eusocial colonies generally have perennial lifecycles and distinct morphological differences between castes such as queens and workers. Winston and Michener (1977) added the criterion that highly eusocial bees are swarm founding. These definitions for bees have been co-opted for all eusocial Hymenoptera (Wilson 1971). However, there are some issues with these definitions when expanded to all social hymenopterans. Some colonies with annual lifecycles have distinct size differences between reproductive and worker castes such as Bombus (this was even pointed out by Michener (1969)), and Vespula (Jeanne 2003). In addition, there are differences between ovarian development. Ant colonies are initiated by single or multiple foundresses (Hölldobler and Wilson 1990). Of the three characteristics (lifecycle, caste morphology, and founding) the one key evolutionary innovation that truly divides the social Hymenoptera in terms of colony structure is the presence of an annual or a perennial lifecycle. The transition from an annual to a perennial lifecycle has generally required a change in food storing behavior which in turn changed the flow of nutrients in a colony. In some cases the level of communication had to change as well. I will define communication as a process that involves the flow of information from a sender to a receiver through signals, information that has been acted upon by natural selection for the purpose of conveying information. In contrast, information can also flow through cues which have not evolved specifically to convey information. Animals can use both signals and cues which I will group together under the general term ―information flow.‖ However, the level in which these two factors change will depend on the climate the colony exists in. Annual colonies persist for one active season. A single foundress initiates the colony, cares for the initial work force, and then remains on the nest once the first cohort of workers emerges. The colony ends with the production of reproductives that eventually leave the colony to mate. The gynes produced by the colony will eventually initiate a new colony (Wilson 1971). In many cases, the gynes go through diapause before doing so. Thus, the colony goes through a distinct growth phase, followed by a final reproductive phase. All of the nutrients entering the colony go towards producing individuals (Hunt and Nalepa 1994). Perennial species will be defined as a species whose workers and reproductives persist in a nest from year to year. Perennial colonies can be initiated by swarms (West-Eberhard 1982, Jeanne 1991b) or individuals (Hölldobler and Wilson 1990). These colonies will also begin with a growth phase followed by a reproductive phase. However, the colony will persist post reproduction and continue to grow and reproduce during the following active season (Oster and Wilson 1978). The incoming resources of these colonies are divided between the production of individuals and colony survival. Thus, the ultimate flow of nutrients through a perennial colony is different than an annual colony. Three factors have likely contributed to the evolution of perennial life histories: the ability to produce long-term food stores, a reduction of direct communication between the foraging force and the brood and an increase in the efficiency of the foraging force. In this chapter, I will review both the food storage and how it ties to nutrient and information flow between workers and demonstrate how changes in both of these traits facilitated the evolution of perennial life histories in social Hymenoptera. I will then examine the evolution of the perennial life history in bees, wasps, and ants.

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The Role of Food Storage and Communication in the Evolution…

3

FORMS OF FOOD STORAGE

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There are several mechanisms for long-term food storage in social Hymenoptera. Some are internal and others are external in respect to the body of the workers. It is also important to recognize that stores come in two forms: energy foods such as carbohydrates and lipids, and nitrogenous sources such proteins. The former is used to maintain the work force and the latter is primarily used for colony growth. The major groups of perennial Hymenoptera have evolved three distinct food-preserving strategies. There are two forms of external food storage, termed here as canning and caching, in addition to internal food storage. Canning: This method of food preservation, employed by social bees and some social wasps, is akin to pickling and curing techniques used by humans. The food is made inedible to microorganisms. In bees, the food is also sealed in cells or pots made from wax, adding another layer of protection for the food. Within Apidae, Euglossini store the food in cells with their brood but the other three social tribes of bees Apini, Bombini, and Meliponini store nectar in the form of honey in separate vessels (Noll 2002). Honey is a very concentrated solution of glucose, fructose (produced from the hydrolysis of sucrose), and other nutrients found in the nectar gathered (Morse 1972). The concentration of sugars in Apis honey is so high that it acts as a desiccant preventing microorganisms from growing in the stores (Morse 1972). However, in stingless bee honeys, sugar concentrations are generally not sufficiently high to prevent microorganism growth (Roubik 1989). In some Trigona honey pots, certain bacteria live in the honey and may chemically prevent the invasion of other microbes (Machado 1971, Gilliam et al. 1985). Most species of bees also store pollen (Michener, 2007) as a protein source (Herbert and Shimanuki 1978). Necrophagous Trigona necrophaga do not store pollen, but instead produce pots filled with a yellowish liquid (Camargo and Roubik 1991) which is a hypopharyngeal proteinaceous secretion. However, the necrophagous bee, Trigona hypogea, appears to combine hypopharyngeal secretions with its honey instead of separating the two (Serrão et al. 1997). There is growing evidence that many species of wasps produce honey as food stores. Temperate and tropical wasp colonies have been noted to store honey solutions for the colony (Wheeler 1908, Rau 1928, Schwarz 1929, Cooper 1993, Kojima 1996, Telleria 1996, Hunt et al. 1998, Guimaráes et al. 2008) or wintering gynes (Strassmann 1979). Wasp honey has similar properties as bee honey but the mechanism used to produce it is still unknown (Hunt et al. 1998). Wasps do not appear to have any form of external long-term protein storage. Telleria (1996) found small amounts of pollen and insect pieces in the honey of Polybia scutellaris but this was probably a byproduct of foraging for nectar. There are a few reports of Polybia occidentalis nests having large numbers of alate termites and ants stuffed in cells (Richards 1978, Jeanne and Taylor 2009). How long these stores last has yet to be investigated. Caching: In this form of storage, seeds are gathered, in some cases dried, and stored in granaries (Tschinkel 1998, Wilson 2003). Dried seeds are another food product that microorganisms find difficult to colonize (Kremer et al. 1984). The seed itself is an adaptation to protect the embryo from the natural environment and provide the embryo with all of the energy it needs to sprout (Raven et al. 1999). Seeds tend to be high in both lipids and proteins

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Timothy M. Judd

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(Kelrick and Macmohan 1985, Judd 2006) providing the ants with a rich source of energy and nitrogen (Judd 2006). For an organism trying to store food, the seed is a perfect nutritious, long-term food storage device. Seed-harvesting has evolved independently several times within Formicidae (Hölldobler and Wilson 1990). These seed-storing ants gather large quantities of seeds. Estimates suggest that the ant Pogonomyrmex can remove up to 10% of the total seed production of the plant community in one year. The seeds gathered comprise of 50% of the seeds of their preferred plant (Whitford 1978). Seed preference has been documented in many seed-harvesting ants. Size, mass and odor have all been attributed to seed preference (Detrain and Pasteels 2000, Reyes-Lopez and Fernandez-Haeger 2001, Valentim et al. 2007). A less explored possibility is that some seeds are more nutritious than others. Judd (2006) found that the preferred seed of Pheidole ceres, sagebrush, has a higher percentage of protein per seed than any other seeds it collects. In this case, the ants preferred the most nutritious seeds. However, whether this preference is due to ants cuing in on seed nutritional content remains to be seen. Nicolai et al. (2007) presented Pogonomyrmex barbatus with seeds varying in protein and carbohydrate content in the fall and the spring. The ants did not choose seeds based on nutritional value. Their results suggests that ants may not be able to determine the nutritional content of individual seeds. It is possible that seed harvesters evolved in close association with certain species of plants. Internal stores: Food can be stored in several places inside the hymenopteran body. Fat bodies, hemolymph, and muscles all have been shown to be potential food storage locales (Wheeler and Martinez 1995, Nation 2002). In some dimorphic ants the majors store a higher percentage of lipids in hypertrophied fat bodies than minors (Wheeler 1994), thus acting as repletes for the colony (Wilson 1974, Stradling 1987, Lachaud et al. 1992, Tschinkel 1993). Even without repletes, ant colonies can persist for some time on their internal stores (Rueppell and Kirkman 2005). One could not review internal food stores without mentioning the honey pot ants, Myrmecocystus (Rissing 1984). These ants have special workers whose abdomens expand well beyond their normal size. Storage proteins are the main mechanism for storing protein internally in wasps (Hunt et al. 2003), bees (Evans and Wheeler 1999), and ants (Martinez and Wheeler 1994, Rosell and Wheeler 1995, Wheeler and Martinez 1995, Lim et al. 2005). However, these proteins were noticeably absent in adults of Vespula maculifrons and adult workers of Dolichovespula maculata (Hunt et al. 2003). In addition, the use of storage proteins has been found the solitary wasp, Monobia quadridens (Hunt et al. 2003), suggesting it is a storage mechanism inherited from solitary ancestors and possibly lost in Vespinae. Individuals of annual colonies use internal stores as well. Gynes of Polistes metricus have higher lipid content in fat bodies than workers (Toth et al. 2009) and gynes have high levels of sodium when overwintering (Judd et al. 2010). There is even evidence of workers and male Polistes having temporary internal storage abilities (Judd et al. 2010). Polistes foundresses have been shown increase their levels of protein in the spring (Judd et al. 2010). The aforementioned repletes in ants may have retained the storage abilities of overwintering gynes. It would be interesting to see if the same genetic mechanisms regulate food storage in repletes and the gynes. Comparing the mechanisms: When the three food storage methods are compared, clearly canning takes more time than the other two methods. First, nectar should be broken down into Social Insects: Structure, Function, and Behavior : Structure, Function, and Behavior, edited by Emily M. Stewart, Nova Science Publishers,

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its monomer sugars. Bees have an enzyme, invertase, which hydrolyzes disaccharides into monosaccharides (Michener 1974). The enzyme used by wasps has yet to be identified (Hunt 2007). Second, water is removed from the solution to increase the concentration (Seeley 1997). Finally, the cell is sealed. Caching on the other hand takes time but drying seeds does not require constant attention. The food is already packaged; it just needs to be collected. Therefore, foragers are free to collect more seeds. In some granivorous ants, such as Pheidole, special castes have evolved to mill seeds (Sempo and Detrain 2004). Internal food stores are probably the most efficient in terms of time. The only requirement is some internal mechanism allowing for food storage. However, the number of individuals in the colony limits this form of storage. Plus, there is an additional cost to carrying around the extra weight (Duncan and Lighton 1994). In comparison, canning and seed storage are more advantageous as the potential for storage is limitless.

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FOOD STORAGE AND WORKER ALLOCATION The evolution of long-term food storage required an ability to allocate a certain percentage of the incoming resources to storage. Both canners and cachers store products of plant reproduction, pollen, nectar and seeds, which coincide with the optimal time for colonies to grow and reproduce. Even colonies that rely on internal stores should collect the nutrient during these optimal times. Thus, food-storing Hymenoptera should collect food for the colony‘s immediate needs and accumulate sufficient food for long-term stores. As a result, the overall flow of nutrients and information should be organized differently in colonies that store food long-term verses colonies that do not (Figure 1). Colonies need a mechanism that separates immediate stores from those shunted for long-term storage. Two possible mechanisms that would allow for this would be to 1) use separate strategies for longterm and short-term storage and 2) separate the foragers from food storage. Smith (2007) provides evidence that there are separate mechanisms regulating long and short-term storage. He found that colonies of the harvester ant Pogonomyrmex badius that were starved for two months relied solely on their internal stores rather than using their seed caches. This result suggests that the colonies have two separate mechanisms regulating the use of short and long-term stores. Further studies have revealed that the larvae are not fed from the seed caches (Smith and Suarez 2010). In seed caching ants, the collection of seeds may be strictly used for long-term storage whereas the internal lipid reserves may be used to as a buffer during the active season of the colony. Many seed caching ants also use repletes (Wilson 1974, Lachaud et al. 1992, Tschinkel 1998) or store different nutrients for immediate use than long-term use (Judd 2006). In honeybees, stingless bees, and perennial Epiponii, the nectar foraging force is generally removed from the food storing process. Foragers return to the colony and usually unload to a food-receiving caste that processes the nectar (Hunt et al. 1987, Seeley 1995, Cepeda 2006). This is an important adaption because honey production is time consuming. A food-receiving caste allows for a greater turnaround in by the foraging force (Anderson and Ratnieks 1999). This caste may also serve as an intermediate point between nurses and foragers (Figure 1). Thus, nectar foragers are generally not in direct contact with nurses. The separation of nectar foragers from nurses is important because the nutritional state of nurses

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reflect the immediate needs of the colony (larvae, queen, and themselves). The lack of communication between nectar foragers and nurse caste means the foraging force is not directly influenced by the colony‘s immediate nutritional needs. This would allow for some food to be shunted for long-term storage. Thus, foragers are left to maximize food intake. Annual bees and wasps lack the food-storer caste (Noll 2002, Jeanne and Taylor 2009) and thus the workers can be influenced by the colony‘s immediate needs.

Figure 1. Flow of nutrients in annual and perennial colonies. Solid lines indicate paths that occur in all colonies and dotted lines indicate paths that occur in some colonies. In annual colonies queens will initially feed the larvae. Once colonies produce workers, foragers will interact with the adults and the larvae. In some species adults will occasionally take food from foragers and feed other colony mates. In perennial colonies, foragers hand food off to a receiver caste. In some species the receiver will pass food to a nurse caste while in other species the receiver feeds other colony members. In some cases, reproductives will intercept incoming foragers (Judd 2005). Thus, foragers in perennial colonies are at least one step removed from the larvae and the food stores.

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In both forms of external stores (canning and caching) the foragers are unable to assess the total amount of food stored in the colony. Thus, seed collection in ants and nectar collection in perennial bees and Epiponii is uncapped allowing for the colony to take in more food than the colony‘s immediate needs. These colonies are able to take advantage of the periods in which food availability is high.

INFORMATION FLOW IN A HYMENOPTERAN COLONY The composition of individuals in a colony changes throughout the year. As a result, the needs of a colony as a whole change as well. In addition, natural fluctuations in resource availability can affect the flow of nutrients into the colony. Three types of information can affect the flow of nutrients: 1) The nutritional needs of individuals, 2) where the food should be distributed and 3) where the food source is located. Of the three categories, only the first two types of information need to exist for a colony to operate. The latter type of information may add to the efficiency of colony operation.

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Information about what to acquire: Foragers are faced with the decision as to what types of foods are needed by the colony. Colonies‘ needs change, and foragers need a reliable indicator by which to determine the needs of the colony. Foragers can acquire this information from other colony-mates or by assessing their own nutritional state. However, if colonies maintain long-term food stores then it may actually more adaptive if the foragers are unaware of the colony‘s nutritional state. Foragers will continue to forage for food, even if the colony is temporarily satiated, in the absence of information about their colony‘s immediate needs. Forager-nonforager communication: Because nutritional fluctuations are caused by the intermittent production of larvae and sexuals, it seems reasonable to predict that larvae may convey their needs to the workers. Worker-larvae communication does seem to occur in annual colonies. Polistes fuscatus workers appear to communicate with larvae through the use of nest vibrations (Gamboa and Dew 1981, Downing and Jeanne 1985, Savoyard et al. 1998). Wasp larvae also give food secretions to adults. Thus, it is possible for communication to occur directly through the use of pheromones or indirectly as workers directly assess the quality of the larval saliva (Hunt 1991, 2007). In large perennial colonies, larvae-forager communication is less likely because the larvae and foragers rarely interact (Figure 1). An intermediate caste usually feeds the larvae. In these colonies, communication between foragers and an intermediate caste would be more likely to occur. In honeybees, the level of pollen foraging is controlled through communication between nurse bees and pollen foragers (Camazine 1993). The nurse bees are the individuals that distribute the protein to the larvae. In addition to this process, nurse bees also give a small amount of food to pollen foragers. If the food is high in protein or amino acids then the number of pollen foragers is reduced (Camazine 1993, Seeley, 1997). In this case, the larvae are indirectly influencing the workers as they drain the nurse bees‘ reserves. Interestingly the flow of nectar is regulated by the interaction of foragers and food-storers. Thus, the nectar foragers and nurse bees do not interact. Honeybee nectar foragers able to

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influence the numbers of food-storer bees through the tremble dance and increase the foraging force through the waggle dance. In this manner the honey bee colony is able to increase its food production rate in times of high nectar influx (Seeley 1993, Seeley et al. 1996). The primary food that is needed for a honeybee colony to overwinter is honey. The addition of a food-storer caste prevents nectar foragers from tuning into the colony‘s immediate carbohydrate needs. In pollen foraging the workers are influenced by the nurse bees, meaning that the intake of nitrogen (protein) is regulated by the colony‘s immediate needs. Indeed, recent studies have shown that the development of pollen foragers is different than nectar foragers (Page et al. 2006) providing evidence of a genetic mechanism that separates short term and long term nutrient regulation in Apis mellifera. In perennial Epiponii, foragers hand off food, water, and pulp to a receiving caste (Hunt et al. 1987). These receivers malaxate the prey, produce honey, and feed larvae leaving the forager free forage again (Jeanne 1986). In Polybia occidentalis, there is some evidence that the youngest workers focus on inside tasks such as brood care and the middle aged adults are found on the nest surface, where the receivers are located, suggesting different individuals are responsible for reception and brood care (Jeanne 1991a). As with honey bees, this work force structure would further separate foragers from the brood care, honey production, and information concerning the colony‘s immediate nutritional needs. Foragers seem to be able to activate other foragers (O'Donnell 2001). The age of onset of receiving behavior varies in P. occidentalis which suggests there may be signals to regulate this caste as well. There is mixed evidence for larva-worker communication in ants. Brian (1977) found evidence that Myrmica rubra larvae influenced the foraging behavior of workers. Sorenson et al. (1985) showed that Solenopsis invicta larvae deprived of lipids increased the colony‘s intake of oil. In the latter study, however, it appeared that the nurse ants might be involved in driving this behavior rather than the larvae (Sorenson et al. 1985). In a different experiments, Cassill &Tschinkel (1995) and Judd (2005) observed that larvae did not directly influence forager behavior in S. invicta and Pheidole ceres respectively. These results are consistent with the hypothesis that larvae of perennial colonies are not interacting with the foragers. It is very likely that a similar phenomenon was happening in the study by Brian (1977). Self-Assessment: Instead of relying on signals from colony-mates, foragers could cue into their own nutritional stores to assess their colony‘s needs. In a colony, non-foragers are sinks for whatever nutrients they need. Larvae are generally protein sinks whereas reproductive adults are carbohydrate sinks. Thus, the nutrients flow from the gatherers to these sinks. Foragers are the last individuals in a colony to be satiated. They only lose those nutrients that are in demand and as a result have a reliable cue in which to determine what nutrients are needed. However, this process can only function if the species depends solely on internal food stores. Self-assessment probably does not occur in bees or perennial Epiponii because they store nutrients in excess leaving the forager without a reliable internal cue. Nectar foragers are always ―empty‖ so will continue to forage (O'Donnell and Jeanne 1995). Ants store food internally, so self-assessment is a viable option for foragers. Several lines of evidence support the hypothesis that ant foragers can self-assess. Workers of Leptothorax albipennis are more likely to forage when they were low in lipids (Blanchard et al. 2000, Robinson et al. 2009). Judd (2006) found that the food preferences of Pheidole ceres matched the nutritional states of the ants. When they were low on a particular nutrient they tended to forage for it. In addition, P. ceres workers will not forage if satiated on

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carbohydrates (Judd 2005). Sorenson et al. (1985) reported that foragers preferred nutrients that are lacking in nurse ants. Presumably, the workers themselves were low in these foods. Thus, unlike bees, ants might use their own nutritional state to make foraging decisions rather than rely on information through food exchange. However, the evidence supporting this conclusion is correlative and causation has yet to be directly tested. Communication between foragers: Although mechanisms are in place that determine what food types are needed, annual colonies evidently lack a mechanism for workers to recruit others to a food source at a specific location (Jeanne et al. 1995) but do have the ability to mobilize foragers. Odor of nectar sources can be passed on to other recruits in bumble bees (Dornhaus and Chittka 2004a, Nieh and Renner 2008, Molet et al. 2009) and yellow jackets (Overmyer and Jeanne 1998, Jant and Jeanne 2005). In addition to odor information, some species are able to alert other potential foragers that food is available outside the colony through the use of pheromones (Dornhaus et al. 2003, Granero et al. 2005), excited motions (Dornhaus and Chittka 2001, 2004a), and perhaps even convey thermal information about food quality (Mapalad et al. 2008). In contrast, many species with perennial colonies show some level of directed recruitment, the ability to pass on the location of a food source. Ants show communication ranging from tandem running in some smaller ant species (Hölldobler and Wilson 1990), chemical trails (Hölldobler and Wilson 1990) in larger colonies, and permanent trunk trails (Traniello 1989, Salzelemann 1992, Lopez et al. 1994). Bees have a range of communication mechanisms (Michener 1974). Honey bees (von Frisch 1967) and several stingless bee species (Nieh 1998, Jarau et al. 2000, Jarau et al. 2002, Jarau et al. 2003, Nieh et al. 2003a, Nieh et al. 2003b, Jarau et al. 2004, Nieh 2004, Hrncir et al. 2006, Barth et al. 2008, Schorkopf et al. 2009) have recruitment behaviors, either inside the nest or outside the nest (such as odor trails or bees following nestmates, piloting), that direct colony members to food sources. In general, such communication processes facilitate quick exploitation of food sources (Phillips et al. 1978, Biesmeijer and Slaa 2004). Ants and social bees are very strong competitors, due partially to the sheer numbers they can quickly send to a food source (Seeley 1987, Traniello 1987, 1989). Perennial wasps (Jeanne and Taylor 2009) and some stingless bees species (Nieh 2004) do not have directed recruitment but these are all tropical species. Therefore the pattern that emerges here is that perennial hymenoptera in fluctuating climates, which for the remaining portion of the chapter will be defined as any climate in which food is not available all year round (temperate, wet-dry, and high altitude climates), all have directed recruitment where as perennial species in non-fluctuating environments may or may not have directed recruitment. The distribution and patchiness of food sources can play a role in the utility of directed recruitment. For example, the honey bee waggle dance provided a selective advantage to colonies, allowing them to increase in weight in an environment with patchily distributed food sources (Sherman and Visscher 2002). However, honey bee colonies in areas with less patchily distributed resources did not benefit as much from the waggle dance (Dornhaus 2002, Dornhaus and Chittka 2004b). Therefore individual search and discovery of resources without communication of resource location can be sufficient in certain environments (Biesmeijer and Slaa 2004, Nieh 2004, Jeanne and Taylor 2009). In fluctuating climates, there is a limited window in which to gather the necessary resources to sustain a colony to the next season while in non-fluctuating environments this pressure is relaxed. Therefore, in order to

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maintain a perennial existence in a fluctuating environment, it may be necessary to have a directed recruitment system.

PERENNIAL COLONIES IN FLUCTUATING AND NON-FLUCTUATING CLIMATES

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Energy storage, protein (nitrogen) storage, food-receiving castes, and directed recruitment all have a potential role in promoting transition from an annual to a perennial life history. However, the necessity of some of these traits in maintaining a perennial life history may vary depending on the climate the colony is adapted to. Fluctuating environments require the colony to be able to sustain itself through a harsh season while this is not the case in nonfluctuating environments. Thus, in the latter climate some of the selective pressures that might prevent the evolution of perennial life histories are relaxed. 1. Energy storage: All perennial colonies have evolved some mechanism for energy storage regardless of the climate. Bees and perennial Epiponii store honey while ants utilize repletes and/or seed caches. Therefore, it appears the ability to store large levels of energy externally or internally is necessary for maintaining perennial colonies in any climate (Table 1). The levels of the stores would increase as you move from non-fluctuating to fluctuating climates. Colonies in border climates should show intermediate levels of energy stores as compare to the levels in the two extremes (Table 1). 2. Protein storage: The evidence thus far suggests that protein storage is only necessary in fluctuating environments (Table 1). Presumably, protein stores allow the colony to get a head start on brood production after the harsh season. Bees use external pollen stores while ants use internal stores. However, the level of protein has not been monitored in perennial wasps. It would be interesting to see if storage proteins exist in species of Polistinae other than Polistes, especially the perennial Epiponii. 3. Food-receiving caste: The existence of the food-receiving caste could have facilitated the evolution of food storage in two ways. First, it would have separated the forager from any cues concerning its colony‘s immediate needs. Second, the food-receiver allows the forager to have a faster turnaround time at the nest which would increase colony food intake (Ratnieks and Anderson 1999). Thus, the foodstorer caste is probably essential for all perennial colonies (Table 1). 4. Directed recruitment: As discussed above, directed recruitment appears in perennial colonies living in fluctuating environments but not necessarily in non-fluctuating environments. Thus, directed recruitment is probably essential for perennial colonies to exists in fluctuating environments where food is more patchy and ephemeral but not non-fluctuating environments (Table 1).

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Table 1. Summary of the predicted necessity of food storage, food-receiving castes and directed recruitment for the evolution of a perennial life history in social Hymenoptera in several climates

Fluctuating Climates (Temperate, Wet/dry, & High altitude) Border Climates (Interface between Fluctuating and nonfluctuating climates) Non-Fluctuating Climates (Tropical rainforest)

Storage of Energy Food Essential

Storage of N Food Essential

Food-Receiving Caste Essential

Directed Recruitment Essential

Essential

Not essential

Essential

Not essential

Somewhat Essential

Not essential

Essential

Not essential

The following scenarios propose how the four traits may have played a role in the evolution of perennial life histories in bees wasps and ants respectively.

