Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances [1 ed.] 9781624175015, 9781612096346

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Copyright © 2011. Nova Science Publishers, Incorporated. All rights reserved. Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and Patrick

Copyright © 2011. Nova Science Publishers, Incorporated. All rights reserved. Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

BOTANICAL RESEARCH AND PRACTICES

POLLINATION

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

MECHANISMS, ECOLOGY AND AGRICULTURAL ADVANCES

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Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

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Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

BOTANICAL RESEARCH AND PRACTICES

POLLINATION MECHANISMS, ECOLOGY AND AGRICULTURAL ADVANCES

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NICHOLE D. RASKIN AND

PATRICK T. VUTURRO EDITORS

Nova Science Publishers, Inc. New York Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

Copyright © 2012 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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Library of Congress Cataloging-in-Publication Data Pollination : mechanisms, ecology and agricultural advances / editor: Nichole D. Raskin and Patrick T. Vuturro. p. cm. Includes bibliographical references and index. ISBN:  (eBook) 1. Pollination. I. Raskin, Nichole D. II. Vuturro, Patrick T. QK926.P595 2011 571.8'642--dc22 2011003553

Published by Nova Science Publishers, Inc. †New York Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

CONTENTS Preface Chapter 1

Chapter 2

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Chapter 3

vii Pollination Strategies: The Point of View of Plants - Personal Experiences and Review of the Literature Guido Flamini Pollination Mechanisms in Passiflora Species: The Common and the Rare Flowers - Ecological Aspects and Implications for Horticulture M. T. Amela García and P. S. Hoc Pollen Tube Growth and Ovule Abortion in Olea Europaea (Oleaceae): A Case of Ovule Selection? Julián Cuevas, Luis Rallo and Hava F. Rapoport

Chapter 4

Application of Airborne Pollen Data to Agronomical Research H. García-Mozo

Chapter 5

2n Pollen Formation: 40 Cytological Mechanisms of Nuclear Meiotic Restitution Nataliya V. Shamina

Chapter 6

Orbicules in Relation to the Pollination Modes B. G. Galati, M. M. Gotelli, S. Rosenfeldt, J. P. Torretta and G. Zarlavsky

Index

Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

1

33

57 73

85 149

165

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PREFACE This new book presents topical research in the study of the mechanisms, ecology and agricultural advances of pollination. Topics discussed include pollination strategies; the pollination mechanisms in common and rare flowers of the Passiflora species; pollen tube growth and ovule abortion in OleaEuropaea (Oleaceae); the application of airborne pollen data to agronomical research and orbicules in relation to pollination modes. Chapter 1 – Pollinators provide an essential service to both natural and agricultural ecosystems. More than 80% of crop production, that is the majority of fruit, vegetable, oil plant, protein plant, nut and spices depend on insect pollination. It is known that the color of a flower is the first and foremost cue for pollinator's attraction, but the scent of a flower also plays a major role in attracting pollinating insects. Other plants, such as some orchids, use deceptive pollination methods producing hormonal-like substances. Studies of floral scents and of their patterns within a single flower are important to better understand the chemical bases of plant-animal relationships and pollination ecology. Furthermore, they may reveal new scented molecules that could be of value to both the food industry and perfumery. Besides volatiles, nutrients may also be used by plants as a reward for pollination services. Sugars, amino acids and secondary metabolites are contained in nectar, a liquid secretion produced by dedicated structures within the flower and highly appreciated by insects and other pollinators. Chapter 2 – Passionvines have flowers with the following basic architecture: 5 sepals, 5 petals, a corona formed by concentric cycles (radii, pali, operculum, limen) and an androgynophore that bears 5 anthers, the ovary, 3 styles and 3 stigmas. Self-pollination may be achieved but some species are self-incompatible, so pollen vectors are required. The different relative sizes and orientation of the floral pieces of the various Passiflora species have implications on which visitors will pollinate; to perform pollination, they must have the adequate size to contact both anthers and stigmas in the same or in successive visits to different flowers. Pollen removal (from the anthers) and deposition (in the stigmas) is carried out by means of different parts of the body of the different pollinators, depending on their size and behaviour. The anthers are dehiscent and the stigmas are receptive as soon as the flower opens until it closes. The styles move throughout anthesis: they tilt down to the anthers and uplift afterwards. Thus, three floral stages occur: in the first and the third, only the anthers can be contacted by

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Nichole D. Rskin and Patrick T. Vuturro

the legitimate visitors while in the second, both the anthers and the stigmas are placed in the way of the pollinators. The style movements succeed in all the studied species. However, in some species, in a proportion of the flowers the styles remain upright since the flowers open. These flowers are not able to receive pollen, neither by the pollinators nor by themselves, so they are functionally staminate. In fewer species, the dehiscence of the anthers does not happen in some flowers, so they are functionally pistillate. Finally, the three types of flowers may coexist in the same plant. This brings about the simultaneous occurrence of pollen donorreceptor flowers and only pollen donors, pollen donor-receptor and only receptor flowers or the three types of flowers in a single plant, respectively, leading to the corresponding functionally andromonoecious, gynomonoecious or trimonoecious systems. Certain floral traits seem to be associated with the absence of styles movements, such as a less developed gynoecium, minor-sized and nectarless flowers. In this chapter, an update of the recorded aspects at the moment as well as original data are discussed, taking in account the ecological interpretations of style movements, analysing the possible causes of the incidence of the less frequent flowers and considering the implications for fruit production in this edible fruited genus, some species of which are grown commercially. Chapter 3 – A mature olive tree may produce as many as 500,000 flowers, but only 1-2% of them become fruit. Female reproductive success is further diminished since olive flowers contain four ovules, but only one becomes a seed. We studied the roles of pollen tube growth and ovule abortion in determining this single-seeded pattern. To differentiate between the effects of syngamy and seed growth on ovule abortion we compared the onset and progression of ovule senescence in fruiting and fruiting-prevented plants, in fertilized versus unfertilized flowers, and in plants with different rates of seed growth using aniline blue fluorescence. The single-seeded fruit condition in olive is the result of only one pollen tube reaching the ovary and syngamy occurring therefore in only one ovule per flower. The syngamy in one ovule is accompanied by the degeneration of the other three ovules. Rather than a passive process, the onset and progress of unfertilized ovule degeneration of unfertilized ovules are hastened by the growth of the fertilized ovule. The effect of the fertilized ovule is more pronounced within the same ovary, but is also noted in other unfertilized flowers of the same plant. The Effective Pollination Period was shortened in flowers growing near developing fruitlets as a result of the reduced ovule longevity, suggesting that fertilized flowers may reduce the chance of syngamy in other flowers by this mechanism. These results are discussed in regard to ovule and fruitlet competition, dispersal mode and evolutionary forces underlying the observed seed/ovule ratio. Chapter 4 – Pollination is only one of the many events comprising the plant development cycle; however, it is extremely important for yield where seed is required. Although successful fertilization depends on a number of environmental and endogenous factors, including climate and plant nutritional status, a sufficient quantity of pollen must reach the receptive stigma in order to enhance fertilization potential. Aerobiological research focuses on the airborne dispersal of biological particles, including pollen grains from anemophilous plants. Airborne pollen data are currently used for various purposes in agricultural research. One major use is as a source of advance information concerning variations in the final fruit harvest of wind-pollinated species. This application, first introduced in the field of plant pathology in

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Preface

ix

the 1940s, was further developed in the 1970s in French studies of vineyard yield; more recently, it has been successfully tested both in crops and in non-crop forest species such as oak or birch. Nowadays, aerobiological research into the influence of pollen emission on final fruit production takes into account a number of other variables, including weather-related factors and phytopathological data; it also uses new, computerized statistical tools to obtain more precise information on agricultural yield and phytopathological risks. Chapter 5 – The illustrated catalogue of meiotic division abnormalities, preferably cytoskeleton aberrations in karyo- and cytokinesis, leading to 2n gametes formation in plants; includes 40 meiotic restitution mechanisms in pollen mother cells. Chapter 6 – Orbicules or Ubisch bodies are corpuscles of sporopollenin that appear in the anther locule during pollen grain development. Their size ranges from 0.14 µm to 20 µm. They present different shapes with a smooth or ornamented surface. Orbicules often form aggregates and sometimes have a plaque-like appearance. Ultrastructurally, they may present a central core with different degree of transparency to electrons. Those that do not have a central core are observed completely solid. Orbicules are resistant to acetolisis, autofluorescent when irradiated with ultraviolet light and have the same reaction to colorants that the exine of pollen grains. Their presence is generally associated with a tapetal membrane in species with secretor type tapetum and with a peritapetal membrane in species with intermediate or plasmodial type tapetum. Although the shed of orbicules out of the anther along with the pollen grains is cited, they are usually attached to the inner surface of the locule when the anther opens. Investigations suggest that orbicules appear in approximately 80 families of Angiosperms and Gimnosperms. It is not certain whether orbicules are not developed in the rests of the families or are just not informed. Researches on ontogeny and ultrastructure of orbicules are rare. However, their tapetal origin and their simultaneous formation with the pollen grain wall are well established. The systematic value of orbicules is known and considered in a few families, such as Loganiaceae, Gentianaceae, Apocynaceae, Rubiaceae and Oxalidaceae. Evolutionary studies on these bodies or on its relationship with the different modes of pollination are lacking. Even though orbicules are so common among angiosperms, their function is unknown and only speculations are made. On this report a review on orbicules is made and an analysis of their presence, ontogeny and morphology is presented. Our aim is to supply information that will help understand orbicules function. Therefore, the orbicules morphology in relation with the pollination mode is studied.

Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

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In: Pollination Editors: N. D. Raskin and P. T. Vuturro

ISBN: 978-1-61209-634-6 ©2012 Nova Science Publishers, Inc.

Chapter 1

POLLINATION STRATEGIES: THE POINT OF VIEW OF PLANTS -PERSONAL EXPERIENCES AND REVIEW OF THE LITERATURE Guido Flamini Dipartimento di Scienze Farmaceutiche, Sede di Chimica Bioorganica e Biofarmacia, Pisa, Italy

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ABSTRACT Pollinators provide an essential service to both natural and agricultural ecosystems. More than 80% of crop production, that is the majority of fruit, vegetable, oil plant, protein plant, nut and spices depend on insect pollination. It is known that the color of a flower is the first and foremost cue for pollinator's attraction, but the scent of a flower also plays a major role in attracting pollinating insects. Other plants, such as some orchids, use deceptive pollination methods producing hormonal-like substances. Studies of floral scents and of their patterns within a single flower are important to better understand the chemical bases of plant-animal relationships and pollination ecology. Furthermore, they may reveal new scented molecules that could be of value to both the food industry and perfumery. Besides volatiles, nutrients may also be used by plants as a reward for pollination services. Sugars, amino acids and secondary metabolites are contained in nectar, a liquid secretion produced by dedicated structures within the flower and highly appreciated by insects and other pollinators.

Pollination is the process by which the pollen grains, containing the male gametes, are transferred from the anthers of a flower to the stigma of another (or of the same) flower. This process permits the fertilization of the ovules and sexual reproduction in plants. Even if the role of pollen was hypothesized since ancient times (Cox, 1988), the first modern study that evidenced sexual reproduction in plants was the “De sexu plantarum epistola” (=Letter about sexuality in plants) by the German botanist Camerarius, published in

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Guido Flamini

1694. It was a milestone, not only for botanists, but also for the whole biology (Zarsky and Tupy, 1995). Thus, pollination is a prerequisite for fertilization and seeds development. However, pollen can travel up to the stigma in many different ways. Apart from self-pollinating plants, the pollen can be transported both by abiotic and biotic vectors. Among the formers, wind plays a major role. Wind-pollinated plants produce small and light pollen grains, often containing 1-3 air-filled bladders, or sacci, that confer aerodynamics to the grain, thereby increasing its dispersal distance (Schwendemann et al., 2007). Generally, anemophily is common in gymnosperms. However, in some angiosperm families (i.e. Poaceae) and in some environments (i.e. some deciduous forests), it is the dominant method of pollination (Whitehead, 1969). It is considered the ancestral state in gymnosperms, whereas in angiosperms it is a derived condition because of pollinator limitation or changes in the abiotic environment (Culley et al., 2002). Sometimes, in angiosperm, wind pollination appears in combination with animal pollination (ambophily), with relative frequencies varying from mostly wind pollinated species to mainly animal pollinated ones. Ambophily could be an adaptation to local environments that vary spatially in conditions that favor either wind or biotic pollination (Culley et al., 2002). Wind pollination is less efficient than animalpollination because of its random nature: large amounts of pollen are required to increase the probability of gametes encounter and the process may produce metabolically wasteful results. In fact, successful capture is limited, in part, by the very small dimensions of pollen grains and of female targets. Furthermore, the pollen grains tend to follow the wind as it flows around female structures, avoiding capture. To improve the wind-pollination efficiency, electrostatic forces play an important role, particularly for smaller pollen grains (Bowker and Crenshaw, 2007). The other main abiotic vector of pollen is water (hydrophily). Pollen can be transported both above and beneath the water surface (Ackerman, 2000). Underwater pollination represents the extreme adaptation to the aquatic environment. In this type of pollination the flower is submersed and the pollen is released in water, and both pollen and stigma are functionally wet during pollination (Philbrick and Les, 1996). In most terrestrial angiosperms, transfer of pollen to stigma is disturbed by water. Furthermore, hydration is one of the major causes of pollen germination on the stigma. The reproductive structures of hydrophilous plants resemble in many aspects those of anemophilous ones, such as loss or reduction of corolla, high pollen production and low ovule number per ovary (Philbrick and Les, 1996). For most submersed plants, pollen is elongated and can reach remarkable lengths, up to 5 mm. This shape has been interpreted as ―search vehicles‖ (Cox, 1993). When flowers are at water surface level, pollen moves on a two-dimensional space, so a better pollination efficiency can be reached. However, plants live in a world populated by many other living organisms and constantly interact with them: the animals. The most common animal biotic pollen vectors are insects, but also birds, bats and some other small vertebrates can play an important role. The topic of animal pollination is very complicated, mainly because of the many complex interactions between plants and animals. These interactions can vary in nature, being both visual and chemical. In addition, the scope of these signals can be different: they may be directed to mutualist attraction or they may serve as a defense against pests and herbivores. Mutualism is a biological interaction between two different species of organisms in which both parties benefit from the association. One of these benefits for plants is pollination. Even if beneficial

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Pollination Strategies

3

for both the organisms, it should be noted that a plant and an animal have different needs: the former prefers to minimize the production of pollen and its dispersion to different species and to reduce waste of energy for nectar production. On the contrary, animals desire to conserve energy spending more time on flowers with a large reward, reducing pollination efficiency. Possibly, if the reward is insufficient, they can even forage on other species. According to some authors, pollination ecology is the result of a coevolutionary process between plants and pollinators that led to suites of convergent floral traits to adapt plants to certain pollen vectors, the so-called ‗pollination syndrome‘ (Delpino, 1873; Stebbins, 1974; Johnson et al., 1998; Pellmyr, 2002). These interactions would act as selective forces that lead to phenological adjustments. This is particularly true in obligate mutualisms, in which each of two partner species can survive and/or reproduce only if it successfully locates the other (Law et al., 2001). Nevertheless, highly specialized interactions may be harmful to plants: it could lead to set no seed at all if the pollinator is absent in any one season because of disease or adverse climate, a tragic situation for an annual plant and very detrimental for a perennial one. For all these reasons, it seems more likely that, in general, the coevolution of plants and their pollinators has followed a middle way, with groups of pollinators coevolving with groups of flowers, rather than one-on-one interactions. These broad interactions could constitute a pollination syndrome (Glover, 2007). As previously mentioned, animal pollination service is not given gratis. In return for pollen transfer, plants provide food to their pollinators in the form of nectar and pollen, highly desirable food sources because of their richness in sugars, proteins and other essential elements (Schoonhoven et al., 2005; Chacoff et al., 2006). While collecting nectar and pollen, animals inadvertently brush against the fertile parts of the flower, causing the transfer of pollen from stamens to their bodies and then to stigmas. Plants therefore need to advertise their rewards to attract pollinators. It is known that the color of a flower is the first and foremost cue for pollinators‘ attraction, but the scent of a flower also plays a major role in attracting pollinating insects (Dobson, 1994; Dobson and Bergstrom, 2000b). Probably, visual signals are more effective than olfactory ones for long-distance signaling (Schaefer et al., 2004). The visual signal of the colored corollas of the flowers is generally enhanced by the contrast with the green leafy background. This is also supported by recent studies on insect vision (Kevan et al., 1996). In nature, flowers show a wide range of colors, each with its distinctive hue, brightness and saturation. While not an absolute rule, each animal prefers a limited kind of color, i.e. honey bees seem to favor flowers that to us appear to be yellow or blue. It is important to note that animals may recognize colors in a different way than humans. In particular, they can perceive electromagnetic radiations outside the human visible spectrum, mainly in the UV frequencies. This can explain why, when food is in short supply, honey bees sometimes also visit the flowers of red poppies, although they are almost insensitive to red. In fact, in the petals of the poppies, some UV-absorbing flavonoids are contained (Arnold et al., 2008). In the case of flies, flowers are often cream or yellow; butterflies have good color vision, so the flowers they visit are usually brightly colored, with reds and yellows often predominant. Since color vision is not very important for beetles, most of the flowers they visit are scarcely apparent, often greenish or white; the same is true for moth-pollinated flowers, since most moths are active at night (Glover, 2007). In the case of vertebrates, these animals have good color vision. Flowers pollinated by birds are usually red, often with contrasting yellow marks. On the contrary, bats are color-blind, so flower color is irrelevant for their attraction, and usually are white to cream or greenish (Glover, 2007). The

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importance of colored flowers is demonstrated by the discrimination of albino variants by pollinators that results in a lower seed-set (Waser and Price, 1981, 1985). The color of an object, including flowers, depends on the electromagnetic radiations that it reflects. Alternatively, it may be caused by the interference of light reflected from complex surfaces (iridescence). The color of a flower is usually due to the presence of particular pigments synthesized by the plant. These pigments can be classified in three main classes: carotenoids, flavonoids and betalains. The most common types are carotenes and xanthophylls for carotenoids, anthocyanins, aurones, chalcones, flavonols and proanthocyanidins for flavonoids, and betacyanins and betaxanthins for betalains. Carotenoids are C-40 isoprenoids that give yellow to orange colors, but they are not very important pigments for flowers, being more distributed in fruits and in chloroplasts. Betalains also have a limited distribution in flowers being restricted to the suborder Chenopodiniae within the order Caryophyllales (Clement and Mabry, 1996). They are glycoside indole derivatives biosynthesized from tyrosine. Betacyanins are responsible for red to purple colors, while betaxanthins for yellow to orange ones. Probably, flavonoids are the most important flower pigments. All of them absorb in the near UV (340 to 380 nm) and some even in the visible (520 to 550 nm) spectrum, so they can be perceived by all animals, including those that extend their vision in the UV region. Among flavonoids, anthocyanins are the most important pigments that contribute to the color of flowers, providing most of the pink, orange, red, violet and blue colors. Chalcones, aurones (collectively known as anthochlor flavonoids) and some flavonols contribute sometimes to the yellow color. The remaining flavonoids are usually colorless to man, but may appear visible to some insects. For all the above pigments, the different number and type of substituents linked to the main skeleton cause a different hue or color intensity. However, the production of a single pigment, or even of a single class of pigments, is an extremely rare case in nature. Typically, a number of pigments are synthesized by the plant and these associations can show striking effects on the resulting color. Apart from new hues deriving from the simple mixing of colors, even colorless substances, when associated with pigments, may exert substantial modification of the original color (co-pigmentation). This behavior explains the almost infinite shades that a relatively few number of pigments, anthocyanins in particular, may impart to flowers. Often there is also the formation of colored supramolecular complexes between several molecules of the same compound, via specific - stacking interactions of the flat aromatic surfaces (Goto and Kondo, 1991; Ellestad, 2006). Co-pigmentation and supramolecular complexes explain why anthocyanins are colored at the physiological pH: in this way, the colored flavylium cation is stabilized with respect to the corresponding uncolored hemiacetal or chalcone forms that should be present at cellular pH values (Santos et al., 1993; Ferreira da Silva et al., 2005). Furthermore, some metal ions, mainly Al3+ and Fe3+, can form colored complexes (adducts) when in the molecule of the pigment a catecholate moiety is present. This event has been demonstrated i.e. for the Himalayan blue poppy (Meconopsis grandis), Phacelia campanularia and Hydrangea macrophylla (Toyama-Kato et al., 2003; Mori et al., 2006; Yoshida et al., 2006). Apart from all the above reasons and from easily predictable concentration effects, the intensity of the color of a flower can also be affected by the shape of the epidermal cells, particularly those facing pollinators. It has been observed that in a conical shaped cell the light is reflected into the cell, enhancing the intensity of pigmentation and, consequently, their visibility (Noda et al., 1994).