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Bees Food storage exists at some level in all nesting bees (Michener 1974). Solitary bees such as the trap-nest bees and burrowing bees create series cells containing pollen, nectar and a single egg (Krombein 1967, Michener 2007). The behavior of storing food with brood exists in social Euglossini bees as well. One important step in the evolution of food storage in social bees was the separation of brood from food stores (Noll 2002). This behavior was accompanied by provisioning of larvae by adults. With this separation in place, the food stores were no longer restricted to the amount of brood and set the stage for the evolution of long-term food storage. Bombini, Apini, and Meliponini all store the food in separate vessels (Noll 2002). Of the three aforementioned tribes, Bombini is the only non-perennial bee. Unlike bumble bees, stingless bees and honeybees have a food-storing caste (Noll 2002), further separating the nectar forager from the nurse caste. Although Bombus can alert others that food has been located (Dornhaus and Chittka 2001, Dornhaus et al. 2003, Mapalad et al. 2008, Nieh and Renner 2008) and other bees can learn the odor of a food source (Dornhaus and Chittka 2004a, Kitaoka and Nieh 2009, Molet et al. 2009), there is no transfer of location information (Dornhaus & Chittka 2001). The lack of directed recruitment between foragers probably prevents bumblebee colonies from persisting from year to year because they are unable to gather enough resources to survive in a fluctuating climate. Apini and many Meliponini, on the other hand, are perennial and have both long-term food storage and methods to transmit locations of food sources (Seeley 1995, Nieh 2004). Some meliponines lack directed recruitment mechanisms but they are in nonfluctuating climates. Directed recruitment may not be necessary for them to obtain enough food (Slaa et al. 1997, Breed et al. 2002, Biesmeijer and Slaa 2004, Nieh 2004). In bees, the evolution of a food-receiving caste may have allowed bees to make the jump to a perennial lifestyle and in some species the evolution of directed recruitment probably

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facilitated this change in life-history. The phylogenetic relationship between the four tribes of social bees is still not completely resolved. However, many studies suggest that perennial (advance) eusocial behavior evolved multiple times (Koulianos et al. 1999, Noll 2002). Given the scenario presented here, this conclusion would not be too difficult to imagine. Bees were preadapted to store pollen and nectar. Three of the four tribes of social bees store the food separate from the brood. All three have mechanisms to alert colony mates to the presence of food. Thus, a major change that would have facilitated a transition to a perennial life history would have been the evolution of a food-receiving caste.

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Wasps Unlike bees, solitary nesting wasps generally provision the cells with particular prey items (Krombein 1967, Evans and O'Neill 2007). The only exception to this rule is the pollen wasps, but no social pollen wasps exist (Gess 1996). Prey items do not make viable long-term storage vessels. They can decay if the prey perishes or the prey will metamorphose into an adult. Unlike bees, continuous larval provisioning by adults was essential to the evolution of social behavior in wasps (Hunt 2007). Within wasps, only Polistinae and Vespinae have perennial representatives. I will discuss these two subfamilies separately. It is interesting to note that all of the wasps that have been recorded to produce honey are all members of Polistinae. There is an absence of evidence that Vesipinae produces honey at all (Simon and Benton 1968, Spradbery 1971, MacDonald and Matthews 1976, MacDonald et al. 1980, Makino 1982, Ross and Matthews 1982, Reed and Akre 1983, Ross and Visscher 1983, Starr and Jacobson 1990, Greene 1991, Matsuura 1991, Ratnieks and Miller 1993, Landolt et al. 2009). Simon and Benton (1968) found a colony of Vespula maculifrons persisting through the winter (mostly reproductives) in Pennsylvania. The authors report that there were no signs of food storage but there were brood in the cells. Other studies that looked at Vespula colonies that survived through a mild winter did not report honey deposits in cells (Ross and Matthews 1982, Ross and Visscher 1983, Ratnieks and Miller 1993, Landolt et al. 2009). Therefore it is possible that honey storage evolved in Polistinae. The basal genera of Polistinae, Polistes, Mischocyttarus, and Ropalidia produce honey droplets in comb late in the colony season. Epiponii such as Brachygastra (Schwarz 1929, Cooper 1993, Sugen and McAllen 1994) and Polybia (Jeanne 1991b, Telleria 1996, Hunt et al. 1998, Hrncir et al. 2007) have been reported to have large levels of honey stores. Hunt (2007) proposed that primitively eusocial wasps, with worker and gyne phenotypes, have similarities to bivoltine solitary wasp life cycles. This suggest that the developmental pathways may be similar in both systems because every other generation persists through a harsh season in which food availability would be non-existent. This reasoning also suggests that wasps evolved in a fluctuating climate. Given that wasps evolved in this type of climate, the evolution of honey storage could have matched the movement of Polistinae to milder climates. Once the social wasps extended into non-fluctuating climates, the pressures of the fluctuating climates were relaxed allowing colonies to get larger. In Polistinae, this colony enlargement may have been accompanied by more extensive carbohydrate storage. Thus, honey storage may have originated as a mechanism to allow gynes to persist into the early part of the harsh season in fluctuating climates. This behavior could have allowed colonies in

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border climates to persist through the entire harsh season as seen by Polistes annularis (Strassmann 1979) and Ropalidia romandi (Kojima 1996). With the pressures of the harsh seasons reduced, polistine colonies in non-fluctuating environments could have grown larger and increased their level of food storage. The Epiponii all have a food receiving caste that can feed the larvae or store food (Jeanne 1991a). Thus, the evolution of perennial life in Epiponii probably occurred in non-fluctuating and mild border climates. The transition from an annual to perennial life history could have been facilitated by the evolution of the receiver caste and the ability to store honey. The lack of directed recruitment and continuous long-term protein stores probably prevented the perennial Epiponii from persisting in fluctuating environments. There are two major patterns found in the Epiponii that match the model outlined in Table 1. First, the hypothesis that directed recruitment is necessary for perennial colonies in fluctuating climates is supported by the fact that no known species of Epiponii appears to have developed directed recruitment and persists in such a climate (Gadagkar 1991, Jeanne 1991b). Naumann (1970) reported a dance performed by returning foragers in Protopolybia pumila but there is no evidence this actually recruit s other foragers to a food source (Jeanne 2009). Directed recruitment has yet to be found in any wasp (Jeanne 2009). Perennial Epiponii are found in non-fluctuating climates and border climates (Table 1). In these climates, directed recruitment is not necessary for the maintenance of a perennial life history. The hypothesis that energy storage is necessary for a perennial life history is further supported by observations that levels of honey storage are more prevalent in border climates than non-fluctuating climates (Hunt et al. 1987, Jeanne 1991b, Hunt et al. 1998). In addition, the honey stores are at their largest when the harsher aspect of the season exists. In nonfluctuating climates food is available all year round and the pressures that require food storage are relaxed and the food stores do not have to be large. Most Vespinae have annual lifecycles regardless of the climate (Greene 1991, Matsuura 1991). Some large nesting Vespula colonies can persist for more than one season in warmer or sheltered climates but not colder ones (Greene 1991). V. vulgaris colonies have been observed to forage throughout the winter (Gambino 1986). Thus, several species of Vespula show colony perennial life histories in border climates but remain annual in fluctuating climates. In the perennial colonies, the gynes do not go into diapause but instead return to a nest and lay eggs. However, it is not clear that these colonies persist after the second year (Greene 1991). This is a sharp contrast to many perennial Epiponii which remain active for years (Jeanne 1991b). The low numbers of perennial colonies in Vespinae is probably due to limited food storage, lack of a food receiving caste and minimal recruitment. There is some cursory evidence for internal energy storage in overwintering colonies of perrenial Vespula. Simon and Benton (1968) noted that the internal nest temperature of an over-wintering V. maculifrons colony was higher than the ambient temperature suggesting that the wasps were regulating temperature. This would require internal stores but this was not directly tested. Thus, internal energy stores may be sufficient to maintain a colony in the border climates of Vespula but not in the fluctuating climates. Unlike Epiponii, Vespinae do not have a specific food-receiving caste. Food may or may not be handed off to other wasps by returning foragers (Greene 1991, Jeanne and Taylor 2009). Workers tend hand off food to other workers in large Vespula colonies more often because the chances of running into a recipient is higher (Jeanne 2003). In the Vespinae, there is some evidence of forager communication but no directed recruitment. V. germanica has been shown to use cues from returning foragers to locate a food source (Overmyer and Jeanne 1998) and local

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enhancement (D'Adamo et al. 2000, D'Adamo et al. 2003, D'Adamo and Lozada 2005). Vespa mandarina engage in group attacks on other hymenopteran colonies. This may be coordinated by pheromones (Matsuura 1991). However most of the nutrients gained from these are not stored. Overall, it appears that the lack of external food stores and a food receiver caste has probably prevented most of the Vespinae from adapting a perennial life history. The large numbers in the Vespula colonies in border climates may be able to provide enough internal energy stores and mimic a food-receiving caste such that they can persist for a second year.

Ants

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In contrast to bees and wasps, all ants have perennial life histories (Hölldobler and Wilson 1990). In fact, most ant colonies will take over a year to mature enough to reproduce (Hölldobler and Wilson 1990, Bourke and Franks 1995). This is a sharp contrast to what is seen in wasp and many bee species. Ants either cache seeds or rely solely on their internal stores to survive long-term. Ant species that rely on internal stores tend to have a replete caste (Wheeler 1994). As with the wasp relatives, the tropical ants are probably under less pressure to store food than temperate ants. Most ants use some form of directed recruitment either tandem running or recruitment pheromones (Hölldobler and Wilson 1990). Ants generally have a food-receiving caste. Therefore, ants have all of the requirements that would facilitate the maintenance of perennial life histories. One interesting question is why ants can persist on stored foods while wasps (apart from the gynes) cannot in fluctuating environments. One possibility is that ant workers are wingless, which suggests the energy requirements per adult individual are less than a flying wasp. The addition of directed recruitment and repletes may further increase an ant colony‘s ability to store energy relative to wasps.

CONCLUSION When examining the evolution of the perennial life history it is clear that climate can influence the relative importance of various traits. Energy storage and the presence of a foodreceiving caste seems to be universal in all perennial colonies. The existence of protein storage and directed recruitment appears to be essential in fluctuating climates but not in nonfluctuating climates (Table 1). This pattern holds true for Apidea, Vespidae, and Formicidae. Jeanne (2003) reviewed the many ways in which insect societies have been divided. He adopted the terms ―simple‖ and ―complex‖ societies from Bourke (1999) and broadened them to fit all insect societies. Simple societies consist of smaller colonies in which there is reproductive conflict between workers and reproductives and a dominance hierarchy. In complex societies the conflict is minimal, there are no dominance hierarchies, and the queen produces pheromones to maintain colony cohesion (Jeanne 2003). He further pointed out that in wasps, complex societies are represented by the swarm-founding wasps while the simple societies are initiated by a single foundress. One could make a similar argument for bees (Michener 1969). What drives the distinction between complex and simple societies? One possibility is the change the flow of nutrients through a colony. The introduction of a foodreceiving caste would change the dynamics of a colony. No longer are all workers in direct

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contact with larvae and queens are dependent on a specific caste for nourishment (Figure 1). This separation also limits the effectiveness of dominance behavior in queens which could reduce direct queen-worker conflict. Long term food storage would select for larger colonies to improve food intake. Indeed colonies of perennial Epiponii are larger in the border climates (Jeanne 1991a). Jeanne (2003) proposed that the reason the large Vespula colonies never moved to the complex societies was the lack of worker specialization. This would correlate with the lack of a distinct food-storing caste presented in this chapter. Jeanne (2003) proposed the swarm might have been the defining adaptation to allow for the transition to complex societies. I suggest that the existence of external stores and a food receiving caste were additional driving forces that allowed the swarm-founding wasps and bees to become complex and possibly perennial. Ants, considered complex societies (Bourke 1999, Jeanne 2003), represent a different type of perennial colony. They are founded by single individuals and colonies are slow to mature (Bourke and Franks 1995). Ants have energy and protein storage, food-receiving castes, and directed recruitment. Therefore, swarming is not essential for colonies to be complex or perennial. Perhaps it is the lack of external food stores and the lack of defined food-receivers that prevented the large Vespula colonies to become complex societies. Not all Epiponii are perennial (Jeanne 1991b) , thus, this tribe may provide some insight into the evolution of the perennial life history once more species are carefully examined, especially colonies with a food-receiving caste.

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REFERENCES Anderson, C. & Ratnieks, F. L. W. (1999). Task partitioning in insect societies. I. effect of colony size on queueing delay and colony ergonomic efficiency. The American Naturalist, 154, 521-535. Barth, F. G., Hrncir, M. & Jarau, S. (2008). Signals and cues in the recruitment behavior of stingless bees (Meliponini). Journal of Comparative Physiology A, 194, 313-327. Biesmeijer, J. C. & Slaa, E. J. (2004). Information flow and organization of stingless bee foraging. Apidologie, 35, 143-157. Blanchard, G. B., Orledge, G. M., Reynolds, S. E. & Franks, N. R. (2000). Division of labour and seasonality in the ant Leptothorax albipennis: worker corpulence and its influence on behaviour. Animal Behaviour, 59, 723-738. Bourke, A. F. G. (1999). Colony size, social complexity and reproductive conflict in social insects. Journal of Evolutionary Biology, 12, 245-257. Bourke, F. G. & Franks, N. R. (1995). Social Evolution in Ants. Princeton University Press, Princeton. Breed, M. D., Stocker, E. M., Baumgartner, L. K. & Vargas, S. A. (2002). Time-place learning and the ecology of recruitment in a stingless bee, Trigona amalthea (Hymenoptera, Apidae). Apidologie, 33, 251-258. Brian, M. V. & Abbott, A. (1977). The control of food flow in a society of the ant Myrmica rubra L. Animal Behaviour, 25, 1047-1055. Camargo, J. M. F. & Roubik, D. W. (1991). Systematics and bionomics of the apoid obligate necrophages: the Trigona hypogea group (Hymenoptera: Apidae; Meliponinae). Biological Journal of the Linnean Society, 44, 13-39.

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Camazine, S. (1993). The regulation of pollen foraging by honeybees: how foragers assess the colony's need for pollen. Behavioral Ecology and Sociobiology, 32, 265-272. Cassill, D. L. & Tschinkel, W. R. (1995). Allocation of liquid food to larvae via trophallaxis in colonies of the fire ant, Solenopsis invicta. Animal Behaviour, 50, 801-813. Cepeda, O. I. (2006). Division of labor during brood production in stingless bees with special reference to individual participation. Apidologie, 37, 175-190. Cooper, M. (1993). A social wasp which stores nectar. Sphecos, 26, 8. D'Adamo, P., Corley, J. & Sackmann, P. L. M. (2000). Local enhancement in the wasp Vespula germanica are visual cues all that matter? Insectes Sociaux, 47, 289-291. D'Adamo, P. & Lozada, M. (2005). Conspecific and food attraction in the wasp Vespula germanica (Hymenoptera : Vespidae), and their possible contributions to control. Annals of the Entomological Society of America, 98, 236. D'Adamo, P., Lozada, M. & Corley, J. (2003). Conspecifics enhance attraction of Vespula germanica (Hymenoptera : Vespidae) foragers to food baits. Annals of the Entomological Society of America, 96, 685. Detrain, C. & Pasteels, J. M. (2000). Seed preferences of the harvester ant Messor barbarus in a Mediterranian mosaic grassland (Hymenoptera: Formicidae). Sociobiology, 35, 3548. Dornhaus, A. (2002). When do dances make a difference? Significance of honey bee recruitment depends on foraging distance. Entomologia Generalis, 26, 93-100. Dornhaus, A., Brockmann, A. & Chittka, L. (2003). Bumble bees alert to food with pheromone from tergal gland. Journal of Comparative Physiology A, 189, 47-51. Dornhaus, A. & Chittka, L. (2001). Food alert in bumblebees (Bombus terrestris): possible mechanisms and evolutionary implications. Behavioral Ecology and Sociobiology, 50, 570-576. Dornhaus, A. & Chittka, L. (2004a). Information flow and regulation of foraging activity in bumble bees (Bombus spp.). Apidologie, 35, 183-192. Dornhaus, A. & Chittka, L. (2004b). Why do honey bees dance? Behavioral Ecology and Sociobiology, 55, 395-401. Downing, H. A. & Jeanne, R. L. (1985). Communication of status in the social wasp Polistes fuscatus (Hymenoptera, Vespidae). Zeitschrift Fur Tierpsychologie-Journal of Comparative Ethology, 67, 78-96. Duncan, F. D. & Lighton, J. R. B. (1994). The burden within: the energy cost of load carriage in the honeypot ant, Myrmecocystus. Physiological Zoology, 67, 190-203. Evans, H. E. & O'Neill, K. M. (2007). The Sand Wasps Natural History and Behavior. Harvard University Press, London. Evans, J. D. & Wheeler, D. E. (1999). Differential gene expression between developing queens and workers in the honey bee, Apis mellifera. Proceedings of the National Acadamy of Science USA, 96, 5575-5580. Gadagkar, R. (1991). Belonogaster, Mishocyttarus, Parapolybia and independent-founding Ropalidia. In: K. Ross, & R. Mathews, (eds) The Social Biology of Wasps. Comstock Publishing Associates, Ithaca, 149-190. Gambino, P. (1986). Winter prey collection at a perennial colony of Paravespula vulgaris (L.) (Hymenoptera: Vespidae). Psyche, 93, 331-340. Gamboa, G. J. & Dew, H. E. (1981). Intracolonial communication by body oscillations in the paper wasp, Polistes metricus. Insectes Sociaux, 28, 13-26.

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17

Gess, S. K. (1996). The Pollen Wasps: Ecology and Natural History of the Masarinae. Harvard University Press, Cambridge, MA. Gilliam, M., Buchmann, S. L. & Lorenz, B. J. (1985). Microbiology of the larval provisions of the stingless bee, Trigona hypogea, an obligate necrophage. Biotropica, 17, 28-31. Granero, A. m., Sanz, J. M. G., Gonzalez, F. J. E., Vidal, J. l. M., Dornhaus, A., Ghani, J., Serrano, A. R. & Chittka, L. (2005). Chemical compounds of the foraging recruitment pheromone in bumblebees. Naturwissenschaften, 92, 371_374. Greene, A. (1991). Dolichovespula and Vespula. In: Ross, K.,Mathews, R., (eds) The Social Biology of Wasps. Comstock Publishing Associates, Ithaca, 263-308. Guimaráes, D. L., de Castro, M. M. & Pretzoto, F. (2008). Patterns of honey storage in colonies of the social wasp Mischicyttarus (Hymenoptera, Vespidae). Sociobiology, 51, 655-660. Herbert, E. W. J. & Shimanuki, H. (1978). Chemical composition and nutritive value of beecollected and bee-stored pollen. Apidologie, 9, 33-40. Hölldobler, B. & Wilson, E. O. (1990). The Ants. Harvard University Press, Cambridge, Mass. Hrncir, M., Barth, F. G. & Tautz, J. (2006). Vibratory and airborne-sound signals in bee communication (Hymenoptera). In: S. Drosopoulous, S. & Claridge, M. F., (eds) Insect sounds and communication. Taylor & Francis Group, New York. Hrncir, M., Mateus, S. & Nascimento, F. S. (2007). Exploitation of carbohydrate food sources in Polybia occidentalis: social cues influence foraging decisions in swarm-founding wasps. Behavioral Ecology and Sociobiology, 61, 975-983. Hunt, J. H. (1991). Nourishment and evolution of the social Vespidae. In: K. Ross, & R. Mathews, (eds) The Social Biology of Wasps. Comstock Publishing Associates, Ithaca, 426-450. Hunt, J. H. (2007). The Evolution of Social Wasps. Oxford University Press, New York. Hunt, J. H., Buck, N. A. & Wheeler, D. E. (2003). Storage proteins in vespid wasps: characterization, developmental pattern, and occurrence in adults. Journal of Insect Physiology, 49, 785-794. Hunt, J. H., Jeanne, R. L., Baker, I. & Grogan, D. E. (1987). Nutrient dynamics of a swarmfounding social wasp species, Polybia occidentalis (Hymenoptera, Vespidae). Ethology, 75, 291-305. Hunt, J. H. & Nalepa, C. A. (1994). Nourishment, evolution and insect sociality. In: J. H. Hunt, & C. A. Nalepa, (eds) Nourishment and Evolution in Insect Societies. Westview Press, Boulder, 1-20. Hunt, J. H., Rossi, A. M., Holmberg, N. J., Smith, S. R. & Sherman, W. R. (1998). Nutrients in social wasp (Hymenoptera : Vespidae, Polistinae) honey. Annals of the Entomological Society of America, 91, 466-472. Jant, J. M. & Jeanne, R. L. (2005). German yellow jacket (Vespula germanica) foragers use odors inside the nest to find carbohydrate food sources. Ethology, 111, 641-651. Jarau, S., Hrncir, M., Ayasse, M., Schultz, C., Francke, W., Zucchi, R. & Barth, F. G. (2004). A stingless bee (Melipona seminigra) marks food sources with a pheromone from its claw retractor tendons. Journal of Chemical Ecology, 30, 793-804. Jarau, S., Hrncir, M., Schmidt, V. M., Zucchi, R. & Barth, F. G. (2003). Effectiveness of recruitment behavior in stingless bees (Apidae, Meliponini). Insectes Sociaux, 50, 365374.

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Jarau, S., Hrncir, M., Zucchi, R. & Barth, F. G. (2000). Recruitment behavior in stingless bees, Melipona scutellaris and M. quadrifasciata. I. Foraging at food sources differing in direction and distance. Apidologie, 31, 81-91. Jarau, S., Hrncir, M., Zucchi, R. & Barth, F. G. (2002). Foot print pheromones used to mark food sources by stingless bees. Page 16 In XIV International Congress of IUSSI: the golden jubilee proceedings. Hokkaido University Coop, Hokkaido University, Sapporo, Japan. Jeanne, R. L. (1986). The organization of work in Polybia occidentalis: costs and benefits of specialization in a social wasp. Behavioral Ecology and Sociobiology, 19, 333-341. Jeanne, R. L. (1991a). Polyethism. In: Ross, K.,Mathews, R., (eds) The Social Biology of Wasps. Comstock Publishing Associates, Ithaca, 389-425. Jeanne, R. L. (1991b). The swarm founding Polistinae. In: Ross, K.,Mathews, R., (eds) The Social Biology of Wasps. Comstock Publishing Associates, Ithaca, 191-231. Jeanne, R. L. (2003). Social complexity in the Hymenoptera, with special attention to the wasps. In: T., Kikuchi, N. Azuma, & S. Higashi, (eds) Genes,Genes,Behaviors and Evolution of Social Insects. Proceedings of Proceedings of the XIVth Congress of the IUSSI, Hokkaido University Press, Sapporo, 81-130. Jeanne, R. L. (2009). Vibrational signals in social wasps: A role in caste determination? In: J. Gadau, & J. Fewell, (eds) Organization of Insect Societies–From Genomes to Sociocomplexity. Harvard University Press, Cambridge MA, 241-263. Jeanne, R. L., Hunt, J. H. & Keeping, M. G. (1995). Foraging in social wasps - Agelaia lacks recruitment to food (Hymenoptera, Vespidae). Journal of the Kansas Entomological Society, 68, 279-289. Jeanne, R. L. & Taylor, B. J. (2009). Individual and social foraging in social wasps. In: Jarau, S.,Hrncir, M., (eds) Food Exploitation by Social Insects: Ecological, Behavioral and Theoretical Approaches. CRC Press, Boca Raton, FL, 53-79. Judd, T. M. (2005). The effects of water, season, and colony composition on foraging preferences of Pheidole ceres [Hymenoptera: Formicidae]. Journal of Insect Behavior, 18, 781-803. Judd, T. M. (2006). Relationship between food stores and foraging behavior of Pheidole ceres (Hymenoptera : Formicidae). Annals of the Entomological Society of America, 99, 398406. Judd, T. M., Magnus, R. M. & Fasnacht, M. P. (2010). A nutritional profile of the social wasp Polistes metricus: Differences in nutrient levels between castes and changes within castes during the annual life cycle. Journal of Insect Physiology, 56, 42-56. Kelrick, M. I. & Macmohan, J. A. (1985). Nutritional and physical attributes of seeds of some common sagebrush-stepp plants: some implications for ecological theory and management. Journal of Range Management, 38, 65-69. Kitaoka, T. K. & Nieh, J. C. (2009). Bumble bee pollen foraging regulation: role of pollen quality, storage levels, and odor. Behavioral Ecology and Sociobiology, 63, 501-510. Kojima, J. (1996). Colony cycle of an Australian swarm-founding paper wasp, Ropalidia romandi (Hymenoptera: Vespidae). Insectes Sociaux, 43, 411-420. Koulianos, S., Schmid-Hempel, R., Roubik, D. W. & Schmid-Hempel, P. (1999). Phylogenetic relationshipd within the corbiculate Apinae (Hymenoptera) and the evolution of eusociality. Journal of Evolutionary Biology, 12, 380-384.