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Many flowers do not have a uniform color, but appear mottled. However, this design is not casual, but has a definite role. In fact, if color is one of the most important cues for recognition of flowers at a distance, patterns become important only at close range because the insect spatial resolution is about 100 times poorer than that of humans (Kevan et al., 1996). These patterns may take a variety of forms. Many are visible to the human eye as color contrasts (i.e. red spots on a yellow background, as in the Andean monkey flower, Mimulus luteus), while others, particularly on flowers pollinated by bees, can be detected by insects due to the intense absorption in the UV (Horovitz and Cohen, 1972; Utech and Kawano, 1975; Gronquist et al., 2001). They are known as ‗honey guides‘ or ‗nectar guides‘ and had been first described by Sprengel at the end of the 18th century (Sprengel, 1793). Their role is to guide pollinating insects to the centre of the flower, where the sex organs and the rewards are present. They are useful for both plants and animals because they contribute to the proper orientation of insects for optimal pollen transfer, and accelerate the foraging of the insect, a fact particularly appreciated by those who do not land on flowers and collect the nectar while hovering (Kelber, 2002). Visible honey guides are normally due to local concentration of anthocyanins in particular zones of the flower (Cooley et al., 2008). UV honey guides mainly contain flavonoids (Thompson et al., 1972; Chang, 1997; Sasaki and Takahashi, 2002). Frequently, however various pigments are associated to give nectar guides observable in both the visible and UV regions (Harborne and Smith, 1978; Schlangen et al., 2009). The importance of color in pollination ecology is demonstrated by the changes that follow the pollination and/or fertilization. Some plants, immediately after pollination, lose their flowers that have completed their function in pollen dispersal and reception. On the contrary, other species become less attractive to pollinators by color changes of their petals. Sometimes the color changes are less visible because they occur at the nectar guides only (Gori, 1983; Paracer and Ahmadjian, 2000), or are not evident at all, taking place only in the UV (Silberglied, 1979; van Doorn, 1997). All these changes permit the plant to save energy in terms of biosynthesis of pigments and rewards and, at the same time, they are signals to animals that food is no longer available in these flowers. It has been suggested that this behavior increases pollinator efficiency, as the floral changes will direct the pollinators to flowers that have not yet been visited (van Doorn, 1997). A further advantage of retention of older flowers on a plant is its increased visibility that provides a long-distance attraction for pollinators. It has been observed that some color-changing species that retain their flowers after pollination receive more visits than those from which the older flowers have been removed (Weiss, 1995). Another very important factor for satisfactory pollination is the aspect of the plant, namely the shape of flower, the plant height, and the inflorescence architecture. Even if the concept of pollination syndrome has been questioned, it is likely that a certain correlation between floral traits and available pollinators within a given community of plants might be expected. Beetles have nutritional requirements and feeding behavior different from those of bees or flies or butterflies, so plant shape and rewards must meet these requests. Beetles are quite heavy insects and their mouth is arranged parallel to the body, so they need flowers with a strong structure on which they can safely land and that offer food sources on a flat surface. On the contrary, flowers pollinated by butterflies can have a much more tiny structure and the nectar is produced at the bottom of a long tube-shaped corolla in which the insects insert their long tongues. Plant height seems to play an important role in attracting pollinators, particularly when it is based on sexual deceit or when the plant presents flower

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dimorphism (Ashman et al., 2000; Raguso, 2004a). Some plants that offer pollen as a reward use as attractants visual signals that mimic stamens or anthers, such as Narcissus pseudonarcissus in which the lateral view of the cylindrical shaped part of the corolla resembles an androecium, or the spots on the upper petal of Rhododendron ponticum that simulate stamens, or the yellow patches on the petals of Saxifraga stellaris that allow the flower to maintain its attractiveness, even when the anthers have fallen off (Lunau, 2000). Many plants have their flowers grouped into higher-level units named inflorescences. These structures exhibit an immense diversity of architectures and often they appear as ―simple‖ flower aggregates; sometimes the single florets are so well integrated that the inflorescence resembles a single flower (i.e. capitula in Asteraceae family). The grouping of many small flowers presents some advantages for the plant: the first one is obviously a greater visibility due to a larger display area. Furthermore, it has been observed that multiple flowers permit to reduce pollen waste reducing the amount removed by each pollinator compared to a single large flower (Harder et al., 2004). Moreover, inflorescences can also affect other aspects of insects behavior, such as the number of visited florets. For instance, depending on inflorescence architecture, some insects depart when they encounter a single flower containing too few rewards (i.e. bees on racemes) or only after several empty flowers (i.e. bees on capitula) (Jordan and Harder, 2006). At first sight, it seems advisable that plants should be as attractive as possible to enhance pollen transfer. However, if a plant has too attractive flowers, it is very likely that a pollinator will visit many flowers on the same plant, losing outcross pollen (conspecific pollen of other plants) and accumulating self pollen. When the ratio of self to outcross pollen increases, there is a higher risk of geitonogamy (pollination between flowers on the same plant), and this risk is directly related to the time the insect spends on the plant (Klinkhamer and de Jong, 1993). To avoid geitonogamy, some self-fertile species have developed different strategies, such as heterostyly (floral morphs that differ in the reciprocal lengths of styles and stamens) (Vuilleumier, 1967; Kohn and Barrett, 1992), dioecy (Beach and Bawa, 1980), herkogamy (spatial separation of anthers and stigmas) (Kelly, 1997; Medrano et al., 2005), and dichogamy (temporal separation of male and female sexual functions) (Bertin and Newman, 1993; Bertin, 1993; Klinkhamer and de Jong, 1993). Clearly, apart from dioecy, the most secure method to avoid self-fertilization is self-incompatibility (Takayama and Isogai, 2005). Furthermore, the development of too attractive flowers may cause an increased risk of nectar and pollen predation by non-pollinating animals. Robbers remove nectar through a hole that they pierce in sympetalous corollas, so they do not pollinate flowers. Theft differs from robbing in that no hole is made, but the flower is visited by an animal for which it is not adapted, i.e. a very small insect which is not able to contact the reproductive organs of the flower. Therefore, the final aspect of the flower results from the two conflicting demands of the plant: to advertise its rewards for pollination purposes while protecting them from thieves and robbers. Nevertheless, sometimes, these visitors, mainly the robbers, may inadvertently pollinate plants because with their movements they cause the transfer of pollen to stigmas. This phenomenon has been observed in species such as Anthyllis vulneraria (Navarro, 2000), Corydalis ambigua (Higashi et al., 1988), Tristerix sp. (Graves, 1982) and Fouquieria splendens (Waser, 1979). To describe animals that behave in that way a new term has been coined: robberlike pollinators (Higashi et al., 1988). Plant density can have conflicting effects on pollination: dense flower patches can attract more pollinators, but these flowers may also compete for visits and resources (Elliott and

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Irwin, 2009). In patches with high density of flowers, the classical advertising signs (size, shape, color) become less important because pollinators obtain the higher nutritional rewards with the lower cost of flight between flowers (Molina-Montenegro and Cavieres, 2006). Negative effects may arise via reduced rates of pollinators visits, transfer of improper pollen, or through the loss of appropriate pollen while visiting inappropriate plants (Sih and Baltus, 1987). Some plant species live in communities constituted by different color morphs. It is not easy to explain this polymorphism; it has been proposed that it could reflect multiple and conflicting selection pressures, involving not only pollinators, but also herbivore-protection strategies or local abiotic conditions (Dormont et al., 2010). In the case of Orchis mascula, it was observed that fruit set was low in case of pure white or pure purple populations. On the contrary, when the rare white-flowered morphs were present within purple ones, the latter morph experienced a significantly higher (about fourfold) reproductive success, while the former morphs did not experience variations (Dormont et al., 2010). Authors demonstrated that only the visual cue played an important role; in fact they obtained comparable results even when they placed white ping-pong balls as attractive lures within a population of pure purple-flowered morphs. Not only the perianth of the flower is involved in the attractant function, but sometimes other parts may play an important role. In some species, anthers are large and intensely colored, even when they no longer contain pollen, such as in the Solanum genus. Often their color contrasts with that of perianth, both in the visible and in the UV range: i.e., in the flowers of Zygadenus nuttallii (Liliaceae) the tepals, nectaries and ovaries strongly reflect UV light, while the anthers and filaments do not (Bernhardt, 1996). Colored androecium (and/or pollen) is typical of those plants that offer pollen as reward. However, pollen is a very expensive resource for the plant (Petanidou and Vokou, 1990; Lunau, 2000); furthermore, not all the produced pollen can be displayed for advertisement purposes to avoid losses due to animal eating, runoff, damages from UV exposition, etc. Because of all these problems, plants prefer to display colored anthers or structures that resemble pollen grain on their flower surfaces (Lunau, 2000, #55272_). However, another problem arises from this situation: pollen-eating animals will visit only flowers that produce their food, that is male and hermaphrodite flowers. Consequently, pistillate flowers will not receive visits and they will not be pollinated. To solve this problem, female flowers have evolved structures that mimic pollen and stamens that use as signals in their place. Alternatively, some species develop structures named staminodes: they derive from stamens that have lost the function of pollen production, but can still perform an attractant role because of their color or morphology. In some species such as the kiwi plant (Actinidia chinensis), Saurauia sp. (Actinidiaceae) or in Amborella trichopoda (Amborallaceae), they still produce pollen as a reward for the pollinator, but it is nonviable or completely sterile, a sexual system known as cryptic dioecy (Sampson, 1993; Walker-Larsen and Harder, 2000; Lunau, 2007). While nectar is typically a clear liquid, colored nectar has evolved in some plant families. The most frequent colors are yellow to red, but also green, blue and even brown and black nectars are known. It has been observed that colored nectar is often associated with vertebrate pollination and isolation of the plant, both in islands and in mainland areas isolated by surrounding deserts or by altitude. Colored nectar is considered a visual floral cue to potential pollinators, and in particular yellow-orange-red nectars are associated to the colors of flowers visited by birds. In the case of black nectars, the color could be associated to the most

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frequent fruit color among bird-disperser plants in the tropics. A further point to support this correlation between birds and colored nectars is that most colored nectars contain high amounts of hexoses, the preferred sugars of nectar-feeding birds that lack the enzymes to digest sucrose (Hansen et al., 2007). At present, only two Fremontia species, F. californica and F. mexicana (Sterculiaceae), are known to possess a UV-fluorescent nectar. It has been speculated that this characteristic may be useful to attract bee pollinators (Thorp et al., 1975). Even extra-floral structures may be useful to attract pollinators in concert with or instead of the perianth. It is the case of bracts. They are modified leaves associated to flowers or inflorescences, often brightly colored. It has been observed that the experimental removal of bracts from inflorescences of Salvia viridis (Lamiaceae) significantly reduced honey bee visits (Keasar et al., 2006). In the case of Dalechampia ipomoeifolia (Euphorbiaceae), the dimension of the bract positively correlates with pollination success (Armbruster et al., 2005). The dove tree or handkerchief tree (Davidia involucrata, Nyssaceae) derives its name from the two white, paperlike bracts that surround the base of each flower head. It has been noted that bees prefer to visit flowers with white bracts. Furthermore, bracts have the important function to protect pollen from rain damages (Sun et al., 2008). In nature, plants rarely live in homogenous communities consisting of a single species only but, normally co-occur with some other species. In these communities, different species may share the same pollinators. Sometimes, a plant species that lack rewards imitate another one to attract pollinators, exploiting the attractive floral signals that share with nearby rewarding plants (Batesian floral mimicry). Obviously, this behavior requires quite precise mimicry of the size and color of the floral display of rewarding plants. According to some authors, in a floral mimicry system, such visual signals are more important than scent, at least for bee pollinators (Roy and Raguso, 1997; Galizia et al., 2005; Craig and Johnson, 2008). Mimicry mainly occurs in the Orchidaceae family: more than one-third of orchid species do not provide their pollinators with either pollen or nectar rewards. Among the many studies on this topic, those on Dendrobium infundibulum, Cymbidium insigne, Diuris maculata, Cephalnthera longifolia, Orchis israelitica, Anacamptis morio and Disia ferruginea can be mentioned as examples (Dafni and Ivri, 1981a, 1981b; Kjellsson et al., 1985; Beardsell et al., 1986; Johnson et al., 2003; Johnson, 2008). However, plant species also belonging to other families are involved in mimicry, such as Begoniaceae (Agren and Schemske, 1991), Asclepiadeceae and Verbenaceae (Bierzychudek, 1981) and Apocynaceae (Haber, 1984), among others. Besides Batesian, another type of mimicry has been postulated: Müllerian mimicry. The difference between Batesian and Müllerian mimicry depends on various aspects. In the former, there is a rewarding model and a rewardless mimic. Thus, the mimic parasitizes the successful advertisement of the model enjoying reproductive benefits as long as it remains at lower densities than the model. On the contrary, Müllerian mimicry is a mutualistic one, since both species reward pollinators and benefit each other by sharing a common advertising display, reaching a higher combined flower density (Roy and Widmer, 1999; Benitez-Vieyra et al., 2007). Sometimes mimicry is involuntary: in some Brassicaceae species the infection of a fungus, Puccinia sp., modifies host leaf morphology to produce ‗pseudoflowers‘. Infected plants form elongated stems crowned by dense, flower-like clusters of yellow-colored and scented leaves, covered with a sugar-rich exudate highly attractive to insects. The insects‘ attraction is improved by the volatiles emitted by the pseudoflowers (Raguso and Roy, 1998).

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Clearly, these structures are not intended to be beneficial for the plant, but serve for the sexual reproduction of fungus only (Roy, 1993). However, it has been observed that plant pathogens that attract pollinating insects may affect the reproductive success of other species belonging to the same community of infected plants (Roy, 1994, 1996), just as different flower species can influence each other for pollination, as above described. Other fungi able to induce pseudoflowers are species of the genus Uromyces (Pfunder and Roy, 2000; Ngugi and Scherm, 2006). It is known that the color of a flower is the first and foremost cue for pollinator's attraction, but the scent of a flower also plays a major role in attracting pollinators. Odors can act both at long distances as attraction cues and at short distances as orientation cues among different parts of the flower or among different flowers (Williams, 1983; Dobson et al., 1990; Knudsen and Tollsten, 1991; Dobson and Bergstrom, 2000a). Volatile organic compounds (VOCs) play many important roles in the relationships between plants and environment, ranging from defense and stress response, to thermotolerance, to pollinators‘ attraction (Pichersky and Gershenzon, 2002; Penuelas and Lluisia, 2004; Penuelas et al., 2005; Dudareva et al., 2006). Among the main VOCs emitted by plants, different chemical classes may be found. The most represented are terpenes, phenylpropanoids, the so-called green leaf volatiles, and some sulfur and nitrogen derivatives. Green leaf volatiles are normally woundinduced defensive compounds (Pare and Tumlinson, 1999; Matsui, 2006). The other ones are more important in terms of pollinator attraction. Terpenes owe their name to turpentine, the resin of pine trees. Their molecular skeleton is formed by C5 isoprene units: two in monoterpenes (C10), three in sesquiterpenes (C15), four in diterpenes (C20), six in triterpenes (C30), and eight in carotenoids (C40). However, only compounds up to 15 carbon atoms have a sufficiently high vapor tension that permit their release in the atmosphere. These isoprene units can be combined in different ways, can undergone successive modifications and can be substituted by many different functional groups to produce more than 20,000 different compounds (Tholl, 2006). Their production is controlled by terpene synthase enzymes. Numerous terpene synthases have been characterized: some of them catalyze the formation of just one terpene, but many others have the incredible ability to form complex mixtures of terpenes with high regio- and stereospecificity (Tholl, 2006). Consequently, plants emit very different bouquets of volatiles that give each species a unique and characteristic fragrance. Often, this emission follows a circadian rhythm to attract the most appropriate pollinators at the most appropriate time (Kolosova et al., 2001). In fact, plants tend to emit scents at maximal levels when the flowers are mature for pollination and concomitantly when their pollinators are active. Phenylpropanoids are C6-C3 compounds biosynthesized from phenylalanine. Apart from their role in lignin production, they have roles almost superimposable to those of terpenes, i.e. defensive, antioxidant, anti-UV and signaling (Dixon and Paiva, 1995; Graham and Graham, 1996; Pichersky and Gershenzon, 2002; Raguso, 2003; Reuber et al., 2006; Solecka, 2007). Sulfur and nitrogen-containing volatiles seem to be important for bat, fly and moth attraction (von Helversen et al., 2000; Levin et al., 2001; Piechulla and Pott, 2003; Jurgens et al., 2006). Some other compounds, such as organic acid derivatives, may play important roles in the attraction of pollinators acting as semiochemicals, particularly in sexually deceptive orchids. Volatiles may be released through the membrane of epidermal tissues, where they are biosynthesized, or from some other structures dedicated to their storage, such as glandular trichomes, vacuoles, resin ducts and laticifers. Often, particularly in vacuoles, they are stored

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as precursors in conjugated form, often as glycosides. In this case, for their release, they must first be deconjugated by lytic enzymes. Studies on the exact role of volatiles in pollination are still in their infancy, although in recent years, investigations about this topic are increasing more and more. In particular, the actual question that most needs an answer is not ―why do flowers smell‖, but rather ―why do flowers have different smells?‖ (Raguso, 2004c). Flowers rarely have the same composition of their bouquet of volatiles throughout all the entire cycle of their life, so not only different flowers may have different smells, but also flowers of the same plant species may differ in their odors depending on their phenological phase. Insects are able to distinguish (or can learn to do it) between these complex mixtures and, often, this leads to a species-specific interaction between plant and pollinator. This kind of interaction, in turn, reduces the possibility of pollen loss and/or unsuccessful interspecific pollen transfer. Probably, volatiles have been evolved to deter feeding by herbivores and to fight pathogens. However, they permitted predacious insects foraging for prey on foliage or using flowers as mating arenas, to readily identify the plant species. From here it was only a short step to cross-pollination because of involuntary pollen transfer from a flower to another with the same aroma while searching for food or mating sites (Robacker et al., 1988). So, as already discussed for other floral traits, volatiles may also be regarded as part of the so-called ‗pollination syndrome‘ (sensu Glover, 2007). Again, we can found ‗specialist‘ and ‗generalist‘ emission types (Whitehead and Peakall, 2009). These ‗fragrance syndromes‘ are most evident for flowers pollinated by nocturnal insects, bats and saprophage insects (Knudsen and Tollsten, 1993a, 1995; Dobson et al., 1997; Raguso et al., 2003; Jurgens et al., 2006). Floral scents are mainly emitted by the petals, but other floral parts may play a fundamental role in volatile production and emission. Not many papers have been published about this topic, however qualitative and quantitative different emissions have been documented by different research groups. Probably, these spatial fragrance patterns within the flowers are used by pollinators for orientation on flowers and can guide visitors to floral rewards, indicating that the visual contrasts presented by the flowers may have olfactory parallels (Dobson et al., 1996). Obviously, the correct orientation of the visitor is important to the plant for a more successful pollination. In particular, a very different odor has been perceived even by the human nose between androecium, especially the stamens, and other flower parts (Dobson et al., 1996). Studying Silene latifolia, Dotterl and Jurgens (2005) clearly evidenced a spatial fragrance pattern within the flowers, with lilac aldehydes and alcohols emitted by small anthophores only. Therefore it seems that these chemicals may guide pollinators to the entrance of the floral tube and finally to the nectar, secreted at the base of the filaments. Authors define this as floral scent nectar guides, in analogy to the classic visual guides described above (Dotterl and Jurgens, 2005). The same hypothesis was previously advanced by Flamini et al. (2002b) for Laurus nobilis flowers. A very similar situation was also suggested for Ranunculus acris (Bergstrom et al., 1995) and Clarkia breweri (Raguso and Pichersky, 1999). Lilac aldehydes and alcohols are relatively soluble in nectar, thus authors consider conceivable that these compounds are, besides olfactory cues, also gustatory signals for pollinators. Furthermore, these chemicals impart fragrance to nectar. Scented nectars have been already described for Abelia x grandiflora, Hedychium coronarium, Oenothera primiveris and Agave palmeri (Raguso, 2004b). In a recent study about the volatiles of grapefruit, our research group has examined the nectar produced by the flowers and the SPME analysis showed that it contained a high amount of non-terpene

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compounds (32.1% of total volatiles) and, in particular, three nitrogen derivatives (31.9%) with methyl anthranylate as the main one (23.6%). The main volatile was the monoterpene alcohol linalool (52.5%) (Flamini and Cioni, 2010). However, in all probability, scented nectars are more widespread in nature and further studies on this topic would be desirable. Scented nectars provide an honest signal, facilitating remote detection by nectar-foraging animals, especially when this scent is qualitatively or quantitatively distinct from overall floral scent (Raguso, 2004b). This may explain how bumble bee workers can distinguish nectar-rich flowers from depleted ones without landing (different intensities were also perceived by the human nose) (Heinrich, 1979). Some of the scented compounds detected in nectars seem to have additional functions, such as sorbic acid esters in Agave nectar that can prevent bacterial spoilage: in fact Agave palmeri produces large nectar pools and the flowers remain open for 4-6 days during the warmest and most humid span of the Sonoran desert year, the ideal condition for microbial infestation (Raguso, 2004b). Not all the volatiles of the nectar are attractant, but among them some repellent ones are present. An elegant investigation (Kessler and Baldwin, 2006) has highlighted this aspect in Nicotiana attenuata flowers and nectar. It was rich in nitrogen derivatives, and nicotine repelled all the examined visitors. Other volatiles were both attractant or repellent according to the different pollinator. The presence of these compounds has been correlated with the inhibition of nectar consumption by nectar robbers or less effective pollinators. Furthermore, even pollinators are affected by these volatiles decreasing nectaring time and the volume of nectar removed, but increasing the number of visits. Authors concluded that repellents optimize the number of flower visitors per volume of nectar produced, allowing plants to keep their nectar volumes small. This topic has been recently investigated for facultative and obligate flower visitors (Junker and Bluthgen, 2010). Obligate flower visitors are defined as those that require floral resources for at least part of their life-cycle, while facultative ones occasionally consume floral resources but are not dependent on them. In their meta-analysis, authors concluded that significantly different responses to flower scents were observed: obligate flower visitors were attracted by most of the floral scents, whereas facultative visitors were negatively affected. Again, the idea that attraction due to the fragrance of flowers is a derived character originating from a feeding-deterrent role of these chemicals is confirmed. Our research group has studied a dioecious plant, Laurus nobilis, to verify if there were differences in the emission of male and female whole flowers and to examine the patterns of emission of the various flower parts (Flamini et al., 2002b). It was observed that the volatile bouquets emitted by the whole flowers were almost superimposable, both qualitatively and quantitatively, in male and female plants. This is in good agreement with the observations of Dobson and Bergstrom on other species (2000b). These authors noted that male flowers only emitted -methyl ketones and alcohols, a feature shared by wind-pollinated plants, such as Cycas rumphii (Pellmyr et al., 1991). Authors proposed a connection between -methyl ketones and anemophily, or at least decreased entomophily, as observed in Filipendula vulgaris whose pollen odor was strongly dominated by 2-heptadecanone, having flowers which attract only few insects and considered to be partly or mainly wind-pollinated (Dobson et al., 1996). It can be concluded that these chemicals may serve in pollen defense. Indeed, in Laurus nobilis, -methyl ketones were detected in the pollen only (Flamini et al., 2002b). A further confirm of this trend can be found in the pollen of Rosa rugosa (Dobson et al., 1990).