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Kremer, R. J., Hughes Jr., L. B. & Aldrich, R. J. (1984). examination of microorganisms and deteriation resistance mechanism associated with velvetleaf seed. Agronomy Journal, 76, 745-749. Krombein, K. V. (1967). Trap-Nesting Wasps and Bees. Smithsonian Press, Washington D.C. Lachaud, J. P., Passera, L., Grimal, A., Detrain, C. & Beugnon, G. (1992). Lipid storage by major workers and starvation resistance in the ant Pheidole pallidula (Hymenoptera, Formicidae). In: J. Billen, (ed). Biology and Evolution of Social Insects. Leuven University Press, Leuven, 153-160. Landolt, P. J., Reed, H. C., Landolt, K. N., Sierra, J. M. & Zack, R. S. (2009). The southern yellowjacket, Vespula squamosa (Drury) (Hymenoptera: Vespidae) in Guatemala, Central America. Proceedings of the Entmological Society of Washington, 111, 429-432. Lim, S. P., Chong, K. K. K., Chong, A. S. C. & Lee, C. Y. (2005). Dietary influence on larval

storage proteins of the Pharaoh 's ant, Monomorium pharaonhs (Hymenoptera: Formicidae). Sociobiology, 46, 505-514. Lopez, F., Acosta, F. J. & Serrano, J. M. (1994). Guerilla vs. phalanx strategies of resource capture: growth and structural plasticity in the trunk trail systemof the harveter ant Messor barbarus. Journal of Animal Ecology, 63, 127-138. MacDonald, J. F. & Matthews, R. W. (1976). Nest structure and colony composition of Vespula vidua and V. consobrina (Hymenoptera: Vespidae). Annals of the Entomological Society of America, 69, 471-475. MacDonald, J. F., Matthews, R. W. & Jacobson, R. S. (1980). Nesting biology of the yellowjacket, Vespula flavopilosa (Hymnoptera: Vespidae). Journal of the Kansas Entomological Society, 53, 448-458. Machado, J. O. (1971). Simbiose entre a abelhas sociais brasileiras (Meliponinae, Apidae) e uma espécie de bactéria. Ciência e Culltura, 23, 625-633. Makino, S. (1982). Nest structure, colony composition and productivity of Dolichovespula media media and D. saxonica nipponica in Japan (Hymenoptera, Vespidae) Japanese Journal of Entomology, 50, 212-224. Mapalad, K. S., Leu, D. & Nieh, J. C. (2008). Bumble bees heat up for high quality pollen. The Journal of Experimental Biology, 211, 2239-2242. Martinez, T. & Wheeler, D. E. (1994). Storage Proteins in Adult Ants (CamponotusFestinatus) - Roles in Colony Founding By Queens and in Larval Rearing By Workers. Journal of Insect Physiology, 40, 723-729. Matsuura, M. (1991). Vespa and Provespa. In: Ross, K.,Mathews, R., (eds) The Social Biology of Wasps. Comstock Publishing Associates, Ithaca, 232-262. Michener, C. D. (1969). Comparative social behavior in bees. Annual Review of Entomology, 14, 299-342. Michener, C. D. (1974). The Social Behavior of Bees. Harvard University Press, Cambridge, MA. Michener, C. D. (2007). The Bees of the World, 2nd edn. The Johns Hopkins University Press, Baltimore. Molet, M., Chittka, L. & Raine, N. E. (2009). How floral odours are learned inside the bumblebee (Bombus terrestris) nest. Naturwissenschaften, 69, 213-219. Morse, R. A. (1972). The Complete Guide to Beekeeping. E. P. Dutton and Co., New York. Nation, J. L. (2002). Insect Physiology and Biochemistry. CRC Press, Boca Raton, Fla.

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20

Timothy M. Judd

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Naumann, M. G. (1970). The nesting behavior of Protopolybia pumila in Panama (Hymenoptera, Vespidae). Doctoral Thesis. University of Kansas, Lawrence. Nicolai, N., Cook, J. L. & Smeins, F. E. (2007). Grassland composition affects season shifts in seed preference by Pogonomyrmex barbatus (Hymenoptera: Myrmicinae) in the Edwards Plateau, Texas. Environmental Entomology, 36, 433-440. Nieh, J. C. (1998). The food recruitment dance of the stingless bee, Melipona panamica. Behavioral Ecology and Sociobiology, 43, 133-145. Nieh, J. C. (2004). Recruitment communication in stingless bees (Hymenoptera, Apidae, Meliponini). Apidologie, 35, 159-182. Nieh, J. C., Contrera, F. A. L. & Nogueira-Neto, P. (2003a). Pulsed mass-recruitment by a stingless bee, Trigona hyalinata. Proceedings of the Royal Society of London B, 270, 2791-2196. Nieh, J. C., Contrera, F. A. L., Ramirez, S. & Imperatriz-Fonseca, V. L. (2003b). Variation in the ability to communicate three-dimensional resource location by stingless bees from different habitats. Animal Behaviour, 66, 1129. Nieh, J. C. & Renner, M. A. (2008). Bumble bee olfactory information flow and contactbased foraging activation. Insectes Sociaux, 55, 417-424. Noll, F. B. (2002). Behavioral phylogeny of corbiculate Apidae (Hymenoptera; Apinae), with special reference to social behavior. Cladistics, 18, 137-153. O'Donnell, S. (2001). Worker biting interactions and task performance in a swarm-founding eusocial wasp (Polybia occidentalis, Hymenoptera : Vespidae). Behavioral Ecology, 12, 353. O'Donnell, S. & Jeanne, R. L. (1995). Worker lipid stores decrease with outside-nest taskperformance in wasps - implications for the evolution of age polyethism. Experientia, 51, 749-752. Oster, G. F. & Wilson, E. O. (1978). Caste and Ecology in the Social Insects. Princeton University Press, Princeton. Overmyer, S. L. & Jeanne, R. L. (1998). Recruitment to food by the German yellowjacket, Vespula germanica. Behavioral Ecology and Sociobiology, 42, 17-21. Page, R. E., Scheiner, R., Erber, J. & Amdam, G. V. (2006). The development and evolution of division of labor and foraging specialization in a social insect (Apis mellifera L.). Current Topics in Developmental Biology, 74, 253-286. Phillips, S. A., Jones, S. R. & Claborn, D. M. (1978). Temporal patterns of Solenopsis invicta and native ants of central Texas. In: C. S. Lofgren, & R. K. Vander Meer, (eds) Fire Ants and Leaf-Cutting Ants Biology and Management. Westview Press, Boulder, 114-122. Ratnieks, F. L. W. & Anderson, C. (1999). Task partitioning in insect societies. Insectes Sociaux, 46, 95-108. Ratnieks, F. L. W. & Miller, D. G. (1993). Two polygyne overwintered nests of Vespula vulgaris from California. Psyche, 100, 43-50. Rau, P. (1928). The honey-gathering habits of Polistes wasps. Biological Bulletin, 54, 503519. Raven, P. H., Evert, R. F. & Eichhorn, S. E. (1999). Biology of Plants, 6th edn. W. H. Freeman and Company, New York.

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Reed, H. C. & Akre, R. D. (1983). Nesting biology of a forest yellowjacket Vespula acadica (Sladen) (Hymenoptera: Vespidae), in the Pacific Northwest Annals of the Entomological Society of America, 76, 582-590. Reyes-Lopez, J. L. & Fernandez-Haeger, J. (2001). Some factors determining size-matching in the harvester ant Messor barbarus: food type, transfer activity, recruitment rate and size-range. Insectes Sociaux, 48, 118. Richards, O. W. (1978). The Social Wasps of the Americas, Excluding the Vespinae. British Museaum (Natural History), London. Rissing, W. S. (1984). Replete caste production and allometry in the honey ant, Myrmecocystus mexicanus. Journal of the Kansas Entomological Society, 57, 347-350. Robinson, E. J. H., Richardson, T. O., Sendova-Franks, A. B., Feinerman, O. & Franks, N. (2009). Radio tagging reveals the roles of corpulence, experience and social information in ant decision making Behavioral Ecology and Sociobiology, 63, 627-636. Rosell, R. C. & Wheeler, D. E. (1995). Storage function and ultrastructure of the adult fatbody in workers of the ant Camponotus festinatus (Buckley) (Hymenoptera, Formicidae). International Journal of Insect Morphology & Embryology, 24, 413-426. Ross, K. & Matthews, R. W. (1982). Two polygynous overwintered Vespula squamosa colonies from the Southwestern U.S. (Hymenoptera: Vespidae). Florida Entomologist, 65, 176-185. Ross, K. G. & Visscher, P. K. (1983). Reproductive plasticity in yellowjacket wasps: a polygynous, perennial colony of Vespula maculifrons. Psyche, 90, 179-192. Roubik, D. W. (1989). Ecology and natural history of tropical bees. Cambridge University Press, New York. Rueppell, O. & Kirkman, R. W. (2005). Extraordinary starvation resistance in Temnothorax rugatulus (Hymenoptera, Formicidae) colonies: Demography and adaptive behavior. Insectes Sociaux, 52, 282. Salzelemann, A., Nagnan, P., Tellier, F. & Jaffe, K. (1992). Leaf-cutting ant Atta laevigata (Formicidae:Attini) marks its territory with colony-specific dufour gland secretion. Journal of Chemical Ecology, 18, 183-196. Savoyard, J. L., Gamboa, G. J., Cummings, D. L. D. & Foster, R. L. (1998). The communicative meaning of body oscillations in the social wasp, Polistes fuscatus (Hymenoptera, Vespidae). Insectes Sociaux, 45, 215-230. Schorkopf, D. L., Hrncir, M., Mateus, S., Zucchi, R., Schmidt, V. M. & Barth, F. G. (2009). Mandibular gland secretions of meliponine worker bees: further evidence for their role in interspecific and intraspecific defense and aggression and against their role in food source signalling. Journal of Experimental Biology, 212, 1153-1162. Schwarz, H. F. (1929). Honey wasps. Natural History, 29, 421-426. Seeley, T. D. (1987). The effectiveness of information collection about food sources by honey bee colonies. Animal Behavior: Seeley, T. D. 35, 1572-1575. Seeley, T. D. (1993). The tremble dance of the honey bee: message and meaning. Behavioral Ecology and Sociobiology. Seeley, T. D. (1995). The Wisdom of the Hive. Harvard University press, Mass. Seeley, T. D. (1997). Honey bee colonies are group-level adaptive units. American Naturalist, 150, S22-S41.

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Seeley, T. D., Kuhnholz, S. & Weidenmuller, A. (1996). The honey bee's tremble dance stimulates additional bees to function as nectar receivers. Behavioral Ecology and Sociobiology, 39, 419-427. Sempo, G. & Detrain, C. (2004). Between-species differences of behavioural repertoire of castes in the ant genus Pheidole: a methodological artefact? Insectes Sociaux, 51, 48. Serrão, J. E., Cruz Landim, C. D. & Silva De Moraes, R. L. M. (1997). Morphological and biochemical analysis of the stored and larval food of an obligate necrophagous bee, Trigona hypogea. Insectes Sociaux, 44, 337-344. Sherman, G. & Visscher, P. K. (2002). Honeybee colonies achieve fitness through dancing. Nature, 419, 920-922. Simon, R. P. & Benton, A. W. (1968). Winter activities of Vespula maculifrons. Annals of the Entomological Society of America, 61, 542-544. Slaa, E. J., van Nieuwstadt, M. G. L., Pisa, L. W. & Sommeijer, M. J. (1997). Foraging strategies of stingless bees (Apidae, Meliponinae): the relation between precision of recruitment, competition and communication. Acta Horticulturae, 437, 193-197. Smith, C. R. (2007). Energy use and allocation in the Florida harvester ant, Pogonomyrmex badius: are stored seeds a buffer?. Behavioral Ecology and Sociobiology, 61, 1479-1487. Smith, C. R. & Suarez, A. V. (2010). The trophic ecology of castes in harvester ant colonies. Functional Ecology, 24, 122-130. Sorensen, A. A., M., B. T. & Vinson, S. B. (1985). Trophallaxis by temporal subcastes in the fire ant, Solenopsis invicta, in response to honey. Physiological Entomology, 10, 105111. Sorenson, A. A., Busch, T. M. & Vinson, S. B. (1985). Control of food influx by temporal subcastes in the fire ant Solenopsis invicta. Behavioral Ecology and Sociobiology, 17, 191-198. Spradbery, J. P. (1971). Seasonal changes in the population structure of wasp colonies (Hymenoptera: Vespidae) Journal of Animal Ecology, 40, 501-523. Starr, C. K. & Jacobson, R. S. (1990). Nest structure in Philippine hornets (Hymenoptera, Vespida, Vespa spp.). Japanese Journal of Entomology, 58, 125-143. Stradling, D. J. (1987). Nutritional ecology of ants. In: Slansky Jr., F.,Rodriguez, J. G., (eds) Nutritional Ecology of Insects, Mites, Spiders and Related Invertebrates. John Wiley and Sons, New York, 927-970. Strassmann, J. E. (1979). Honey caches help female paper wasps (Polistes annularis) survive Texas winters. Science, 204, 207-209. Sugen, E. A. & McAllen, R. L. (1994). Observations of foraging, population and nest biology of the Mexican honey wasp, Brachy mellifera (Say) in Texas [Vespidae: Polybiinae]. Journal of the Kansas Entomological Society, 67, 141-155. Telleria, M. C. (1996). Plant resources foraged by Polybia scutellaris (Hym. Vespidae) in the Argentine Pampas. Grana, 35, 302-307. Toth, A. L., Bilof, K. B. J., Henshaw, M. T., Hunt, J. H. & Robinson, G. E. (2009). Lipid stores, ovary development, and brain gene expression in Polistes metricus females. Insectes Sociaux, 56, 77-84. Traniello, J. F. A. (1987). Comparative foraging ecology of north temperate ants: the role of worker size and cooperative foraging in prey selection. Insectes Sociaux, 34, 118-130.

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Traniello, J. F. A. (1989). Foraging strategies of ants. Annual Review of Entomology, 34, 191210. Tschinkel, W. R. (1993). Sociometry and sociogenesis of colonies of the fire ant Solenopsis invicta during one annual cycle. Ecological Monographs, 63, 425-457. Tschinkel, W. R. (1998). Sociometry and sociogenesis of colonies of the harvester ant, Pogonomyrmex badius: worker characteristics in relation to colony size and season. Insectes Sociaux, 45, 385-410. Valentim, C. L., Mota-Andrade, J. V., Teixeira, M. C. & Schoereder, J. H. (2007). Do Atta robusta (Hymenoptera : Formicidae) ants prefer small seeds? Sociobiology, 50, 10511057. von Frisch, K. (1967). The Dance Language and Orientation of Bees. Harvard University Press, Cambridge. West-Eberhard, M. J. (1982). The nature and evolution of swarming in tropical social wasps (Vespidae, Polistinae, Polybiini). In: P. Jaisson, (ed). Social Insects in the tropics. University of Paris XIII Press, Paris, 98-128. Wheeler, D. E. (1994). Nourishment in ants: patterns in individuals and societies. In: J. H., Hunt, J. H. & C. A. Nalepa, (eds) Nourishment and Evolution in Insect Societies. Westview Press, Boulder, 245-278. Wheeler, D. E. & Martinez, T. (1995). Storage proteins in ants (Hymenoptera, Formicidae). Comparative Biochemistry and Physiology B-Biochemistry & Molecular Biology, 112, 15-19. Wheeler, W. M. (1908). Vestigal instincts in insects and other animals. American Journal of Psychology, 29, 1-13. Whitford, W. G. (1978). Foraging by seed-harvesting ants. In: M. V. Brian, (ed). Production Ecology of Ants and Termites. Cambridge University Press, Cambridge, 107-110. Wilson, E. O. (1971). The Insect Societies. The Belknap Press of Harvard University Press, Cambridge, Mass. Wilson, E. O. (1974). The soldier of the ant Camponotus (Colobopsis) fraxinicola as a trophic caste. Psyche, 81, 182-188. Wilson, E. O. (2003). Pheidole in the New World. Harvard University Press, Cambridge, Massachusetts. Winston, M. L. & Michner, C. D. (1977). Dual origin of highly social behavior among bees. Proceedings of the National Acadamy of Science USA, 74, 1135-1137.

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Chapter 2

ISSUES IN THE STUDY OF PROBOSCIS CONDITIONING Charles I. Abramson1*, Michel B.C. Sokolowski2 and Harrington Wells3 1

Oklahoma State University Department of Psychology 116 N. Murray Stillwater, Oklahoma 74078 2 Université de Picardie – Jules Verne Department of Psychology Chemin du Thil 80025 Amiens Cedex 1, France 3 University of Tulsa Department of Biology 304 Oliphant Hall Tulsa, Oklahoma, 74104

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ABSTRACT This paper discusses methodological and theoretical issues associated with proboscis conditioning in neglected pollinators. The case is made that the search for learning curves similar to those found in Apis is hindered by several factors. Among the most important are the lack of consensus on what is classical conditioning and the allure of cognitive explanations of insect learning. However, the problem is fueled by few examples of individual data and no generally accepted learning taxonomies. On the other hand, the possibility exists that the proboscis learning found in Apis is overestimated as a research methodology. The few cases of proboscis conditioning in pollinators not generally used as model systems (not Apis or Bombus) are compared to results found with honey bees. Although generally conditioning is less complete, what is also noteworthy is the spotty occurrence of cases across the pollinator taxa. Finally, suggestions are provided on the type of training variables that should be manipulated in an effort to find learning curves similar to those found in Apis.

Keywords: Proboscis conditioning, classical conditioning, learning, pollinators *

Corresponding author: Charles I. Abramson, Oklahoma State University, Laboratory of Comparative Psychology and Behavioral Biology, Departments of Psychology and Zoology, 116 N. Murray, Stillwater, OK 74078, USA. Email: [email protected]. This project was supported in part by grants from the National Science Foundation (OISE – 1043057 and DBI – 0851651).

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Charles I. Abramson, Michel B. C. Sokolowski and Harrington Wells

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1. INTRODUCTION The honey bee (Apis mellifera) proboscis conditioning technique has provided a wealth of information not only about learning (Mallon, et al., 2003) but also as a bioassay for the effect of agrochemicals (Abramson et al., 1999), questions about repellants and attractants and about food preferences (Abramson et al., 2010). Indeed, what makes this methodology such an ideal system for agricultural questions is the rapid rate at which learning occurs and the high acquisition levels obtained (Abramson, 1994). Apis and Bombus have a long record as being used as model systems for making generalizations about the learning, behavior and physiology of bees and even of insect pollinators. Their importance as managed pollinator systems has been a key component to the green revolution in terms of crop productivity. Relying on a single or even a limited number of pollinator species presents risks, as we have experience recently with Apis mellifera and emergent diseases such as Varroa and CCD. However, only about 10 of approximately 20,000 pollinator bee species are managed for agricultural purposes (Cane, 1997), and so great potential exists to develop new pollinator systems using bees alone (Bosch and Kemp, 2002). We have little knowledge of whether our Apis and Bombus system are really applicable as models of what to expect if using other pollinators. The proboscis conditioning method offers itself as a potential comparative measure for similarities and differences among pollinator species. This paper addresses the difficulty of researchers working with insect pollinators not typically used as model systems to find learning curves generated from the classical conditioning of proboscis extension to approximate those found with honey bees from the genus Apis. It is not uncommon, for example, for learning curves in Apis mellifera to reach almost 100% while for some stingless bees no evidence for proboscis conditioning can be found. The question is why. We would like to note at the outset that there are very few studies on proboscis conditioning in pollinators other than honey bees. We will therefore focus our paper on theoretical and methodological issues in the hope and expectation that it will serve as a guide for future research. The proboscis extension reflex is most easily studied by placing the test insect in some type of restraining device. With Apis mellifera, this can be accomplished by taping them into small metal tubes (Abramson, 1990). Once harnessed, the insect, in theory, readily extend its mouthparts (proboscises) to feed on a sucrose solution (the unconditioned stimulus or US) after the solution has been briefly applied to the antennae, on which sucrose sensitive contact sensillae are found (Minnich, 1932). One or more forward pairings of an odor (the conditioned stimulus or CS) with sucrose feeding increases the frequency of background emissions of proboscis extension to odor. The proboscis extension technique was first described by Frings (1944) and refined over the years by Kuwabara (1957), Takeda (1961), and Bitterman, Menzel, Fietz and Schäfer (1983). Automated versions of the technique are available from Vareschi (1971) and Abramson and Boyd (2001). The virtue of the proboscis extension reflex technique is that the experimenter can control training variables known to influence learning. For example, both conditioned stimulus and unconditioned stimulus durations, as well as interstimulus interval, and intertrial interval, can be precisely controlled with harnessed subjects. This type of variable control is not obtainable with free flying test subjects, even when using artificial flower patches to control the test

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situation (Sanderson et al., 2006; Cakmak et al., 2009). The proboscis extension reflex technique is also better suited for quantitative physiological and biochemical analysis (Menzel, et al., 1991) and it should be noted that the development of the proboscis extension reflex technique permits a comparison between the learning ability of Apis under both ―natural‖ (i.e., unharnessed) and ―unnatural‖ (i.e., harnessed) situations (Menzel and Bitterman, 1983). A further advantage is that subspecies differences in classical conditioning can be examined under very repeatable conditions in any location (Abramson et al., 1997; Abramson et al., 2008). The study of the proboscis extension reflex has led to many areas of fruitful research in Apis mellifera (Menzel and Bitterman, 1983; Kartzev, 1996). Of course, this includes mechanisms of learning such as studies of Pavlovian conditioning (Bitterman et al., 1983; Mercer, 1987; Batson et al., 1992; Buckbee and Abramson, 1997), and discriminative punishment (Smith et al., 1991). However, it has also been an important tool for environmental studies, which include the influence of pesticides on learning (Taylor et al., 1987; Mamood and Waller, 1990; Stone et al., 1997) olfactory discrimination for application of insect attractants and repellants (Getz and Smith, 1987; Smith and Menzel, 1989), and as a rapid bioassay to measure detection of adulterated beeswax (Aquino et al., 1999). Additionally, the proboscis extension reflex has served as a model system for studying the biochemistry of learning and other forms of behavior modification in Apis (Mercer, 1987; Abramson, 1994; Menzel and Muller, 1996).

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II. THEORETICAL ISSUES Before embarking on a research program using the proboscis extension reflex in nonstandard pollinators we believe that it is important for the researcher to be aware of, and frankly to confront, several issues. We may not be voicing the popular opinion but nevertheless we believe we are correct in that these issues must be discussed and considered by researchers interested in extending the proboscis conditioning paradigm to pollinators not typically used as model insect systems.

A. Inconsistencies in Definitions Within the area of learning theory, there is the lack of consensus among researchers as to the definitions of many learning phenomena. We will illustrate this with examples from classical conditioning, and operant conditioning.

A.1. Classical conditioning The definition of classical conditioning is not always consistent among behavioral scientists. It is important to recognize this lack of consistency when evaluating studies of classical conditioning and in designing experiments using proboscis conditioning. Some psychologists, for example, stress that classical conditioning is the learning of relationships between cognitive events and that conditioning cannot be defined in terms of behavioral

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change. Other psychologists stress that it may be incorrect to lump together various disparate procedures under the general category of classical conditioning. Within these extremes are definitions which stress that 1) prior to training the CS must not elicit the response that is to be conditioned, 2) the contingency between the CS and US, or 3) classical conditioning is a procedure for creating a new reflex. On the other hand, a zoologist or entomologist might consider classical conditioning to be the pairing of a ―search image‖ with a sign stimulus or innate motor program. For the physiologist, the CS can be considered solely in terms of the electrical stimulation of afferent fibers. In considering the problem of what is classical conditioning, Gormezano and Kehoe (1975) and Gormezano, Kehoe and Marshall (1983) describe four variations of classical conditioning based on the nature of the conditioned response (CR): 1) Conditioned StimulusConditioned Response (CS-CR), 2) Conditioned Stimulus-Instrumental response (CS-IR), 3) Instrumental Approach Behavior, and 4) Autoshaping. 1. The Conditioned Stimulus-Conditioned Response paradigm is considered to represent the ―pure‖ case of classical conditioning. Here the CS does not elicit the unconditioned response (UR) prior to training, and the CR emerges from the same effector system as the UR. For example, consider a hypothetical experiment in which a novel odor (e.g., peppermint) is presented to a stingless honey bee such as Melipona scutellaris and sucrose (US) is injected into the preoral cavity which elicits a feeding response (UR). Initially, the novel CS odor does not elicit a response. However, following several pairings of odor and sucrose, the bee will extend its feeding apparatus when the odor is introduced (CR) and before the US is presented. 2. The Conditioned Stimulus-Instrumental Response paradigm contains those experimental designs commonly known as ―transfer of control: or classicalinstrumental transfer‖ in which classical conditioning is assessed not directly but by its influence on instrumental or operant responding. Perhaps the most well known example of this design is the conditioned suppression procedure in which a CS is paired, for example, with electric shock (US) and the ability of the CS to suppress on-going behavior is assessed. Consider a harnessed M. scutellaris that receives a novel odor (CS) paired with the presentation of shock 20 times in succession. An hour or so after the pairings the bee is permitted to feed from a large droplet of sucrose. While the bee is drinking, the experimenter presents the odor previously paired with shock. The question of interest is whether the proboscis retracts. If it does, conditioned suppression (assuming appropriate controls) is demonstrated 3. The Instrumental Approach Design is a type of conditioning in which some ―approach behavior‖ to a CS is necessary to receive the US. This procedure is illustrated by general activity to stimuli preceding food (i.e., conditioning of general activity) and either an instrumental runway or maze situation in which movement toward the food source is necessary. For example, a food deprived M. scutellaris might be confined to a small cage and illumination is paired with a feeding. Over several pairings the light begins to increase the general activity of the bee as the bee learns to associate food with light. 4. Autoshaping is related to the CS-CR paradigm with the interesting property that the CR is not from an effector system related to the US. Autoshaping has never been attempted with invertebrates in part because the necessary apparatus has not been

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developed. In the original experiment pigeons learned to approach and peck a colored disk which was turned on before a food hopper was exposed (Brown and Jenkins, 1968). It is important to note that all four categories differ in many ways, including how the CR is measured, the accuracy with which the CS and US are presented in a response independent fashion, the nature of the target response, the amount of control the experimenter has over the training variables, and the degree to which the animal is restrained in the conditioning situation. In their discussions of classical conditioning, Gormezano and Kehoe (1975) consider the CS-CR paradigm the only unambiguous case of classical conditioning. It is no accident that much of what we know about the physiology and biochemistry of the classical conditioning of vertebrates comes from the rabbit nictitating membrane preparation, which uses the CS-CR paradigm (Gormezano, 1984; Byrne, 2003). The proboscis conditioning paradigm is not technically a CS-CR paradigm because the animal must stick out its proboscis to obtain the US. The proboscis is elicited by touching the antenna with sucrose. Thus the delivery of the US is not technically response independent – if the honey bee does not extend its proboscis the experimenter cannot delivery the sucrose. There is the further complication that the proboscis conditioning procedure can be considered a case where two unconditioned stimuli are paired. The first is antennae stimulation and the second is sucrose delivery to the now extended proboscis. It is worth noting that Gormezano and Kehoe (1975) do not consider alpha conditioning an example of classical conditioning. Alpha conditioning is a ―conditioned stimulus‖ that already, prior to training, elicits a small version of the conditioned response. For example, a weak sucrose solution might elicit a partial proboscis extension and a strong sucrose solution a full extension. By pairing the weak solution with the strong solution, a strong proboscis extension will come to be elicited by the weak solution. The issue of alpha conditioning versus classical conditioning was one of the factors that hindered the development of planarians as a model system in the 1960s (Nicolas et al., 2008). We would urge researchers interested in developing a proboscis conditioning procedure with non-standard social pollinators to look at some of the early planarian conditioning studies (Nicolas et al., 2008). It is important to realize that the reverse argument has also been made with respect to alpha conditioning. That is, alpha conditioning might actually represent instrumental conditioning (Razran, 1971). For example, consider a situation in which an experimenter presents a stingless honey bee with a chemosensory CS which elicits, prior to its pairing with a US, a partial proboscis extension. The US is a second chemosensory stimulus that elicits a large proboscis extension, and over the course of several CS-US pairings, the strength of the proboscis extension increases. Is this classical conditioning? Unfortunately, many behavior scientists would say yes. The results can just as easily be interpreted as a case of instrumental conditioning where a response to a stimulus is strengthened by a reward (the second chemosensory stimulus).

A.2. Instrumental and operant conditioning Instrumental and operant behaviors are examples of associative learning in which the behavior of the subject is controlled by the consequences of behavior. Instrumental and

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operant behavior can be explored using a variety of apparatus, which include devises such as running wheels, runways, mazes, shuttle boxes, and lever-press situations. Instrumental and operant behaviors are generally thought to be more complex than classical conditioning. One might roughly characterize the difference by saying that classical conditioning describes how associations between stimuli are made, and instrumental and operant conditioning describe how stimuli are associated with our own motor actions. Classically conditioned behavior emphasizes sensory integration, and instrumental and operant behavior emphasizes motivation: new behaviors are learned in order to obtain or avoid some stimulus. In addition, instrumental and operant behaviors are thought to be more complex than classical conditioning because learning depends on the subjects own behavior and usually requires an obviously new behavior. Despite these differences, instrumental and operant behavior share many properties with classically conditioned behavior. These include extinction, spontaneous recovery, generalization and discrimination. As originally conceived, operant behavior is characterized by the ―goal-directed‖ motor manipulation of the environment (Lee, 1988). In place of the goal-directed modification of behavior that was the hallmark of the Skinnerian system, operant conditioning now generally consists of any behavior sensitive to response-reinforcer contingencies. Thus, operant conditioning is now considered to include such procedures that modifying body position, running against a taxic or kinetic preference, and learning various mazes and runways (Brembs, 2003). We believe that instrumental behavior is not as complex as operant behavior and that there should be a distinction between the two. It is important to note that instrumental conditioning procedures may not constitute operant behavior. A major requirement of operant conditioning has been that species-typical behavior is minimized by interjecting a ―novel‖ behavior such as a lever press, or a non-arbitrary response brought under the control of a discriminative stimulus (or cue) placed between the animal and the animal‘s reception of some consequence. In this way, the experimenter demonstrates that the animal has learned not only how to operate some device but also ―how to use it.‖ One might be more confident that honey bees are engaging in operant behavior if it can be shown that: 1) the operant responses minimize species-typical behavior, 2) some property of the response class such as its rate, force, or interresponse time can be modified, 3) the response no longer occurs when such responses postpone the delivery of reward, and 4) the response can be brought under the control of a cue (i.e., discriminative stimulus). These experiments have not been performed. For example, it would be a simple matter to determine if a honey bee can adjust its running speed (move faster or slower) to obtain some reward.

B. Taxonomies of Learning In addition to problems with the definitions of learning phenomena, it is important to be aware that there is no generally accepted taxonomy of learning. The importance of taxonomy and the relationships among entities is well known. An understanding of the relationships among the different types of learning is equally important. It is interesting to note how much effort was devoted to learning taxonomies in the early learning literature and how discussions of learning taxonomies have all but disappeared from the contemporary learning literature. As

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Bitterman (1962) noted almost 50 years ago, ―Classification is not merely a matter of taste‖ (p. 81). Tulving (1985) describes six ways in which a classification scheme can advance the field of learning. These include providing theoretical structure to the design and analysis of experiments, replacing general categories such as classical and operant conditioning with detailed descriptions of the procedures, and novel procedures and results can be described easily in terms of the amount of deviation from specified categories. For example, it was suggested that those interested in conducting learning research with rattlesnakes (Abramson and Place, 2008) attempt to link their procedure with one of the classification schemes. Several taxonomies have been proposed. Important learning taxonomies to consider are those by Dyal and Corning (1973) and by Gormezano and Kehoe (1975) for classical conditioning, and the model by Woods (1974) for instrumental and operant conditioning. Woods‘ (1974) classification of instrumental conditioning identifies 16 categories of conditioning based on the presence or absence of a discriminative stimulus and the desirability of the reward.

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C. The Reporting of Individual Data Most studies of proboscis conditioning involve the reporting of group data. Many examples can be found in the honey bee literature. Data are often presented as the percentage or proportion of animals responding on each trial. In other cases, data are presented as group means or some learning score. Group data do not give the shape of individual learning curves nor information about the variation among animals (Stepanov and Abramson, 2008). Moreover, the number of bees discarded from a test population is rarely reported. Without such data, it is difficult to know how many animals from a given population do indeed learn. Thus, the reliance on group data could lead to statements about species characteristics that are not reliable or valid (Hirsch and Holliday, 1988). In the case of proboscis conditioning in honey bees, the lack of individual data is particularly striking. It has been shown that behavioral plasticity in honey bees is directly linked to sucrose responsiveness (Scheiner et al., 2004). Some honey bees may achieve high learning scores and others may not. This kind of behavioral variability may be reported only if the researcher looks at individual behavior. We believe that the analysis of individual learning curves is one of the most important methodological contributions of animal behaviorists and remains completely unknown for most contemporary behavioral biologists (Sidman, 1960, Smith et al., 1991). As learning is defined as a change in individual behavior, learning must be detected and verified at the individual level. Single case experimental designs are especially adapted for that purpose. Inter-individual variation and how learning traits are distributed in a population are questions worthy of investigation. Even here, however, the analysis of individual behavioral traits should not be overlooked.

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D. Cognitive Explanations of Learning The trend toward interpreting proboscis conditioning in terms of ―representations,‖ ―cognitions,‖ and other language borrowed from the vocabulary of human information processing may be unwarranted when applied to the learning of insects. It is often overlooked that there are still challenges to the use of cognitive concepts when applied to the learning of vertebrates (Amsel, 1989; Skinner, 1989). Even if it is concluded that some vertebrate species possess cognitive structures, the necessary experiments and replications by independent laboratories have not been performed to determine whether proboscis conditioning also posses such structures. The procedures of blocking, overshadowing, and successive negative contrast, for example, can all be explained without recourse to cognitive constructs (Couvillon and Bitterman, 1984). The same is also true with avoidance learning (Abramson et al., 1988). We view the rise in cognitive interpretations of insect behavior as alarming. Students are no longer being trained in traditional learning methodology and few professors have a grasp of the early learning literature. As surprising as this may sound, the word ―Behavior‖ has all but disappeared from glossaries of introductory textbooks in zoology, biology and psychology (Abramson and Place, 2005). We view the absence of the word behavior as problematic given the rise of cognitive approaches to the study of behavior (Amsel, 1989). Traditional behavioral issues are being tossed aside and all but forgotten by a new generation of students (Abramson, 1994; 1997). It is not a fantasy to imagine that in the future, discussions of such important areas as classical and operant behavior will be distorted and subsequently forgotten. Such a trend has already been documented in the case of texts used for advanced courses in the psychology of learning (Coleman, et al., 2000). Similarly, Sheldon (2002) has shown the inconsistency with which current introductory psychology texts portray operant conditioning and associated issues. Before one can conclude that pollinators not typically used as model insect system have ―cognitive‖ processes, a significant database must be developed based on traditional learning procedures. Clearly, the few invertebrate studies that are available do not warrant any cognitive interpretation. The necessary database is not there. We are not voicing the popular opinion but to argue cognition in an invertebrate because a phenomena appears in a vertebrate can equally suggest that the phenomena in the vertebrate is not based on cognition precisely because it is found in an invertebrate. In the previous section, we have summarized the general issues that a researcher must consider before conducting a proboscis conditioning experiment. These issues include lack of consistency in behavioral definitions, no generally accepted taxonomy of learning, the lack of data on individual organisms, and at least some amount of skepticism in the application of cognitive explanations when applied to insects. We also touched briefly upon the lack of training of students in traditional areas of learning. We will now discuss some of the methodological factors that should be considered in studying proboscis conditioning in neglected social pollinators.

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III. METHODOLOGICAL ISSUES The first author, who has over 20 years experience conducting proboscis conditioning experiments, had the opportunity to conduct a proboscis conditioning experiment with the Uruçu (Melipona scutellaris) honey bee in Brazil, which is a stingless bee. Much to his surprise no effect of conditioning was found despite a robust unconditioned response to honey obtained from the same colony. Also, surprising was that a high molarity sucrose solution was ineffective in eliciting proboscis extension Abramson et al., 1999c). The odors of hexanal, geraniol, citral, Uruçu wax and Africanized honey bee wax were tried as conditioned stimuli, as was stimulation of the antenna with water. In each case, after 12 training trials few conditioned responses were noted. It was quite a surprise! Similar results occur when testing the stingless bee Scaptotrigona depilis (McCabe et al., 2007). They did not show proboscis conditioning. Nevertheless, success in proboscis conditioning has been reported in some species of stingless bees, albeit the degree to which learning occurred was only about half of that of honey bees (McCabe et al., 2007). Although not a neglected insect model, it is worth noting that bumble bees (Bombus terrestris) also show proboscis conditioning; they however require more motivation and longer trial exposure to the CS than do honey bees (Laloi et al., 1999), which agrees with relative ease and success of recruiting bumble bees to feeders from colonies using odors (Wenner and Wells, 1991; Renner and Nieh, 2008). The differences among bee species in proboscis conditioning highlight the need for work with neglected pollinator species as well as the well established model system. Although, we would like to make generalizations about behavioral responses across pollinators using model systems such as honey bees and bumble bees, basic differences appear to exist even when dealing with colonial bee species that are pollinators. An important case in point was shown to be when honey bees base flower choice not on reward quality, but rather on flower color alone (Hill et al., 1997; Hill et al., 2001), which is not observed in bumble bees (Gegear and Laverty, 2004). The few proboscis conditioning studies that have utilized Lepidoptera present a picture that is similar to that observed with hymenopter pollinators. The moths Heliothis virescens (Hartlieb, 1996; Skiri et al., 2005) and Spodoptera littoralis (Fan et al., 1997), as well as the butterfly Agraulis vanillae (Kroutov et al., 1999), all demonstrate proboscis conditioning to various conditioned stimuli. However, the moth Manduca sexta does not show proboscis conditioning. Nevertheless, similar learning can be shown using activation of the cibarial pump using electromylographic activity in M. sexta (Daly and Smith, 2000; Daly et al., 2001). This has in turn been used to look at olfactory learning with respect to neural representation (Ito et al., 2008). Whether results are directly comparable in terms of learning rate or frequency of acquired response is yet to be determined. At least one non-pollinator insect also demonstrate proboscis conditioning. Drosophila melanogaster under harnessed conditions will also show classical conditioning through proboscis extension to CS odors (Chabaud et al., 2006) The importance here is that the process is potentially applicable to a very wide range of insects (not just pollinators), but as noted above is not seen even in all pollinator species, even all social pollinator species. On the other hand, feeding in many insects is not related to proboscis extension and so proboscis conditioning is not a valid approach to learning. Clearly these insects can learn, as seen in

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Charles I. Abramson, Michel B. C. Sokolowski and Harrington Wells

parasitoid wasps (Wackers et al., 2002; Bleeker et al., 2006), but direct comparison of learning to those insects using proboscis conditioning may never be possible. In reviewing the literature on proboscis conditioning of honey bees (Apis) in comparison with pollinators not typically used as model animal system it is readily apparent the proboscis conditioning technique is not as effective. The learning curves for moths and butterflies are all significantly lower than that for Apis (Hartlieb, 1996; Fan et al., 1997; Kroutov et al., 1999; Skiri et al., 2005). The learning curves for ants are also lower (Sauer et al., 2002). The question is why. Some possible answers for this discrepancy in performance is that non-Apis social pollinators are less ―intelligent‖ than Apis and/or less sensitive to environmental contingencies. We readily discount this possibility because of the work done under field conditions suggest that these animals are able to successfully solve problems. The basic reward and cost variables associated with nectar foraging are rapidly learned by nectivores (Wells and Wells, 1986; Laverty and Plowright, 1988; Laverty, 1994; Chittka and Thomson, 1997). Further, floral landscapes are complex not only because of the diversity of flowers with different handling problems and rewards but also due to the changes in rewards offered by alternative flowers over the course of a day (Percival, 1965; Heinrich, 1979). Both bumble bees and honey bees foragers have been shown to rapidly assess reward difference and respond accordingly (Hill et al., 2001; Wells et al., 1992; Chittka, 1998; Chittka et al., 2003). Rapid learning of floral cues associated with specific rewards allows bees to track the changes in rewards, thus improving foraging efficiency by allowing individuals to preferentially visit the current most profitable flower type (Raine et al., 2006a; Raine et al., 2006b). In fact, honey bees and bumble bees appear to handle increased complexity in much the same way (Raine and Chittka, 2007; Cakmak et al., 2009). We also discount this possibility because parametric manipulations of training variables known to influence conditioning have yet to be performed. A similar situation occurred when we searched for classical conditioning in the triatomine Rhodnius prolixus, which is the major vector of Chagas disease in Venezuela. Although classical conditioning was not found, headway was made in identifying several training variables that can be manipulated in future experiments (Abramson et al., 2005). A second possibility is that the data obtained with Apis is not altogether accurate and frankly overestimates conditioning in this species. We would like to say explicitly that the proboscis conditioning in Apis does have problems and that it is often not as easy to condition animals as the literature suggests. It is well known, yet unpublished, in the conditioning community that many laboratories have difficulty conditioning their honey bees (personal experience: the first author is often contacted by researchers who have difficulty conditioning bees). Even if conditioning is readily demonstrated on one day, the next day no bees may condition. In an earlier experiment using a discriminative punishment paradigm we showed that the learning curve of Apis is made up of several different response patterns (Smith et al., 1991) Moreover, if one carefully reviews the literature on proboscis conditioning in Apis or Bombus it is apparent that the techniques differ widely. For example, some laboratories overlap the CS and US, and others use a trace conditioning procedure in which the CS is turned on then off before the US is presented. Similarly, differences exist in experiments with respect to intertrial times: some use a 10 minute intertrial interval, others use 5 minutes. Motivation of test individuals also widely varies, with some depriving their animals of food

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Issues in the Study of Proboscis Conditioning

35

for approximately 24 hours prior to the experiment while others use animals just several hours after harnessing. Materials and methods vary among laboratories as well. Some harness their animals in plastic tubes, metal tubes, or capsules (Toda et al., 2009). Some laboratories leave the head of the honey bee free to move while others restrain the head with metallic pins (Riveros and Gronenberg, 2009). Perhaps most importantly is that some laboratories directly stimulate the mouthparts to elicit proboscis extension and other first touch the antennae and then feed the now extended proboscis. In regards to the latter procedure it might be surprising for some researchers to learn that there is a build-up of sucrose on the antennae which is easily seen with a magnifying glass. One would expect that such a build-up would sensitize the animal and lead to artificially higher learning curves. Perhaps the most unfortunate procedural variation is that some experiments discard data from bees that respond to the CS on the first training trial and before the US is presented. It should be recognized that some of the remaining sample, although not responding on the first trial, might respond on the second, not because of learning but because of sensitization. There are few studies which employ CS only controls which would assess such a possibility. In our view it is not enough to rely on control data generated from a different laboratory. Moreover, bees that do not learn at all are sometimes discarded. In our work with Africanized honey bees the learning curves are typically lower than that found with European honey bees and in some cases approximate those found with moths and butterflies. We do not discard data from any subject. In interpreting these data we have a problem because we do not know whether it is a subspecies effect or actually represents the true learning curve of Apis. Therefore, the lower learning curves seen in moths and butterflies, for example, may actually represented the more accurate level of conditioning (Abramson et al., 1997; Abramson et al., 1999c). Further complicating the issue of obtaining an accurate measure of learning in proboscis conditioning is the fact that tactile and olfactory learning has been showed to be linked to sucrose responsiveness. Pollen foraging bees learn faster than nectar foraging bees, but these differences disappear if we compare two groups of nectar and pollen foragers with similar response thresholds (Scheiner et al., 1999). Moreover, sucrose responsiveness varies among bees (Mujagic and Erber (2009) and influences proboscis conditioning (Scheiner et al., 2003). Variations in learning curves in proboscis conditioning may result from uncontrolled, unknown, or low sucrose responsiveness in observed bees. We believe that it is important for researchers to direct some attention to the development of a proboscis conditioning technique for pollinators not typically used as model insect systems. Such a technique has many potential uses, such as a bioassay for the effect of agrochemicals, the study of food preferences, and for general questions related to the comparative analysis of behavior. Table 1 shows some of the variation in proboscis conditioning techniques used with honey bees. The table is not exhaustive but designed to give the researcher an appreciation of the variation a researcher will encounter in the literature. Differences include type of conditioned stimulus, method delivering the conditioned stimulus, duration of the conditioned stimulus, whether the unconditioned stimulus is conceptualized as a reward or as an unconditioned stimulus, method of harnessing the honey bee, interval between training trials, whether a trace or overlap procedure is used when pairing the conditioned stimulus with the

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Charles I. Abramson, Michel B. C. Sokolowski and Harrington Wells

unconditioned stimulus, and the period of time between harnessing and subsequent use an experiment. Table 1. Variations in Proboscis Conditioning Methodology Delivery of CS 1. Plastic Syringe 2. Valve 3. Automated 4. Constant airflow

5. Glass Capillary 6. Glass Rod 7. Toothpick Duration of CS 1. 2 seconds 2. 4 seconds 3. 5 seconds

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4. 6 seconds 5. Double CS 4 sec and 3 sec, 1 sec overlap Type of CS 1. Cinnamon 2. Pesticide Endosulfan Decis Baytroid Sevin 3. Odor Citral Geraniol 2-hexanal nonanone 2-octanol limonene

(Abramson et al., 2007b) (Guerrieri et al., 2005) (Chandra et al., 2001) (Abramson & Boyd, 2001) (Couvillon et al., 2010) (Hussaini et al., 2007) (Chandra et al., 2000) (Smith & Cobey, 1994) (Takeda, 1961) (Chaline et al., 2005) (Giurfa & Malun, 2004)

(Abramson et al., 2006a) (Giurfa & Malun, 2004) (Chandra et al., 2001) (Chandra et al., 2000) (Couvillon et al., 2010) (Guerrieri et al., 2005) (Smith & Cobey, 1994) (Chaline et al., 2005) (Bonod et al., 2003) (Bitterman et al., 1983) (Hussaini et al., 2007)

(Abramson et al., 2006b) (Abramson et al., 2004) (Abramson et al., 1999b)

(Hussaini et al., 2007)

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Issues in the Study of Proboscis Conditioning Type of CS Odor 1-octanol 1-nonanol eugenol limonene Odor Geraniol 1-hexanol Odor Citral Hydroxycitronellal p-isopropel--ethylhydrocinnamic aldehyde Odor Citral Geraniol Hexanal Octanal 1-hexanol 1-octanol 4. Mechano-sensory 5. Absence of a stimulus 6. Hydrocarbons 7. Organic Pesticide 4% each Thyme Clove Sesame Remaining 88% Water Soybean oil Wintergreen oil Lecithin 8. Citronella 9. Sweet fennel essential oil 10. Wintergreen CS-US Interval 1. 1 second 2. 2 seconds 3. 3 seconds

4. 5 seconds

(Guerrieri et al., 2005)

(Chandra et al., 2001) (Takeda, 1961)

(Smith & Cobey, 1994)

(Giurfa & Malun, 2004) (Abramson et al., in press) (Chaline et al., 2005) (Abramson et al., 2006a)

(Abramson et al., 2006b) (Abramson et al., 2007b) (Abramson et al., 2008)

(Hussaini et al., 2007) (Guerrieri et al., 2005) (Couvillon et al., 2010) (Chaline et al., 2005) (Giurfa & Malun, 2004) (Bonod et al., 2003) (Chandra et al., 2001) (Chandra et al., 2000) (Bitterman et al., 1983) (Takeda, 1961)

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38

Charles I. Abramson, Michel B. C. Sokolowski and Harrington Wells Table 1 (Continued) Intertrial Interval 1. 6-8 minutes 2. 10 minutes 3. 12 minutes 4. 15 minutes 5. 30 minutes

(Smith & Cobey, 1994) (Abramson et al., 2006a) (Guerrieri et al., 2005) (Bitterman et al., 1983) (Hussaini et al., 2007) (Chaline et al., 2005) (Bonod et al., 2003) (Takeda, 1961) (Couvillon et al., 2010)

Type of US 1. 30% Sucrose solution 2. 20-40% Sucrose solution 3 42.8% Sucrose solution (1.25 M) 4. 50% Sucrose solution

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5. 51.3% Sucrose solution (1.5 M)

6. 61.6% Sucrose solution (1.8 M) 7. 1% ethanol, 5% ethanol, 10% ethanol, 20% ethanol in 61.6 % sucrose solution (1.8 M) 8. 3 M Sodium Chloride 9. Pesticide in 2.9 M Sucrose (99.3%) (or distilled water) endosulfan decis baytroid sevin Pesticide in 1.8 M Sucrose (61.6%) Tebufenozide Diflubenzuron Delivery of US 1. Filter Paper 2. Microsyringe 3. Toothpick 4. Glass Syringe

(Hussaini et al., 2007) (Chaline et al., 2005) (Bonod et al., 2003) (Bitterman et al., 1983) (Giurfa & Malun, 2004) (Couvillon et al., 2010) (Guerrieri et al., 2005) (Chandra et al., 2001) (Chandra et al., 2000) (Smith & Cobey, 1994) (Takeda, 1961) (Abramson et al., 2007b) (Abramson et al., 2007a) (Chandra et al., 2001) (Abramson et al., 1999b)