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Pollen, at least in animal-pollinated plants, is perhaps one of the most scented part of the flower. Here it is necessary to repeat one more time that pollen odors evolved as a defense against pathogens and pollen-feeding animals, prior to the development of animal pollination. As flowering plants became dependent on pollinators, there was an increasing selection pressure to include attractant volatiles among the others (Dobson and Bergstrom, 2000b). Because of its invaluableness for plant reproduction, pollen is the one that most has to face the two conflicting pressures: to be protected by overexploitation by pollen-feeding animals and at the same time advertise its presence as desirable food to pollinators. Insect-pollinated plants have the grains of their pollen coated with an oily and often sticky colored substance, the pollenkitt, which cause pollen grains to clump and increase their adhesion to animals. Pollenkitt appears to be the main source of pollen odors, as first hypothesized (Knoll, 1930), and then demonstrated in 60 angiosperm species (Dobson, 1988). These pioneering studies paved the way for the investigations of pollen volatiles, reported in a review by Dobson and Bergstrom (2000b). However, the main difficulty with this study was to collect sufficient amounts of pollen to sample the volatiles in its headspace. In all their investigations, to overcome the limited emission of volatiles, authors needed 50-200 mg of pollen, a quantity not always easily recoverable. Sometimes it was necessary to collect pollen over several days (Dobson et al., 1996). They sampled volatiles by means of an air-flow of 50-60 ml/min, forced by a pump through the pollen packed inside a glass tube. Volatiles were then captured by an adsorbent trap from which they were recovered by solvent extraction. Prior to analysis, a concentration at 40°C of the extractive solution was required. The authors themselves have pointed out the difficulties in collecting quantities of volatiles sufficient for a reliable GC-MS analysis (Dobson et al., 1996). We have obtained noteworthy improvements by using the Head Space-Solid-Phase Microextraction (HS-SPME) technique. The high concentration capability of this method permits using considerably lower amounts of pollen (3 mg instead of 50-200 mg); furthermore, the sampling time is very reduced (15-30 min instead of 24-48 h), minimizing the possibility of sample contamination due to the prolonged forced flow of air required by the former method. The absence of solvents prevents the possible loss of volatiles during concentration of the extractive solution and, finally, the higher concentration capability of this technique permits the identification of a greater number of compounds (Flamini et al., 2002b). The effectiveness of this technique has been demonstrated by further studies on the volatiles from other pollens (Flamini et al., 2003a, 2003b, 2007; Maccioni et al., 2007; Flamini and Cioni, 2010) and by the fact that SPME or other thermal desorption methods have been also adopted by other research groups (Jurgens and Dotterl, 2004; Zhang et al., 2007). Most of the studies agree in reporting that pollen odor is clearly distinct from the whole flower odor, both in the number of volatiles and their relative amounts in each emitted mixture. However, each species has its own characteristic emission and there is no evidence for a general ‗pollen odor‘. The potential advantage to plants in having distinct pollen odors varies with each plant reproductive biology and needs to be evaluated on a species by species basis (Dobson et al., 1996). The main advantage derived from a distinct pollen odor is an increased fitness for the plant through enhanced flower constancy, greater pollen export and more effective pollen transport to stigmas. It has been observed that odors are learned by bees more rapidly than colors and many insects can learn and discriminate odor bouquets that differ in only a single chemical. Thus, even minor differences among floral odors, not necessarily detectable by the human nose, can foster constancy by pollinators (Dobson et al.,

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1996; Schiestl and Schluter, 2009). Often, pollen odors contain species-specific chemicals and/or a high proportion of certain volatiles that may be important in floral recognition and evaluation of rewards availability by pollinators, such as protoanemonin in Ranunculus acris (Bergstrom et al., 1995), -zingiberene in Cistus albidus (Maccioni et al., 2007), phenylacetaldehyde in Chrysanthemum coronarium (Flamini et al., 2003a) and 1-heptadecene in Citrus deliciosa (Flamini et al., 2003b). Similarly, the lack in pollen emission of one or more of the main volatiles produced by other floral parts may be a distinctive sign that can be discriminated by pollinators, such as trans--ocimene not detected in pollen of Ranunculus acris (Bergstrom et al., 1995), or limonene that constitutes up to 62.5% in other flower parts and not produced at all by Citrus limon pollen (Flamini et al., 2007). Multivariate statistical analysis applied to gas-chromatographic data may help to better understand the correlations in the various models of volatile emissions of different plant organs (Flamini et al., 2007; Flamini and Cioni, 2010). An interesting case is that of grapefruit plants (Flamini and Cioni, 2010). In fact, the extensive cultivated genus Citrus exhibits tremendous variation in pollination requirements. Many Citrus cultivars are selfincompatible and require pollinating insects for high fruit yields. The pollination requirements of grapefruit are poorly understood, and the available information is contradictory. According to some studies, cross-pollination is not necessary for fruit set (Roubik, 1995), but according to others, insect-mediated pollination improve fruit production (Burger, 1985; Chacoff and Aizen, 2006, 2007). The multivariate statistical analysis of the emissions from different plant and flower parts at different development stages permitted highlighting many differences but, at the same time, to group the samples on the basis of common similarities. By means of Hierarchical Cluster Analysis (HCA), Principal Component Analysis (PCA) and Multidimensional Scaling (MDS), it was noted that all floral parts were clustered in the same group, with the exception of young flowers whose emission was more similar to vegetative plant parts (i.e. leaves, branches, etc.). The very different emission pattern of this sample with respect to the mature flower may be a signal to indicate to pollinators that the flower is still not receptive (i.e. the flower has not yet accumulated rewards, from the pollinator point of view). At the same time, this behavior can also be related to the production of defense metabolites by the young flower to ensure its survival. In chemical terms, young plants and organs are normally more heavily defended (Bryant et al., 1991). A closer observation evidenced that young flowers and leaf buds emitted monoterpenes active against herbivores and pathogens (Raffa and Smalley, 1995; Müller-Schwarze and Thoss, 2008). Statistical analyses also confirmed the spatial odor gradient within the mature flower, indicating that grapefruit flowers have all the requirements necessary for entomophilous pollination. It would be interesting to know whether the reported self-pollination for some grapefruit plants (Roubik, 1995) have different emission profiles or whether self-pollination is used as a last resource to produce fruits in the absence of pollinator species. If volatiles are important attraction and orientation cues, they become invaluable tools for night-blooming plants. While visual signals are undoubtedly involved in plant-pollinator interactions during day-time, they are not reliable under low-light conditions, hence other senses, such as olfaction, must become important for night pollinators. Unfortunately, nocturnal pollination is extremely less studied than its diurnal counterpart, both because of organizational reasons and the need of dedicated equipment. However, a circadian rhythm has been widely observed in the emission of volatiles. Most plant species show peak emissions

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during the day, while others have their maximum production of volatiles at night (Altenburger and Matile, 1988; Dotterl et al., 2005; Verdonk et al., 2005; Loughrin et al., 2006, among the others). Most of the nocturnal pollinators are probably insects, such as moths and beetles, however the contribution of vertebrates such as bats and rodents should not be ignored. It is difficult to state which chemicals are mainly responsible for night-pollinators attraction. However, it has been noted that moth-pollinated flowers often emit benzenoids (Knudsen and Tollsten, 1993b; Makkholela and Manning, 2006; Okamoto et al., 2008), while the fragrances of bat-pollinated ones are characterized by displeasing, reminiscent of fermenting fruits, or garlic-like scents because of the presence of sulfur compounds (Bestmann et al., 1997; von Helversen et al., 2000). Even if in dim-light conditions colors cannot play a major role, they should not be neglected. Night-pollinated flowers tend to be white, cream or bright yellow to offer a strong contrast to the dark environment, so nocturnal insects can detect them more easily. It has been observed that nocturnal hawkmoths use achromatic intensity contrast for flower detection and recognition in dim light, but also chromatic cues could be important for these insects (Kelber et al., 2003). Fig trees (Ficus sp., Moraceae) are unique in their pollination biology: the ‗fruits‘ are actually inflorescences, named syconia, with the internal walls of a central cavity lined with many tiny florets. At one end, there is a small opening, the ostiole. Pollination is the result of an obligate mutualism between fig plant and female wasps of the subfamily Agaonidae. The insects lay their eggs inside fig flowers, where the larvae feed on some of the developing seeds (Ronsted et al., 2008). Because of the almost complete absence of visual cues, pollinators must be attracted by means of volatile compounds. It has been hypothesized that each fig species emit a particular fragrance to attract its specific pollinator. In the case of Ficus hispida, it has been observed that receptive syconia mainly emit palmitic acid, linalool, -pinene and -terpineol (Song et al., 2001), while Ficus carica emissions are characterized by benzyl alcohol, linalool, linalool oxides, cinnamaldehyde and indole (Gibernau et al., 1997). Further data on tropical figs are reported by Grison-Pigé et al. (2002). The flowers of Olea europaea do not offer nectar as a reward and they are commonly considered anemophilous. Nevertheless, the pollen has intermediate characteristics between typical anemophilous and entomophilous species (Wodehouse, 1935). Furthermore, a few papers report the visits of honey bees to olive flowers, contributing to their pollination (Griggs et al., 1975; McGregor, 1976; Barbier, 1986). Recently, our research group has compared the volatiles emitted by olive flowers of plants not visited by honey bees from different localities of Tuscany (Italy) with those of a group of plants heavily visited by these insects. In the latter group, honey bees collected olive pollen by crushing anthers with their mandibles and then they stored the grains inside their curbiculae. Even if the fruit sets were comparable, the emission of the flowers visited by honey bees was characterized by a higher amount of monoterpenes, both in pollen (59.5-70.4% vs. 13.2-33.8%) and whole flowers (40.4-51.2% vs. 0.3-1.0%). The pollen foraging was confirmed by melissopalynologic analyses on honey samples collected from hives next to the olive orchard (Flamini et al., 2008). Interestingly, volatiles emission and flower color seem to be correlated. Indeed, the combination of scent and visual cues increases the number of visits and degree of foraging activity for many pollinators in rewarding plants (Majetic et al., 2007) and in food-deceptive ones (Kunze and Gumbert, 2001). Probably, biosynthetic pathways that lead to floral pigments share enzymes and metabolic precursors with those that synthesizes volatiles.

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Similar links has been discovered for defense volatiles and pigments. It seems that genes affecting floral pigmentation may have pleiotropic effects on other characters, such as both defensive and odorous volatiles (Fineblum and Rausher, 1997; Lewinsohn et al., 2005; Majetic et al., 2007; Raguso, 2008). For example, light- or white-colored flowers have been reported to emit more benzenoid compounds than other color morphs, and this has been hypothesized to be an adaptation to maximize attraction of night-flying moth pollinators (Raguso et al., 2003). Because volatile aromatic compounds and anthocyanin-derived pigments responsible for flower coloration both originate from the same phenylpropanoid biosynthetic pathway, Zucker et al. (2002) and Majetic et al. (2007) hypothesized that color– scent associations in flowers may result from articular biochemical processes, by which flavonoid precursors of flower pigmentation in blocked biosynthetic pathways may be converted into volatile aromatic compounds. Viola etrusca (Calcaratae section) is strictly endemic to hills and mountains higher than 700 m in South Tuscany (Italy) and its corolla can be completely purple, completely white-yellowish, or speckled with various shades of purple, yellow and white. By means of SPME a completely different pattern of volatiles emission has been observed, with purple flowers that produce mainly (E)--ocimene, limonene and sesquiphellandrene, while yellows ones have 1,8-cineole, sabinene and (E,E)--farnesene among the principal volatiles. Another important difference between the two different color morphs is the production of benzenoids, in particular benzyl acetate, by the yellow ones only (Flamini et al., 2002a). It is known that some Viola species of the Calcaratae section have adapted to moth pollination (Veerman and Van Zon, 1965; Beattie, 1974). Many moths are attracted by benzenoids (Heath et al., 1992; Raguso and Light, 1998), so the emission of these compounds by the only light-colored morphs may be explained by the possibility of twilight or nocturnal pollination, facilitated by the yellow color that better stands out against the dark background. Only a few studies, limited to deceptive pollination in orchids, report the different emission of the single florets within an inflorescence. Most studies deal with changes due to pollination that make the florets less attractive for pollinators (Moya and Ackerman, 1993; Schiestl et al., 1997; Ayasse et al., 2000). In a very recent study, we have studied the emission of the whole capitula and of isolated tubular and ligulate florets of Coleostephus myconis (Asteraceae) (Flamini et al., 2010, in press). The SPME investigation evidenced a precise emission pattern, with marked differences between tubular and ligulate florets. T-cadinol was the main volatile emitted by the former type, while (E)--farnesene was the major constituent released by the latter. Although Asteraceae species are a significant component of almost all terrestrial ecosystems, the pollination biology of relatively few taxa has been examined in detail (Torres and Galetto, 2002). Asteraceae are pollinated by many insect orders, such as Hemiptera, Coleoptera, Lepidoptera, Hymenoptera and Diptera (Cerana, 2004). However, because of the strong structure and the flat shape of their head inflorescence, beetles are the most frequent visiting insects (Glover, 2007). Among the main volatiles emitted by the florets we can find attractants for different pollinators. This large presence of possible semiochemicals in C. myconis is in good agreement with the generalist nature of this family that lacks specialized pollinators (Grombone-Guaratini et al., 2004). Their different distribution in the inflorescence could provide orientation cues to pollinators. Moreover, the role of minor constituents, which further differentiate the emission of florets, cannot be a priori ignored because most semiochemicals have effects at extremely low concentrations.

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Not all the scents emitted by plants are pleasant. Some species, known as sapromyophilous, emit strong unpleasant fetid scents, which mimic carrion or dung. These odors imitate the decaying flesh in which some insects oviposit or feed. It is a deceptive pollination system because animals are not interested in the flowers as such, but they expect to find rotting proteins. When they do not find what they are searching for, they fly away, but inadvertently they transfer pollen between different flowers. To ensure a better pollination, most plants have traps that prevent a too rapid departure of the insect. Among sapromyophilous plants, species from different families, such as Apocynaceae, Araceae, Aristolochiaceae, Iridaceae, Hydnoraceae, Orchidaceae, Rafflesiaceae and Rhamnaceae, can be found. Insects attracted by these species belong mainly to Coleoptera and Diptera orders (Jurgens et al., 2006). Volatiles are mainly sulfur and nitrogen derivatives, such as dimethyl di- and trisulfides in Hydnoraceae (Burger et al., 1988), oligosulfides or amines in some Araceae (Smith and Meeuse, 1966; Borg-Karlsson et al., 1994). Furthermore, some foul smelling short-chain aliphatic acid, such as butyric and isovaleric acids (Kaiser, 1993). In this kind of pollination, however, olfactory cues alone are not sufficient, but they also need attractant visual displays at short distances, as demonstrated by the use of vials emitting oligosulfides that were not attractive to flies (Shuttleworth and Johnson, 2010). In fact, most of these flowers are big, dark-colored, often maculated, resembling a piece of rotting flesh or animal feces (Glover, 2007). To facilitate the dispersal of scent, some species are thermogenic, especially among the Araceae, where thermogenesis is salicylic acid-induced and permits to reach 42-44°C, regardless of ambient temperature (Raskin et al., 1987; Meeuse and Raskin, 1988). Even in these plants there is an organ-specific production of volatiles. These studies have been carried out mainly on Araceae species. In the voodoo lily, Sauromatum guttatum, scarab beetles and dung-flies, after being lured into the floral chamber by foul volatiles, encounter a much more pleasant odor, made up mainly of -pinene and limonene. The heat as well as the smell stimulates the activity and mating behavior of the insects, thereby enhancing pollination (Meeuse and Raskin, 1988; Borg-Karlsson et al., 1994). The spadix inflorescence of many Araceae is the main source of scented volatiles, particularly its upper sterile appendix. Removal of this appendix significantly reduces fruit set in Alocasia odora (Miyake and Yafuso, 2003). Sometimes it is the scent itself that is the reward pollinators are searching for. Some orchids, known as perfume orchids or euglossine orchids, offer no nectar, pollen, or other food rewards to the visiting bees, the so-called euglossine bees or orchid bees (Apidae: Euglossini). These orchids emit a unique blend of volatiles, very attractive for the bees that pollinate the plant. The bees collect as reward the volatiles from the surfaces of the flowers with the capillary brushes of their front tarsi and store them in spongy organs within the enlarged tibia on their hind legs. All the attracted bees are males and, apparently, they use the collected fragrances as sexual perfumes during their courtship. Other orchids that offer volatile compounds as a reward are those included in Bulbophyllum genus. They attract male fruit flies, mainly Bactrocera species, emitting synomone-acting substances such as methyl eugenol, raspberry ketone and zingerone, which the insect uses as a sex pheromone precursor or as a booster to pheromonal components, depending on the fruit fly species. To ensure pollination, the orchid traps the insect by means of a see-saw mechanism that places the fly in the appropriate position to stick pollinaria on its dorsum. The fruit fly spends up to 46 minutes to free itself (Tan and Nishida, 2007).

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Orchids are not the only plants that offer volatiles as a reward for these insects. Among Guttiferae (Clusiaceae), the filaments of the stamens of male flowers of Tovomita macrophylla or staminodes of female ones, produce tiny droplets of a volatile oil. Male Euglossine bees land on stamens or on the large stigmas to collect the oil with their forelegs. When the bees land on the stamens, their whole sternum is dusted with the pollen, which is then transferred to the stigma when the insects land on female flowers (de L. Nogueira et al., 1998). Finally, it has been observed that Euglossa viridissima collects fragrances from the flowers and even vegetative organs of other species, such as Begonia sp., Senna sp., Ocimum basilicum, Solanum sp., etc. (Pemberton and Wheeler, 2006). Plants have developed other non-nutritive rewards to ensure pollinators‘ services. Some species, such as Dalechampia magnistipulata and D. spathulata (Armbruster and Webster, 1979), have flowers that secrete resinous material collected by bees for the construction of their nests. During collection of this material, insects pollinate the flowers. Among angiosperms, only five genera are known to possess flowers that secrete resin. Besides the above Euphorbiaceae, four Clusiaceae genera show this behavior: Clusia, Clusiella, Chrysochlamys and Tovomitopsis (Gustafsson and Bittrich, 2002). In Clusia the resin is produced in schizogenous secretory ducts in the stamens of male flowers or in staminodes of female ones. Being a lipophilic and viscous material it is used by bees as a glue for fixing together sand, wood, twigs and other substances collected for their nest construction (Hochwallner and Weber, 2006). In addition to the mechanical utility, bees derive another benefit from the resin because this material has toxic effects against nest pathogens, particularly on gram-positive bacteria. Both male- and female-derived resins are endowed with this property, so bees collect this material on both flowers, ensuring an effective pollination (Lokvam and Braddock, 1999). Most of these resins are composed by polyisoprenylated benzophenones (de Oliveira et al., 1996; Lokvam et al., 2000). Besides sapromyophily, another very specialized deceptive pollination method is adopted by some non-rewarding orchids, which mimic the female of an insect to stimulate the male to mate with the flower; so-called ‗pseudocopulation‘ behavior. The appearance of the flower with the female insect is very impressive, but what is even more impressive is the type of volatiles emitted: in the bouquet also the sex pheromones of the female insects are included. Ophrys is one of the main orchid genus that uses sexual deception to attract pollinators. Males are lured by the visual display at long distances and then, at closer range, chemical signals from the flowers elicit their sexual behavior and try to copulate with the flower labellum. As a result, the insect eventually touches the gynostemium and the pollinia sticks on the animal, usually on its head. When the insect becomes aware of being deceived, it flies away and during a new pseudocopulation transfers the pollinia to the stigmatic surface of another flower (Ayasse et al., 2003). The extreme specialized mutualism between the orchid and its pollinator is testified by the species-specific visual display of the flower and the corresponding released volatiles endowed with pheromonal activity, peculiar for its pollinator. Sometimes these volatiles are not found elsewhere in plants, such as (-1)hydroxy and (-1)-oxo acids, especially 9-hydroxydecanoic acid (Ayasse et al., 2003). These findings surely give credit to the pollination syndrome theory. Other uncommon rewards offered by a limited number of plant species are waxy secretions and non-volatile oils. The labellum of the Brazilian orchids Maxillaria cerifera and M. brasiliensis produces a waxy secretion collected and stored in the curbiculae of some

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Meliponini bees. Chemical analyses of the labellar secretion permitted to identify as main constituents some tetracyclic triterpenes with cycloartane skeleton (Flach et al., 2004). Vogel (1971) first described the interactions between flower-oil-producing plants and oil-collecting bees. About 2400 species of plants in ten families are known to produce oil, usually instead of nectar. The oil is released as a trichome exudate in flowers of the Iridaceae and Scropulariaceae or is produced by specific epithelial glands, the elaiophores, in the Malpighiaceae, Orchidaceae and Krameriaceae (Vinson et al., 1997). The oil is used by the bees that pollinate the flowers, besides as provision for the larvae, also as a water resistant brood-cell lining (Dotterl and Schaffler, 2007). These very specialized oil-collecting bees do not use their mouths to gather the oil, but instead they absorb the product with the trichomes of their legs or by mopping their hairy abdomens. During collection, they become passively dusted with pollen, which will be transferred to the stigma during successive floral visits, as described in detail by Buchmann (1987) in his comprehensive review about oil flowers. Most of the oils are composed by free -acetoxy fatty acids, free fatty acids and acylglycerols containing -acetoxy fatty acids. Since some hydroxylated fatty acids are reported to possess antibiotic properties, it has been also hypothesized as playing a role in preventing nest infections and spoilage of provisions by microorganisms (Dumri et al., 2008). As mentioned above, some plants are able to produce heat. Besides enhancing scent diffusion, heat may also be an energy reward for insects, particularly in cold climates. Some insects depend on a threshold thoracic temperature to initiate flight at low-air temperatures. Instead of warming up passively by basking in the sun or by contracting the flight muscles, they may also profit from flowers whose temperature is higher than that of the ambient air (Sapir et al., 2006). Most of these flowers are protogynous, i.e. the female parts of the flower mature first and pollination takes place before male parts release pollen. With this strategy, insects bring in pollen from other plants allowing cross-fertilization (Seymour and SchultzeMotel, 1997). In addition to exothermic metabolic reactions, heat can be accumulated from the sun. The flower of some members of at least four families, Asteraceae, Papaveraceae, Ranunculaceae and Rosaceae, are characterized by heliotropism, i.e. they track the sun. This results in a heating of the internal cavities of the flower, up to several degrees above the air temperature. The more a flower is aligned with the sun, the warmer is its interior. Bowlshapes and dark colors contribute positively to the temperature increase. Among the most recent studies, those on the flowers of Ranunculus adoneus (Cooley, 1995; Galen, 2006), Adonis ramosa (Kudo, 1995), Ranunculus acris (Totland, 1996) and Iris species of Oncocyclus Section (Sapir et al., 2006) can be cited. An unusual theory about a possible strategy for pollinator attraction is reported by Warren and James (2008). They tried to answer if flowers wave to attract pollinators. Authors argue that some stalked species are found in extremely windy habitats, exposing flowers to possible damages. A wiser behavior should be a post-flowering stalk elongation, which is common in many species. After a field study on Silene maritima, authors conclude that mobile flowers are visited more frequently by pollinating insects. Although average visit durations were less in mobile flowers, this was more than compensated for by the increased number of visits. Insects are not the only animals that plants use for pollination purposes, but also vertebrates are involved in this process. Among winged animals, birds and bats play an important role as pollen vectors. As mentioned above, ornitophilous flowers are generally

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characterized by red corollas, abundant diluted nectar and lack of scent. Ornitophilous flowers have been adapted to two different types of bird approach: hovering flight and perching. The former approach is almost exclusive of hummingbirds, which collect nectar without landing on the plant. For this reason, many plants pollinated by hummingbirds have hanging flowers. Perching birds, on the contrary, land on stems or branches or, for low herbaceous plants, on the ground. These plants normally have erect flowers (Fenster et al., 2004; Johnson and Nicolson, 2008). Sometimes, plants pollinated by perching birds present structures that facilitate the landing of the animal, such as the sterile inflorescence axis of the South African Cape endemic rat‘s tail (Babiana ringens, Iridaceae). The scope of this appendage is exclusively to provide a perch for pollinating birds. It promotes the plant pollination success by causing the bird to adopt a position ideal for the cross-pollination (Anderson et al., 2005). At least 65 angiosperm families present bird pollination, even if it is notably absent in some of the largest ones. For example, in Asteraceae, only the genus Mutisia is ornitophilous (Cronk and Ojeda, 2008). Bird-pollinated flowers deposit pollen on the head or the back or around the base of the beak of the animal, according to the flower morphology (Wester and Classen-Bockhoff, 2007). The main reward for pollination services is nectar. Normally, this nectar is more diluted and in larger volume than in insect-pollinated plants. Sugar concentration ranges between 10% and 34% and, besides nutritional considerations, it determines nectar viscosity, a very important parameter that affects the ease of uptake by birds. Another hypothesis considers the possibility that nectar dilution discourages theft by non-pollinating insects: nectar with sugar concentration below 18% is not useful for honeybees because of the high energetic cost of evaporating water to produce honey (Percival, 1965; Cronk and Ojeda, 2008). Finally, dilute nectars in these flowers could be a secondary consequence of their having evolved as deep tubular flowers, in fact nectar evaporates (and thereby concentrates) much faster in flowers with small corolla depths (Plowright, 1987). Bat pollination is primarily limited to tropical and subtropical regions and, as already mentioned, chiropterophilous flowers are generally dull in color or white and emit characteristic scents. Basically, these flowers may be subdivided into two groups, depending on the size of the bats that visit them. Bigger bats often pollinate wide pendent flowers, such as those of the baobab (Adansonia digitata, Bombacaceae). Visiting bats hold on by clasping the ball of stamens to their breasts while lapping nectar from the base of the stamen column. The pollen that dusts their breast is then transferred to the stigma of the subsequent flower. Smaller bats visit bell-shaped gamopetalous flowers. To reach the nectar they must enter inside the corolla, dusting with pollen. Bat-pollinated plants need to be strong to support the weight and the flying habits of their pollinators, and generally are trees (Baker, 1961). Besides olfaction, bats can also rely on their echolocation abilities to find the flowers in the dark, especially those already visited in the past. Bell-shaped flowers are those that are better identified because their echoes are clearly identifiable among those from the vegetation background (von Helversen et al., 2003). Some plants are even equipped with ‗acoustic nectar guides‘: the flowers of the vine Mucuna holtonii contain a small concave ‗mirror‘ reflecting most of the energy of the echolocation calls of the bat back into the direction of incidence. Even the pollen dispersal mechanism is quite singular: when the bat lands on the papilionaceous flower and presses its snout into the slit between the wings, the keel burst and the staminal column, being under tension, catapults most of the pollen load onto the bat rump (von Helversen and von Helversen, 1999).