(Abramson et al. 2004)

(Abramson et al., 2008) (Abramson et al., 2006a) (Bitterman et al., 1983) (Couvillon et al., 2010) (Guerrieri et al., 2005) (Giurfa & Malun, 2004) (Smith & Cobey, 1994)

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Issues in the Study of Proboscis Conditioning How is US administered 1. Touch antenna and feed to extended proboscis

2. Directly into Mouth parts 3. Touch tarsi and feed to extended proboscis How is the US Conceptualized 1. As US

2. As Reward/ Reinforcement

Type of Pairing 1. Non-overlap (Trace)

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2. Overlap

(Couvillon et al., 2010) (Abramson et al., 2006b) (Guerrieri et al., 2005) (Bonod et al., 2003) (Chandra et al. 2000) (Smith & Cobey, 1994) (Bitterman et al., 1983) (Giurfa & Malun, 2004) (Abramson & Boyd, 2001) (Takeda, 1961)

(Abramson et al., 2008) (Hussaini et al., 2007) (Chaline et al., 2005) (Guerrieri et al., 2005) (Giurfa & Malun, 2004) (Bonod et al., 2003) (Chandra et al., 2001) (Bitterman et al., 1983) (Bitterman et al., 1983) (Takeda, 1961)

(Abramson et al., 2006b) (Takeda, 1961) (Hussaini et al., 2007) (Chaline et al., 2005) (Guerrieri et al., 2005) (Giurfa & Malun, 2004) (Bonod et al., 2003) (Chandra et al., 2000) (Bitterman et al., 1983)

Are Bees dropped from Data Set (Couvillon et al., 2010) (Guerrieri et al., 2005) (Bitterman et al., 1983) Capture of Animals 1. Day before

2. Same day

(Couvillon et al., 2010) (Chandra et al., 2001) (Chandra et al., 2000) (Abramson et al., 1999b) (Smith & Cobey, 1994) (Bitterman et al., 1983) (Hussaini et al., 2007) (Guerrieri et al., 2005)

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40

Charles I. Abramson, Michel B. C. Sokolowski and Harrington Wells Table 1 (Continued) Capture of Animals 2. Same day (Giurfa & Malun, 2004) (Bonod et al., 2003) (Chandra et al. 2000) (Bitterman et al., 1983) Number of Training Days 1. 1 Day 2. 2 Days How Anesthetized 1. Ice Water Bath

2. Placed in Freezer 3. Ether 4. Cold-Anesthetized

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How are they restrained 1. Metal tubes

2. Plastic tubes 3. Straws 4. Glass tubes Number of Training Trials 1. 5 2. 6 3. 8 4. 10 5. 12

(Abramson et al., 2007) (Giurfa & Malun, 2004) (Smith & Cobey, 1994) (Takeda, 1961)

(Abramson et al., 2007) (Hussaini et al., 2007) (Guerrieri et al., 2005) (Smith & Cobey, 1994) (Chandra et al., 2000) (Giurfa & Malun, 2004) (Takeda, 1961) (Bitterman et al., 1983)

(Abramson et al., 2007b) (Hussaini et al., 2007) (Guerrieri et al., 2005) (Bitterman et al., 1983) (Couvillon et al., 2010) (Chaline et al., 2005) (Bonod et al., 2003)

(Hussaini et al., 2007) (Guerrieri et al., 2005) (Giurfa & Malun, 2004) (Bitterman et al., 1983) (Smith & Cobey, 1994) (Abramson et al., 2006)

Pre-screening of Responders, Length of Time before Experiment 1. 10 minutes 2. 15 minutes 3. 30 minutes

(Hussaini et al., 2007) (Abramson et al., 2006) (Giurfa & Malun, 2004) (Chandra et al., 2001) (Chandra et al., 2000)

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Issues in the Study of Proboscis Conditioning How selected as subjects 1. Workers selected at random from the apiary 2. Only bees with vigorous proboscis extension 3. Foraging bees captured from hives 4. Bees collected immediately after emergence from combs 5. Bees collected at random from the top box of the hive 6. Bees without pollen loads 7. Bees caught at the entrance of the hive

41

Abramson et al., 1999 Abramson et al. 2006 Abramson et al. 2007b Bonod et al., 2003 Chaline et al., 2005 Couvillon et al., 2010 Bitterman et al., 1983 Giurfa & Malun, 2003 Guerrieri et al., 2005 Hussaini et al., 2007 Smith & Cobey, 1994

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A. Suggestions In this section we provide suggestions on what to manipulate in the search for a reliable proboscis extension procedure. We would urge that journals publishing insect behavior papers be prepared to publish experiments describing negative results, since a true picture will not emerge without them. Further, techniques and conditioning methods must be standardized so that learning curves can be accurately compared. Finally, we would urge the reader to consult the comparative psychological literature – especially results from the 1940, 1950s and 1960s on invertebrate and vertebrate learning. We believe that this literature is all but forgotten yet still has much to recommend it regarding issues in cognition. Many of the suggestions on how to perform proboscis extension learning experiments with invertebrates can be found in Abramson (1994). In summary they are: 1. Try a variety of conditioned stimuli and vary the intensity. 2. Try a variety of unconditioned stimuli. 3. Keep the CS-US interval as short as possible. Be prepared to use a test trial procedure if a very short ISI is used. 4. Use a relatively long intertrial interval. In our experiments we use 10 minutes. In this way the bee is not over stimulated. Our experience suggests that if a bee is over stimulated it will fail to retract its proboscis or stop extending it. A long intertrial interval will also help the animal to discriminate the olfactory CS from background odors. 5. Vary the type of pairing. If an overlap procedure is used the CS is presented, and a few moments later, the US is also presented. Both the CS and US terminate at the same time. The alternative is to use a non-overlap procedure where the CS is presented, turned off and then the US is presented. 6. If one conditioning paradigm does not work try another. The conditioned suppression paradigm is certainly underutilized. We have used it as a bioassay in addition to

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Charles I. Abramson, Michel B. C. Sokolowski and Harrington Wells proboscis extension in Apis. We would also suggest investigating pseudoconditioning. One method is to present several USs followed by a CS presentation (Abramson et al., 2006). Pseudoconditioning should not be considered merely a control in classical conditioning experiments. The behavior modifications produced by pseudoconditioning are as important as those produced by classical conditioning and deserve additional attention by behavioral scientists (Wickens and Wickens, 1942; Razran, 1971; Abramson and Buckbee, 1995). This behavior change is perfectly lawful and again underutilized. Alpha conditioning should also be tried in which a weaker stimulus is paired with a stronger stimulus. One way to conceptualize alpha conditioning is US-US conditioning. Although not generally considered a case of classical conditioning, it represents learning when appropriate controls are used such as a group receiving unpaired presentations of the two stimuli and a discrimination group in which the two weaker USs are used – only one of which is paired with the stronger US. We would also like to make the point that when using an unpaired or discrimination control the experimenter should use a pseudorandom sequence of ABBA BAAB where A may represent the CS and B the US in an unpaired control or A may represent the CS+ (paired with the US) and CS- (not paired with the US). The intertrial interval in this case should be half what is used in the paired animals. In our experiments we use a 10 minute intertrial interval for paired animals and a 5 minute intertrial interval for unpaired animals (or between the CS+ and CS- in a discrimination experiment). It is obvious that the 5 minute ITI will keep the time between CSs approximately 10 minutes. If a 10 minute ITI is used for unpaired animals the time between CS presentations is approximately 20 minutes and any difference between paired and unpaired animals may not be the result of learning but time in the apparatus. 7. Try learning paradigms other than those associated with classical conditioning. If the harnessed insect reliably elicits a proboscis extension response to the US there is no reason to restrict learning studies to classical conditioning. The instrumental technique of punishment can be used in which, for example, proboscis extension to a sucrose US is followed by shock or bitter tasting solution. Habituation can also be used in which proboscis extension elicited by a US is not followed by a feeding. Be aware, however, that this procedure may be more properly classified as punishment because under natural conditions proboscis extension once elicited is followed by food. It should also be pointed out that such simple situations such as punishment and habituation can easily be made more complex by conducting the experiment in a context. For example, proboscis extension elicited by sucrose stimulation can be punished against a background of cinnamon odor, and not when the background is a second odor. Habituation can also be studied in context. 8. Vary the method of harnessing. The results of a study measuring stress as revealed by heat shock protein HSP 70 indicate that Apis mellifera is not stressed when harnessed. It would certainly be important to know whether harnessing of neglected social pollinators induces stress. If so, the effect of stress might contribute to the lower levels of learning in proboscis conditioning studies (Hranitz et al., 2010) 9. Conduct inhibitory learning experiments. Finding low levels of excitatory conditioning does not mean no learning has occurred. It is possible that some type of

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Issues in the Study of Proboscis Conditioning

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inhibitory learning may have been acquired. In conditioning studies on Rhodnius prolixus, we were not able to find evidence of excitatory conditioning but inhibitory learning was found (Abramson et al., 2005). 10. We also suggest that researchers interested in beginning the search for successful proboscis conditioning in neglected social pollinators first use unautomated procedures. A CS odor cartridge can easily be constructed out of a 20 cc syringe and readily directed to any part of the bee during preliminary or pilot experiments. The US can be presented on strips of filter paper or microsyringe (Abramson, 1990); if successful conditioning is found automated procedures can be developed. 11. Our final suggestion is to measure sucrose thresholds prior to training and work with insects with the highest sucrose sensitivity.

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CONCLUSIONS Proboscis conditioning represents a potential methodology for making comparisons of insect pollinators across taxa. Its advantage is that theoretically it should be applicable to insect pollinators which are both social and non-social, and as evolutionarily diverse as Hymenoptera and Lepidoptera. Obviously basic questions regarding learning can be asked with this technique, but perhaps more importantly the response of pollinator species to agrochemical, repellants and attractants, and plant/food preferences can be studied at diverse locations under the same controlled conditions. While the potential for comparison is great there are important theoretical and methodological issues that cloud comparison across species of current and future experiments. These include how broadly to interpret results in terms of theoretical learning models, which also has roots in how narrowly to restrict definitions of learning models such as classical conditioning. In addition, questions remain on interpretation of results bases on different intertrial and interstimulus interval length, and which experimental controls are essential. Not withstanding the theoretical and methodological issues, proboscis conditioning studies on non-model pollinator systems are rare. This in itself presents problems with generalizations, but the findings thus far present a picture that is much more complex than one might expect either from Apis as a model system or from phylogenetics.

REFERENCES Abramson, C. I. (1990). Invertebrate Learning: A Laboratory Manual and Source Book Washington, D. C.: American Psychological Association, Abramson, C. I. (1994). A Primer of Invertebrate Learning: The Behavioral Perspective Washington, D.C.: American Psychological Association. Abramson, C. I. (1997). Where have I heard it all before: Some neglected issues of invertebrate learning,‖ In Comparative Psychology of Invertebrates: The Field and Laboratory Study of Insect Behavior (G. Greenberg, & E. Tobach, eds.), 55-78, New York: Garland Publishing.

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Abramson, C. I., Aquino, I. S., Ramalho, F. S. & Price, J. M. (1999). Effect of insecticides on learning in the Africanized honey bee (Apis mellifera L.). Archives of Environmental Contamination and Toxicology, 37, 529-535. Abramson, C. I., Aquino, I. S., Silva, M. C. & Price, J. M. (1997). Learning in the Africanized honey bee: Apis mellifera L. Physiology & Behavior, 62, 657-674. Abramson, C. I., Aquino, I. S. & Stone, S. M. (1999c). Failure to find proboscis conditioning in one-day old Africanized honey bees (Apis mellifera L.) and in adult Uruçu honey bees (Melipona scutellaris). International Journal of Comparative Psychology, 12, 242-262. Abramson, C. I., Armstrong, P. M., Feinman, R. A. & Feinman, R. D. (1988). Signaled avoidance in the eye withdrawal reflex in the green crab, Journal of the Experimental Analysis of Behavior, 50, 483-492. Abramson, C. I. & Boyd, B. J. (2001). An automated apparatus for conditioning proboscis extension in honey bees (Apis mellifera L.), Journal of Entomological Science, 36, 78-92. Abramson C. I. & Buckbee, D. A. (1995). Pseudoconditioning in earthworms (Lumbricus terrestris): Support for nonassociative explanations of classical conditioning phenomena through an olfactory paradigm. Journal of Comparative Psychology, 109, 90-397. Abramson, C. I., Giray, T., Mixson, T. A., Nolf, S. L., Wells, H., Kence, A. & Kence, M. (2010). Proboscis conditioning experiments with honey bees (Apis mellifera caucasica) with butyric acid and DEET mixture as conditioned and unconditioned stimuli. Insect Science, In Press). Abramson, C. I., Mixson, T. A., Cakmak, I., Place, A. J. & Wells, H. (2008). Pavlovian conditioning of the proboscis extension reflex in harnessed foragers using paired vs. unpaired and discrimination learning paradigms: Tests for differences among honeybee subspecies in Turkey, Apidologie, 39, 428-435. Abramson, C. I., Nolf, S. L., Mixson, A. & Wells, H. (in press).Can honey bees learn the removal of a stimulus as a conditioning cue? Ethology. Abramson C. I. & Place, A. J. (2005). A note regarding the word ―Behavior‖ in glossaries of introductory textbooks and encyclopedia. Perceptual and Motor Skills, 101, 568-574. Abramson C. I. & Place, A. J. (2008). Learning in rattlesnakes: Review and analysis. In The Biology of Rattlesnakes (W. K., Hayes, K. R., Beaman, M. D. Cardwell, & S. P. Bush, eds.), 123-142. Loma Linda, CA: Lomo Linda University Press. Abramson, C. I., Romero, E. S., Frasca, J., Fehr, R., Lizano, E. & Aldana, E. (2005). The psychology of learning: A new approach to study behavior of Rhodnius prolixus Stal under laboratory conditions. Psychological Reports, 97, 721-731. Abramson, C. I., Singleton, J. B., Wilson, M. K., Wanderley, P. A., Ramalho, F. S. & Michaluk, L. M. (2006). The effect of an organic pesticide on mortality and learning in Africanized honey bees (Apis mellifera L.) in Brasil. American Journal of Environmental Science, 2, 37-44. Abramson, C. I., Squire, J., Sheridan, A. & Mulder, P. G. (2004). The effect of insecticides considered harmless to honey bees: prpboscis conditioning studies using the insect growth regulators tebufenozide and diflubenzuron. Environmental Entomology, 33, 378388. Abramson, C. I., Wanderley, P. A., Wanderly, M. J., Silva, J. C. & Michaluk, L. M. (2007b). The effect of essential oils of sweet fennel and pignut on mortality and learning in Africanized honey bees. Neotropical Entomology, 36, 828-835.

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Abramson, C. I., Wells, H. & Bozic, J. (2007a). A social insect model for the study of ethanol induced behavior: the honey bee. (R. Yoshida, ed.), 197-218. Hauppauge, NY: Nova Science Publishers, Inc. Amsel, A. (1989). Behaviorism, Neobehaviorism, and Cognitivism in Learning Theory: Historical and Contemporary Perspectives. Hillsdale, New Jersey: LEA. Aquino, I. S., Abramson, C. I. & Payton, M. E. (1999). A rapid bioassay for detection of adulterated beeswax,‖ Journal of Entomological Science, 34, 265-272. Batson, J. D., Hoban, J. S. & Bitterman, M. E. (1992). Simultaneous conditioning in honeybees (Apis mellifera). Journal of Comparative Psychology, 97, 107-119. Bitterman, M. E. (1962). Techniques for the study of learning in animals: Analysis and classification. Psychological Bulletin, 59, 81-93. Bitterman, M. E., Menzel, R., Fietz, A. & Schäfer, S. (1983). ―Classical conditioning of proboscis extension in honeybees (Apis mellifera),‖ Journal of Comparative Psychology, 97, 107-119. Bleeker, M. A. K., Smid, H. M., Steidle, L. M., Kruidhof, H. M., van Loon, J. J. A. & Vet, L. E. M. (2006). Differences in memory dynamics between two closely related parasitoid wasp species,‖ Animal Behaviour, 71, 1343-1350. Bonod, I., Sandoz, J., Loublier, Y. & Pham-Delegue, M. (2003). Learning and discrimination of honey odours by the honey bee. Apidologie, 34, 147-159. Bosch, J. & Kemp, W. P. (2002). Developing and establishing bee species as crop pollinators: the example of Osmia spp. (Hymenoptera: Megachilidae) and fruit trees. Bulletin of Entomological Research, 92, 3-16. Brembs, B. (2003). Operant conditioning in invertebrates, Current Opinion in Neurobiology, 13, 710-717. Brown, P. & Jenkins, H. M. (1968). Autoshaping of the pigeon‘s key-peck. Journal of the Experimental Analysis of Behavior, 11, 1-8. Buckbee D. A. & Abramson, C. I. (1997). Identification of a new contingency-based response in honey bees (Apis mellifera) through revision of the proboscis extension conditioning paradigm. Journal of Insect Behavior, 10, 479-491. Byrne, J. H. (2003). Learning & memory 2nd ed. New York: MacMillan. Cakmak, I., Sanderson, C., Blocker, T. D., Pham, L. L., Checotah, S., Norman, A. A., Harader-Pate, B. K., Reidenbaugh, R. T., Nenchev, P., Barthell, J. F. & Wells, H. (2009). Different solutions by bees to a foraging problem,‖ Animal. Behaviour, 77, 1273-1280. Cane, J. H. (1997). Ground-nesting bees: the neglected pollinator resource for agriculture Acta Horticulturae, 437, 309-324. Chabaud, M. A., Devaud, J. M., Pham-Delegue, M. H., Preat, T. & Kaiser, L. (2006). Olfactory conditioning of proboscis activity in Drosophila melanogaster. Journal of Comparative Physiology, 192, 1335-1348. Chaline, N., Sandoz, J., Martin, S. J., Ratnieks, F. L. & Jones, G. R. (2005). Learning and discrimination of individual cuticular hydrocarbons by honeybees. Chemical Senses, 30, 327-335. Chandra, S. B., Hosler, J. S. & Smith, B. H. (2000). Heritable variation for latent inhibition and its correlation with reversal learning in honeybees. Journal of Comparative Psychology, 144, 86-97.

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Charles I. Abramson, Michel B. C. Sokolowski and Harrington Wells

Chandra, S. B., Hunt, G. J., Cobey, S. & Smith, B. H. (2001). Quantitative trait loci associated with reversal learning and latent inhibition in honeybees. Behavior Genetics, 31, 275-285. Chittka, L. (1998). Sensorimotor learning in bumblebees: long term retention and reversal training, Journal of Experimental Biology, 201, 515-524. Chittka, L., Dyer, A. G., Bock, F. & Dornhaus, A. (2003). Bees trade off foraging speed for accuracy. Nature, 424, 388. Chittka L. & Thomson, J. D. (1997). Sensori-motor learning and its relevance for task specialization in bumble bees. Behavioral Ecology and Sociobiology, 41, 385-398. Coleman, S. R., Fanelli, A. & Gedeon, S. (2000). Psychology of the Scientist: LXXII. Coverage of classical conditioning is textbooks in the psychology of learning, 1952-1995. Psychological Reports, 86, 1011-1027. Couvillon P. A. & Bitterman, M. E. (1984). The overlearning-extinction effect and successive negative contrast in honeybees (Apis mellifera), Journal of Comparative Psychology, 98, 100-109. Couvillon, M. J., DeGrandi-Hoffman, G. & Gronenberg, W. (2010). Africanized honeybees are slower learners than their European counterparts. Naturwissenschaften, 97, 153-160. Daly, K. C. & Smith, B. H. (2000). Associative olfactory learning in the moth Manduca sexta, Journal of Experimental Biology, 203, 2025-2038. Daly, K. C., Chandra, S. C., Durtschi, M. L. & Smith, B. H. (2001). The generalization of an olfactory-based response reveals unique but overlapping odour representations in the moth Manduca sexta. Journal of Experimental Biology, 204, 3085-3095. Dyal, J. A. & Corning, W. C. (1973). Invertebrate learning and behavioral taxonomies. In Invertebrate Learning: Vol. 1. Protozoans through Annelids (W.C. Corning, J. A. Dyal, & A. O. D. Willows, eds.), 1-48. New York: Plenum. Fan, R. J., Anderson, P. & Hansson, B. S. (1997). Behavioural analysis of olfactory conditioning in the moth Spodoptera littoralis (Boisd.) (Lepidoptera: Noctuidae). Journal of Experimental Biology, 200, 2969-2976. Frings, H. (1944). The loci of olfactory end-organs in the honeybee. Journal of Experimental Zoology, 97, 123-134. Gegear R. J. & Laverty, T. M. (2004). Effects of a colour dimorphism on the flower constancy of honey bees and bumble bees. Canadian Journal of Zoology, 82, 587-593. Getz, W. M. & Smith, K. B. (1987). Olfactory sensitivity and discrimination of mixtures in the honey bee, Apis mellifera. Journal of Comparative Physiology, 160, 239-245. Giurfa, M. & Malun, D. (2004). Associative mechanosensory conditioning of the proboscis extension reflex in honeybees. Learning and Memory, 11, 294-302. Gormezano, I. (1984). The study of associative learning with CS-CR paradigms. In Primary Neural Substrates of Learning and Behavioral Change (D. L. Alkon & J. Farley, eds.), 524. Cambridge: Cambridge University Press. Gormezano, I. & Kehoe, E. J. (1975). Classical conditioning: Some methodologicalconceptual issues. In Handbook of Learning and Cognitive Processes, Vol. 2. Conditioning and Behavior Theory (W. K. Estes, ed.), 143-179. Hillsdale, New Jersey: LEA. Gormezano, I., Kehoe, E. J. & Marshall, B. S. (1983). Twenty years of classical conditioning research with the rabbit,‖ In Progress in Psychobiology and Physiological Psychology, Vol. 10 (J. M. Sprague, & A. N. Epstein, eds.), 197-275. New York: Academic Press.

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Issues in the Study of Proboscis Conditioning

47

Guerrieri, F., Lachnit, H., Gerber, B. & Giurfa, M. (2005). Olfactory blocking and odorant similarity in the honeybee. Learning and Memory, 12, 86-95. Hartlieb, E. (1996). Olfactory conditioning in the moth Heliothis virescens, Naturwissenschaften, 83, 87-88. Heinrich, B. (1979). Bumblebee Economics. Cambridge, MA: Harvard University. Hill, P. S., Hollis, J. & Wells, H. (2001). Flower constancy to colour in honeybees (Apis mellifera ligustica) when interfloral distances are varied, Animal Behavior, 62, 729-737. Hill, P. S., Wells, P. H. & Wells, H. (1997). Spontaneous flower constancy and learning in honey bees as a function of colour. Animal Behavior, 54, 615-627. Hirsch J. & Holliday, M. (1988). A fundamental distinction in the analysis and interpretation of behavior, Journal of Comparative Psychology, 102, 372-377. Hranitz, J. M., Abramson, C. I. & Carter R. P. (in press). Ethanol increases HSP 70 concentration in honey bees (Apis mellifera L.) brain tissue. Alcohol. Hussaini, S. A., Komischke, B., Menzel, R. & Lachni, H. (2007). Forward and backward second-order Pavlovian conditioning in honeybees. Learning and Memory, 14, 678-683. Ito, I., Ong, R. C., Raman, R. & Stopfer, M. (2008). Sparse odor representation and olfactory learning, Nature Neuroscience, 11, 177-1183. Kartzev, V. M. (1996). Local orientation and learning in insects. In Russian Contributions to Invertebrate Behavior (C. I. Abramson, Z. P. Shuranova, & Y. M. Burmistrov, eds.), 177212. Westport, CT.: Praeger. Kroutov, V., Mayer, M. S. & Emmel, T. C. (1999). Olfactory conditioning of the butterfly Agraulis vanilla (L.) (Lepidoptera, Nymphalidae) to floral but not host-plant odors, Journal of Insect Behavior, 12, 833-843. Kuwabara, M. (1957). ―Bildung des bedingten reflexes von pavlous typus bei der honigbiene, Apis mellifera [Establishment of Pavlovian conditioned reflexes in honeybees],‖ Journal of the Faculty of Science, Hokkaido University, Zoology, 13, 458464. Laloi, D., Sandoz, J. C., Picard-Nizou, A., Marchesi, A., Pouvreau, A., Tasei, J. N. Poppy, G. & Pham-Delegue, M. H. (1999). Olfactory conditioning of the proboscis extension in bumble bees. Entomologia Experimentalis et Applicata, 90, 123-129. Laverty, T. M. (1994). Bumble bee learning and flower morphology. Animal Behaviour, 47, 31-545. Laverty T. M. & Plowright, R. C. (1988). Flower handling by bumblebees: a comparison of specialists and generalists, Animal Behaviour, 36, 733-740. Lee, V. L. (1988). Beyond Behaviorism. Hillsdale, New Jersey: LEA. Mallon, E. B., Brockmann, A. & Schmid-Hempel, P. (2003). Immune response inhibits associative learning in insects. Proceedings of the Royal Society B, 270, 2471-2473. Mamood A. N. & Waller, G. D. (1990). Recovery of learning responses by honeybees following a sublethal exposure to permethrin. Physiological Entomology, 15, 55-60. McCabe, S. I., Hartfelder, K., Santana, W. C. & Farina, W. M. (2007). Odor discrimination in classical conditioning of proboscis extension in two stingless bee species in comparison to Africanized honeybees. Journal of Comparative Physiology A., 193, 1089-1099. Menzel, R and Bitterman, M. E. (1983). Learning by honeybees in an unnatural situation. In Neuroethology and Behavioral Physiology: Roots and Grouping Points, (F. Huber, & M. E. Markl, eds.), 206-215. Berlin: Springer-Verlag.

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48

Charles I. Abramson, Michel B. C. Sokolowski and Harrington Wells

Menzel, R. & Muller, U. (1996). Learning and memory in honey bees - From behavior to neural substrates, Annual Review of Neuroscience, 19, 379-404. Menzel, R., Hammer, M., Braun, G., Mauelshaugen J. & Sugawa, M. (1991). Neurobiology of learning and memory in honeybees. In The Behavior and Physiology of Bees, (L. J. Goodman, & R. C. Fisher, eds.), 323-353. Slough, UK: C.A.B. International. Mercer, A. (1987). Biogenic amines in the bee brain. In Neurobiology and Behavior of Honeybees (R. Menzel, & A. Mercer, eds.), 244-252, Berlin: Springer-Verlag. Minnich, D. E. (1932). The contact chemoreceptors of the honey bee Apis mellifera Linn. Journal of Experimental Zoology, 61, 375-393. Mujagic, S. & Erber, J. (2009). Sucrose acceptance, discrimination and proboscis responses of honey bees (Apis mellifera L.) in the field and the laboratory. Journal of Comparative Physiology A., 195, 325-339. Nicolas, C., Abramson, C. I. & Levin, M. (2008). Analysis of behavior in the planarian model. In Planaria: A Model for Drug Action and Abuse (R. B. Raffa, & S. M. Rawls, eds.), 83-94. Austin, TX: R.G. Landes Co. Percival, M. S. (1965). Floral Biology, New York: Pergamon Press Ltd.. Raine N. E. & Chittka, L. (2007). ―Pollen foraging: learning a complex motor skill by bumblebees (Bombus terrestris),‖ Naturwissenschaften, 94, 459-464. Raine, N. E., Ings, T. C., Dornhaus, A., Saleh, N. & Chittka, L. (2006) Adaptation, genetic drift, pleiotropy, and history in the evolution of bee foraging behavior, Advances in the Study of Behavior, 36, 305-354. Raine, N. E., Ings, T. C., Ramos-Rodrıguez, O. & Chittka, L. (2006). Intercolony variation in learning performance of a wild British bumblebee population (Hymenoptera: Apidae: Bombus terrestris audax). Entomologia Generalis, 28, 241-256. Razran, G. (1971). Mind in Evolution: An East-West Synthesis of Learned Behavior and Cognition. Boston: Houghton Mifflin. Renner, M. A. & Nieh, J. C. (2008). Bumble bee olfactory information flow and contactbased foraging activation. Insectes Sociaux, 55, 417-424. Riveros, A. J. & Gronenberg, W. (2009). Olfactory learning and memory in the bumblebee Bombus occidentalis. Naturwissenschaften, 96, 851-856. Sanderson, C. E., Orozco, B. S., Hill, P. S. M. & Wells, H. (2006). Honeybee (Apis mellifera ligustica) response to differences in handling time, rewards, and flower colours, Ethology, 112, 937-946. Sauer, D. L., Abramson, C. I. & Lawson, A. (2002). Exploratory studies of classical conditioning of the preoral cavity in harnessed carpenter ants (Camponotus pennsylvanicus). Psychological Reports, 90, 1037-1050. Scheiner, R., Barnert, M. & Erber, J. (2003). Variation in water and sucrose responsiveness during the foraging season affects proboscis extension learning in the honey bees. Apidologie, 34, 67-72. Scheiner, R., Erber, J. & Page, R. E. (1999). Tactile learning and the individual evaluation of the reward in honey bees (Apis mellifera L.). Journal of Comparative Physiology A., 185, 1-10. Scheiner, R., Page, R. E. & Erber, J. (2004). Sucrose responsiveness and behavioral plasticity in honey bees (Apis mellifera). Apidologie, 35, 133-142. Sheldon, J. P. (2002). Operant conditioning concepts in introductory psychology textbooks and their companion web sites. Teaching of Psychology, 29, 281-285.

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49

Sidman, M. (1960). Tactics of scientific research: evaluating experimental data in psychology. New York: Basic Books, Inc. Skinner, B. F. (1989). Recent Issues in the Analysis of Behavior. Columbus, Ohio: Merrill. Skiri, H. T., Stranden, M., Sandoz, J. C., Menzel, R. & Mustaparta, H. (2005). Associative learning of plant odorants activating the same or different receptor neurons in the moth Heliothis virescens. Journal of Experimental Biology, 208, 786-796. Smith, B. H. & Cobey, S. (1994). The olfactory memory of the honeybee. Journal of Experimental Biology, 195, 91-108. Smith B. H. & Menzel, R. (1989). The use of electromyogram recordings to quantify odourant discrimination in the honey bee, Apis mellifera. Journal of Insect Physiology, 35, 369-375. Smith, B. H., Abramson, C. I. & Tobin, T. R. (1991). Conditioned withholding of proboscis extension in honey bees (Apis mellifera) during discriminative punishment. Journal of Comparative Psychology, 105, 345-356. Stepanov I. I. & Abramson, C. I. (2008). The application of an exponential mathematical model for 3- arm radial maze learning. Journal of Mathematical Psychology, 52, 309319. Stone, J. C., Abramson, C. I. & Price, J. M. (1997). Task dependent effects of dicofol (Kelthane) on learning in the honey bee (Apis mellifera). Bulletin of Environmental Contamination and Toxicology, 58, 177-183. Takeda, K. (1961). Classical conditioned response in the honeybee,‖ Journal of Insect Physiology, 6, 168-179. Taylor, K. S., Waller, G. D. & Crowder, L. A. (1987). Impairment of a classical conditioning response of the honeybee (Apis mellifera L) by sublethal doses of synthetic pyrethroid insecticides. Apidologie, 18, 243-252. Toda, N. R. T., Song, J. & Nieh, J. C. (2009). Bumblebees exhibit the memory spacing effect. Naturwissenschaften, 96, 1185-1191. Tulving, E. (1985). On the classification problem in learning and memory. In Perspectives on Learning and Memory (L. Nilsson, & T. Archer, eds.), 67-94. Hillsdale, New Jersey: LEA Vareschi, E. (1971). Duftunterscheidung bei den honigbiene: Einzellzel-ableitungen und verhaltensreaktionen [Odor discrimination by the honeybee: Single cell recording and behavior reaction], Zeitschrift für Vergleichende Physiologie, 75, 143-173. Wackers, F. L., Bonifay, C. & Lewis, W. J. (2002). Conditioning of appetitive behavior in the Hymenopteran parasitoid Microplitis croceipes. Entomologia Experimentalis et Applicata, 103, 135-138 Wells H. & Wells, P. H. (1986). Optimal diet, minimal uncertainty and individual constancy in the foraging of honey bees, Apis mellifera, Journal of Animal Ecology, 55, 881-891. Wells, H., Hill, P. S. & Wells, P. H. (1992). Nectivore foraging ecology: Rewards differing in sugar type. Ecological Entomology, 17, 280-288. Wenner A. M. & Wells, P. H. (1991). Anatomy of a Controversy. New York: Columbia University Press. Wickens D. D. & Wickens, C. D. (1942). Some factors related to pseudoconditioning. Journal of Experimental Psychology, 31, 518-526. Woods, P. J. (1974). A taxonomy of instrumental conditioning. American Psychologist, 29, 584-596.

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In: Social Insects: Structure, Function, and Behavior Editor: Emily M. Stewart

ISBN: 978-1-61761-466-8 © 2011 Nova Science Publishers, Inc.

Chapter 3

REGULATION OF REPRODUCTIVE STATES AND CONTROL OF SEX OF EGGS BY REPRODUCTIVE FEMALES IN EUSOCIAL HYMENOPTERA Ken Sasaki Department of Applied Bioscience, Human Information Systems, Kanazawa Institute of Technology, Hakusan, Ishikawa, Japan

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ABSTRACT In Hymenoptera, sex of an egg is determined by egg fertilizations on a haplodiploidy sex determination system. Female reproductive organs in eusocial species have unique functions to regulate ovarian development and control the sex of an egg. Egg fertilizations could be controlled by muscle activities of the spermathecal pump and/or secretion of the spermathecal glands. Activities of the spermathecal pump are regulated by neuronal systems and may inhibit release of sperm for unfertilized male eggs. Secretions of the spermathecal glands could activate spermatozoa from the spermathecal receptacle and therefore, lack of spermathecal gland secretions may cause unfertilization of eggs. In highly eusocial species, especially honeybees, queens can control the egg fertilization in response to comb cell types for oviposition. They can make a choice of the cell types and behaviorally control the egg sex ratios in the colonies. Workers in honeybees can develop reproductive organs in the absence of a queen and lay only unfertilized eggs. They select preferentially male comb cells for the oviposition to produce larger males for mating competition between males. I introduce general structure and function of female reproductive organs for controlling egg fertilizations, physiological processes for regulating reproductive states and behavioral controls of the egg sex ratio in colonies in Hynemoptera.

INTRODUCTION In most species in Hymenoptera, fertilized eggs develop into females, while unfertilized eggs do parthenogenetically into males. Such manner of sex determination is widely known

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Ken Sasaki

as haplodiploidy. The haplodiploidy sex determination system had been observed by Dzierzon (1845) in honeybees, and then Whiting (1943) proposed an explanation of this genetic mechanism based on his discovery of diploid males among inbred progenies of the parasitoid Bracon hebetor. In his model of complementary sex determination (CSD), heterozygototes at a multi-allelic sex determining locus develop as females, whereas hemizygotes and homozygotes as males. The single-locus complementary sex determination has been shown to be the case for over 30 species (Crozier and Pamilo, 1996). Recently, the single-locus nature of CSD in honeybees was confirmed by molecular biological studies and the single-locus CSD gene (csd) has been identified (Beye et al., 2003; Hasselmann et al., 2008). If csd is heterozygous it produces an active form of the protein it codes for, which then causes feminizer gene (fem) to produce the female form of fem mRNA (Gempe et al., 2009). In some species in Hymenoptera, however, the prolonged inbreeding does not lead to production of diploid males. The sex determination in such non-typical species is explained by multi-locus sex determination (Crozier, 1971) or by genomic imprinting sex determination with a switching gene turned off in eggs but on in sperm (Beukeboom, 1995; Dobson and Tanouye, 1998). Current empirical evidence and theoretical studies suggest that the singlelocus complementary sex determination should be the ancestral mechanism of sex determination in Hymenoptera, but that alternative mechanisms, multi-locus sex determination or genomic imprinting, have evolved in inbreeding species because of high cost of producing diploid males. Haplodiploidy provides females with a physiological mechanism for the maternal manipulation of the sex of eggs. Mothers can potentially control the sex of eggs by controlling the egg fertilization in order to enhance their fitness at the oviposition. Some endoparasitic wasp females manipulate the sex ratio of eggs laid into hosts in response to the host quality including their body size and instar (Charnov et al., 1981). The selective advantage of the sex ratio manipulation by mothers can be explained by host-size model (Charnov et al., 1981) and local mate competition theory (Hamilton, 1967). The former predicts that if one sex could gain relatively more benefit by being large in the host, the sex should be produced preferentially in larger hosts. The latter proposes that under the subdivided population structure in which the mating occurs among offspring of limited females, the production of female-biased broods decreases competition among sons while increasing the number of potential mates for the remaining sons. In eusocial species in Hymenoptera, frequency-dependent sex ratio theory (Fisher, 1930), inclusive fitness theory (Hamilton, 1964), and parent-offspring conflict (Trivers, 1974) have been combined for studies on sex ratio conflict between queens and workers in a colony. Since haplodiploidy results in asymmetries of genetic relatedness among colony members, the asymmetries cause colony members to have different inclusive fitness, which leads to a parent-offspring conflict over relative allocation of resources in the two sexes (Trivers and Hare, 1976; Pamilo, 1991). Because queens are equally related to both sexes of reproductives reared in their colony, their fitness is maximized by an even sex ratio investment. In contrast, because workers are more closely related to their sisters than to their brothers, their inclusive fitness is maximized by an increased investment in female brood. Consequently, the population investment sex ratio is predicted to be even under queen control, and to be femalebiased under worker control. Despite its theoretical interest and some empirical observations on the sex ratio in colonies or a population, very few studies in eusocial species have investigated the potential

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Regulation of Reproductive States and Control of Sex of Eggs…

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ability of queens to adjust the sex of eggs. Since the egg sex ratios controlled by queens could contribute to the final sex ratio in colonies, a clarification of the mechanisms at behavioral and neuro-endorine levels must be important. The controls of egg sex ratios by queens result from multi-level manipulations: a control of sperm discharge in the reproductive organs and a control of egg-laying behaviors associated with the egg fertilization. Since some detailed studies on the reproductive organs and egg-laying behaviors have been reported in honeybee queens, I introduce the mechanisms for the sex ratio manipulation by females, especially honeybee queens, in this chapter. Workers in many eusocial species have a potential to lay eggs in the absence of a queen. They can control the reproductive states depending on the colony circumstances and produce only unfertilized male eggs because of unmating. The worker reproductions could contribute to male egg production in a colony and seem to be based on similar mechanisms to regulate the development and action of reproductive organs in queens. Since the transition of reproductive states from infertile workers to reproductive individuals can help to understand general mechanisms of reproductive maturations and oviposition in females, this topic is also discussed.

MORPHOLOGY AND FUNCTION OF FEMALE REPRODUCTIVE ORGANS

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The internal organs of reproduction in females vary in structure from species to species in insects. There is usually a pair of ovaries each of which opens into a lateral oviduct. The two lateral oviducts unite in a single median common oviduct (Figure 1A, B). The common oviduct opens into the vagina (genital chamber). The spermatheca storing spermatozoa connects the dorsal vagina via a spermathecal duct (Figure 1A). A few spermatozoa are discharged from the spermatheca into the vagina during egg fertilization.

(a) Reproductive Tract Oviduct Mature eggs produced from a pair of ovaries are stored in the lateral oviducts until the queen begins an oviposition. Muscles of the lateral oviducts were attached on the outer surface of the connective tissue, but could not make a strong force to distort eggs because of their wide expandable tracts in the lateral oviducts. In contrast to the thin layer of muscles surrounding the lateral oviducts, muscles of the common oviduct were well developed in a relatively thicker layer in honeybee queens (Figure 1B) (Laidlaw, 1944; Snodgrass, 1956; Ruttner, 1961; Sasaki and Obara, 2002). The muscle fibers in the common oviduct are attached to the connective tissue with the inner cuticle of the tract from the dorsal to lateral region, but not to the connective tissue of the ventral floor of the tract, and terminate on the median antecosta of sternum VII like the other muscles of the tract (Laidlaw, 1944; Ruttner, 1956, 1961; Camargo and Mello, 1970). This structure can move the inner cuticle of the tract inside and squeeze the egg with a strong force of the semi-circular muscle when the muscles contract with an egg within (Sasaki and Obara, 2002). The well-developed semi-circular

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muscles are found in reproductive tracts in other eusocial species in Hymenoptera including ants (Sasaki, unpublished).

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Vagina The valve-fold projects into the lumen of the vagina like a tongue-shaped roll just behind the caudal end of the median oviduct in honeybee queens (Figure 1A, C). There are small numbers of muscle fibers attached to the ventral outer layer of the valve-fold in the vagina which originate from the median antecosta of sternum VII (Ruttner, 1956; Camargo and Mello, 1970). The valve-fold would be raised normally and fixed posteriorly during copulation (Ruttner, 1956), which may caused by the action of the muscle attached to the vale-fold in the vagina. It has been reported that the size of valve-fold is larger in younger queens than old queens (Fyg, 1966), but the functional roles of the valve-fold are still unknown.

Figure 1. Structure of the reproductive organ in a honeybee queen. A: a longitudinal section of the reproductive tract with spermatheca, B: a horizontal section of the reproductive tract, C: a vertical section of the vagina, D: a vertical section of the spermathecal pump, CO: common oviduct, LO: lateral oviduct, Sm: semi-circular muscle, SP: spermatozoa, SpD: spermathecal duct, SpG: spermathecal gland, SpP: spermathecal pump, SpPM: spermathecal pump muscle, SpR: spermathecal receptacle, TAG: terminal abdominal ganglion, Va: vagina, VF: valve-fold in the vagina.

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(b) Spermatheca and Spermathecal Glands The spermatheca has functions to store and nourish the spermatozoa until they are used to fertilize the eggs. The spermatozoa can be stored in the spermatheca for 2-3 years in honeybee queens and more than 10 years in ant queens (Hӧlldobler and Wilson, 1990). As solitary species, the spermatheca in the honeybee queens consists of a receptacle for storing spermatozoa, spermathecal glands for producing sperm-activating secretion, and a spermathecal pump controlling sperm release into the vagina for egg fertilization (Figure 1A, D).

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Receptacle The receptacle walls in honeybee queens consist of a single epithelial layer of cells resting on the basement membrane over which ramifies an elaborate tracheal system (Poole, 1970; Dallai, 1975). There are no muscles surrounding the receptacle of the spermatheca. The thickness of the epithelial cells of receptacle increases with age in queens (Poole, 1970). The spermathecal epithelium does not seem to be involved in exocrine secretion related to nutrition of long-term stored spermatozoa. However, the ultrastructure points to ion transport functions and to an engagement in the maintenance of an adequate physicochemical environment ensuring the viability of the spermatozoa (Dallai, 1975; Kressin et al., 1996). The epithelium maintains large concentration gradients of inorganic ions, generates an electrical positive potential in receptacle and produces an alkaline condition in spermathecal fluid in comparison to hemolymph (Gessner and Gessner, 1976). The epithelium of the spermathecal wall transports K+ (and possibly HCO3- or OH-) actively into the receptacle, but handles Na+ passively. The higher concentration of K+ and alkaline condition in receptacle seems to be required to maintain the spermatozoa alive over longer periods of time (Gessner and Gessner, 1976). Spermathecal Gland The spermathecal gland opens into the unobstructed sperm duct where it joins the capsule (Figure 1D). The secretions of the spermathecal gland have positive effects on sperm viability (den Boer et al., 2009) and are thought to activate the spermatozoa before fertilization (Flanders, 1939; Snodograss, 1956). For an egg to be fertilized as it passes outward through the vagina, a few of the sperm need to have been activated previously. Therefore, secretions of the spermathecal gland are thought to serve both to transport the sperm and to activate it. The epithelium of the spermathecal gland of honeybee queens is composed of two types of cells: secretory cells and cuticologenic cells (Dallai, 1972). The secretory cells are engaged in the secretion of a polysaccharide containing proteins which are then poured into a central cavity. An efferent duct, organized by the cuticologenic cells, accomplishes the task of conveying the secretion from the central cavity of the secretory cells to the lumen of the gland. Musclular coats are absent outer layer of the spermathecal gland. Recently, the comparisons of proteomic profiles between the secretion of spermathecal glands and the fluid of spermathecal receptacle in honeybee queens have been reported (den Boer et al., 2009; Baer et al., 2009). The profile of proteins from spermathecal fluid is very similar to the secretions of the spermathecal gland, suggesting that the spermathecal glands are main contributors to the spermathecal fluid proteome. The proteins detected from the spermathecal

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gland secretion contain enzymes of energy metabolism and antioxidant defense (Collins et al., 2004). It remains, however, how these proteins act on the spermatozoa alive over longer periods of time with low mobility in the receptacle and activate the spermatozoa with high mobility in the spermathecal duct before fertilization.

Spermathecal Pump The spermathecal pump is a bend in the sperm duct and store temporally a few of sperm and secretions of spermathecal glands into the reservoir in the duct (Figure 1D and Figure 2A). The walls of the sperm duct at the bend are supplied with muscles which operate to change the angle of the bend (Bresslau, 1905; Snodograss, 1956) (Figure 2). The compressor muscles surround the sperm duct and attach both sides of cuticle of the reservoir. The extensor muscles attach the cuticle of the reservoir and the sperm duct. The contractions of compressors and extensors distort the cuticle of the bend and transport the activated spermatozoa to the sperm duct for releasing it into the vagina. The ultrastructure of these muscle fibers is similar to those fibers observed in the visceral muscles of other insects (Dallai, 1975). This finding may be useful for exploring the neuroactive substances regulating the spermathecal pump muscles.

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CONTROL OF EGG FERTILIZATION Females in solitary species in Hymenoptera can control the sex of offspring at the oviposition in response to external cues to enhance their fitness (Charnov et al., 1981). The mechanisms by which females respond to external cues to control egg fertilization remain unknown, although there are some studies on an advantage of sex adjustment by females. Hymenopterous females generally control sperm discharge by muscle activity of a spermathecal pump (Gerber and Klostermeyer, 1970) and by secretion in the spermathecal gland (Flanders, 1939, 1950). The former plays a role in keeping the duct closed and in opening it to release sperm into the vagina (Figure 2), while the latter increases activity of spermatozoa, thus raising the pressure in the spermathecal pump. Production of the spermathecal gland secretion may change through a year or nutritional states of females, which influence the egg fertilization. The histology of the spermathecal pump and spermathecal gland in Hymenoptera has been investigated by many researchers (Bresslau, 1905; Dallai, 1972; Dallai, 1975; Wheeler and Krutzsch, 1994; Pabalan et al., 1996). However, few studies have concentrated on the mechanisms for the sperm discharge during egg transports from the oviduct to the vagina.

(a) Neural Loops for Sperm Discharge It is thought that sperm discharges into the vagina from the spermatheca may be caused by the mechanical stimuli expanding the wall of vagina by an egg during ovulation. In locusts, there are several studies demonstrating the neural loop from sensory systems in the genital chamber to motor systems in the spermathecal duct (Okelo, 1979; Clark and Lange, 2000, 2001). In Hymenoptera, there are several studies determining the nerve innervations to

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the common oviduct, vagina and spermathecal pump (Fyg, 1943; Ruttner, 1961), but few studies demonstrating the neural loops for sperm discharge have been reported. In honeybees, queens can control the sperm discharge depending on the comb cell sizes; larger cells for unfertilized eggs (males) and smaller cells for fertilized eggs (females). When they are allowed to lay eggs in an open place where no comb cells for oviposition are available, the most eggs are fertilized, suggesting that the egg fertilization occurs automatically in response to an egg passage though the vagina (Sasaki and Obara, 1999). This experimental set-up is abnormal for the queens in respect to oviposition in three ways; (1) absence of cell inspection, (2) no ventral bending of abdomen and (3) no insertion of abdomen into cells. The ventral bending of the abdomen and insertion of the abdomen into cells are not necessarily required for the release of sperm. Queens might be stimulated to prevent egg fertilization by the exact size of male cells or other unknown cues including chemical substances in male cells. Actions of the spermathecal pump during sperm transfer into spermatheca after copulation in honeybee queens provide a hint to understand the neural loop for opening the spermathecal pump. Ruttner and Koeniger (1971) removed the spermathecal glands from young unmated queens and allowed the queens to naturally mated or artificially inseminated. In the queens operated, only a small number of spermatozoa reached the spermatheca. Paralysis of the skeletal muscles by poison (microbracon) in the queen‘s abdomen caused a reduction of sperm entering the spermatheca. When intact queens were inseminated with damaged spermatozoa, the spermatheca remained almost empty. Thus, the transfer of spermatozoa out of the oviducts into the spermatheca is a complex process in which the muscles of the queen, as well as the fluid of the spermathecal glands and the individual movements of the spermatozoa take part.

Figure 2. Sperm release mechanisms by action of spermathecal pump muscles in honeybee queens. A: muscle distribution of spermathecal pump (Bresslau, 1905), B: cuticular distortion by spermathecal pump muscles during sperm release. Secretions of the spermathecal glands and sperm from the receptacle are mixed in the reservoir in the spermathecal duct (arrows in the left figure). By the contractions of spermathecal pump muscles, the cuticle of the sperm duct distorts as directions indicated by arrow heads and the activated sperm are released to the vagina (arrows in the right figure). Points in the figures indicate the cuticular connections with the pump muscles. Comp: compressor muscles, Ext: extensor muscle, Res: a reservoir in the spermathecal duct, SpD: spermathecal duct, SpGD: a duct from the spermathecal glands, SpR: receptacle in the spermatheca.

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Figure 3. Mechanisms for cleansing of unused sperm left in the vagina in honeybee queens. A: a reproductive tract during cleansing of sperm left in the vagina, B: a micropyle at anterior tip of an egg taken by SEM, scale: 10 m, C: Nuclei of sperm cells stained by Feulgen staining on the posterior end of the laid egg indicated in a dashed line circle, scale: 50 m.