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There are conflicting points of view about the importance of non-flying animals, particularly mammals, for plant pollination. Often it is thought to be incidental, during irregular visits. For some plants, however, non-flying mammals may well act as the sole pollinators, particularly in the case of some of the plants visited by lemurs in Madagascar (Goldingay et al., 1991; Carthew and Goldingay, 1997). In recent years, the mutualism between plant and reptiles has been re-evaluated and lizards are now considered among the most common pollinators, especially on islands. These animals have been ignored as pollinators possibly because most are regarded as being carnivorous. However, many lizards have a broad diet, which can include nectar and pollen (Olesen and Valido, 2003). Generally pollen adheres to different parts of lizard bodies, according to plant and reptile species (Godinez-Alvarez, 2004). The main attractant feature implemented by these plants is a colored nectar, as observed for some Mauritian species (Hansen et al., 2006). Summarizing, to achieve the primary purpose of sexual reproduction, plants adopt various effective strategies to permit the pollen to reach the stigma, preferably on a different plant of the same species. Paraphrasing Raguso (2004a), we can say that flowers act as sensory billboards. To better understand these signals, an integrated multidisciplinary approach is essential.

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REFERENCES Ackerman, J. D. (2000). Abiotic pollen and pollination: ecological, functional, and evolutionary perspectives. Plant Syst. Evol, 222, 167-185. Agren, J., and Schemske, S. W. (1991). Pollination by deceit in a neotropical monoecious herb, Begonia involucrata. Biotropica, 23, 235-241. Altenburger, R., and Matile, P. (1988). Circadian rhythmicity of fragrance emission in flowers of Hoya carnosa R. Br. Planta, 174, 248-252. Anderson, B., Cole, W. W., and Barrett, S. C. H. (2005). Specialized bird perch aids crosspollination. Nature, 435, 41-42. Armbruster, W. S., Antonsen, L., and Pelabon, C. (2005). Phenotypic selection on Dalechampia blossoms: hones signaling affects pollination success. Ecology, 86, 33233333. Armbruster, W. S., and Webster, G. L. (1979). Pollination of two species of Dalechampia (Euphorbiaceae) in Mexico by Euglossine bees. Biotropica, 11, 278-283. Arnold, S., Savolainen, V., and Chittka, L. (2008). FReD: the floral reflectance spectra database. Nature Precedings, http://dx.doi.org/10.1038/npre.2008.1846.1. Ashman, T. L., Swetz, J., and Shivitz, S. (2000). Understanding the basis of pollinator selectivity in sexually dimorphic Fragaria virginiana. Oikos, 90, 347-356. Ayasse, M., Schiestl, F. P., Paulus, H. F., Ibarra, F., and Franke, W. (2003). Pollinator attraction in a sexually deceptive orchid by means of unconventional chemicals. Proceedings of the Royal Society B, 270, 517-522. Ayasse, M., Schiestl, F. P., Paulus, H. F., Lofstedt, C., Hansson, B., Ibarra, F., and Francke, W. (2000). Evolution of reproductive strategies in the sexually deceptive orchid Ophrys

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sphegodes: how does flower-specific variation of odor signals influence reproductive success? Evolution, 54, 1995-2006. Baker, H. G. (1961). The adaptation of flowering plants to nocturnal and crepuscular pollinators. Quarterly Review of Biology, 36, 64-73. Barbier, E., (1986). La pollinisation des cultures. Edmond. Beach, J. H., and Bawa, K. S. (1980). Role of pollinators in the evolution of dioecy from distyly. Evolution, 34, 1138-1142. Beardsell, D. V., Clements, M. A., Hutchinson, J. F., and Williams, E. G. (1986). Pollination of Diuris maculata R Br (Orchidaceae) by floral mimicry of the native legumes Daviesia spp. and Pulteria scabra R Br. Aust. J. Bot, 34, 165-173. Beattie, A. J. (1974). Floral evolution in Viola. Annals of the Missouri Botanical Garden, 61, 781-793. Benitez-Vieyra, S., Hempel de Ibarra, N., Wertlen, A. M., and Cocucci, A. A. (2007). How to look like a mallow: evidence of floral mimicry between Turneraceae and Malvaceae. Proceedings of the Royal Society B, 274, 2239-2248. Bergstrom, G., Dobson, H. E. M., and Groth, I. (1995). Spatial fragrance patterns within the flowers of Ranunculus acris (Ranunculaceae). Plant Systematics and Evolution, 195, 221–242. Bernhardt, P. (1996). Anther adaptations in animal pollination. In W. G. D'Arcy, and R. C. Keating, The anther: form, function and phylogeny (pp. 192-220). Cambridge: Cambridge University Press. Bertin, R. I. (1993). Incidence of monoecy and dichogamy in relation to self-fertilization in angiosperms. American Journal of Botany, 80, 557-560. Bertin, R. I., and Newman, C. M. (1993). Dichogamy in angiosperms. The Botanical Review, 59, 112-152. Bestmann, H. J., Winkler, L., and von Helversen, O. (1997). Headspace analysis of volatile flower scent constituents of bat-pollinated plants. Phytochemistry, 46, 1169-1172. Bierzychudek, P. (1981). Asclepias, Lantana, and Epidendrum: a floral mimicry complex? Biotropica, 13, 54-58. Borg-Karlsson, A. K., Englund, F. O., and Unelius, C. R. (1994). Dimethyl oligosulphides, major volatiles released from Sauromatum guttatum and Phallus impudicus. Phytochemistry, 35, 321-323. Bowker, G. E., and Crenshaw, H. C. (2007). Electrostatic forces in wind-pollination-Part 2: simulation of pollen capture. Atmospheric Environment, 41, 1596-1603. Bryant, J. P., Kuropat, P. J., Reichardt, P. B., and Clausen, T. P. (1991). Controls over the allocation of resources by woody plants to chemical antiherbivore defense. In R. T. Palo, and C. T. Robbins, Plant Defenses Against Mammalian Herbivory (pp. 83). CRC Press. Buchmann, S. L. (1987). The ecology of oil flowers and their bees. Annual Review of Ecology and Systematics, 18, 343-369. Burger, B. V., Munro, Z. M., and Visser, J. H. (1988). Determination of plant volatiles 1: analysis of the insect-attracting allomone of the parasitic plant Hydnora africana using Grob-Habich activaterd charcoal traps. Journal of High Resolution Chromatography and Chromatography Communications, 11, 496-499. Burger, D. W. (1985). Pollination effects on fruit production of 'Star Ruby' grapefruit (Citrus paradisi Macf.). Scientia Horticulturae, 25, 71-76.

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Carthew, S. M., and Goldingay, R. L. (1997). Non-flying mammals as pollinators. Tree, 12, 104-108. Cerana, M. M. (2004). Flower morphology and pollination in Mikania (Asteraceae). Flora, 199, 168-177. Chacoff, N. P., and Aizen, M. A. (2006). Edge effects on flower-visiting insects in grapefruit plantations bordering premontane subtropical forest. Journal of Applied Ecology, 42, 1827. Chacoff, N. P., and Aizen, M. A. (2007). Pollination requirements of pigmented grapefruit (Citrus paradisi Macf.) from Northwestern Argentina. Crop Science, 47, 1143-1150. Chacoff, V. R., Aizen, M. A., and Galetto, L. (2006). Nectar concentration and composition of 26 species from the temperate forest of South America. Annals of Botany, 97, 413-421. Chang, C. S. (1997). Flavonoid chemistry of Weigela (Caprifoliaceae) in Korea. Journal of Plant Research, 110, 275-281. Clement, J. S., and Mabry, T. J. (1996). Pigment evolution in the Caryophyllales: a systematic overview. Botanica Acta, 109, 360-367. Cooley, A. M., Carvallo, G., and Willis, J. H. (2008). Is floral diversification associated with pollinator divergence? Flower shape, flower colour and pollinator preference in Chilean Mimulus. Annals of Botany, 101, 641-650. Cooley, J. R. (1995). Floral heat rewards and direct benefits to insect pollinators. Annals of the Entomological Society of America, 88, 576-579. Cox, P. A. (1988). Hydrophilous pollination. Annual Review of Ecology and Systematics, 19, 261-280. Cox, P. A. (1993). Water-pollination in plants. Scientific American, 269, 68-74. Craig, I. P., and Johnson, S. D. (2008). mimics and magnets: the importance of color and ecological facilitation in floral deception. Ecology, 89, 1583-1595. Cronk, Q., and Ojeda, I. (2008). Bird-pollinated flowers in an evolutionary and molecular context. Journal of Experimental Botany, 59, 715-727. Culley, T. M., Weller, S. G., and Sakai, A. K. (2002). The evolution of wind pollination in angiosperms. Trends in Ecology and Evolution, 17, 361-369. Dafni, A., and Ivri, Y. (1981a). Floral mimicry between Orchis israelitica Baumann and Dafni (Orchidaceae) and Bellevalia flexuosa Boiss. (Liliaceae). Oecologia, 49, 229-232. Dafni, A., and Ivri, Y. (1981b). The flower biology of Cephalanthera longifolia (Orchidaceae)-Pollen imitation and facultative floral mimicry. Plant Systematics and Evolution, 137, 229-240. de L. Nogueira, P. C., Marsaioli, A. J., Amaral, M. D. C. E., and Bittrich, V. (1998). The fragrant floral oils of Tovomita species. Phytochemistry, 49, 1009-1012. de Oliveira, C. M. A., Porto, A. M., Bittrich, V., Vencato, I., and Marsaioli, A. J. (1996). Floral resin of Clusia spp.: chemical composition and biological function. Tetrahedron Letters, 37, 6427-6430. Delpino, F. (1873). Uteriori osservazioni e considerazioni sulla dicogamia nel regno vegetale. Atti della Società Italiana di Scienze Naturali, 16, 151-349. Dixon, R. A., and Paiva, N. L. (1995). Stress-induced phenylpropanoid metabolism. The Plant Cell, 7, 1085-1097. Dobson, H. E. M. (1988). Survey of pollen and pollenkitt lipids-chemical cues to flower visitors? American Journal of Botany, 75, 170-182.

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Pollination Strategies

23

Dobson, H. E. M. (1994). Floral volatiles in insect biology. In E. A. Bernays, Insect-plant interactions (pp. 47–81). Boca Raton: CRC Press. Dobson, H. E. M., Arroyo, J., Bergstrom, G., and Groth, I. (1997). Interspecific variation in floral fragrances within the genus Narcissus (Amaryllidaceae). Biochemical Systematics and Ecology, 25, 685-706. Dobson, H. E. M., and Bergstrom, G. (2000a). The ecology and the evolution of pollen odors. Plant Systematics and Evolution, 222, 63–87. Dobson, H. E. M., and Bergstrom, G. (2000b). The ecology and the evolution of pollen odors. Plant Systematics and Evolution, 222, 63–87. Dobson, H. E. M., Bergstrom, G., and Groth, I. (1990). Differences in fragrance chemistry between flower parts of Rosa rugosa Thunb. (Rosaceae). Israel J. Bot, 39, 143–156. Dobson, H. E. M., Bergstrom, G., and Groth, I. (1996). Pollen advertisement: chemical contrasts between flower and pollen odors. American Journal of Botany, 83, 877–885. Dormont, L., Delle Vedove, R., Bessiere, J. M., Hossaert-McKey, M., and Schatz, B. (2010). Rare white-flowered morphs increase the reproductive success of common purple morphs in a food-deceptive orchid. New Phytologist, 185, 300-310. Dotterl, S., and Jurgens, A. (2005). Spatial fragrance patterns in flowers of Silene latifolia: lilac compounds as olfactory nectar guides? Plant Systematics and Evolution, 255, 99109. Dotterl, S., and Schaffler, I. (2007). Flower scent of floral oil-producing Lysimachia punctata as attractant for the oil-bee Macropis fulvipes. Journal of Chemical Ecology, 33, 441-445. Dotterl, S., Wolfe, L. M., and Jurgens, A. (2005). Qualitative and quantitative analyses of flower scent in Silene latifolia. Phytochemistry, 66, 203-213. Dudareva, N., Negre, F., Nagegowda, D. A., and Orlova, I. (2006). Plant volatiles: recent advances and future perspectives. Critical Reviews in Plant Sciences, 25, 417-440. Dumri, K., Seipold, L., Schmidt, J., Gerlach, G., Dotterl, S., Ellis, A. G., and Wessjohann, L. A. (2008). Non-volatile floral oils of Diascia spp. (Scrophulariaceae). Phytochemistry, 69, 1372-1383. Ellestad, G. A. (2006). Structure and chiroptical properties of supramolecular flower pigments. Chirality, 18, 134-144. Elliott, S. E., and Irwin, R. E. (2009). Effects of flowering plant density on pollinator visitation, pollen receipt, and seed production in Delphinium barbeyi (Ranunculaceae). American Journal of Botany, 96, 912-919. Fenster, C. B., Armbruster, W. S., Wilson, P., Dudash, M. R., and Thomson, J. D. (2004). Pollination syndromes and floral specialization. Annual Review of Ecology and Systematics, 35, 375-403. Ferreira da Silva, P., Lima, J. C., Freitas, A. A., Shimizu, K., Macanita, A. L., and Quina, F. H. (2005). Charge-transfer complexation as a general phenomenon in the copigmentation of anthocyanins. Journal of Physical Chemistry A, 109, 7329-7338. Fineblum, W. L., and Rausher, M. D. (1997). Do floral pigmentation genes also influence resistance to enemies? The W locus in Ipomoea purpurea. Ecology, 78, 1646-1654. Flach, A., Dondon, R. C., Singer, R. B., Koehler, S., Amaral, M. D. C. E., and Marsaioli, A. J. (2004). The chemistry of pollination in selected brazilian maxillariinae orchids: floral rewards and fragrance. Journal of Chemical Ecology, 30, 1045-1056.

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Guido Flamini

Flamini, G., and Cioni, P. L. (2010). Odour gradients and patterns in volatile emission of different plant parts and developing fruits of grapefruit (Citrus paradisi L.). Food Chemistry, 120, 984-992. Flamini, G., Cioni, P. L., Maccioni, S., and Baldini, R. (2010). Essential oil composition and in vivo volatile emission by different parts of Coleostephus myconis capitula. Natural Product Communications, Flamini, G., Cioni, P. L., and Morelli, I. (2002a). Analysis of the essential oil of the aerial parts of Viola etrusca from Monte Labbro (South Tuscany, Italy) and in vivo analysis of flower volatiles using SPME. Flavour Fragr. J, 17, 147–149. Flamini, G., Cioni, P. L., and Morelli, I. (2002b). Differences in the fragrances of pollen and different floral parts of male and female flowers of Laurus nobilis. Journal of Agricultural and Food Chemistry, 50, 4647–4652. Flamini, G., Cioni, P. L., and Morelli, I. (2003a). Differences in the Fragrances of Pollen, Leaves, and Floral parts of Garland (Chrysanthemum coronarium) and Composition of the Essential Oil from Flowerheads and Leaves. Journal of Agricultural and Food Chemistry, 51, 2267–2271. Flamini, G., Cioni, P. L., and Morelli, I. (2003b). Use of solid-phase micro-extraction as a sampling technique in the determination of volatiles emitted by flowers, isolated flower parts and pollen. Journal of Chromatography A, 998, 229–233. Flamini, G., Marcucci, S., Pinzauti, M., Conidi, M., Filippi, L., Bedini, G., and Felicioli, A. (2008). Possibile attività pronuba svolata dall'ape (Apis mellifera: Hymenoptera, Apoidea) nell'impollinazione dll'olivo (Olea europaea). In Codice Armonico 2008 (pp. 89-93). Rosignano Solvay (Livorno): ETS Flamini, G., Tebano, M., and Cioni, P. L. (2007). Volatiles emission patterns of different plant organs and pollen of Citrus limon. Analytica Chimica Acta, 589, 120-124. Galen, C. (2006). Solar-furnaces or swamp coolers: costs and benefits of water use by solartracking flowers of the alpine snow buttercup, Ranunculus adoneus. Oecologia, 148, 195201. Galizia, C. G., Kunze, J., Gumbert, A., Sachse, S., Markl, C., Menzel, R., and Borg-Karlsson, A. K. (2005). Relationship of visual and olfactory signal parameters in a food-deceptive flower mimicry system. Behavioral Ecology, 16, 159-168. Gibernau, M., Buser, H. R., Frey, J. E., and Hossaert-McKey, M. (1997). Volatile componds from extracts of figs of Ficus carica. Phytochemistry, 46, 241-244. Glover, B. J., (2007). Understanding flowers and Flowering, an integrated approach. New York: Oxford University Press. Godinez-Alvarez, H. (2004). Pollination and seed dispersal by lizards: a review. Revista Chilena de Historia Natural, 77, 569-577. Goldingay, R. L., Carthew, S. M., and Whelan, R. J. (1991). The importance of non-flying mammals in pollination. Oikos, 61, 79-87. Gori, D. F. (1983). Post-pollination phenomena and adaptive floral changes. In C. E. Jones, and R. J. Little, Handbook of experimental pollination biology (pp. 31-49). New York: Van Nostrand Reinhold. Goto, T., and Kondo, T. (1991). Structure and molecular stacking of anthocyanins-flower color variation. Angewandte Chemie, 30, 17-33.

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25

Graham, T. L., and Graham, M. Y. (1996). Signaling in soybean phenylpropanoid responses (dissection of primary, secondary, and conditioning effects of light, wounding, and elicitor treatments). Plant Physiology, 110, 1123-1133. Graves, G. R. (1982). Pollination of a Tristerix nistletoe (Loranthaceae) by Diglossa (Aves, Thraupidae). Biotropica, 14, 316-317. Griggs, N. H., Hartmann, H. T., Bradley, M. V., Iwakiri, B. T., and Whisler, J. E., (1975). The olive pollination in California. Grison-Pigé, L., Bessiere, J. M., and Hossaert-McKey, M. (2002). Specific attraction of figpollinated wasps: role of volatile compounds released by tropical figs. Journal of Chemical Ecology, 28, 283-295. Grombone-Guaratini, M. T., Solferini, V. N., and Semir, J. (2004). Reproductive biology in species of Bidens L. (Asteraceae). Scientia Agricola, 61, 185-189. Gronquist, M., Bezzerides, A., Attygalle, A., Meinwald, J., Eisner, M., and Eisner, T. (2001). Attractive and defensive functions of the ultraviolet pigments of a flower (Hypericum calycinum). Proceedings of the National Academy of Sciences of the United States of America, 98, 13745-13750. Gustafsson, M. H. G., and Bittrich, V. (2002). Evolution of morphological diversity and resin secretion in flowers of Clusia (Clusiaceae): insights from ITS sequence variation. Nordic Journal of Botany, 22, 183-203. Haber, W. A. (1984). Pollination by deceit in a mass-flowering tropical tree Plumeria rubra L. (Apocynaceae). Biotropica, 16, 269-275. Hansen, D. M., Beer, K., and Muller, C. B. (2006). Mauritian coloured nectar no longer a mystery: a visula signal for lizard pollinators. Biology Letters, 2, 165-168. Hansen, D. M., Olesen, J. M., Mione, T., Johnson, S. D., and Muller, C. B. (2007). Coloured nectar: distribution, ecology, and evolution of an enigmatic floral trait. Biological Reviews, 82, 83-111. Harborne, J. B., and Smith, D. M. (1978). Anthochlors and other flavonoids as honey guides in the compositae. Biochemical Systematics and Ecology, 6, 287-291. Harder, L. D., Jordan, C. Y., Gross, W. E., and Routley, M. B. (2004). Beyond floricentrism: the pollination function of inflorescences. Plant Species Biology, 19, 137-148. Heath, R. R., Landolt, P. J., Dueben, B., and Lenczewski, B. (1992). Identiication of floral compounds of night blooming jessamine attractive to cabbage looper moths. Environmental Entomology, 21, 854-859. Heinrich, B. (1979). Resource heterogeneity and patterns of movement in foraging bumblebees. Oecologia, 40, 235-245. Higashi, S., Ohara, M., Arai, H., and Matsuo, K. (1988). Robber-like pollinators: overwintered queen bumblebees foraging on Corydalis ambigua. Ecological Entomology, 13, 411-418. Hochwallner, H., and Weber, A. (2006). Flower development and anatomy of Clusia valerioi, a Central American spcesies of Clusiaceae offering floral resin. Flora, 201, 407-418. Horovitz, A., and Cohen, Y. (1972). Ultraviolet reflectance characteristicsin flowers of crucifers. American Journal of Botany, 59, 706-713. Johnson, S. D. (2008). Evidence for Batesian mimicry in a butterfly-pollinated orchid. Biological Journal of the Linnean Society, 53, 91-104. Johnson, S. D., Craig, I. P., Nilsson, L. A., and Agren, J. (2003). Pollination success in a deceptive orchid is enhanced by co-occurring magnet plants. Ecology, 84, 2919-2927.

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Guido Flamini

Johnson, S. D., Linder, H. P., and Steiner, K. E. (1998). Phylogeny and radiation of pollination systems in Disa (Orchidaceae). American Journal of Botany, 85, 402-411. Johnson, S. D., and Nicolson, S. W. (2008). Evolutionary associations between nectar properties and specificity in bird pollination systems. Biology Letters, 4, 49-52. Jordan, C. Y., and Harder, L. D. (2006). Manipulation of bee behavior by inflorescence architecture and its consequences for plant mating. American Naturalist, 167, 496-509. Junker, R. J., and Bluthgen, N. (2010). Floral scents repel facultative flower visitors, but attract obligate ones. Annals of Botany, 105, 777-782. Jurgens, A., and Dotterl, S. (2004). Chemical composition of anther volatiles in Ranunculaceae: genera-specific profiles in Anemone, Aquilegia, Caltha, Pulsatilla, Rannculus, and Trollius species. American Journal of Botany, 91, 1969-1980. Jurgens, A., Dotterl, S., and Meve, U. (2006). The chemical nature of fetid floral odours in stapeliads (Apocynaceae-Asclepiadoideae-Ceropegieae). New Phytologist, 172, 452-468. Kaiser, R., (1993). The scent of orchids. Amsterdam: Elsevier. Keasar, T., Pollak, G., Arnon, R., Cohen, D., and Shmida, A. (2006). Honesty of signaling and pollinator attraction: the case of flag-like bracts. Israel Journal of Plant Sciences, 54, 119-128. Kelber, A. (2002). Pattern discrimination in a hawkmoth: innate preferences, learning performance and ecology. Proceedings of the Royal Society B, 269, 2573-2577. Kelber, A., Balkenius, A., and Warrant, E. J. (2003). Colour vision in diurnal and nocturnal hawkmoths. Integrative and Comparative Biology, 43, 571-579. Kelly, L. M. (1997). A cladistic analysis of Asarum (Aristolochiaceae) and implication for the evolution of herkogamy. American Journal of Botany, 84, 1752-1765. Kessler, D., and Baldwin, I. T. (2006). Making sense of nectar scents: the effect of nectar secondary metabolites on floral visitors of Nicotiana attenuata. Plant Journal, 49, 840854. Kevan, P., Giurfa, M., and Chittka, L. (1996). Why are there so many and so few white flowers? Trends in Plant Science, 1, 280-284. Kjellsson, G., Rasmussen, F. N., and Dupuy, D. (1985). Pollination of Dendrobium infundibulum, Cymbidium insigne (Orchidaceae) and Rhododendron lyi (Ericaceae) by Bombus eximius (Apidae) in Thailand: a possible case of floral mimicry. Journal of Tropical Ecology, 1, 289-302. Klinkhamer, P. G. L., and de Jong, T. J. (1993). Attractiveness to pollinators: a plant's dilemma. Oikos, 66, 180-184. Knoll, F. (1930). Über Pollenkitt und Bestäubungsart. Zeitschrift fur Botanik, 23, 609-675. Knudsen, J. T., and Tollsten, L. (1991). Floral scent and intrafloral scent differentiation in Monoses and Pyrola (Pyrolaceae). Plant Systematics and Evolution, 177, 81–91. Knudsen, J. T., and Tollsten, L. (1993a). Trends in floral scent chemistry in pollination syndromes: floral scent composition in moth-pollinated taxa. Botanical Journal of the Linnean Society, 113, 263-284. Knudsen, J. T., and Tollsten, L. (1993b). Trends in floral scent chemistry in pollination syndromes: floral scent composition in moth-pollinated taxa. Botanical Journal of the Linnean Society, 113, 263-284. Knudsen, J. T., and Tollsten, L. (1995). Floral scent in bat-pollinated plants: a case of convergent evolution. Botanical Journal of the Linnean Society, 119, 45-57.

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27

Kohn, J. R., and Barrett, S. C. H. (1992). Experimental studies on the functional significance of heterostyly. Evolution, 46, 43-55. Kolosova, N., Gorenstein, N., Kish, C. M., and Dudareva, N. (2001). Regulation of circadian methyl benzoate emission in diurnally and nocturnally emitting plants. The Plant Cell, 13, 2333-2347. Kudo, G. (1995). Ecological significance of flower heliotropism in the spring ephemeral Adonis ramosa (Ranunculaceae). Oikos, 72, 14-20. Kunze, J., and Gumbert, A. (2001). The combined effect of color and odor on flower choice behavior of bumble bees in flower mimicry systems. Behavioral Ecology, 12, 447-456. Law, R., Bronstein, J. L., and Ferriere, R. (2001). On mutualists and exploiters: plant-insect coevolution in pollinating seed-parasite systems. Journal of Theoretical Biology, 212, 373-389. Levin, R. A., Raguso, R. A., and McDade, L. A. (2001). Fragrance chemistry and pollinator affinities in Nyctaginaceae. Phytochemistry, 58, 429–440. Lewinsohn, E., Sitrit, Y., Bar, E., Azulay, Y., Ibdah, M., Meir, A., Yosef, W., Zamir, D., and Tadmor, Y. (2005). Not just colors-carotenoids degradation as a link between pigmentation and aroma in tomato and watermelon fruit. Trends in Food Science and Technology, 16, 407-415. Lokvam, J., and Braddock, J. F. (1999). Anti-bacterial function in the sexually dimorphic pollinator rewards of Clusia grandiflora (Clusiaceae). Oecologia, 119, 534-540. Lokvam, J., Braddock, J. F., Reichardt, P. B., and Clausen, T. P. (2000). Two polyisprenylated benzophenones from the trunk latex of Clusia grandiflora (Clusiaceae). Phytochemistry, 55, 29-34. Loughrin, J. H., Hamilton-Kemp, T. R., Andersen, R. A., and Hildebrand, D. F. (2006). Circadian rhythm o volatile emission from flowers of Nicotiana sylvestris and N. suaveolens. Physiologia Plantarum, 83, 492-496. Lunau, K. (2000). The ecology and evolution of visual pollen signals. Plant Syst. Evol, 222, 89-111. Lunau, K. (2007). Stamens and mimic stamens as compoents of floral colour patterns. Botanische Jahrbucher fur Systematik, Pflanzengeschichte und Pflanzengeographie, 127, 13-41. Maccioni, S., Baldini, R., Cioni, P. L., Tebano, M., and Flamini, G. (2007). In vivo volatiles emission and essential oils from different organs and pollen of Cistus albidus from Caprione (Eastern Liguria, Italy). Flavour and Fragrance Journal, 22, 61-65. Majetic, C. J., Raguso, R. A., Tonsor, S. J., and Ashman, T. L. (2007). Flower color-flower scent associations in polymorphic Hesperis matronalis (Brassicaceae). Phytochemistry, 68, 865-874. Makkholela, T., and Manning, J. C. (2006). First report of moth pollination in Struthiola ciliata (Thymelaeaceae) in southern Africa. South African Journal of Botany, 72, 597603. Matsui, K. (2006). Green leaf volatiles: hydroperoxide lyase pathway of oxylipin metabolism. Current Opinion in Plant Biology, 9, 274-280. McGregor, S. E., (1976). Insect pollination of cultivated crop plants. Medrano, M., Herrera, C. M., and Barrett, S. C. H. (2005). Herkogamy and mating patterns in the self-compatible daffodil Narcissus longispathus. Annals of Botany, 95, 1105-1111.

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Guido Flamini

Meeuse, B. J. D., and Raskin, I. (1988). Sexual reproduction in the Arum, lily family, with emphasis on thermogenecity. Sexual Plant Reproduction, 1, 3-15. Miyake, T., and Yafuso, M. (2003). Floral scents affect reproductive success in fly-pollinated Alocasia odora (Araceae). American Journal of Botany, 90, 370-376. Molina-Montenegro, M. A., and Cavieres, L. A. (2006). Effect of density and flower size on the reproductive success of Nothoscordum graminum (Alliaceae). Gayana Botanica, 63, 93-98. Mori, M., Kondo, T., Toki, K., and Yoshida, K. (2006). Structure of anthocyanin from the blue petals of Phacelia campanularia and its blue flower color development. Phytochemistry, 67, 622-629. Moya, S., and Ackerman, J. D. (1993). Variation in the floral fragrance of Epidendrum ciliare (Orchidaceae). Nordic Journal of Botany, 13, 41-47. Müller-Schwarze, D., and Thoss, V. (2008). Defense on the rocks: low monoterpenoid levels in plants on pillars without mammalian herbivores. Journal of Chemical Ecology, 34, 1377-1381. Navarro, L. (2000). Pollination ecology of Anthyllis vulneraria subsp. vulgaris (Fabaceae): nectar robbers as pollinators. American Journal of Botany, 87, 980-985. Ngugi, H. K., and Scherm, H. (2006). Mimicry in plant-parasitic fungi. FEMS Microbiology Letters, 257, 171-176. Noda, K. I., Glover, B. J., Linstead, P., and Martin, C. (1994). Flower colour intensity depends on specialized cell shape controlled by a Myb-related transcription factor. Nature, 369, 661-664. Okamoto, T., Kawakita, A., and Kato, M. (2008). Floral adaptations to nocturnal moth pollination in Diplomorpha (Thymelaeaceae). Plant Species Biology, 23, 192-201. Olesen, J. M., and Valido, A. (2003). Lizards as pollinators and seed disperser: an island phenomenon. Trends in Ecology and Evolution, 18, 177-181. Paracer, S., and Ahmadjian, V., (2000). Symbiosis. New York: Oxford University Press. Pare, P. W., and Tumlinson, J. H. (1999). Plant volatiles as a defense against insect herbivores. Plant Physiology, 121, 325-332. Pellmyr, O. (2002). Pollination by animals. In C. M. Herrera, and O. Pellmyr, Plant-animal interactions, an evolutionary approach (pp. 157-184). Oxford: Blackwell. Pellmyr, O., Tang, W., Groth, I., Bergstrom, G., and Thien, L. B. (1991). Cycad cone and angiosperm floral volatiles:inferences for the evolution of insect pollination. Biochemical Systematics and Ecology, 19, 623-627. Pemberton, R. W., and Wheeler, G. S. (2006). Orchid bees don't need orchids: evidence from the naturalization of an orchid bee in Florida. Ecology, 87, 1995-2001. Penuelas, J., and Lluisia, J. (2004). Plant VOC emissions: making use of the unavoidable. Trends in Ecology and Evolution, 19, 402-404. Penuelas, J., Lluisia, J., Asensio, D., and Munne-Bosch, S. (2005). Linking isoprene with plant thermotolerance, antioxidants and monoterpene emissions. Plant, Cell and Environment, 28, 278-286. Percival, M., (1965). Floral Biology. Oxford: Pergamon Press. Petanidou, T., and Vokou, D. (1990). Pollination and pollen energetics in Mediterranean ecosystems. American Journal of Botany, 77, 986-992.

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Pfunder, M., and Roy, B. A. (2000). Pollinator-mediated interactions between a pathogenic fungus, Uromyces pisi (Pucciniaceae), and its host plant, Euphorbia cyparissias (Euphorbiaceae). American Journal of Botany, 87, 48-55. Philbrick, T., and Les, D. H. (1996). Evolution of aquatic angiosperm reproductive systems. Bioscience, 46, 813-826. Pichersky, E., and Gershenzon, J. (2002). The formation and function of plant volatiles: perfumes for pollinator attraction and defense. Current Opinion in Plant Biology, 5, 237243. Piechulla, B., and Pott, M. B. (2003). Plant scents-mediators of inter- and intraorganismic communication. Planta, 217, 687-689. Plowright, R. C. (1987). Corolla depth and nectar concentration: an experimental study. Canadian Journal of Botany, 65, 1011-1013. Raffa, K. F., and Smalley, E. B. (1995). Interaction of pre-attack and induced monoterpene concentrations in host conifer defense against beetle-fungal complexes. Oecologia, 102, 285-295. Raguso, R. A. (2003). olfactory landscapes and deceptive pollination: signal, noise and convergent evolution in floral scent. In G. J. Blomquist, and R. G. Vogt, Insect pheromone biochemistry and molecular biology: the biosynthesis and detection of pheromones and plant volatiles (pp. 631-650). Amsterdam: Elsevier. Raguso, R. A. (2004a). Flowers as sensory billboards: progress towards an integrated understanding of floral advertisement. Current Opinion in Plant Biology, 7, 434-440. Raguso, R. A. (2004b). Why are some floral nectars scented? Ecology, 85, 1486-1494. Raguso, R. A. (2004c). Why do flowers smell? The chemical ecology of fragrance-driven pollination. In R. T. Cardé, and J. G. Millar, Advances in Insect Chemical Ecology (pp. 151-178). Cambridge: Cambridge University Press. Raguso, R. A. (2008). Start making scents: the challenge of integrating chemistry into pollination ecology. Entom. Exper. Appl, 128, 196-207. Raguso, R. A., Levin, R. A., Foose, S. E., Holmberg, M. W., and McDade, L. A. (2003). Fragrance chemistry, nocturnal rhythms and pollination "syndromes" in Nicotiana. Phytochemistry, 63, 265-284. Raguso, R. A., and Light, D. M. (1998). Electroantennogram responses of male Sphinx perelegans hawkmoths to floral and 'green-leaf volatiles'. Entom. Exper. Appl, 86, 287293. Raguso, R. A., and Pichersky, E. (1999). A day in the life of a linalool molecule: chemical communication in a plant-pollinator system. Part 1: linalool biosynthesis in lowering plants. Plant Species Biology, 14, 95-120. Raguso, R. A., and Roy, B. A. (1998). 'Floral' scent production by Puccinia rust fungi that mimic flowers. Molecular Ecology, 7, 1127-1136. Raskin, I., Ehmann, A., Melander, W. R., and Meeuse, J. D. B. (1987). Salicylic acid: a natural inducer of heat production in Arum lilies. Science, 25, 1601-1602. Reuber, S., Bornman, J. F., and Weissenbock, G. (2006). Phenylpropanoid compounds in primary leaf tissues of rye (Secale cereale). Light response of their metabolism and the possible role in UV-B protection. Physiologia Plantarum, 97, 160-168. Robacker, D. C., Meeuse, B. J. D., and Erickson, E. H. (1988). Floral aroma. Bioscience, 38, 390-398.

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Ronsted, N., Weiblen, G. D., Clement, W. L., Zerega, N. J. C., and Savolainen, V. (2008). Reconstructing the phylogeny of figs (Ficus, Moraceae) to reveal the history of the fig pollination mutualism. Symbiosis, 45, 45-56. Roubik, D. W. (1995). Pollination of cultivated plants in the tropics. FAO Agricultural Service Bulletin, Roy, B., and Widmer, A. (1999). Floral mimicry. A fascinating yet poorly understood phenomenon. Trends in Plant Science, 418, 325-330. Roy, B. A. (1993). Floral mimicry by a plant pathogen. Nature, 362, 56-58. Roy, B. A. (1994). The effects of pathogen-induced pseudoflowers and buttercups on each other's insect visitation. Ecology, 75, 352-358. Roy, B. A. (1996). A plant pathogen influences pollinator behavior and may influence reproduction of nonhosts. Ecology, 77, 2445-2457. Roy, B. A., and Raguso, R. A. (1997). Olfactory versus visual cues in a floral mimicry system. Oecologia, 109, 414-426. Sampson, F. B. (1993). Pollen morphology of the Amborellaceae and Hortoniaceae (Hortonioideae:Monimiaceae). Grana, 32, 154-162. Santos, H., Turner, D. L., Lima, J. C., Figueiredo, P., Pina, F. S., and Macanita, A. L. (1993). Elucidation of the multiple equilibria of malvin in aqueous solution by one- and twodimensional NMR. Phytochemistry, 33, 1227-1232. Sapir, Y., Shmida, A., and Ne'eman, G. (2006). Morning floral heat as a reward to the pollinators of the Oncocyclus irises. Oecologia, 147, 53-59. Sasaki, K., and Takahashi, T. (2002). A flavonoids from Brassica rapa flower as the UVabsorbing nectar guide. Phytochemistry, 61, 339-343. Schaefer, H. M., Schaefer, V., and Levey, D. J. (2004). How plant-animal interactions signal new insights in communication. Trends in Ecology and Evolution, 19, 577-584. Schiestl, F. P., Ayasse, M., Paulus, H. F., Erdmann, D., and Franke, W. (1997). Variation of floral scent emission and postpollination changes in individual flowers of Ophrys sphegodes subsp. sphegodes. Journal of Chemical Ecology, 23, 2881-2895. Schiestl, F. P., and Schluter, P. M. (2009). Floral isolation, specialized pollination, and pollinator behavior in orchids. Annual Review of Entomology, 54, 425-446. Schlangen, K., Miosic, S., Castro, A., Freudmann, K., Luczkiewicz, M., Vitzthum, F., Schwab, W., Gamsjager, S., Musso, M., and Halbwirth, H. (2009). Formation of UVhoney guides in Rudbeckia hirta. Phytochemistry, 70, 889-898. Schoonhoven, L. M., van Loon, J. J. A., and Dicke, M., (2005). Insect-plant biology. Oxford: Oxford University Press. Schwendemann, A. B., Wang, G., Mertz, M. L., McWilliams, R. T., Thatcher, S. L., and Osborn, J. M. (2007). Aerodynamics of saccate pollen and its implication for wind pollination. American Journal of Botany, 94, 1371-1381. Seymour, R. S., and Schultze-Motel, P. (1997). Heat-producing flowers. Endeavour, 21, 125129. Shuttleworth, A., and Johnson, S. D. (2010). The missing stink: sulphur compounds can mediate a shift between fly and wasp pollination systems. Proceedings of the Royal Society B, published on line doi: 10.1098/rspb.2010.0491, Sih, A., and Baltus, M. S. (1987). Patch size, pollinator behavior, and pollinator limitation in catnip. Ecology, 68, 1679-1690.

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31

Silberglied, R. E. (1979). Communication in the ultraviolet. Annual Review of Ecology, Evolution, and Systematics, 10, 373-398. Smith, B. N., and Meeuse, J. D. (1966). Production of volatile amines and skatole at anthesis in some arum lily species. Plant Physiology, 41, 343-347. Solecka, D. (2007). Role of phenylptopanoid compounds in plant responses to different stress factors. Acta Physiologiae Plantarum, 19, 257-268. Song, Q., Yang, D., Zhang, G., and Yang, C. (2001). Volatiles from Ficus hispida and their attractiveness to fig wasps. Journal of Chemical Ecology, 27, 1929-1942. Sprengel, C. K., (1793). Das entdeckte Geheimniss del Natur im Bau und in der Befruchtung der Blumen. Berlin: Stebbins, G. L., (1974). Flowering plants: evolution above the species level. Cambridge (USA): Harvard University Press. Sun, J. F., Gong, Y. B., Renner, S. S., and Huang, S. Q. (2008). Multifunctional bracts in the dove tree Davidia involucrata (Nyssaceae: Cornales): rain protection and pollinator attraction. American Naturalist, 171, 119-124. Takayama, S., and Isogai, A. (2005). Self-incompatibility in plants. Annual Review of Plant Biology, 56, 467-489. Tan, K. H., and Nishida, R. (2007). Zingerone in the floral synomone of Bulbophyllum baileyi (Orchidaceae) attracts Bactrocera fruit flies during pollination. Biochemical Systematics and Ecology, 35, 334-341. Tholl, D. (2006). Terpene synthases and the regulation, diversity and biological roles of terpene metabolism. Current Opinion in Plant Biology, 9, 297-304. Thompson, W. R., Meinwald, J., Aneshansley, D., and Eisner, T. (1972). Flavonols: pigments responsible for ultraviolet absorption in nectar guide of flower. Science, 177, 528-530. Thorp, R. W., Briggs, D. L., Estes, J. R., and Erickson, E. H. (1975). Nectar fluorescence under ultraviolet irradiation. Science, 189, 476-478. Torres, C., and Galetto, L. (2002). Are nectar sugars composition and corolla tube length related to the diversity of insects that visit Asteraceae flowers? Plant Biology, 4, 360-366. Totland, O. (1996). Flower heliotropism in an alpine population of Ranunculus acris (Ranunculaceae): effects on flower temperature, insect visitation, and seed production. American Journal of Botany, 83, 452-458. Toyama-Kato, Y., Yoshida, K., Fujimori, E., Haraguchi, H., Shimizu, Y., and Kondo, T. (2003). Analysis of metal elements of hydrangea sepals at various growing stages by ICP-AES. Biochemical Engineering Journal, 14, 237-241. Utech, F. H., and Kawano, S. (1975). Spectral polymorphisms in angiosperm flowers determined by differential ultraviolet reflectance. Journal of Plant Research, 88, 9-30. van Doorn, W. G. (1997). Effects of pollination on floral attraction and longevity. Journal of Experimental Botany, 48, 1615-1622. Veerman, A., and Van Zon, J. C. J. (1965). Insect pollination of pansies (Viola spp.). Entom. Exper. Appl, 8, 123-134. Verdonk, J. C., Haring, M. A., van Tunen, A. J., and Schuurink, R. C. (2005). ODORANT1 regulates fragrance biosynthesis in Petunia flowers. The Plant Cell, 17, 1612-1624. Vinson, S. B., Williams, H. J., Frankie, G. W., and Shrum, G. (1997). Floral lipid chemistry of Byrsonima crassifolia (Malpighiaceae) and use of floral lipids by Centris bees (Hymenoptera: Apidae). Biotropica, 29, 76-83.

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Vogel, S. (1971). Olproduzierende Blumen, die durch olsammelnde Bienen bestaubt werden. Naturwissenschaften, 58, 58-69. von Helversen, D., Holderied, M. W., and von Helversen, O. (2003). Echoes of bat-pollinated bell-shaped flowers: conspicuous for nectar-feeding bats? Journal of Experimental Biology, 206, 1025-1034. von Helversen, D., and von Helversen, O. (1999). Acoustic guide in bat-pollinated flower. Nature, 398, 759-760. von Helversen, O., Winkler, L., and Bestmann, H. J. (2000). Sulphur-containing "perfumes" attract flower-visiting bats. Journal of Comparative Physiology A, 186, 143-153. Vuilleumier, B. S. (1967). The origin and evolutionary development of heterostyly in the angiosperms. Evolution, 21, 210-216. Walker-Larsen, J., and Harder, L. D. (2000). The evolution of staminodes in angiosperms: patterns of stamen reduction, loss, and functional re-invention. American Journal of Botany, 87, 1367-1384. Warren, J., and James, P. (2008). Do lowers wave to attract pollinators? A case study with Silene maritima. Journal of Evolutionary Biology, 21, 1024-1029. Waser, N. M. (1979). Pollinator availability as a determinant of flowering time in ocotillo (Fouquieria splendens). Oecologia, 39, 107-121. Waser, N. M., and Price, M. V. (1981). Pollinator choice and stabilizing selection for flower color in Delphinium nelsonii. Evolution, 35, 376-390. Waser, N. M., and Price, M. V. (1985). The effect of nectar guides on pollinator preference: experimental studies with a montane herb. Oecologia, 67, 121-126. Weiss, M. R. (1995). Floral color change: a widespread functional convergence. American Journal of Botany, 82, 167-185. Wester, P., and Classen-Bockhoff, R. (2007). Floral diversity and pollen transfer mechanism in bird-pollinated Salvia species. Annals of Botany, 100, 40-421. Whitehead, D. R. (1969). Wind pollination in the angiosperms: evolutionary and environmental considerations. Evolution, 23, 28-35. Whitehead, M. R., and Peakall, R. (2009). Integrating floral scent, pollination ecology and population genetics. Functional Ecology, 23, 863-874. Williams, N. H. (1983). Floral fragrances as cues in animal behaviour. In C. E. Jones, and R. J. Little, Handbook of Experimental Pollination Biology (pp. 50–72). New York: Academic Press. Wodehouse, R. F., (1935). The fundamentals of fruit production. New York: Mc. Graw-Hill. Yoshida, K., Kitahara, S., Daisuke, I., and Kondo, T. (2006). Ferric ions involved in the flower color development of the Himalayan blue poppy, Meconopsis grandis. Phytochemistry, 67, 992-998. Zarsky, V., and Tupy, J. (1995). A missed anniversary: 300 years after Rudolf Jacob Camerarius' "De sexu plantarum epistola". Sexual Plant Reproduction, 8, 375-376. Zhang, X., Chen, T., Zhang, H., and Gao, W. (2007). Headspace solid-phase microextractiongas chromatography-mass spectrometry for analysis of volatile components from pollen pine. Zhongyaocai, 30, 1521-1525.

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Chapter 2

POLLINATION MECHANISMS IN PASSIFLORA SPECIES: THE COMMON AND THE RARE FLOWERS -ECOLOGICAL ASPECTS AND IMPLICATIONS FOR HORTICULTURE M. T. Amela García and P. S. Hoc Depto. de Biodiversidad y Biología Experimental, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, PROPLAME-PHRIDEP (CONICET), Argentina

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ABSTRACT Passionvines have flowers with the following basic architecture: 5 sepals, 5 petals, a corona formed by concentric cycles (radii, pali, operculum, limen) and an androgynophore that bears 5 anthers, the ovary, 3 styles and 3 stigmas. Self-pollination may be achieved but some species are self-incompatible, so pollen vectors are required. The different relative sizes and orientation of the floral pieces of the various Passiflora species have implications on which visitors will pollinate; to perform pollination, they must have the adequate size to contact both anthers and stigmas in the same or in successive visits to different flowers. Pollen removal (from the anthers) and deposition (in the stigmas) is carried out by means of different parts of the body of the different pollinators, depending on their size and behaviour. The anthers are dehiscent and the stigmas are receptive as soon as the flower opens until it closes. The styles move throughout anthesis: they tilt down to the anthers and uplift afterwards. Thus, three floral stages occur: in the first and the third, only the anthers can be contacted by the legitimate visitors while in the second, both the anthers and the stigmas are placed in the way of the pollinators. The style movements succeed in all the studied species. However, in some species, in a proportion of the flowers the styles remain upright since the flowers open. These flowers are not able to receive pollen, neither by the pollinators nor by themselves, so they are functionally staminate. In fewer species, the dehiscence of the anthers does not happen in some flowers, so they are functionally pistillate. Finally, the three types of flowers may coexist in the same plant. This brings about the simultaneous occurrence of

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M. T. Amela García and P. S. Hoc pollen donor-receptor flowers and only pollen donors, pollen donor-receptor and only receptor flowers or the three types of flowers in a single plant, respectively, leading to the corresponding functionally andromonoecious, gynomonoecious or trimonoecious systems. Certain floral traits seem to be associated with the absence of styles movements, such as a less developed gynoecium, minor-sized and nectarless flowers. In this chapter, an update of the recorded aspects at the moment as well as original data are discussed, taking in account the ecological interpretations of style movements, analysing the possible causes of the incidence of the less frequent flowers and considering the implications for fruit production in this edible fruited genus, some species of which are grown commercially.