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(b) Regulation of Spermathecal Action by Biogenic Amines Neural regulation of the spermatheca may be involved in the release of biogenic amines. In locusts, the spermathecal ducts are innervated by dorsal unpaired median (DUM) neurons (Okelo, 1979; Clark and Lange, 2000; da Silva and Lange, 2008). These neurons synthesize both tyramine and octopamine and release mainly octopamine. Both tyramine and octopamine increase the frequency and basal tonus of spermathecal contractions in a dose-dependent manner, with octopamine having a lower threshold (da Silva and Lange, 2008). These phenolamines also act on the muscle of reproductive tracts and associate the ovulation in Orthoptera (Kalogianni and Theophilidis, 1993; Lange, 2009). In honeybees, tyramine has a role of acceleration of ovarian development in the reproductive workers (Sasaki and Harano, 2007; Thompson et al., 2007). It is possible that both tyramine and octopamine have roles of the regulation of motor systems in reproductive tracts and spermathecal pump, and probably activities of spermathecal gland secretions in honeybees.

(c) Cleansing of Spermatozoa Left in the Vagina Honeybee queens can control egg fertilization accurately depending on comb cell types (Koeniger, 1970; Sasaki et al., 1996; Ratnieks and Keller, 1998). However, the mechanism by which unused sperm left in the vagina is cleared off so as not to fertilize subsequent eggs

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mistakenly is unclear (Michener, 1974). Sasaki and Obara (1999) suggested that the spermatozoa left in the vagina should be trapped on the posterior end of eggs and pushed out (Figure 3). Since the micropyle of an egg is located at the anterior end (Retnakaran and Percy, 1985; Sasaki and Obara, 2002), an egg should be exposed to the fresh semen on the micropyle to be fertilized, or to the vaginal liquid that contain no sperm to be not. It is, therefore, likely that the posterior end first manner of egg passage through the vagina constitutes the mechanism of cleansing the vagina of unused semen, thereby ensuring accurate control over sex determination of the eggs to be laid by the queen.

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(d) Initiation of Development in Unfertilized Eggs For many sexually reproducing animals the final stages of egg development are initiated by changes accompanying sperm entry. However, it is clear that such mechanisms cannot operate in insects that have the ability to reproduce parthenogenetically. In most species in Hymenoptera, fertilized eggs develop into females whilst unfertilized eggs develop parthenogenetically into males. Unfertilized eggs of various Hymenopterans do not develop unless they receive a specific stimulus which induces parthenogenesis (Salt, 1965; King and Rafai, 1973). The development of unfertilized eggs in Parasitica, including Ichneumonoidea, is activated by mechanical stresses encountered during passage through the female‘s ovipositor (King and Rafai, 1973; Went and Krause, 1973, 1974; Went, 1982; Vinson and Jang, 1987). In most Aculeata, including honeybees, however, eggs are laid through the ovipositional pore without being mechanically distorted in the ovipositor, since this has lost its egg-laying function and has been converted into a sting. The development of unfertilized, mature eggs in honeybees can be activated by mechanical stresses (Sasaki et al., 1997; Sasaki and Obara, 2002) as the eggs in Ichneumonoidea. These could be encountered during passage through the reproductive organs, especially the common oviducts surrounded by welldeveloped semi-circular muscles (Figure 1B and Figure 3).

BEHAVIORAL CONTROL OF SEX RATIO OF EGGS Behavioral controls of egg fertilization during oviposition have been reported in several solitary species in Hymenoptera (Gerber and Klostermeyer, 1970; Charnov et al., 1981). In these species, times for oviposition between fertilized and unfertilized eggs differ (Gerber and Klostermeyer, 1970; Cole, 1981; Ueno, 1995). These controls of sex of eggs are based on the external stimuli indicating quality or quantity of resource (e.g. host or pollen) and space available for growing the offspring (Charnov et al., 1981). In social species, queens perform short-term and long-term switches of sex of eggs. The former is a temporal regulation of egg fertilization by action of the spermathecal pump or secretion of the spermathecal glands in response to external stimuli (e.g. different types of cells for oviposition). The latter is a seasonal or nutritional regulation of egg fertilization. Queens using both switching mechanisms to control the egg sex ratio in the colonies are found in limited species in highly social bees. Rather, queens with the long-term switches of egg fertilizations seem to be usual in eusocial species including ants, social wasps and other social bees.

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(a) Short-Term Switches of Sex of Eggs Short-term switches for controlling sex of eggs are frequently reported in solitary species in Hymenoptera (Charnov et al., 1981). Trap nesting bees and wasps where the mother places food plus an egg in a cell constructed inside a crevice (e.g. a hollowed-out twig). The size of the crevice places constraints on the cell size, the amount of food packed in and consequently the size of resulting adult. In these species, females tend to lay an unfertilized egg in a larger diameter tunnel (Charnov et al., 1981). Similar sex controls of eggs depending on cell size are observed in honeybees. Honeybee queens can control the sex of eggs on the basis of different sizes of comb cells available for oviposition; males in larger cells and females in smaller ones. This cell size dimorphism likely arose or was maintained through selection on males for large body size, as small worker-sized males experience a lower reproductive success (Berg et al., 1997). The dimorphism of comb cell size between male and worker is various among species in a genus of Apis (Ruttner, 1988; Crane, 1990). The difference in size between worker and male cells is larger in A. florea (drone cell / worker cell = 1.50-1.59) than A. mellifera (1.31-1.38), and less pronounced in A. cerana (1.08-1.17) than in A. mellifera. No difference in the size between worker and male cells has been detected in A. dorsata (almost 1.0). The variation of the dimorphism of cell size is due to the different optimal sizes of males and for competition among males and workers for their tasks in each species. Before an ovipositing honeybee queen deposits an egg in any cell, she first briefly inspects the cell by thrusting her head and forelegs into it. Several studies insisted that during this cell inspection the queen discriminates between male and worker cells (Flanders, 1950; Koeniger, 1970). Flanders (1950) suggested that the sensory inputs from the antennae during cell inspection caused secretion in the spermathecal gland. Koeniger (1970) reported that the queen‘s forelegs are primarily important in recognition of male cells during cell inspection, this observation being based upon the experiments using queens with their forelegs amputated or with flags on intact forelegs to prevent them from inspecting cells. Sasaki and Obara (1999) suggested that queens laid fertilized eggs when sensory inputs from the cells are lacking and that external cues are necessary to initiate production of unfertilized male eggs. The external cues for discrimination between male and worker cells have been unknown.

(b) Long-Term Switches of Sex of Eggs In some bees and ants with highly organized social systems, the sex ratio of offspring and the proportion of reproductives in colonies could be manipulated by workers, because they have the ability to adjust the amount of food to either their sisters or brothers during their developmental stage. In such colonies, workers can distinguish the sex of larvae, provide preferential care for either sex of their broods, or eliminate male larvae (Aron et al., 1995; Keller et al., 1996; Sasaki et al., 2004). Several studies have hypothesized that workers determine the allocation of investment in male and female reproductives by referring to such factors as the genetic relatedness among colony members (Boomsma and Grafen, 1991), local mate or resource competition (Frank, 1987a, b), and resource availability (Nonacs, 1986). A few studies in social species have investigated the potential ability of queens to adjust the sex of eggs (Aron et al., 1995; Keller et al., 1996; Sasaki et al., 1996). In the Argentine ant

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Irdomyrmex humile and the fire ant Solenopsis invicta, the adult sex ratio is influenced both by the queen that adjusts the sex ratio of her eggs during oviposition and by the workers that eliminate male larvae. In monogynous species, queens may exert some control over the investment proportion in the reproductives. They control the number of haploid and diploid eggs laid and the workers are obliged to work within this constraint. It is expected that queens might also be able to adjust the number of female reproductives during the reproductive season by regulating the number of male eggs. Further, queens might have the ability to determine when to produce male reproductives or non-reproductives for colony growth and maintenance. It has been reported in several species of ants that queens determine the initial sex ratio in response to environmental factors such as seasonal change, temperature and the degree of colony growth (Göswald and Bier, 1955; Hölldobler and Wilson, 1990; Hasegawa, 1992; Aron et al., 1994). Similar manipulation of the initial sex ratio has been reported in honeybees. Honeybee colonies produce males and infertile female workers throughout most of the active broodrearing season, with a few queens produced before colony fission. Workers might be responsible for the sex ratio adjustment in the colony by adjusting the number of male, queen and worker cells that they build (Allen, 1963; Free and William, 1975; Page, 1981; Page et al., 1993). Workers, depending on the season, could change the amount of care invested in male broods (Page and Metcalf, 1984). Queens are also in a position where they could exert initial sex ratio control. Queens could recognize the type of cell during their cell inspection and adjust the sex of eggs laid in each cells (Koeniger, 1970). Harbo (1976) observed that virgin queens showed a preference for oviposition in male cells. In the paper, however, the numbers of male and worker cells prepared by workers were not given. Does the queen actually select either male or worker cells for her own benefit? Alternatively, is the queen forced to accept any type of cell that is prepared by the workers, and is she unable to make a choice as to the sex of the eggs that she lays? Sasaki et al. (1996) examined the queens‘ selections between male and worker cells by using special flames arranging male and worker cell zones in a checkered pattern (Figure 4). In this experiment, queens allowed to select either male or worker cells for oviposition and could select male or worker cells depending on season (Sasaki et al., 1996). The queen‘s production of male eggs may be regulated by negative feedback. Studies supporting the mechanisms found that queens who had recently laid male eggs subsequently produced fewer male eggs than queens who had recently laid only worker eggs (Sasaki et al., 1996; Wharton et al., 2007). Nutritional factors seem to influence the egg sex ratios in honeybees (Sasaki and Obara, 2001). Our study examined the interaction of season and food availability on the queen‘s egglaying decisions. When colonies are continuously supplied with food throughout the year, queens lay more male eggs in spring and summer than in fall. Additionally, while the queen lays few male eggs in the fall independent of food condition, in the late spring she appears to produce a higher proportion of male eggs during good food conditions than during poor food conditions. This means that queens produce more male eggs in the reproductive season and fewer in other seasons of the year independently of the nutritional status of the colonies. It is not clear, however, whether the queens adjust the egg sex ratios on the basis of cues of food from workers directly or those originating from, for example, altered worker activity due to changed temperature or CO2 concentration in the nest.

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Figure 4. Queen‘s oviposition on the checkered frame. A: chromosomes in a haploid egg laid in a male cell, B: chromosomes in a diploid egg laid in a worker cell, C: an example of a sequence of oviposition by a queen on the checkered frame.

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REGULATION OF REPRODUCTIVE STATES IN FEMALES In colonies of eusocial species in Hymenoptera, queens inhibit workers‘ reproduction behaviorally and chemically. In primitive eusocial species including paper wasps and bumble bees, queens attack workers by antennal crashing, biting and attempting to sting. The queens‘ dominant behaviors are thought to cause the inhibition of ovarian development in workers. In highly eusocial species, queens do not show the dominant behaviors and secret queen pheromones (queen substances). The pheromones have functions to display the existence of a queen in the colony and inhibit the reproduction by workers. With the evolution from primitively to highly eusociality, the queens‘ behavioral inhibitions of worker reproduction may change the chemical inhibitions using queen pheromone.

(a) Behavioral Changes of Workers for Reproduction In primitively eusocial species, workers have relatively higher reproductive potentials in comparison to those in highly eusocial species. In paper wasps, the reproductive potential of workers is correlated with the ranks of dominance (Paradi, 1996; Hunt, 2007). Several dominant workers lay unfertilized male eggs in a queenright colony. However, these male eggs may be removed by a queen and other workers before hatching of eggs, likely as queen and worker policing in Polistes chinensis (Saigo and Tsuchida, 2004). In queenless colonies, workers of the wasps with developed ovaries could succeed in the production of male individuals (Miyano, 1986).

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Workers in many species of highly eusocial species do not ordinary develop ovaries and exhibit egg laying behaviors. The absence of queens or larvae causes a low concentration of queen or larval pheromones in the colony, inducing development of ovaries in workers. Without exception, reproductive workers can not copulate and therefore they can lay only unfertilized male eggs. A colony in which workers are reproductive individuals becomes extinct if no female individuals are reproduced. Unless their sons copulate with a new queen from a different colony, the reproductive workers cannot enhance reproductive success. Although the transition of workers from helpers into reproductive individuals is a tactical change in reproduction, it is defined as a last resort under circumstances in which the colony cannot be reconstructed. Observation of the behaviors in honeybee workers with developed ovaries shows that cooperative relationships among individuals become combative, with some individuals losing their hair attacked by others or bitten on the wings (Sakagami, 1954; Evers and Seeley, 1986). Once a reproductive worker has developed ovaries, it starts secreting pheromones similar to those secreted by queens (Velthuis et al., 1965), suppressing the development of ovaries by other workers, allowing a few reproductive workers to monopolize egg laying. A behavioral sequence of oviposition in the reproductive workers is similar to that in queens. Reproductive workers, laying only male eggs, inspect comb cells as queens do and mostly lay eggs in male cells (Page and Erickson, 1988). In our experiments, reproductive workers select mainly male cells in the checkered frames when empty male cells are largely available (Figure 5A). However, when the empty male cells are limited, proportions of male cells used for oviposition gradually decrease (Figure 5B). This means that reproductive workers may select basically male cells for the oviposition and be obliged to lay unfertilized eggs in worker cells when empty male cells in the upper parts of area on the frame (these areas are maintained better condition for brood rearing) are not available.

Figure 5. Cell selections for laying unfertilized eggs by reproductive workers. A: an example of numbers of cells used for oviposition on the checkered frame, B: a negative correlation between total number of cells used and proportion of male cells used. Social Insects: Structure, Function, and Behavior : Structure, Function, and Behavior, edited by Emily M. Stewart, Nova Science Publishers,

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(b) Hormonal Factors Involving the Transition of Reproductive States in Workers

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Transition of reproductive states in workers occurring behaviorally is detected as a change in brain and nervous system activity. Environmental changes such as queenless condition materialize as changes of the frequency of queen‘s dominant behaviors or the concentration of queen pheromones in colonies. These changes could be detected by sensory organs and induce immediate neural activity via neurotransmitters, or sustainable activity or change in the humoral environment via release of neuromodulators or neurohormones. Another possible process is direct effects of queen pheromones on neural tissues via oral intakes of neuroactive substances in the pheromones (Beggs et al., 2007; Beggs and Mercer, 2009). This possibility has been reported in honeybees, but a question whether such neuroactive substances in queen pheromones exist in other highly eusocial species is still controversial.

Primitively Eusocial Species Gonadotropic effects of dopamine have been reported in primitively eusocial species. High dopamine concentration has been detected in reproductive workers‘ brain in the bumble bee Bombus terrestris (Bloch et al., 2000) and paper wasp Polistes chinensis (Sasaki et al., 2007). In P. chinensis, the brain levels of dopamine are correlated with not only ovarian development, but also egg laying behaviors among reproductive individuals with welldeveloped ovaries. Oral application of dopamine to queenless isolated workers in P. chinensis causes an acceleration of ovarian development (Sasaki et al., 2009). In Polistes females, juvenile hormone also shows gonadotropic effects (Bohm 1972; Röseler et al. 1980), suggesting neuro-endocrine interactions between brain dopamine and hemolymph juvenile hormone. Such an interaction has been suggested in male honeybees. In male honeybees, application of a juvenile hormone analog enhances brain dopamine levels (Harano et al., 2008). Both juvenile hormone and dopamine enhance the behavioral activities for mating in males (Giray and Robinson, 1996; Tozetto et al., 1997; Akasaka et al., 2010). Honeybees In female honeybees, juvenile hormone is believed to have lost its reproductive function (West-Eberhard, 1996; Hartfelder, 2000). Instead, juvenile hormone plays a role as a pacemaker hormone that regulates the division of labor among workers (Robinson and Vargo 1997; Page and Amdam, 2007). During transition from normal workers to reproductive individuals, tyramine, dopamine and dopamine metabolites (norepinephrine and N– acetyldopamine) increase in concentration in the brain (Harris and Woodring, 1995; Sasaki and Nagao, 2001, 2002). Recently, it has been reported that the queen mandibular pheromone decreases the levels of brain dopamine in workers and inhibits the expression of AmDOP1 gene mRNA, but not AmDOP2 and AmDOP3 gene mRNA (Beggs et al., 2007). In reproductive workers, the brain tyramine concentration rises first followed by the dopamine concentration (Sasaki and Nagao, 2002). When workers were experimentally transferred from a queenless colony to a queenright colony, the individuals stopped ovarian development, becoming intermediate reproductive workers. In such workers, the brain tyramine concentration is maintained at a value between normal and reproductive workers (Sasaki and

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Nagao, 2002). Tyramine injections into the head cause brain dopamine concentrations to rise (Sasaki and Harano, 2007). Continued tyramine or dopamine oral intake for 10–17 days causes the acceleration of ovarian development (Dombroski et al., 2003; Sasaki and Harano, 2007). It remains unknown whether the acceleration of ovarian development are caused by direct tyramine and/or dopamine effects on the ovaries or indirect effects through other gonadotropins. A direct effect of tyramine on ovarian development or ovulation is possible in the reproductive workers, because of down-expression of a tyramine receptor in ovaries when the ovarian development is inhibited (Thompson et al., 2007). Behavioral effects of phenolamines in honeybee workers may be different between queenright and queenless colonies. In queenright colonies, transition to foraging in workers involves changes in brain levels of octopamine (Taylor et al., 1992; Wagener-Hulme et al., 1999). Octopamine treatments accelerate the transition to foraging (Schulz and Robinson, 2001; Fussnecker et al., 2006). It seems that queenright workers convert tyramine into octopamine in the brains during aging, because the levels of tyramine in the brains decrease from 10 to 17 days of age (Sasaki and Nagao, 2002). In queenless colonies, tyramine levels in the brains of queenless bees are relatively greater than in queenright bees (Sasaki and Nagao, 2002). It is plausible that the high levels of tyramine in the brain of queenless workers may inhibit their foraging behavior (Schulz and Robinson, 2001; Fussnecker et al., 2006) and encourage them to stay in the nest for the transition to reproductive workers.

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CONCLUSION Queens in eusocial Hymenoptera can manipulate the sex of eggs by egg fertilization depending on season or colony condition. This ability has been maintained from ancestral solitary species. Sperm discharges could be controlled by actions of a spermathecal pump and/or spermathecal glands. The actions of these spermathecal tissues may be regulated by neuroactive substances including biogenic amines. Initiations of development in unfertilized eggs by mechanical distortions has been also maintained from ancestral species, but the portion providing mechanical stimulus in reproductive tracts may be converted from the ovipositor in Parasitica to the common oviduct in Aculeata. Queens in several eusocial species including honeybees can behaviorally control the sex ratio of eggs. They control egglaying behaviors associated with the egg fertilization (e.g., male-worker cell selections). Workers have an ability to change the reproductive states and produce unfertilized eggs under a special colony condition likely as in the absence of a queen. The transitions of workers to reproductive individuals are caused by neuro-endocrine mechanisms. The mechanisms may work for reproductive maturation in not only workers but also queens. Future studies confirming the mechanisms on the molecular levels should be done.

REFERENCES Akasaka, S. Sasaki, K., Harano, K. & Nagao, T. (2010). Dopamine enhances locomotor activity for mating in male honeybees (Apis mellifera L.). J. Insect Physiol., 56, 11601166.

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66

Ken Sasaki

Allen, M. D. (1963). Drone production in honeybee colonies (Apis mellifera L.). Nature, 199, 789-790. Aron, S., Passera, L. & Keller, L. (1994). Queen-worker conflict over sex ratio: A comparison of primary and secondary sex ratios in the Argentine ant, Iridomyrmex humilis. J. Evol. Biol., 7, 403-418. Aron, S., Vargo, E. & Passera, L. (1995). Initial and secondary sex ratios in monogyne colonies of the fire ant. Anim. Behav., 49, 749-757. Baer, B., Eubel, H., Taylor, N. L., O‘Toole, N. & Millar, H. (2009). Insights into female sperm storage from the spermathecal fluid proteome of the honeybee Apis mellifera. Genom. Biol., 10, R67. Beggs, K. T., Glendining, K. A., Marechal, N. M., Vergoz, V., Nakamura, I., Slessor, K. N. & Mercer, A. R. (2007). Queen pheromone modulates brain dopamine function in worker honey bees. Proc. Natl. Acad. Sci. USA, 104, 2460-2464. Beggs, K. T. & Mercer, A. R. (2009). Dopamine receptor activation by honey bee queen pheromone. Curr. Biol., 19, 1206-1209. Berg, S., Koeniger, N., Hoeniger, G. & Fuchs, S. (1997). Body size and reproductive success of drones (Apis mellifera L.). Apidologie, 28, 449-460. Beukeboom, L. W. (1995). Sex determination in Hymenoptera: a need for genetic and molecular studies. BioEssays, 17, 813-817. Beye, M., Hasselmann, M., Fondrk, M. K., Page, R. E. & Omholt, S. W. (2003). The gene csd is the primary signal for sexual development in the honeybee and encodes an SR-type protein. Cell, 114, 419-429. Bloch, G., Simon, T., Robinson, G. E. & Hefetz, A. (2000). Brain biogenic amines and reproductive dominance in bumble bees (Bombus terrestris). J. Comp. Physiol. A, 186, 261–268. Bohm, M. K. (1972). Effects of environment and juvenile hormone on ovaries of the wasp, Polistes metricus. J. Insect Physiol., 18, 1875-1883. Boomsma, J. J. & Grafen, A. (1991). Colony-level sex ratio selection in the eusocial Hymenoptera. J. Evol. Biol., 3, 383-407. Bresslau, E. (1905). Der Samenblasengang der Bienekӧnigin (Studien über den Geschlechtsapparat und die Fortpflanzung der Bienen. I.). Zool. Anz., 29, 299-323 (in German). Camargo, J. M. F. & Mello, M. L. S. (1970). Anatomy and histology of the genital tract, spermatheca, spermathecal duct and glands of Apis mellifera queens (Hymenoptera: Apidae). Apidologie, 1, 351-373. Charnov, E. L., Los-den Hartogh, R. L., Jones, W. T. & van den Assem, J. (1981). Sex ratio evolution in a variable environment. Nature, 289, 27-33. Clark, J. & Lange, A. B. (2000). The neural control of spermathecal contractions in the locust, Locusta migratoria. J. Insect Physiol., 46, 191-201. Clark, J. & Lange, A. B. (2001). Evidence of a neural loop involved in controlling spermathecal contractions in Locusta migratoria. J. Insect Physiol., 47, 607-616. Cole, L. R. (1981). A visible sign of a fertilization action during oviposition by an Ichneumonid wasp, Itoplectis maculator. Anim. Behav., 29, 299-300. Collins, A. M., Williams, V. & Evans, J. D. (2004). Sperm storage and antioxidative enzyme expression in the honey bee, Apis mellifera. Insect Mol. Biol., 13, 141-146.

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Regulation of Reproductive States and Control of Sex of Eggs…

67

Crane, E. (1990). Bees and beekeeping. Oxford: Heinemann Newnes. Crozier, R. H. (1971). Heterozygosity and sex determination in haplodiploidy. Am. Nat., 105, 399-412. Crozier, R. H. & Pamilo, P. (1996). Evolution of social insect colonies. Oxford: Oxford Univ. Press. da Silva, R. & Lange, A. B. (2008). Tyramine as a possible neurotransmitter / neuromodulator at the spermatheca of the African migratory locust, Locusta migratoria. J. Insect Physiol., 54, 1306-1313. Dallai, R. (1972). Fine structure of the spermathecal gland of Apis mellifera. Redia, 53, 413425. Dallai, R. (1975). Fine structure of the spermtheca of Apis mellifera. J. Insect Physiol., 21, 89-109. den Boer, S. P. A., Boomsma, J. J. & Baer, B. (2009). Honey bee males and queens use glandular secretions to enhance sperm viability before and after storage. J. Insect Physiol., 55, 538-543. Dobson, S. L. & Tanouye, M. A. (1998). Evidence for a genomic imprinting sex determination mechanism in Nasonia vitripennis (Hymenoptera; Chalcidoidea). Genetics, 149, 233-242. Dombroski, T. C. D., Simões, Z. L. P. & Bitondi, M. M. G. (2003). Dietary dopamine causes ovary activation in queenless Apis mellifera workers. Apidologie, 34, 281-289. Dzierzon, J. (1845). Gutachtern über die von Herrn Direktor Stӧhr in erstern und zweiten Kapitel des General-Gutachtens aufgestellten Fragen. Eichstädter Bienenzeitung, 1, 109113, 119-121 (in German). Evers, C. A. & Seeley, T. D. (1986). Kin discrimination and aggression in honey bee colonies with laying workers. Anim. Behav., 34, 924-944. Fisher, R. A. (1930). The genetical theory of natural selection. Oxford: Oxford Univ. Press. Flanders S. E. (1939). Environmental control of sex in hymenopterous insects. Ann. Ent. Soc. Am., 39, 11-26. Flanders, S. E. (1950). Control of sex in the honeybee. Sci. Month., 70, 237-240. Frank, S. A. (1987a). Individual population sex allocation pattern. Theor. Popul. Biol., 31, 4774. Frank, S. A. (1987b). Variable sex ratio among colonies of ants. Behav. Ecol. Sociobiol., 20, 195-201. Free, J. B. & Williams, I. H. (1975). Factors determining the rearing and rejection of drones by the honeybee colony. Anim. Behav., 23, 650-675. Fussnecker, B. L., Smith, B. H. & Mustard, J. A. (2006). Octopamine and tyramine influence the behavioral profile of locomotor activity in the honey bee (Apis mellifera). J. Insect Physiol., 52, 1083-1092. Fyg, W. (1943). Experimentelle Untersuchungen über den Eilegeakt der Bienenkӧnigin. Mitt. Schweiz. Ent. Ges., 18, 493-521 (in German). Fyg, W. (1966). On structure and function of the valve-fold of the queen honeybee (Apis mellifera L.). Z. Bienenforschung, 8, 256-266 (in German with English summary). Gempe, T., Hasselmann, M., Schiøtt, M., Hause, G., Otte, M. & Beye, M. (2009). Sex determination in honeybees: two separate mechanisms induce and maintain the female pathway. PLoS Biology, 7, e1000222.