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INTRODUCTION Passion vines have hermaphrodite flowers with the following basic architecture (Figure 1): 5 sepals, 5 petals, a corona formed by different kinds of pieces arranged in various cycles (radii, pali, operculum, limen) and 5 anthers and a gynoecium with 3 styles and stigmas that stand over an androgynophore. The architectural differences among Passiflora species flowers lie on the total flower size and on the relative sizes of each piece (especially the androgynophore length and the corona pieces) and the floral pieces orientation (especially the radii and anthers), which have implications in the kind (involving size and behaviour) of visitors that will carry out pollination. Other floral traits that vary among species are flower colour, odour and anthesis period. The floral cycles perform movements throughout anthesis situating the reproductive cycles in different positions; based on those changes, three floral stages were defined as follows. In the bud, the styles are upright, the staminal filaments are adnate to the styles and the anthers are parallel to the filaments and facing inwards. As soon as the sterile cycles separate and recurve, the anthers make a spin over their filaments while these bend until they reach a perpendicular plane with respect to the floral axis. Thus, the anther face changes to an outward position (extrorse), remaining in different orientations (Figs. 3 and 4, Table 1), depending on the pollination syndrome. This constitutes the first floral stage (Figure 4, A, B), along which the styles progressively bend, bringing the stigmas towards the level of the anthers. Stage 2 is defined as the time during which the stigmas are at the same level of the anthers, or even lower (Figure 4, C). During the third stage (Figure 4, D), the styles go back to the former position, while the stamens descend and the corona, petals and sepals incurve, closing the flower. Anthers are dehiscent and stigmas are receptive throughout the whole anthesis; so the flowers are not dicogamous. Due to the styles‘ movements, the stigma surface and the pollen are not available (in the way of the pollinators) at the same time during stages 1 and 3 (hercogamy), but they are both available during stage 2, so the hercogamy is temporal, or, what is equivalent, there is functional dicogamy. To carry out pollination, a visitor must have the adequate size and/or behaviour to contact both or either an anther and a stigma in the same or in successive visits to different flowers. Pollen removal (from the anthers) and pollen deposition (on the stigmas) can only be performed by a single pollinator when the stigmas had bent down to the level of the anthers. Pollen removal and deposition is carried out by means of the dorsal thorax of hymenopterans (in the melittophilous species), different parts of the head of hummingbirds (in the

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ornitophilous ones), on the head and neck of bats (in the quiropterophilous ones), on the mouthparts and antennae of sphingids (in the sphingophilous ones). The bees land on the larger pieces of the corona (radii); the rest of the animals hover while they feed. The style movements have been reported for all the species in which the floral biology has been studied (Knuth, 1908; Janzen, 1968; Sazima and Sazima, 1978; Corbet and Willmer, 1980; Girón Vander-Huck, 1984; Kay, 2001; AmelaGarcía, 1999 and others). In spite of this, it has been detected that in certain species a proportion of the flowers do not bend the styles in any moment of the anthesis, remaining straight as when the flowers open. These flowers are not able to receive pollen from the legitimate visitors, i.e., the pollinators, so they become functionally staminate. Other floral traits like a minor size of the gynoecium, minor sized and nectarless flowers seem to be associated with the lack of styles movement. In some species, the anthers do not open in a few flowers, so these flowers become functionally pistillate. The occurrence of non-bending styles flowers, non-dehiscent anthers flowers, or both of them concomitantly with hermaphrodite flowers makes the sexual system functionally andromonoecious, gynomonoecious or trimonoecious, respectively.

Figure 1. Floral architecture. Longitudinal section of flower of Passiflora mooreana showing the floral parts. Drawing by Amela García, M. T. Scale bar = 10 mm. a, androgynophore; ann, annulus; ant, anther; c, cup; ch, nectar chamber; f, staminal filament; l, limen; n, nectary; o, operculum; of, operculum filaments; ov, ovary; p, pali; r, radii; st, stigma; sty, style; t, trochlea.

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M. T. Amela García and P. S. Hoc

Figure 2. Flowers of Passiflora exhibiting different pollination syndromes. A, C, ornitophily; B, melittophily; D, sphingophily. A and C, photographs leg. by J. P. Torreta and O. R. Di Iorio, respectively.

All these related features have not been studied in detail and future research should take them into account, as well as others, such as the possibility of underdeveloped ovules associated with non-bending styles flowers, in which case, the gynoecium would be a pistillode and the system would be addressed as truly andromonoecious. Here, a compilation of original data is presented and it is discussed with the reported aspects at the moment. The ecological significance of the styles‘ movements are interpreted. Furthermore, the traditional concepts of hercogamy and dicogamy are discussed.

MATERIALS AND METHODS In this chapter, the terminology employed and revised by Tillet (1988) is used for flower morphology, without taking into account their origin, but rather their function. The terms used for sex expression in flowers is that of Wyatt (1983); although sexuality is a property of the gametophytes and not of the flowers that bear them, we prefer to call the flowers hermaphrodites when they bear the structures in which both gametophytes form, as it is a conservative denomination.

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Figure 3. Flower opening and anther orientation. A, sepals and petals separating, the anthers face the floral axis except one already spinning; B, 3 anthers spinning (right), 2 anthers facing downwards (left); C, anthers facing outwards; D, anthers facing downwards; a bee landed on the radii and licking. A-B, P. caerulea; C, P. misera; D, P. foetida.

Most of the observations described here refer to species studied in Argentina (Amela García, 1999 and unpublished data), from subgenus Passiflora, supersect. Passiflora: P. caerulea, P. mooreana (sect. Stipulata), P. palmatisecta (sect. Passiflora), P. foetida, P. chrysophylla (sect. Dysosmia) and subg. Decaloba, supersect. Decaloba: P. misera, P. urnaefolia (sect. Decaloba), P. capsularis (sect. Xerogona) and supersect. Cieca: P. suberosa (sect. Cieca), according to the classification of MacDougal and Feuillet (2004). The field and laboratory methodology applied is described in detail in Amela García (1999), and in Amela García and Hoc (1997, 1998). All the photographs were taken by Amela García, M. T., except when quoted as a different author.

RESULTS 1. Floral Architecture and Function The base of the flower is constituted by the hypanthium; this is formed by the floral cup and the floral tube. The first may acquire, mainly, the form of a plate, cup or urn, of varying

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depth, depending on the species; it is the region compressed between the apex of the pedicel and the base of the androgynophore, up to the insertion of the operculum (excluding it) (Figure 1); there, in some species, the tube originates (Figure 2, A), which ends in the insertion of the perianth; the tube may be not developed or reduced only to an edge in the cup. The sepals and petals are inserted on the outer edge. In contrast with most Angiosperms, the sepals contribute to the visual attraction when the flower is open, as they form together with the petals the outline of a star when the flower is observed from the front or above (Figure 2, B), at least in the species in which they are not deflexed (Figs. 2C, 3C, 5C). The corona is found more internally, comprising the pieces that are between the perianth and the androgynophore. The corona is formed by the radii (sing. radius), the pali (sing. palus), the operculum, the nectar chamber, the annulus, the nectary, the limen and the trochlea. The radii are filaments of varying width, length and position (alone and in group), depending on the species; they are arranged in one or more cycles; the conjunct is called nimbus. It has different functions, depending on the pollination syndrome (see section ―Floral architecture variants‖). The pali are shorter filaments, frequently with a wider apex, arranged in one or more cycles, forming a fence (sepimentum) that apparently deters the access to the nectar to non-pollinating visitors. The operculum is a membrane, smooth or with radial fan-like foldings, curved towards the androgynophore, that reaches the upper edge of the limen, constituting an impediment for access to the nectar by nectar thieves and, in the pendant flowers, for the nectar to drop; the smooth operculum fits the limen by means of a crease, constituting a barrier more difficult to transpose than the folded operculum (which only leans on the limen), reinforced sometimes with filaments that originate from its edge. The nectar chamber consists in the cavity formed by the floral cup, closed by the operculum and the limen. The annulus is a ring that emerges from the inner surface of the floral cup that may divide the nectar chamber in an antechamber and an inner chamber. The nectary covers the inner surface of the nectar chamber, not necessarily the annulus. The limen is a membrane, generally placed in the base of the androgynophore or a bit up, over this, where the operculum sits. The trochlea is an annular bulk in the base of the androgynophore, over the limen; it is supposed to restrict the access to the nectar to non-legitimate visitors and reinforce the androgynophore base against the pushing of the bees when they lick (Figure 3, D). The androgynophore separates the androecium and the gynoecium from the reward and, in that way, reduces the damages that the visitors could provoke.

2. Floral Pieces Movements, Floral Stages and Implications in Pollination Among other authors, the floral movements were described by Masters (1871) and MacDougal (1994), although not completely. The movements described in this section correspond to most Passiflora spp. flowers; variants of them are detailed in section 4 (―Floral architecture variants and their relation to pollinators size and behavior‖). In the bud, the anthers filaments are parallel to the floral axis and the anthers are extrorse. When anthesis begins (Figure 3, A), the sepals, the petals and the nimbus expand until each cycle remains campanulate, perpendicular to the floral axis or deflexed, depending on the species (Table 1). The anthers spin on their insertion in their filaments while these filaments, originally parallel to the floral axis, descend backwards and place forming an acute angle with the floral axis; in this way, the anthers remain facing outwards (Figure 3, C);in other species, the filaments

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descend even more, till they rest almost perpendicular to the floral axis; in these species, a spin of 90o takes part between the filament and the connective, changing the anthers from a radial position to a tangential one; thus, the dehiscent face rests facing towards the corona, forming a parallel plane to this one (Figure 3, D). In some species, the anthers rest in an intermediate position between the two ones described (Table 1). The styles, which are parallel to the floral axis in the bud, tilt progressively to reach the same level of the anthers (Figure 4, A-C). The styles‘ movements are useful to define three floral stages (Figure 4). Stage 1 (Figure 4, A-B). The anthers spin over their filaments and these bend from their vertical position to a horizontal one. Progressive bending of the styles until they reach the level of the anthers. Stage 2 (Figure 4, C). The stigmas remain just over the level of the anthers, among or below these (depending on the species). The styles do not always move synchronously, so it must be considered that the flower is in this stage when at least one of the stigmas is in the described position, i.e., plausible of receiving pollen from a legitimate pollinator. Stage 3 (Figure 4, D). Progressive lifting of the styles up to their former position in the bud, parallel to the floral axis. The anthers‘ filaments curve downwards. The radii, the corolla and the calix incurve, thus closing the flower, which does not open again. Although this stage consists in the progressive closing of the flower, it must be taken into account as a third stage, as it is visited and sometimes there is pollen left in the anthers, depending on the visits received in the former stages. The described movements have been observed in P. caerulea, P. mooreana, P. palmatisecta, P. foetida, P. chrysopylla, P. capsularis and P. suberosa. In a few species (P. misera and P. urnaefolia), the lifting of the styles or the bending of the anther filaments does not always occur (Figure 6, C). The stigmas are receptive during the whole anthesis in most of the species in which this state was measured (Amela García, 1999), but in some of them they react more intensely during stage 2, which is in accordance with the stigma‘s position in this instance (capable of receiving pollen from the pollinators). Similar results were obtained by Akamine and Girolami (1959) in P. edulis, who employed different techniques, and by Souza et al. (2004), who measured a decrease of the percentage of receptivity at the beginning of the evening, when flowers must have been changing to stage 3 (although the stage‘s changes were not recorded by these authors). Considering the possibility of contact with the legitimate pollinators in their way to the reward, floral stages 1, 2 and 3 are functionally staminate (pollen donor), hermaphrodite and functionally staminate, respectively. Nevertheless, the second stage could be deemed mainly functionally pistillate (pollen receptor); the pollen offered in this stage depends on the removal done by the visitors during stage 1. If fertilization occurs, all the floral pieces persist (on the contrary, only the bracts) and the ovary begins to increase its size generally at the following day of anthesis. The lifting of the stigmas and the descent of the anthers during stage 3 would reduce the possibility of self-pollination, except in the species of subg. Passiflora, supersect. Passiflora, sect. Dysosmia, in which the styles, after lifting, curve upwards the upper third of their length (Figure 6, D); nevertheless, fruits from self-pollination only were obtained from P. foetida but not from P. chrysophylla (Amela García, 1999). The lifting of the styles, besides, must diminish the possibility of damage or pollen clogging to the stigmas once they have been

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pollinated, especially by some of the pollen robbers or thieves that remain near the anthers or even inside the flowers while these close or when they are already closed.

Figure 4. Styles movements throughout anthesis. A-B, stage 1; C, stage 2; D, stage 3; A,C, D, anthers facing downwards; B, anthers facing outwards. Drawing by P. S. Hoc based on P. caerulea (A, C, D) and P. capsularis (B).

The unique moments when a flower can self-pollinate is when it opens (as the anthers are dehiscent and facing the stigmas and they turn outwards on their filaments afterwards) or when the stigmas descend to the level of the anthers and place among them. In both instances, only the edges of the stigmas can be impregnated with self-pollen. In the cases when during stage 3 the stamens do not descend, there could be a possibility of self-pollination if the anthers contact the edges of the stigmas when these uplift, but the lack of stamens descending in this stage occurs in presumptive self-incompatible species (AmelaGarcía, 1999); besides in the third stage little pollen is left on the anthers if pollinators have visited the flowers assiduously, even fewer in the anthers edges. In the species of Passiflora, the transfer of autogamous and geitonogamous pollen is restricted to the removal that the visitors have performed during stage 1 (for both cases), and, in the case of geitonogamy, to the synchrony in the change of stages between the flowers of a single plant (that is not total) and the quantity of flowers that are visited in each arrival to a

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plant. According to Janzen (1968), the pollinators would mainly transfer pollen that has been deposited on their bodies during the previous visits to flowers in stage 1 of co-specific plants; although some visitors remove it, in the species of Xylocopa a line of pollen remains in the center of the thorax dorsum (Amela García, 1999), a fact that was also observed by Corbet and Willmer (1980). Webb and Lloyd (1986) interpreted erroneously which happens in Passiflora, as they included the genus in a subclass of hercogamy that does not consider the styles‘ movements, nor how the contact with the pollinators occurs; but the genus neither can be placed in another of the subclasses that they defined. As in all of them, they consider that the pollinators contact both reproductive cycles in the same visit. Due to the spatial separation between the anthers and the stigmas during stage 1 (hercogamy), and that the stigmas (although they are receptive) remain out of the way of the pollinators that visit the flowers in this stage (contacting only the anthers), both reproductive cycles are not presented effectively at the same time. With the ulterior movement of the styles, and the location of the stigmas at the same level of the anthers, hercogamy disappears, so, there would be a ―temporal‖ hercogamy (only during stage 1, and again in stage 3). Although during the whole anthesis the stigmas are receptive and the anthers expose the pollen, considering that during stage 2 the stigma receptivity is greater, that in some cases the stigmas descend bellow the level of the anthers and that these are almost empty if the flower has been visited, stage 2 is mainly pistillate; there would be, then, a tendency to dicogamy (stage 1 = staminate; stage 2 = pistillate). The proper Webb and Lloyd (1986) stated that, in general, dicogamy and hercogamy are not equivalent alternatives, and frequently occur together. Lloyd and Webb (1986) commented that, in the flowers of some species that present the pollen first, the stigmas may situate nearer the way of the pollinator than where the anthers have presented, diminishing in that mode the interference with autogamous pollen; this occurs sometimes in Passiflora when the stigmas place below the level of the anthers (Janzen, 1968; Amela García, 1999).

3. The Approach to the Flowers: Floral Orientation Flowers of most Passiflora species are solitary, derived from a higher numbered inflorescence (Ulmer and MacDougal, 2004). Reduction of the peduncle has led to a unique stalk in the majority of the species. The peduncle and pedicel from a continuous unit only interrupted by the abscission layer and the bracts, usually three. What matters for pollination biology is the length of the whole unit, which contributes to the exposition of the flower outside the foliage, so peduncle plus pedicel will be addressed with a unique term (―stalk‖) from here and on. The flowers may appear erect, patent or pendant (Figure 5, A-C). Each species has a typical gravitational orientation, apparently measured by geotropic responses of the buds (MacDougal, 1994). The floral stalk curves with respect to the eventual orientation of the vine carrying branch (more or less horizontal or vertical), in such a way that the flower maintains its proper position. For example, in erect flowers, when the branches are situated more or less perpendicular to the ground, an upwards curvature of the apex of the stalk is observed (Figure 5, D); this orientation is readily observed in the buds (Figure 5, D). When the ovary begins to grow, the stalk changes the curvature and the fruit becomes pendant (Figure 5, D).

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Figure 5. Orientation of Passiflora flowers. A, erect flower; B, pendant flower; C, patent flower; D, progressive change of the curvature of the stalk from the bud to the fruit in an erect flower. A, P. capsularis; B, P. alata; C, P. palmatisecta; D, P. urnaefolia.

4. Floral Architecture Variants and their Relation with Pollinator Size and Behaviour The comparisons with out-groups on the base of the floral morphology and the classic theories of pollination syndromes suggest that the ancestor of the genus was pollinated by insects, probably hymenopterans (Faegri and Pijl, 1976, cited in MacDougal, 1994). Among the genus, melittophily is the most common syndrome (MacDougal, 1994). Ornithophily appeared independently in various lines and the records of pollination by wasps and bats are isolated (MacDougal, 1994). Vogel (l990) suggested pollination by lepidopterans for one species and it is suspected for 2 more (Amela García, 1999).

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Figure 6. Rare flowers or unusual position of floral parts. A, twisted styles (right); B, non-dehiscent anthers; C, non-lifting styles during stage 3; D, lifted styles in stage 3 with the upper third of their length curved downwards. A, P. caerulea; B-C, P. misera; D, P. foetida.

These syndromes usually manifest in the genus as follows: Melitophily (Figure 2, B). Anthesis is diurnal. The flower‘s size ranges from small to large. The visual and olfactory attraction, as well as the surface where the bees land (Fig. 3, D, Table 1), are provided by the radii, which are blue or purplish with white stripes, scented and long. The dehiscent side of the anthers face the radii (Figure 3, D; Figure 4, A, C); the hypanthium is dish or cup shaped (constituted only by the cup, the floral tube is not developed). Generally speaking, two variants can be considered: 1) erect or patent flowers, with the radii extended and the nimbus forming a platform perpendicular to the floral axis; 2) pendant flowers, with the radii fringed in their distal part and the nimbus campanulate (from where the hymenopterans climb). In the erect flowers, the bees land on the outer edge of the radii and walk a bit till they place their mouth parts between the limen and the operculum, separating them; opening in that way allows access to the nectar. After liking, they walk backwards and fly away. In the pendant flowers, the bees may visit legitimately in two ways: 1) entrance and exit of the flower in the same position, 2) the bee changes the body position inside the flower (Varassin and Gomes da Silva, 1998). The different sized melittophilous flowers are pollinated by different sized-bees (Amela García and Hoc, 2001). The length and disposition of the radii conditions the bees of what size could land (Table 1).

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Table 1. Structural characteristics of the hypanthium, radii and other corona pieces involved in the access to the nectar in Passiflora spp. with different pollination systems

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Passiflora sp. (pollinators)

Anther orientation

Hypanthium shape

P. caerulea (large bees)

tangential

P. mooreana (large bees)

Nectar barrier

Radii orientation

operculum

pali

campanulate

smooth, with filaments

short

tangential

campanulate

smooth, with filaments

short

P. palmatisecta (sphingids)

tangential

tubular

smooth, without filaments

absent

P. foetida (medium sizedbees)

tangential

campanulate

smooth, without filaments

short

P. chrysophylla (medium sizedbees)

tangential

dish-shaped

smooth, without filaments

short

P. misera (medium sizedbees)

radial

campanulate

folded

Long

P. capsularis (butterflies)

radial

campanulate

folded

absent

P. suberosa (wasps and bees)

intermediate

dish-shaped, shallow

folded

short

P. urnaefolia (large bees)

intermediate

campanulate

folded

short

Ornitophily. Anthesis is diurnal. The flowers are large. The orientation is erect, patent or pendant. The visual attraction is given by the scarlet sepals and petals (Figure 2, A, C), the radii are short and parallel to the floral axis (thus preventing access to the abundant nectar against the thieves) or reduced (tubercular) without odour (Tillet, 1988). The anthers face outwards (Figure 2, A), the androgynophore is longer than in the melittophilous flowers

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(Figure 2, C) and the hypanthium is tubular or urceolate (constituted by the cup and the floral tube, of varying length, depending on the species). The length of the hypanthium conditions if the flowers will be visited by short or long-billed hummingbirds; flowers with short hypanthiums may be visited either by short or long-billed hummingbirds but flowers with long hypanthiums must be visited by long-billed hummingbirds (Christensen, 1998). These birds contact the reproductive organs with different parts of the head (throat, cheeks, forehead), not the bill (Girón Vander-Huck, 1984; Christensen, 1998). Quiropterophily. Few species show this syndrome (Sazima and Sazima, 1978; Kay, 2001). Most of the anthesis takes part at night. The long stalks make the flowers stand out from the foliage. The flowers are large. The radii are short and completely (P. penduliflora) or more or less (P. mucronata) parallel to the floral axis. The faint odour is fruity (P. mucronata) or musty (P. penduliflora). Most flower parts are greenish (P. penduliflora) or whitish (P. mucronata). P. mucronata erect flowers become asymmetric during their aperture (the anthers and the stigmas remain grouped in semicircle and facing outwards of the plant); instead, P. penduliflora pendant flowers bend the anthers and stigmas a bit, facing the ground. The firm and flexible stalk supports the bats‘ visits (Sazima and Sazima, 1978). Wasp and wasp and bee-pollinated flowers. Anthesis is diurnal. The flowers pollinated by wasps are more or less erect, the radii are not conspicuously colour-banded and the odour is similar to musk or escatol (MacDougal, 1994). In the small P. suberosa flowers, pollinated by wasps and bees (Kosnitchke and Sazima, 1997), the scant radii are incurved at their middle, resting as spider legs seen from the side (Table 1) and the sepals might contribute to support the pollinating insects while they lick (Amela García, 2008). Psicophily. It is suspected in P. jorullensis (Vogel, com. pers.) and in P. capsularis (Amela García, 1999). The suspicion is based on the size of the flowers, the length of the androgynophore, the width of the access to the nectar and the orange coloration (Vogel, com. pers.), the outward orientation of the anthers (Figure 5, A; Table 1), the campanulate nimbus (Figure 5, A; Table 1), diurnal anthesis and the presence of butterfly scales on the flowers (Amela García, 1999). The syndrome could not be confirmed by the observations of pollinators for any of the species by now; only thieves have been recorded for P. capsularis (Amela García, 1999). Phalaenophyly. Pollination by moths is suspected in P. palmatisecta (Amela García, 1999) taking into account the following traits: anthesis before dawn till mid-morning, patent or erect flowers (Figure 5, C), developed floral tube, sepals and petals curved backwards (Figure 5, C), radii apparently not forming a landing surface but instead a highly dissected image in front view with a darker centre that might function as a nectar guide (Figure 2, D), narrow access to the nectar, the stigmas never descend below the anthers (Figure 5, C), whitish colour, intense citric odour. In spite of this, no pollinators could be registered until now.

5. Structural Flower Class and Pollen Transfer FaegriandPijl (1979) included Passifloraamong the "dish-bowl" flower type, with the reproductive organs elevated over the plane formed by the perianth and the nimbus. Endress (1996) included the genus in a more precise classification: among the flowers with canalized access to the reward, which is arranged in a circle (―roundabout flowers‖). Thus, Endress

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(1996) considered the species of Passiflora in which the visitors land on the nimbus, placing the body radially oriented to the flower and moving on it around the androgynophore, walking laterally, describing a circle, separating the operculum and the limen at different points when they probe for nectar; while doing so, the pollination is carried out nototribically (Figure 3, D). So do the hymenopterans. This ―several testes from different points from a circle‖ mode of visit could be applied for hummingbirds, who in fact sometimes lick in a similar way (moving apart from the flower and returning again to a different point), although it is not so frequent and exhaustive, as it depends on the relative position of the flower on the plant, i.e., if the flower is accessible from different sides for these hovering, licking animals. The bees, instead, may fly over the flower and go to the reward from any side, regardless the proximity of the foliage. To get all the nectar in a single bout, the researcher must separate the operculum from the limen in different points. In a equivalent manner, the hymenopterans that access the nectar must not be able to extract all the nectar from a single place. The circular location of the reward compels the pollinator to approach the flower from different points, thus contacting more than an anther and/or a bent stigma.

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6. Anthesis, Flowering Period, Inter-Plant Distance: Attraction and Resource Availability Each individual flower usually lasts from a morning to a day, depending on the species (Amela García, 1999). During the course of the anthesis, the three floral stages succeed, and each single flower does not open again. Meteorological conditions influence the duration of the anthesis, delaying it when it is cloudy, cold, wet, and/or rainy or when the flowers are in the shade (Amela García and Hoc, 1997; Amela García, 1999). In this way, the probability of receiving visits from pollinators increases, as the frequency of them diminishes during such conditions. The passion vines have a long flowering period but the daily production of flowers is scarce (monotonous blooming or steady state). Faegri and Pijl (1979) interpreted that the genus had gregarious flowering. This happens in the species with bigger specimens, such as P. mooreana, P. caerulea and P. urnaefolia (Amela García, 1999), but the opening of flowers per day is minor in the species with smaller specimens, such as P. lutea (Neff and Rozen, 1995) and P. palmatisecta, P. foetida, P. chrysophylla, P. capsularis, P. misera and P. suberosa (Amela García, 1999). In some opportunities, species of the second group can develop specimens of greater size (like P. misera), so they should be included in the first group. The frequency of visits was higher in the species of the first group (Amela García, 1999), surely conditioned, at least in part, by the greater attraction and resource availability. Passion vines grow rather sparsely. Nonetheless, sometimes, a conspicuous group is found. This depends mainly on the distribution performed by the dispersal animals of the zoochorous seeds and the proper conditions for the establishment of the seedlings. The duration of the anthesis being less than 24 hours is common in species with flowers pollinated by trap liners (Endress, 1996). The tropical taxa grow rather sparsely, so they require pollinators capable of long distance travels; these perform daily travels that include predictable resources, previously localized, that utilize regularly during an extended period (Endress, 1996); this coincides with the monotonous flowering of Passiflora. Among the

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trap-liners, species of Ptiloglossa, Bombus, Centris, Xylocopa (Janzen, 1971), sphingids, birds and bats are encountered (Endress, 1996). The pollination by this type of animals constitutes an effective mode of xenogamy. Many self-incompatible species of other genera are pollinated by this type of flyers (Endress, 1996). Passiflora is an eminently tropical genus, so it is probable that co-adaptation has occurred with this type of pollinators, mainly in the self-incompatible species, the majority of which are pollinated by Xylocopa: P. mooreana and P. caerulea (Amela García, 1999), P. edulis f. flavicarpa (Akamine and Girolami, 1959; Corbet and Willmer, 1980; Sazima and Sazima, 1989), by hummingbirds: P. vitifolia (Snow, 1982) and by bats: P. mucronata (Sazima and Sazima, 1978; Kay, 2001). P. foetida, P. chrysophylla, P. misera and P. urnaefolia also receive pollinators that exhibit this behavior (Amela García, 1999). In general, species of the minor sized plants group are partially or totally self-compatible (Amela García, 1999) while species of the larger sized plants group are self-incompatible (Amela García, 1999). In Argentina, Passiflora species bloom usually between the middle of August till March; depending on the weather, flowers may be encountered some months later, even in May (Amela García, 1999). The maximum production of flowers occurs between September and December. At least in the more conspicuous vines (P. caerulea, P. mooreana), the production is slow at the beginning but it increases soon and remains stable for 3 or 4 months, then it decreases. This would function as an attention grabber for the trap-liners, so they remain attracted the rest of the season.

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7. Rare Flower Types 7.1. Non-Bending Styles’Flowers Non-bending styles‘ flowers have been reported in Passiflora from time to time (Harms, 1925, Snow and Gross, 1980, La Rosa, 1984, cited in MacDougal, 1994). This last author suspects that this sexual system might be more frequent in this genus, which includes more than 525 species (MacDougal and Feuillet, 2004), but lacks detailed observation. Lack of styles‘ movements has been detected in P. quadrangularis and P. pinnatistipula (Knuth, 1904 cited in Gottsbergeret al., 1988), P. edulis (Akamine and Girolami, 1957; Free, 1970), P. foetida, P. incarnata and P. alata (Gottsberger et al., 1988; May and Spears, 1988 and Varassin et al., 2001), respectively, in P. exsudans, P. oxacensis and P. sicyoides (MacDougal, 1994), in P. mooreana, P. caerulea and P. chrysophylla (Amela García, 1999), in P. cincinnata (Aponte and Jáuregui, 2002) and in P. misera (this chapter). The relative amount of these flowers is minor than the normal ones in most of the species in which they were counted (Akamine and Girolami, 1959; Gottsberger et al., 1988; Amela García, 1999; Aponte and Jáuregui, 2002). This ratio varies between plants (Ruggiero et al., 1976), along the shoot (MacDougal, 1994) and throughout the flowering season (May and Spears, 1988). Andromonoic species may have constant or labile proportions of male and hermaphrodite flowers, depending on the predictable and efficient service of pollinators or on the uncertainty of these at the beginning of the flowering season, correspondingly, and the optimal allocation of limited reproductive resources (May and Spears, 1988).

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The upright styles of most of the functionally andromonoic species discovered do not move at all from their initial position, but the ―upright‖ ones of P. cincinnata bend a bit and return to the previous position or diminish the degree of curvature afterwards (Aponte and Jáuregui, 2002). This intermediate type of flower was also observed by Akamine and Girolami (1957), Free (1970), Ruggiero et al. (1976) and Aponte and Jáuregui (2002) in P. edulis, in which the styles bend until they form an angle of 45º with the floral axis, not the 90º one of the common flowers. This might be a misinterpretation, at least of the last author for this species, as she registered the styles‘ movements only for 3 hours and the anthesis lasts more (Corbet and Willmer, 1980), so perhaps those flowers would have bent the styles completely later. Moreover, styles‘ movements‘ speed is influenced by meteorological conditions, experiencing a lag when it is cloudy, cold and/or rainy (Amela García and Hoc, 1997). Anyway, partial stylar movements have also been registered in P. mooreana (Amela García and Hoc, 1998). These functionally staminate flowers produce fertile pollen, as evidenced when it was used to artificially pollinate hermaphrodite flowers (Akamine and Girolami, 1957).

7.1.1. Case Study 1: P. caerulea The occurrence of upright flowers and their fate was studied from October to December 1997, in the natural population of P. caerulea growing in the campus of Buenos Aires University. Sixty-six flowers of 7 plants were tagged, the styles movements registered during the day of anthesis and the product of open pollination (flower or fruit) recorded. Most of the styles that did not descend remained upright, parallel to the floral axis, as previously described; but in some flowers, the styles that did not descend were twisted (Figure 6, A). This may be a teratogenic phenomenon, or a strategy that prevents the bending of the styles. Bending-styles‘ and upright-styles‘ flowers coexisted in a same plant, but not all the plants bore upright-styles‘ flowers. Although the movements of the styles could be registered for all the flowers tagged, the product of some flowers were lost, so the set of data are presented separated in Table 2. As it can be seen, the proportion of flowers that bended the styles was substantially greater than the ones with upright styles (Table 2, a); if the flowers in which the products could not be recorded are discarded, the minor sample gives a minor percentage of upright styles‘ flowers (Table 2, b). This low percentage is constituted by 1 flower that set fruit (Table 2, b), which reached maturity. The unique possibility of pollination in the upright-styles‘ flowers is at the flower opening, when it can be impregnated with the pollen of its dehiscent anthers (as described above). P. caerulea is highly self-incompatible, and produces self-fruits in very low percentages, which are mainly seedless (Amela García and Hoc, 1997). Nevertheless, the origin of the pollen on the stigma might have been hardly from another source (exogenous pollen dropped from the body of a pollinator flying over the flower while passing pollen to the hind legs or from pollen deposited by means of unusual behavior of a floral visitor). Other flowers that did not bend the styles, although the product could not be recorded, had pollen on their stigmas. Even based on a low number to make conclusions, the unique flower of P. caerulea that did not bend the stigmas had pollen on them and set fruit, so it was not female sterile. May and Spears (1988), by means of artificial pollinations, detected two physiological flower types among morphologically normal flowers that did not bend the styles: ones that set fruit

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(although in less proportion than hermaphrodites) and ones that did not set fruit, depending on the period of the flowering season during which pollination took place. They attributed this phenomenon to a seasonal component. On the other hand, the upright styles‘ flowers of P. edulis were reported as female sterile by Free (1970) and Ruggiero et al. (1976) when handpollinated, and the upright styles‘ flowers of P. foetida drop after flowering (Gottsbergeret al. 1988). These observations do not contradict those of May and Spears (1988) but they evidence that studies involving the whole flowering season are needed. Table 2. Fruit production of bending and upright styles‟ flowers in a population of P. caerulea in Ciudad Universitaria, Buenos Aires Number of tagged flowers (number of plants) Bending styles (% flowers) Upright styles (% flowers)

66 (7) 92 8

a. Occurrence of flowers with upright and bending styles.

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Number of tagged flowers with recorded products (number of plants) Bending styles flowers (%) Fruits from bending styles flowers (%) Upright styles flowers (%) Fruits from upright styles flowers (%) b. Fruit set in upright and bending styles‘ flowers.

54 (7) 98 92 2 100

7.2. Non-Dehiscent Anthers Flowers In some species, a proportion of the flowers do not open the anthers (Amela García, 1999). In each single flower, no anther opens (P. capsularis) or not all the anthers open at the same time (P. urnaefolia and P. suberosa) (Amela García, 1999). The amount of these flowers is less than the functionally hermaphrodite ones. The late and asynchronous pollen exposition in some flowers of P. urnaefolia and P. suberosa probably are due to the high humidity and long exposition to shade in the forest environments where they were studied, as the dehiscence is produced when the endothecium cell walls dehydrate (Valla, 1999). In P. suberosa, the anthers were not always dehiscent when the flowers opened but they finally did; sometimes they dehisced at the end of stage 1 or during stage 2, when humidity was lower than 67 %, temperature was higher than 17-25 °C and the flowers began to be exposed to the sun (Amela García, 2008). In P. urnaefolia, the anthers were not always dehiscent when the flowers began to open, even in some flowers in stage 2 they were still closed if the flowers were not exposed to the sun, and not all the anthers open simultaneously. But in P. capsularis, in some flowers (more frequently in a patch than in another of two studied ones) the anthers did not open at all (although they had pollen), in spite of all the flowers were exposed to the same microclimatic conditions; in these cases, a physiological factor must be implied. In some of those non-dehiscent anthers‘ flowers, odor was not perceived (Amela García, 1999). In these flowers, with the non-dehiscent anthers until stage 2, there would not be a tendency to dicogamy, i.e., there would not be functional dicogamy, as the stigmas and the pollen are exposed at the same time. In this case, the probability that the pollinators transfer

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autogamous pollen is greater because this pollen has not been removed in the previous stage. On the contrary, the anthers of P. misera, P. palmatisecta, P. caerulea, P. mooreana, P. foetida and P. chrysophylla, species that were studied in sites where they receive sun during the entire day, the anthers were dehiscent as soon as the flowers began to open (Amela García, 1999). Besides, the ambient temperature where P. urnaefolia was growing was rather lower than in the rest of the sites. It is probable that the high relative ambient humidity where the plants of P. capsularis were had also influenced the lack of anthers‘ aperture during the whole anthesis, although it rests to be determined why the aperture occurred only in some of the flowers of a single day. Anyway, regardless of the cause that prevents the anthers‘ dehiscence, the lack of this makes the flowers functionally pistillate. This provokes the concomitantly occurrence of pollen donor-receptor and only receptor flowers in a single plant, in which case the system is functionally gynomonoecious.

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7.3. Case Study 2: Non-Bending Styles’Flowers and Non-Dehiscent Anthers’Flowers: P. misera The occurrence of flower types in this species was recorded in a specimen growing in a garden in Buenos Aires province during January and February 2010. The specimen occupied a space of approximately 2 m high and 5 m width. Daily, 5 ± 2 flowers open (n = 12 days). The flower types that were produced are reported in Table 3. Aberrant androecium implies:     

Anthers do not spin over their filament (partially or completely) Anthers do not open: no anther opens (partially or at all) or only some anthers open Stamens do not bend down to a horizontal plane Stamens‘ filaments shorter than normal ones Stamens‘ filaments lack anther

This fact may occur in some of the stamens of a flower, not in all of them, and aberrant androecium may occur in combination with lack of style movement. For example, in one of the flowers that the anthers did not open, they did not spin over their filament, and a style (together with its corresponding stigma) was contracted like a blower (this style bent a bit but not as much as the rest). Some flowers (in which the androecium was aberrant) were smaller than the normal ones. The teratology consisted in a flower with the ovary laterally bent, one of the stamens‘ filaments shorter than the rest and one of the styles shorter that the other two; surprisingly, this was the unique style of the three that bent. The occurrence of these flower types, especially the teratologic ones, may be due to their belonging from a single isolated specimen without genetic flow of this self-incompatible species (Amela García, unpublished data), which flowers regularly every year but does not produce fruits. When the three types of flowers coexist in the same plant, the system becomes functionally trimonoecious.

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Table 3. Flower types in a specimen of P. misera growing in a garden in Merlo, Buenos Aires province

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Tagged flowers (n) Bending styles (% flowers) Upright styles (% flowers) Aberrant androecium (% flowers) Teratologic flowers (% flowers)

30 64 10 23 3

7.4. Floral Traits Associated with Lack of Styles Movements The gynoecia of the upright-styles‘ flowers in certain species from section Pseudodysosmia (subgenus Decaloba) are not fully developed, yellowish and smaller (MacDougal (1994). In P. caerulea, the stigma colour may be yellowish-green, violet, deep yellowish brown (Amela García, 1999) or creamy green to murrey (Deginani, 2001). Some flowers that have the stigmas both small and green are suspected of not descending the styles, but some of these stigmas descended; flowers with upright styles may have the stigmas of ―normal‖ colour, i.e., brown. So the stigma colour seems to be more associated with intra-plant variation rather than with style movement capacity, although it seems more frequent that non-descending stigmas are green and smaller. The same was observed in P. mooreana (Amela García and Hoc, 1998). Some of the flowers in which the styles did not descend had the stigmas small and of different colour than the usual flowers, but other non-bending styles‘ flowers had them of normal appearance. Moreover, these differences have no relation with intrafloral colour change, as the stigma colour remains constant throughout anthesis; it only had been observed to change in one species, and when the flowers had closed (Amela García and Hoc, 1998). In P. caerulea, not always the three stigmas of a single flower have the same morphology, i.e., one stigma may be normal and the rest atrophied. The lack of complete correlation between stigma size and/or colour with the lack of stylar movements reinforces the suspicion of May and Spears (1988) that a physiological factor is involved. Some of the flowers of P. foetida that did not bend the styles had smaller ovaries than hermaphrodites, some apparently atrophied (Gottsberger et al., 1988). P. incarnata produces upright styles‘ flowers with atrophied ovaries and styles at the beginning of the flowering season (May and Spears, 1988). In the species in which the anatomy of the styles was compared between upright and descending styles‘ flowers, the flowers that bend the styles had more aerenchyma in the medium and upper part of the styles; this tissue was mainly localized in the bending ―face‖ of the styles and these styles were of greater diameter than the ones of upright styles‘ flowers (Aponte and Jáuregui, 2002); the larger amount of this kind of tissue and its distribution is supposed to increase the flexibility and to favor the nastic-like movement (Aponte and Jáuregui, 2002). In the course of a study of the floral biology of P. chrysophylla at El Palmar National Park in Entre Ríos province, Argentina (Amela García, 1999), a few smaller flowers were recorded among the population, with smaller nectar chamber, nectarless, although scented, whose styles did not descend but the stigmas were receptive (Amela García, 1999). The

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nectarless but scented flowers might contribute to long distance attraction with a minor cost for the plant, as they are deceptive. The less proportion of those flowers with respect to the nectar-producing ones would constitute an effective imitative system, because the imitator does not surpass the number of the model (Faegri and Pijl, 1979). In monoic species, unisexual flowers are small (Jong et al., 2008), and among unisexual flowers, female flowers are larger than male flowers (Delph, 1996). The minor sized and upright styles‘ flowers of P. chrysophylla function as males. Flowers of smaller size than the normal one have also been found among a population of P. caerulea growing naturally at El Palmar National Park (Amela García, unpublished data). It remains to be determined if these flowers bend the styles or not.

7.5. Pollination and Fertilization Problems. Implications for Horticulture The economic importance of the passion fruit stands on the production of seeds, as it is the pulp that covers them (arile), the main product consumed. The arile is only developed when the ovule is fertilized. So it is important that the plant set fruits and that those fruits contain seeds. Low fruit set has been recorded more than once (Akamine and Girolami, 1957; May and Spears, 1988 and cites therein; Sazima and Sazima, 1989). Sometimes the ovary grows but few or no seeds are developed inside (Akamine and Girolami, 1957, 1959; Amela García and Hoc, 1997; 1998). This may be due to reception of incompatible pollen, as this fact was observed in self-incompatible species (Akamine and Girolami, op. cit.; Amela García and Hoc, op. cit.) or pollen limitation, such as low frequency of pollinators due to lack of nesting sites or competition by nectar robbers (Sazima and Sazima, 1989). Increasing non-bending styles‘ flowers during the flowering season decreases the potential number of flowers to set fruit. If artificial pollination is desired, at least a portion of the upright flowers would set fruit but they are not morphologically recognizable. So effort in hand pollination of all the flowers, regardless their stylar movements, would be employed if that were the decision to be made. Manual work on passion fruit culture seems common. A wax film covers each fruit after collection in order to increase preservation (Vilela, com. pers.).

CONCLUSION Although some of the species in the genus are partially or totally self-compatible (Amela García, 1999), the temporal hercogamy and the tendency to the functional dicogamy generated by the styles‘ movements favor the genetic material exchange in the population. Different types of flowers occurring along the flowering season have been reported for Phaseolus (Hoc, unpublished data), but in this case the difference stands in that the flowers are chasmogamous or cleistogamous at the beginning and at the end of the flowering period. The following considerations must be taken in account for future research concerning the different flower types:

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

  

53

In some bending-styles‘ flowers the styles are left twisted when they lift; this must not be mistaken with non-bending-styles‘ flowers that have the styles twisted as soon as the flowers open. So observations throughout the whole anthesis are necessary to study this phenomenon More exhaustive studies, dealing with the whole flowering season and in natural populations, are necessary to assess the occurrence of these ―abnormal‖ flowers and their physiological performance More species must be studied in this aspect Artificial pollinations are necessary to discover the physiological capacity of the completely and partially non-bending styles flowers Stigma receptivity must be tested

ACKNOWLEDGMENTS To the Consejo Nacional de Investigaciones Científicas y Técnicas, for the doctoral fellowships and the grant BID-Conicet Nº 548 with which this study (part of the Ph. D. thesis of M. T. Amela García), was financed.

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REFERENCES Akamine, E. K.andGirolami, G. (1957). Problems in fruit set in yellow passion fruit. Hawaii Farm Sci., April, 3-5. Akamine, E. K. and Girolami, G. (1959). Pollination and fruit set in the yellow passion fruit. Hawai Agricultural Experiment Station, Tech. Bull., 39, 1-44. Amela García, M.T. (1999). Biología floral y sistema reproductivo de especies nativas de Passiflora (Passifloraceae) de la Argentina: Ph. D. thesis. Buenos Aires University. Amela García, M. T. and Hoc, P. S. (1997). Floral biology and reproductive system of Passiflora caerulea (Passifloraceae). Beitr. Biol. Pflanzen, 70, 1-20. Amela García, M. T. and Hoc, P. S. (1998). Aspectos de la biología floral y el sistema reproductivo de Passiflora mooreana (Passifloraceae). Darwiniana, 35, 9-27. Amela García, M. T. and Hoc, P. S. (2001). Pollination of Passiflora: do different pollinators serve species belonging to different subgenera? Acta Hort., 561, 71-74. Amela García, M. T. (2008). Breeding system and related floral features of Passiflora suberosa (Passifloraceae) under natural and experimental conditions in Argentina. Bol. Soc. Argent. Bot., 43, 83-93. Aponte, Y. and Jáuregui, D. (2002). Estructura morfoanatómica de los estilos y estigmas en Passiflora edulis f. flavicarpa Degener y Passiflora cincinnata Mast. Anales de Botánica Agrícola, 9, 35-47. Christensen, A. B. (1998). Passionflowers and their birds, bees and butterflies. The ecology and evolution of pollination and herbivory in Passiflora. Ph. D. thesis. Univ. of Aarhus, Denmark. Corbet, S. A. and Willmer, P. G. (1980). Pollination of the yellow passionfruit: nectar, pollen and carpenter bees. J. Agric. Sci., 95, 655-666.

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Deginani, N. G. (2001). Las especies argentinas del género Passiflora (Passifloraceae). Darwiniana, 39, 43-129. Delph, L. F. (1996). Flower size dimorphism in plants with unisexual flowers. In: D. G. Lloyd, and S. C. H. Barret (eds.), Floral biology: Studies on floral evolution in animalpollinated plants (1st. ed., pp. 217-237). NY: Chapman and Hall. Endress, P. K. (1996). Diversity and evolutionary biology of tropical flowers (2nd. ed.). Cambridge: Cambridge Univ. Press. Faegri, K. andPijl, L.van der. (1979). The principles of pollination ecology (3rd. ed).Oxford: PergamonPress. Free, J. B. (1970). Insect pollination of crops. London: Academic Press. Girón Vander-Huck, M. (1984). Biología floral de tres especies de Passiflora: Graduation thesis. Medellín, Univ. de Antioquía. Gottsberger, G., J. M. F. Camargo and I. Silberbauer-Gottsberger. (1988). A bee-pollinated tropical community: the beach dune vegetation of Ilha de Sao Luís, Maranhao, Brazil. Bot. Jahrb. Syst., 109, 469-500. Janzen, D. H. (1968). Reproductive behaviour in the Passifloraceae and some of its pollinators in Central America. Behaviour, 32, 33-48. Janzen, D. H. (1971). Euglossine bees as long-distance pollinators of tropical plants. Science, 171, 203-205. Jong de Tom J., Avi S. and Frank, T. (2008). Sex allocation in plants and the evolution of monoecy. Ecology Research, 10, 1087-1109. Kay, E. (2001). Observations on the pollination of Passiflora penduliflora. Biotropica, 33, 709-713. Koschnitzke, C. and Sazima, M. (1997). Biologia floral de cinco espécies de Passiflora L. (Passifloraceae) em mata semidecídua. Revta. Brasil. Bot., 20, 119-126. Knuth, P. (1908). Handbook of flower pollination. 2. Oxford: Clarendon Press. Lloyd, D. G. and Webb, C. J. (1986). The avoidance of interference between the presentation of pollen and stigmas in angiosperms. I. Dichogamy. New Zealand Journal of Botany, 24, 135-162. MacDougal, J. M. (1994). Revision of Passiflora Subgenus Decaloba Section Pseudodysosmia (Passifloraceae). Ann. Arbor. Mich. American Soc. Plant Taxon. Syst. Bot. Mon., 41. MacDougal, J. M. and Feuillet, C. (2004). Systematics. In: T. Ulmer and J. M. MacDougal (Eds.), Passiflora: passionflowers of the world (1st. ed., pp. 27-31). Cambridge, Timber Press. Masters, M. T. (1871). Contributions to the natural history of the Passifloraceae. Trans. Linn. Soc., 27, 593-645. May, P. G. and Spears, E. E. (1988). Andromonoecy and variation in phenotypic gender of Passiflora incarnata (Passifloraceae). Amer. J. Bot., 75, 1830-1841. Neff, J. L. and J. G. Rozen. (1995). Foraging and nesting biology of the bee Anthemurgus passiflorae (Hymenoptera: Apoidea), descriptions of its immature stages, and observations on its floral host (Passifloraceae). Amer. Mus. Novitates, 3138, 1-19. Ruggiero, C., Banzatto, D. A. and Sánchez-Lam, A. (1976). Studies on natural and controlled pollination un yellow passion fruit (Passiflora edulis f. flavicarpa Deg.). Acta Hort., 57, 121-124.