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Copyright © 2011. Nova Science Publishers, Incorporated. All rights reserved.

68

Ken Sasaki

Gerber, H. S. & Klostermeyer, E. C. (1970). Sex control by bees: A voluntary act of egg fertilization during oviposition. Science, 167, 82-84. Gessner, B & Gessner, K. (1976). Inorganic ions in spermathecal fluid and their transport across the spermathecal membrane of the queen bee, Apis mellifera. J. Insect Physiol., 22, 1469-1474. Giray, T. & Robinson, G. E. (1996). Common endocrine and genetic mechanisms of behavioral development in male and worker honey bees and the evolution of division of labor. Proc. Natl. Acad. Sci. USA, 93, 11718-11722. Gösswald, K. & Bier, K. (1955). Beeinflussung des Geschlechtsverhältnisses durch Temperatureinwirkung bei Formica rufa L.. Naturwissenschaften, 42, 133-134 (in German). Hamilton, W. D. (1964). The genetical evolution of social behaviour, I. J. Theor. Biol., 7, 116. Hamilton, W. D. (1967). Extraordinary sex ratios. Science, 156, 477-488. Harano, K., Sasaki, K., Nagao, T. & Sasaki, M. (2008). Influence of age and juvenile hormone on brain dopamine level in male honeybee (Apis mellifera): Association with reproductive maturation. J. Insect Physiol., 54, 848-853. Harbo, J. R. (1976). The effect of insemination on the egg-laying behavior of honey bees. Ann. Entomol. Soc. Am., 6, 1036-1038. Harris, J. W. & Woodring, J. (1995). Elevated brain dopamine levels associated with ovary development in queenless worker honey bees (Apis mellifera). Comp. Biochem. Physiol., 111C, 271-270. Hartfelder, K. (2000). Insect juvenile hormone: from status quo to high society. Braz. J. Med. Biol. Res., 33, 157-177. Hasegawa, E. (1992). Annual life cycle and timing of male-egg production in the ant Colobopsis nipponicus (Wheeler). Insect. Soc., 39, 439-446. Hasselmann, M., Gempe, T., Schiøtt, M., Nunes-Silva, C. G., Otte, M. & Beye, M. (2008). Evidence for the evolutionary nascence of a novel sex determination pathway in honeybees. Nature, 454, 519-522. Hölldobler, B. & Wilson, E. O. (1990). The Ants. Cambridge, Massachusetts: Belknap Press. Hunt, J. H. (2007). The Evolution of social wasps. Oxford: Oxford Univ. Press. Kalogianni, E. & Theophilidis, G. (1993). Centrally generated rhythmic activity and modulatory function of the oviductal dorsal unpaired median (DUM) neurons in two orthopteran species (Calliptamus sp. and Decticus albifrons). J. Exp. Biol., 174, 123-138. Keller, L., Aron, S. & Passera, L. (1996). Internest sex-ratio variation and male brood survival in the ant Pheidole pallidula. Behav. Ecol., 7, 292-298. King, P. E. & Rafai, J. (1973). A potential mechanism for initiating the parthenogenetic development of eggs in a parasitoid Hymenopteran, Nasonia vitripennis (Walker) (Pteromalidae). Entomologist, 106, 118-120. Koeniger, N. (1970). On the natural disposition of the queen for distinguishing worker bee cells from male cells. Apidologie, 1, 115-142 (in German with English summary). Kressin, M., Sommer, U. & Schnorr, B. (1996). The spermathecal epithelium of the honey bee queen (Apis mellifera): ultrastructure, age-dependent alternation, and cellular junctions. Anat. Histol. Embryol. 25, 31-35 (in German with English summary). Laidlaw, H. H. (1944). Artificial insemination of the queen bee (Apis mellifera L.): Morphological basis and results. J. Morphol., 74, 429-465.

Social Insects: Structure, Function, and Behavior : Structure, Function, and Behavior, edited by Emily M. Stewart, Nova Science Publishers,

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Regulation of Reproductive States and Control of Sex of Eggs…

69

Lange, A. B. (2009) Tyramine: From octopamine precursor to neuroactive chemical in insects. Gen. Comp. Endocrinol., 162, 18-26. Michener, C. D. (1974). The social behavior of the bees. Cambridge, Massachusetts: Harvard Univ. Press. Miyano, S. (1986). Colony development, worker behavior and male production in orphan colonies of a Japanese paper wasp, Polistes chinensis antennalis (Hymenoptera: Vespidae). Res. Popul. Ecol., 28, 347-361. Nonacs, P. (1986). Ant reproductive strategies and sex allocation theory. Q. Rev. Biol., 61, 121. Okelo, O. (1979). Mechanisms of sperm release from the receptaculum seminis of Schistocerca vaga Scudder (Orthoptera: Acrididae). Int. J. Invert. Reprod., 1, 121-131. Pabalan, N., Davey, K. G. & Packer, L. (1996). Comparative morphology of spermathecae in solitary and primitively eusocial bees (Hymenoptera; Apoidea). Can. J. Zool., 74, 802808. Page, R. E. (1981). Protandrous reproduction in honey bees. Environ. Entomol., 10, 359-362. Page, R. E. & Metcalf, R. A. (1984). A population investment sex ratio for the honey bee (Apis mellifera L.). Am. Nat., 124, 680-702. Page, R. E. & Erickson, E. H. (1988). Reproduction by worker honey bees (Apis mellifera L.). Behav. Ecol. Sociobiol., 23, 117-126. Page, R. E., Fondrk, M. K. & Robinson, G. E. (1993). Selectable components of sex allocation in colonies of the honeybee (Apis mellifera L.). Behave. Ecol., 4, 239-245. Page, R. E. & Amdam, G. V. (2007). The making of a social insect: developmental architectures of social design. BioEssays, 29, 334-343. Pamilo, P. (1991). Evolution of colony characteristics in social insect. II. Number of reproductive individuals. Am.Nat., 138, 412-433. Paradi, L. (1996). Polistes: analysis of a society. In: S. Turillazzi, & J. West-Eberhard, (Eds.), Natural history and evolution of paper-wasps: 1-17, Oxford: Oxford Univ. Press. Poole, H. K. (1970). The wall structure of the honey bee spermatheca with comments about its function. Ann. Ent. Soc. Am., 63, 1625-1628. Ratnieks, F. L. W. & Keller, L. (1998). Queen control of egg fertilization in the honey bee. Behav. Ecol. Sociobiol., 44, 57-61. Retnakaran, A. & Percy, J. (1985). Fertilization and special modes of reproduction. In: G. A., Kerkut, & L. I. Gilbert, (Eds.), Comprehensive insect physiology biochemistry and pharmacology, 231-293, Oxford: Pergamon Press. Robinson, G. E. & Vargo, E. (1997). Juvenile hormone in adult eusocial Hymenoptera: gonadotropin and behavioral pacemaker. Arch. Insect Biochem. Physiol., 35, 559-583. Röseler, P. F., Röseler, I. & Strambi, A. (1980). The activity of corpora allata in dominant and subordinated females of the wasp Polistes gallicus. Insect. Soc., 27, 97-107. Ruttner, F. (1956). Zur Frage der Spermaübertrangung bei der Bienenkӧnigin. Insect. Soc., 3, 351-359 (in German with English summary). Ruttner, F. (1961). Die Innervation der Fortpflanzungsorgane der Honigbiene (Apis mellifica L.). Z. Bienenforschung, 5, 253-266 (in German with English summary). Ruttner, F. & Koeniger, G. (1971). The filling of the spermatheca of the honey bee queen: active migration or passive transport of the spermatozoa. Z. Vergl. Physiologie, 72, 411422 (in German with English summary).

Social Insects: Structure, Function, and Behavior : Structure, Function, and Behavior, edited by Emily M. Stewart, Nova Science Publishers,

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70

Ken Sasaki

Ruttner, F. (1988). Biogeography and taxonomy of honeybees. Springer-Verlag, Berlin. Saigo, T. & Tsuchida, K. (2004). Queen and worker policing in monogynous and monoandrous colonies of a primitively eusocial wasp. Proc. R. Soc. Lond. B (suppl.), 271, S509-S512. Sakagami, S. F. (1954). Occurrence of an aggressive behaviour in queenless hives, with considerations on the social organization of honeybee. Insect. Soc., 1, 331-343. Salt, G. (1965). Experimental studies in insect parasitism XIII. The haemocytic reaction of a caterpillar to eggs of its habitual parasite. Proc. R. Soc. Lond. B., 162, 303-318. Sasaki, K., Satoh, T. & Obara, Y. (1996). The honeybee queen has the potential ability to regulate the primary sex ratio. Appl. Entomol. Zool., 31, 247-254. Sasaki, K., Sobajima, H., Satoh, T. & Obara, Y. (1997). Activation in vitro of unfertilized egg development in honeybee queens. Naturwissenschaften, 84, 74-76. Sasaki, K. & Obara, Y. (1999). Honeybee queens lay fertilized eggs when no comb cells for oviposition are available. Zool. Sci., 16, 735-737. Sasaki, K. & Obara, Y. (2001). Nutritional factors affecting the egg sex ratio adjustment by a honeybee queen. Insect. Soc., 48, 355-359. Sasaki, K. & Nagao, T. (2001). Distribution and levels of dopamine and its metabolites in brains of reproductive workers in honeybees. J. Insect Physiol., 47, 1205-1216. Sasaki, K. & Nagao, T. (2002). Brain tyramine and reproductive states of workers in honeybees. J. Insect Physiol., 48, 1075-1085. Sasaki, K. & Obara, Y. (2002). Egg activation and timing of sperm acceptance by an egg in honeybees (Apis mellifera L.). Insect. Soc., 49, 234-240. Sasaki, K., Kitamura, H. & Obara, Y. (2004). Discrimination of larval sex ratio and timing of male brood elimination by workers in honeybees (Apis mellifera L.). Appl. Entomol. Zool., 39, 393-399. Sasaki, K. & Harano, K. (2007). Potential effects of tyramine on the transition to reproductive workers in honeybees (Apis mellifera L.). Physiol. Entomol., 32, 194-198. Sasaki, K., Yamasaki, K. & Nagao, T. (2007). Neuro-endocrine correlates of ovarian development and egg-laying behaviors in the primitively eusocial wasp (Polistes chinensis). J. Insect Physiol., 53, 940-949. Sasaki, K., Yamasaki, K., Tsuchida, K. & Nagao, T. (2009). Gonadotropic effects of dopamine in isolated workers of the primitively eusocial wasp, Polistes chinensis. Naturwissenschaften, 96, 625-629. Schulz, D. J. & Robinson, G. E. (2001). Octopamine influences division of labor in honey bee colonies. J. Comp. Physiol. A, 187, 53-61. Snodgrass, R. E. (1956). Anatomy of the honey bee. London: Cornell Univ. Press. Taylor, D. J., Robinson, G. E., Logan, B. J., Laverty, R. & Mercer, A. R. (1992). Changes in brain amine levels associated with the morphological and behavioral development of the worker honeybee. J. Comp. Physiol. A, 170, 715-721. Thompson, G. J., Yockey, H., Lim, J. & Oldroyd, B. P. (2007). Experimental manipulation of ovary activation and gene expression in honey bee (Apis mellifera) queens and workers: testing hypotheses of reproductive regulation. J. Exp. Zool., 307A, 600-610. Tozetto, S. O., Rachinsky, A. & Engels, W. (1997). Juvenile hormone promotes flight activity in drones (Apis mellifera carnica). Apidologie, 28, 77-84. Trivers, R. L. (1974). Parent-offspring conflict. Am. Zool., 14, 249-264.

Social Insects: Structure, Function, and Behavior : Structure, Function, and Behavior, edited by Emily M. Stewart, Nova Science Publishers,

Copyright © 2011. Nova Science Publishers, Incorporated. All rights reserved.

Regulation of Reproductive States and Control of Sex of Eggs…

71

Trivers, R. L. & Hare, H. (1976). Haplodiploidy and the evolution of the social insects. Science, 191, 249-263. Ueno, T. (1995). Abdominal tip movements during oviposition by two parasitoids (Hymenoptera: Ichneumonidae) as an index of predicting the sex of depositing eggs. Appl. Entomol. Zool., 30, 588-590. Velthuis, H. H. W., Verheijen, F. J. & Gottenbos, A. J. (1965). Laying worker honey bee: similarities to the queen. Nature, 207, 1314. Vinson, B. S. & Jang, H. (1987). Activation of Campoletis sonorensis (Hymenoptera: Ichneumonidae) eggs by artificial means. Ann. Ent. Soc. Am., 80, 486-489. Wagener-Hulme, C., Kuehn, J. C., Schlz, D. J. & Robinson, E. G. (1999). Biogenic amines and division of labor in honey bee colonies. J. Comp. Physiol. A, 184, 471-479. Went, D. F. & Krause, G. (1973). Normal development of mechanically activated, unlaid eggs of an endoparasitic Hymenopteran. Nature, 244, 454-455. Went, D. F. & Krause, G. (1974). Egg activation in Pimpla turionellae (Hym.). Naturwissenschaften, 61, 407-408. Went, D. F. (1982). Egg activation and parthenogenetic reproduction in insects. Biol. Rev., 57, 319-344. West-Eberhard, M. J. (1996). Wasp societies as microcosms for the study of development and evolution. In: S. Turillazzi, & J. West-Eberhard, (Eds.), Natural history and evolution of paper-wasps: 290-317, Oxford: Oxford Univ. Press. Wharton, K. E., Dyer, F. C., Huang, Z. Y. & Getty, T. (2007). The honeybee queen influences the regulation of colony drone production. Behav. Ecol., 18, 1092-1099. Wheeler, D. E. & Krutzsch, P. H. (1994). Ultrastructure of the spermatheca and its associated gland in the ant Crematogaster opuntiae (Hymenoptera, Formicidae). Zoomorphology, 114, 203-212. Whiting, P. W. (1943). Multiple alleles in complementary sex determination of Habrobracon. Genetics, 28, 365-382.

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In: Social Insects: Structure, Function, and Behavior Editor: Emily M. Stewart

ISBN: 978-1-61761-466-8 © 2011 Nova Science Publishers, Inc.

Chapter 4

THE GLOBAL EMPIRE OF AN INVASIVE ANT

1

Eiriki Sunamura1, Hironori Sakamoto2, Shun Suzuki1, Koji Nishisue1, Mamoru Terayama1 and Sadahiro Tatsuki1

Graduate School of Agricultural and Life Sciences, University of Tokyo, Bunkyo-ku, Tokyo 113-8657, Japan 2 Graduate School of Environmental Sciences, Hokkaido University, Kita-ku, Sapporo 060-0808, Japan

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ABSTRACT Social insects have obtained their prosperity by cooperation among individuals. This can be applied particularly to the success of invasive ants, which form unusual social structure called supercolonies, within which individuals can move freely among physically separated nests, and thereby gain high population densities to dominate indigenous ants. Native to South America, the Argentine ant Linepithema humile has been unintentionally introduced into many parts of the world during the last 150 years. Although it is well known that the introduced Argentine ant populations form much larger and fewer supercolonies than the native populations, the relationship among beyond-ocean populations has been poorly understood. Recent studies, however, are uncovering the behavioral, chemical and genetic relationships among introduced Argentine ant populations worldwide. Individuals from the dominant supercolonies around the world have very similar cuticular hydrocarbon profiles (nestmate recognition cue), and do not show aggressive behavior toward each other, when artificially put into contact. The supercolonies constitute the largest cooperative unit ever known. Their genetic closeness suggests a common introduction pathway. Considering historical records, descendants of the most ancient introduced population have spread to many parts of the world, without losing memory of their roots. In this chapter, we introduce the nestmate recognition system and mechanism of supercolony expansion in invasive ants, with the global empire of Argentine ants as an example.

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Eiriki Sunamura, Hironori Sakamoto, Shun Suzuki et al.

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INTRODUCTION Social animals have obtained their prosperity by evolving cooperative behavior among individuals. This can be applied particularly to invasive alien ants. About 150 species of ‗tramp‘ ants have been brought out of their native ranges by human activities, with some becoming invasive and causing damage to ecosystems, economic resources and human wellbeing [McGlynn 1999]. The most damaging species are shown in Table 1. Many invasive ants, including these species, are unicolonial, and form large networks of numerous, mutually cooperative (non-aggressive) nests, called ‗supercolonies‘ [Holway et al. 2002]. A mature supercolony can extend over several hundred meters, and sometimes exceeds thousands of kilometers [Tsutsui et al. 2000]. The most extreme case is the Argentine ant, which forms an intercontinental, global-scale supercolony [Sunamura et al. 2009a; van Wilgenburg et al. 2010]. Compared to most native ants whose colonies are composed of one or a limited number of nests and territorial aggression among closely located nests are common, invasive ants that form supercolonies can invest more on reproduction, because of the lower cost associated with territorial aggression [Holway et al. 1998; Holway 1999]. With the numerical advantage gained by this high reproductive ability, invasive ants overwhelm native ants and almost completely displace them in the infested area [Holway et al. 2002]. The direct displacement of arthropod fauna including native ants causes indirect negative impacts on many other taxa related to them [Holway et al. 2002; Lach 2003; Ness and Bronstein 2004; Krushelnycky et al. 2005; Lach and Thomas 2008; Sunamura et al. in press]. Sometimes the impact can be so huge that the landscape is altered. In the tropical rainforest of Christmas Island, the displacement of the red land crab by the yellow crazy ant has turned the open forest floor into a bush with a dense and diverse cover and thick litter layer [O‘Dowd et al. 2003]. The high density of invasive ants also leads to serious problems for human life. Invasive ants cause agricultural damage by protecting aphids, scale insects and mealybugs from their natural enemies, or by stinging farmers and livestock [Wetterer and Porter 2003; Pimentel et al. 2005; Wetterer 2007; Sunamura et al. in press]. In addition, they become nuisance pests that invade structures with high frequency and sanitary pests by biting or stinging [e.g., Rhoades et al. 1971]. In one sense, ―ant mega-colony takes over world‖ [Walker 2009]. In this chapter we review the nestmate recognition of invasive ants and the process of the global supercolony formation by Argentine ants.

NESTMATE RECOGNITION SYSTEM Many ant species use cuticular hydrocarbons as nestmate recognition cue [Howard and Blomquist 2005; Martin and Drijfhout 2009]. Cuticular hydrocarbon profile (compounds and their relative abundance) differs among both species and conspecific colonies. When an individual encounters another conspecific, it detects the cuticular hydrocarbons of the opponent by its antennae, and compares the profile with the experience-based neural template of the nestmates [Ozaki et al. 2005]. When they match, the individual recognizes the opponent as a nestmate, but when they vary, it recognizes the opponent as a foreigner and attacks it or runs away. All of the hydrocarbon compounds present on the ant exoskeleton are not necessarily involved in nestmate recognition, but relative proportion of multiple

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The Global Empire of an Invasive Ant

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components is important. Cuticular hydrocarbons can be derived from both genetic and environmental (e.g., diet and nest material) factors [e.g., Crosland 1989; Beye et al. 2004; Sorvari et al. 2008; van Zweden et al. 2009]. Relative contribution of genetic and environmental factors varies from species to species. Hydrocarbons are shared and homogemized among colony members via grooming [Soroker et al. 1994; Meskali et al. 1995]. Chemicals other than hydrocarbons may be used as nestmate recognition cue, but knowledge on such chemicals is currently scarce. Invasive ants may also use cuticular hydrocarbons for nestmate recognition. Direct evidence for the use of hydrocarbons has been obtained in Argentine ants [Liang and Silverman 2000], and similarity within or disparity between supercolonies have been reported in some other species [Errard et al. 2005; Cremer et al. 2008; Fournier et al. 2009]. Relative influence of genetic and environmental factors to cuticular hydrocarbon profile also varies among invasive ant species. For instance, in Argentine ants and big headed ants, genetically close populations originated from single introductions maintain similar hydrocarbon profiles even after dispersing over several thousand kilometers of diverse environmental condition for decades [Tsutsui et al. 2000; Fournier et al. 2009]. This suggests that environmental factors do not strongly affect cuticular hydrocarbon profile of these species in the field. A similar pattern was reported in little fire ants, in which clonal reproduction occurs [Errard et al. 2005; Fournier et al. 2005]. On the contrary, in garden ants, variation in cuticular hydrocarbon profile and the resultant aggression is observed even among populations presumably originated from a common source population [Cremer et al. 2008; Ugelvig et al. 2008]. In this species, effect of environment on cuticular hydrocarbon profile may be relatively strong. In short, an invasive ant population with lower genetic variation and lower susceptibility to environment for hydrocarbons may be able to form larger supercolonies. As a unique case, whether supercolonies are formed or not depends on a single gene or linkage group in fire ants [Keller and Ross 1998]. Table 1. Representative invasive alien ant species and their distributions. All of them but the garden ant are listed among the world’s 100 worst invasive species by the IUCN (International Union for Conservation of Nature) [Lowe et al. 2000] Species Argentine ant Linepithema humile

Native range South America

Big-headed ant Pheidole megacephala

Africa or Asia?

Garden ant Lasius neglectus Little fire ant Wasmannia auropunctata Red imported fire ant Solenopsis invicta Yellow crazy ant Anoplolepis gracilipes

Middle East? Middle and South America Middle and South America Africa or Asia?

Introduced range Africa, Asia, Australia, Europe, North America, Oceanic Islands (Atlantic and Pacific) Africa, Australia, North America, South America, Oceanic Islands (Indian and Pacific) Europe Africa, Australia, North America, Oceanic Islands (Pacific) Asia, Australia, North America Africa, Asia, Australia, Oceanic islands (Indian and Pacific)

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GLOBAL-SCALE SUPERCOLONY OF ARGENTINE ANTS Argentine ants are native to the Parana River drainage of Argentina, Brazil, Uruguay and Paraguay [Wild 2004, 2007]. In the native habitat, frequent disturbance, namely flooding, occurs [LeBrun et al. 2007]. This might have been important for the evolution of supercolony in Argentine ants. Unicolonial species, in which queens walk out of their natal nests accompanied by workers and establish new nests, can rapidly recover from frequent disturbance and thus are more adaptive to such environment than typical ant species in which new queens disperse for a long distance by nuptial flight and establish new nests without the aid of workers [Tsuji and Tsuji 1996; Nakamaru et al. 2007]. Interestingly, the native range of Argentine ants overlaps those of other invasive unicolonial ants such as Solenopsis fire ants and little fire ants [LeBrun et al. 2007]. With the development of human commerce, Argentine ants have been unintentionally introduced to many parts of the world during the last 150 years [Suarez et al. 2001; Wetterer et al. 2009]. Their opportunistic nesting behavior [Vega and Rust 2001] and polygyny [Keller et al. 1989] may enhance the opportunity of human-mediated dispersal of nest fragment, and the probability of inclusion of reproductive queens in it [Suarez et al. 2008]. First recorded from Madeira somewhere between 1847 and 1858, Argentine ants landed Europe (Portugal and France), North America (Louisiana and California), and Africa (South Africa) during 1890-1910, Central America (Mexico and Bermuda) and Australia (Victoria, Western Australia, New South Wales, and Tasmania) around 1940-1950, and finally Asia (Japan) in 1993 [Suarez et al. 2001; Wetterer et al. 2009]. Within each continent, country or island, Argentine ants have rapidly expanded their distribution via human-mediated jump dispersal, sometimes at a rate of > 100 km/yr [Suarez et al. 2001; Ward et al. 2005; Okaue et al. 2007; Blight et al. 2009; Pitt et al. 2009; Roura-Pascual et al. 2009], although their unaided dispersal is rather slow (colony budding on foot;