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Sazima, M. and Sazima, I. (1978).Bat pollination of the Passion flower, Passiflora mucronata in Southeastern Brazil. Biotropica, 10, 100-109. Sazima, I. and Sazima, M. (1989).Mamangavas e irapuás (Hymenoptera, Apoidea): visitas, interacoes e consequencias para polinizaçao do maracujá (Passifloraceae). Rvta. Bras. Entom., 33, 109-118. Snow, A. A. (1982). Pollination intensity and potential seed set in Passiflora vitifolia. Oecologia, 55, 231-237. Souza, M. M., Pereira, T.N.S., Viana, A.P., Pereira, M.G., Texeira do Amaral Júnior, A. and Madureira, H. C. (2004). Flower receptivity and fruit characteristics associated to time of pollination in the yellow passion fruit Passiflora edulis Sims f. flavicarpa Degener (Passifloraceae). Scientia Horticulturae, 101, 373-385. Tillet, S. S. (1988). PassionisPassifloris II. Terminología. Ernstia, 48, 1-40. Ulmer, T. and MacDougal, J. M. (Eds.) (2004). Passiflora: passionflowers of the world (1st. ed.). Cambridge, Timber Press. Valla, J. J. (1999). Botánica: Morfología de las plantas superiores (13 ed.). Bs. As.: Hemisferio Sur. Varassin, I. G. and Gomes da Silva, A. (1998). A melitofilia em Passiflora alata Dryander (Passifloraceae) em vegetação de Restinga. Rodriguesia, 50, 5-18. Varassin, I. G.. Trigo, J. R. and Sazima, M. (2001). The role of nectar production, flower pigments and odour in the pollination of four species of Passiflora (Passifloraceae) in south-eastern Brazil. Bot. J. Linn. Soc., 136, 139-152. Vogel, S. (1990). Radiación adaptativa del síndrome floral en las familias neotropicales. Bol. Acad. Nac. Cs. Córdoba, Argentina, 59, 5-29. Webb, C. J. and Lloyd, D. G. (1986). The avoidance of interference between the presentation of pollen and stigmas in angiosperms. II. Herkogamy. New Zealand J. Bot., 24, 163-178. Wyatt, R. (1983). Pollinator-plant interactions and the evolution of breeding systems. In: L. Real (ed.), Pollination biology (1st. ed., pp. 51-95). N.Y.: Academic Press.

Reviewed by Gabriel Bernardello Universidad Nacional de Córdoba

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Chapter 3

POLLEN TUBE GROWTH AND OVULE ABORTION IN OLEA EUROPAEA (OLEACEAE): A CASE OF OVULE SELECTION? Julián Cuevas1, Luis Rallo2 and Hava F. Rapoport3 1

Departamento de Producción Vegetal, Universidad de Almería, Almería, Spain 2 Departamento de Agronomía, Universidad de Córdoba, Córdoba, Spain 3 Instituto de Agricultura Sostenible, CSIC, Córdoba, Spain

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ABSTRACT A mature olive tree may produce as many as 500,000 flowers, but only 1-2% of them become fruit. Female reproductive success is further diminished since olive flowers contain four ovules, but only one becomes a seed. We studied the roles of pollen tube growth and ovule abortion in determining this single-seeded pattern. To differentiate between the effects of syngamy and seed growth on ovule abortion we compared the onset and progression of ovule senescence in fruiting and fruiting-prevented plants, in fertilized versus unfertilized flowers, and in plants with different rates of seed growth using aniline blue fluorescence. The single-seeded fruit condition in olive is the result of only one pollen tube reaching the ovary and syngamy occurring therefore in only one ovule per flower. The syngamy in one ovule is accompanied by the degeneration of the other three ovules. Rather than a passive process, the onset and progress of unfertilized ovule degeneration of unfertilized ovules are hastened by the growth of the fertilized ovule. The effect of the fertilized ovule is more pronounced within the same ovary, but is also noted in other unfertilized flowers of the same plant. The Effective Pollination Period was shortened in flowers growing near developing fruitlets as a result of the reduced ovule longevity, suggesting that fertilized flowers may reduce the chance of syngamy in other flowers by this mechanism. These results are discussed in regard to



E-mail: [email protected].

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Julián Cuevas, Luis Rallo and Hava F. Rapoport ovule and fruitlet competition, dispersal mode and evolutionary forces underlying the observed seed/ovule ratio.

Keywords: ovule abortion, olive, Olea europaea, seed pattern, seed/ovule ratio

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INTRODUCTION Olive (Olea europaea L.) is a long-lived tree crop extensively cultivated in the Mediterranean Basin for its oil and fruit since ancient times. Its wild relatives, the oleaster forms, are a common component of the Mediterranean vegetation. The olive tree typically has a low fruit/flower ratio, as occurs in many other flowering plants (Stephenson, 1981). Cultivated olives are partially self-incompatible, but even after cross pollination they undergo massive flower and fruitlet abscission by as many as 98% of the initial flowers (Lavee, 1986). Selective abortion of the smaller fruitlets triggered by seed and fruit growth is responsible for such low fruit set (Cuevas et al., 1995). Female reproductive success in olive is further diminished by a low seed/ovule ratio. Olive flowers contain four ovules, but only one becomes a seed. The seed/ovule ratio of a given species seems to be linked to its breeding system and life history (Wiens, 1984), but in other cases it is also related to the fruit dispersal syndrome (Casper and Wiens, 1981; Augspurger and Hogan, 1983; Augspurger, 1986; Ganeshaiah and Uma Shaanker, 1988; Lange et al., 1993; Tybirk, 1993). Drupe fruits represent an extreme case of seed packaging in which single seeds are dispersed in an indehiscent stony endocarp. This single dispersal structure for each flower implies the abortion of all ovules except that which becomes the seed. The selection of that ovule can occur before anthesis, as in plum (Thompson and Liu, 1973), cherry (Eaton, 1959) and peach (Harrold, 1935), or after anthesis as happens in olive (King, 1938). The olive pistil is composed of a relatively large stigma, a short style and a round ovary formed by two fused carpels each containing two unitegmic, tenuinucellate, anatropous ovules (King, 1938). The four ovules with Polygonum type embryo sac development occupy symmetrical positions in the ovary, and are equally sized and viable at anthesis (Cuevas, 1992). Syngamy, however, usually takes place in only one ovule (King, 1938; Altamura et al., 1982). Thus in olive, syngamy determines the functional ovule, i.e. the ovule that will become a seed (Mogensen, 1975), while its lack dictates ovule degeneration. At present, however, we do not know how syngamy in the surplus ovules is precluded and if degeneration of those unfertilized ovules is a passive consequence of the absence of syngamy, or if, on the contrary, its onset and progress are hastened by the developing functional ovules within the common ovary and nearby.

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PLANT MATERIALS AND METHODS Pollen Tube Growth Measurements Pollen adhesion, pollen germination, variations in pollen tube number along the pistil, percentage of fertilized flowers and the proportion of ovules in which syngamy occurred were recorded every other day from 1 to 15 d after anthesis in random samples of 15-20 hand cross-pollinated flowers of two small trees of the cultivar ‗Arbequina‘. The plants, genetically identical clones, equally managed and pollinated in an identical manner, were respectively kept in growth chambers at 25 C, 65-95% relative humidity and 14/10 hr day length and 20 C, 70-95% relative humidity and 14/10 hr day length (referred to as 25 C and 20 C chambers). Pollen of ‗Picual‘ was used for cross-pollination, that was performed using a camel hair paintbrush. Viability of ‗Picual‘ pollen was confirmed by fluorescein diacetate test (Pinillos and Cuevas, 2008) before application. Pollen adhesion for each flower was estimated by the number of pollen grains adhering in three different 0.1 mm2 areas of the stigma surface. Pollen germination was determined by the proportion of those pollen grains forming a pollen tube. Pollen tube growth was followed in squashed pistils stained with 0.1% Aniline Blue solution in 0.1 N K3PO4 using a Nikon epifluorescence microscope with a high-pressure mercury lamp (HBO 100) and the following filter combination: EX330-380, DM400 and BA420. To assess ovule penetration by the pollen tube the ovules were first dissected from the ovaries. Syngamy was considered to occur when a pollen tube penetrated the ovule, and a flower was considered fertilized when that occurred in at least one ovule.

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Ovule Viability Assessment Ovule viability was estimated by the aniline blue fluorescence (ABF) method, which has proven useful in detecting early symptoms of ovule senescence (Pimienta and Polito, 1982; Stösser and Anvari, 1982). ABF detects callose deposition in the nucellus or integuments of the ovules by fluorescence. Fluorescent, senescent ovules are not penetrated by pollen tubes (Martínez-Téllez and Crossa-Raynaud, 1982; Stösser and Anvari, 1982; Vishnyakova, 1991). Ovule degeneration in this study was assessed by observing ovules stained with 0.1% Aniline Blue solution in 0.1 N K3PO4 and illuminated as described above for pollen tube growth measurement. Viable olive ovules analyzed by ABF exhibit a light yellow fluorescence limited to the micropylar area (Figure 1). In contrast, senescent ovules show a progressively more intense and generalized fluorescence (Figures 2-4). To facilitate the study of the factors implicated in ovule degeneration in olive we developed a scoring system based on the intensity and extent of fluorescent areas in the ovules (Cuevas et al., 1994). Although numerical scores can only approximate a continuous process such as ovule degeneration, it permitted us to distinguish among different degrees of ovule degeneration and thus infer information regarding the timing and factors implicated in such degeneration. The scores range from 0 to 3, and represent successive stages of ovule degeneration as follows:

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Score 0: Viable ovule. Orange-colored ovule with a fluorescent-yellowish focus in the micropylar area. Occasionally with scattered fluorescent points in vascular bundles (Figure 1).

Figures 1-4. Ovules stained with aniline blue and illuminated with UV light. Ovules were dissected from the ovary to facilitate observations. Arrows locate the micropylar area. 1. Viable ovule. Orangecolored ovule with a fluorescent-yellowish focus in micropylar area. Abortive ovule degeneration score 0. 2. Early stage of ovule degeneration. The fluorescent micropylar area expands and the vascular bundles begin to form a fluorescent ring. Abortive ovule degeneration score 1. 3. Intermediate stage of ovule degeneration. The intensity of fluorescence increases and extends over the entire ovule surface. The fluorescent ring of vascular bundles is more pronounced. Abortive ovule degeneration score 2. 4. Advanced stage of ovule degeneration. The fluorescence is brightest and occupies the entire ovule surface. Abortive ovule degeneration score 3. All x 154. Scale bar=0.1 mm.

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Score 1: Senescent ovule, early stage. The fluorescent micropylar area expands and fluorescence sometimes also reaches a newer small focus in the chalazal zone. The vascular bundles begin to form a fluorescent ring (Figure 2). Score 2: Senescent ovule, intermediate stage. The intensity of fluorescence increases and extends over the entire ovule surface. The fluorescent ring of vascular bundles is more pronounced (Figure 3). Score 3: Senescent ovule, advanced stage. The fluorescence is brightest and occupies the entire ovule surface (Figure 4). In ovules with very advanced degeneration it is possible that the fluorescence disappears and the ovule becomes blackish.

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Fruiting Versus Fruiting-Prevented Plants To determine the fruit set effect on ovule abortion, four genetically identical and equally treated trees of the cultivar 'Arbequina' were placed in the 20 C (two plants) and 25 C (two plants) growth chambers described above. For one tree in each chamber, pollination and fruit set were prevented by emasculating and bagging every flower immediately prior to anthesis. All flowers of the second tree were hand cross-pollinated at anthesis and on the subsequent 23 days as described above, and then allowed to set fruit. To keep a record of its age, every flower was marked the day of anthesis using different colored threads. Additionally, we assessed ovule viability in samples of 15-20 flowers (60-80 ovules) every 2 days from 1 to 19 days after anthesis and again at 22 days after anthesis. Ovule viability was represented as the percentage of viable, 0-scored, ovules. The fertilized ovules were excluded from the calculation. To estimate the influence of temperature on itself on ovule senescence we compared the process in unpollinated flowers from fruiting-prevented plants in both chambers. This trial served two important additional purposes. First, unpollinated flowers from the plants in which fruiting was prevented indicated the potential ovule longevity; in combination with fertilization time and stigma receptivity, this parameter determines the effective pollination period (EPP) (Williams, 1965) duration. Second, the fruiting plants bearing a mix of fertilized and unfertilized flowers resemble natural conditions and provide information regarding the effects of fertilization in reducing ovule longevity, and indirectly EPP, and a possible relation to fruitlet abscission and fruit set levels.

Fertilized Versus Non-Fertilized Flowers To determine the effect of fertilization and seed growth on ovule abortion we compared the ovule viability and ovule degeneration rate between fertilized and unfertilized, but pollinated, flowers of the fruiting plants described above. As in the prior experiment a flower was considered fertilized when at least one ovule was penetrated by a pollen tube. Ovule viability for each group of flowers was calculated every two days from 1 to 15 days after anthesis as indicated above. The ovule degeneration rate was deduced from the abortive ovule degeneration scores for successive days. Sample size within each group varied due to fertilization level.

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Plants with Similar Level of Fertilization and Different Functional Ovule Growth Rates We compared functional ovule growth rates at different temperatures in plants with similar fertilization percentage in order to separate the effects of ovule growth from the effects of fertilization on ovule abortion. With this aim, we determined the abortive ovule degeneration score every two days from 1 to 15 days after anthesis in random samples of 1520 hand cross-pollinated flowers of two plants kept in the 25 C and 20 C growth chambers. Next, correlation tests by previous logit transformation of the abortive ovule senescence scores (OS) at both temperatures were performed to determine the degree of association with the percentage of fertilization (%F) found, and with the functional ovule length (FOL) measured at each sampling date. Fertilization and abortive ovule degeneration scores were assessed as above. Functional ovule length was measured every two days under the microscope to the nearest 0.01 mm using an ocular micrometer.

RESULTS

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Seed Pattern Determinants Pollen tube behavior was similar at 20 C and 25 C, although some quantitative differences were found. Pollen adhesion averaged 53±6 pollen grains per 0.1 mm2 of stigma surface of which 55±3% germinated at 25 C, whereas 39±5 pollen grains adhered per 0.01 mm2 and 35±4% germination were found at 20 C. Since stigma area in olive flowers may reach 2.5-3 mm2 (Griggs et al., 1975), adhesion and germination values are equivalent to about 1,300 pollen grains attached to the stigma surface and more than 700 germinated at 25 C, and around 1,000 pollen grains adhered at the stigma and 350 germinated at 20 C. Massive pollen tube growth in the stigma could be observed at both temperatures as soon as 1 day after anthesis. Although detailed counts were not made, more than one hundred pollen tubes were often observed growing in the upper part of squashed stigmas. The number of pollen tubes decreased drastically at both temperatures when the pollen tubes reached the upper part of the style and usually only one pollen tube reached the ovary (Figure 5). Many of the pollen tubes appeared to be arrested with their apex swollen, deformed and strongly ABF stained (Figure 6). A slightly slower pollen tube growth was measured at 20 C. Ovule penetration by a pollen tube was recorded by 3 days after anthesis at 25 C and two days later at 20 C (Table 1). The percentage of fertilized flowers fluctuated around 40-60% until 13 days after anthesis, when at both temperatures a drastic increase was observed, likely associated with the preferential abscission of unfertilized flowers (Table 1). No further differences were found between temperatures for either pollen tube behavior in the style or seed pattern. Consequent to the drastic reduction in pollen tube number in the transmitting tissue of the style, very few pollen tubes were present at the base of the style. Thus in 80% of the flowers only one pollen tube was present at the style-ovary junction, with fewer than five pollen tubes in the remaining 20%. Syngamy took place in only one ovule in 95% of the flowers, and syngamy in two or three ovules was observed only in four and two flowers, respectively. Only viable,

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0-scored, ovules were fertilized. In no case was an ovule that showed senescence to any degree observed being penetrated by a pollen tube.

Fruiting Versus Fruiting-Prevented Plants

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Ovule longevity was reduced in the flowers of both fruiting plants. The percentage of viable ovules diminished 6-8 days earlier and declined more rapidly in the fruiting plant than in the fruiting-prevented plant at 25 C as well as that at 20 C (Figure 7). In flowers from the fruiting plant at 25 C, no ovules other than the fertilized ones remained viable 15 days after anthesis. In contrast, at that date 73% of the ovules remained apparently viable in flowers from the plant in which fruit set was prevented. The same trend was observed at 20 C, at which temperature 22% of the ovules from the fruiting plant were viable at 15 days after anthesis, compared to 77% in the non-fruiting plant (Figure 7). Likewise, only fertilized flowers were present on the fruiting trees beyond 15 days after anthesis whereas a high number of unfertilized flowers remained on the non-fruiting trees at 15 days after anthesis and even later. Based on observations carried out at harvest time, no flowers set fruit in unpollinated plants, whereas 3.55% of flowers set fruit at 25 C and 3.88% at 20 C.

Figures 5-6. Pollen tube growth in pistil squashed and observed with aniline blue fluorescence. Sg=Stigma, St=Style. 5. Progressive decrease in the number of pollen tubes (arrow) in the style. Only one pollen tube reaches the style base. The ovary has been removed for image clarity. X 89. Scale bar=0.1 mm. 6. Detail of pollen tubes. Note the swollen, deformed and deeply stained apex of arrested pollen tubes. X 356. Scale bar=25 μm.

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Cuevas et al.

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Figure 7. Percentage of viable ovules in pollinated flowers from a fruiting plant (solid line) and unpollinated flowers from a fruiting-prevented plant (dashed line) at successive times at 25 C and 20 C. Ovule degeneration was assessed with aniline blue fluorescence as described. N=60-80 ovules. According to the nature of the data and the fitting to the equations, sigmoid (Gompertz) curves were assigned. The curve equations were solved by means of the three-point method.

Cuevas et al. Figure 8. Percentage of viable ovules excluding the developing seed of fertilized flowers (solid line) and of unfertilized flowers (dashed line) on the same plants at 25 C and 20 C. Ovule degeneration was assessed with aniline blue fluorescence. Sample size within each group varied with the fertilization level and was between 15-52 ovules except * and ** where N=8 and N=4 respectively. According to the nature of the data and the fitting to the equations, sigmoid (Gompertz) curves were assigned, except for unfertilized flowers curve at 20 C which did not fit well to any sigmoid equation. The curve equations were solved by means of the three-point method.

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Cuevas et al.

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Figure 9. Abortive ovule degeneration score excluding the developing seed for fertilized flowers (solid line) and for unfertilized flowers (dashed line) at 25 C. Only 0-scored ovules were considerd viable. 1, 2 and 3 express increasing degrees of ovule degeneration assessed with aniline blue fluorescence. Sample size within each group varied with the fertilization level and was between 15-52 ovules except * and ** where N=8 and N=4, respectively. According to the nature of the data and fitting to the equations, sigmoid (Gompertz) curves were assigned. The curve equations were solved by means of the three-point method.

Ovule viability patterns were similar at the two tested temperatures. Loss of ovule viability in unpollinated flowers from the fruiting-prevented plants began 13 days after anthesis at 25 C and until 15 days after anthesis at 20 C (Figure 7). Subsequently, there was almost no difference between temperatures in the time course of ovule senescence in fruitingprevented plants (Figure 7, dashed lines). Since fertilization was detected as soon as 3 and 5 days after anthesis (for 25 and 20 C chamber plants, respectively) and stigma receptivity was not limiting (it prolonged beyond sampling date in virgin flowers), EPP can be estimated as 10 days at both temperatures.

Fertilized Versus Non-Fertilized Flowers No clear differences were found in the percentage of viable ovules between fertilized and non-fertilized flowers at 25 C (Figure 8). Clearer differences were found by comparing the abortive ovule degeneration score (Figure 9). Significant differences were found in this parameter between fertilized versus unfertilized flowers starting at 9 days after anthesis (Mann-Whitney U-test; U=187, p=0.01), indicating that the rate of the abortion process was accelerated in the unfertilized ovules sharing the ovary with a developing seed. Differences between ovules from fertilized and unfertilized flowers were higher at 20 C (Figure 8). However, the temporal pattern for unfertilized flowers seems somehow erratic and did not fit to any sigmoid curve (Figure 8).

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Table 1. Fertilized flowers (%), abortive ovule degeneration score and functional ovule length (mm) variation at different times for fruiting plants growing at 20 C and 25 C Days after anthesis

Fertilized flowers (%) Abortive ovule degeneration score Functionala ovule length (mm)

Temp. (C) 20 25 20 25 20 25

1 0 0 0.0 0.0 0.57 0.56

3 0 28 0.2 0.1 0.57 0.67

5 33 60 0.1 0.3 0.58 0.78

7 33 38 0.1 0.5 0.68 1.10

9 60 53 0.3 1.1 0.84 1.42

11 53 63 0.6 1.9 0.97 1.58

13 73 87 1.2 2.0 1.12 3.08

15 87 93 2.2 2.7 1.47 2.94

a

: Before syngamy, the largest ovule. At both temperatures the onset of abortive ovule degeneration () was immediately preceded by the start of functional ovule growth.

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Plants with a Similar Level of Fertilization and Different Functional Ovule Growth Rates At both temperatures the onset of abortive ovule degeneration occurred immediately after the beginning of seed growth (Table 1). However, as might be expected, FOL and %F were highly correlated (r=0.90 at 20 C and r= 0.87 at 25 C) making the separation of their effects on ovule abortion difficult. Although higher for functional ovule size, the correlation coefficients between OS and FOL versus %F were not significantly different either at 20 C (r=0.96 versus r=0.80; p=0.17), nor at 25 C (r=0.89 versus r=0.83; p=0.62). Partial correlations fixing alternatively one of the proposed explanatory variables were then performed. At 20 C, partial correlation between OS and FOL was highly significant (r=0.93; p=0.0007), whereas correlation between OS and %F did not reach statistical significance (r=0.59; p=0.12). At 25 C, the values were r=0.64 (p=0.09) for partial correlation between OS and FOL, and r=0.35 (p=0.40) for correlation between OS and %F.

DISCUSSION The relation between pollen tube behavior and seed pattern in olive was clearly evident. The one-seeded pattern in olive is fixed by the number of pollen tubes reaching the ovary. After intense pollen tube competition in the style, the numerous pollen tubes were reduced to only one entering the ovary (Figure 5) and therefore syngamy took place in only one ovule. An essentially identical control of single-seeded pattern occurs in Macadamia (Sedgley, 1983). In some flowers (around 20%) of this study several, but always less than five, pollen tubes (from hundreds growing in the upper stigma) were observed gaining access to the ovary, leading to an even lower percentage (5%) of flowers with more than one (sometimes two, sometimes three) fertilized ovule. These results emphasize the role of the transmitting tissue of the style as a key checkpoint for pollen selection, although they also suggest that pollen competition may continue to a lower extent in ovary tissues. The same pollen tube

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attrition dynamic have been observed by the first author in a broad range of olives genotypes in unrelated experiments carried out in both growth chambers and in the field. Although syngamy was usually limited to one ovule per flower, degeneration of the unfertilized ovules was not a passive process; on the contrary, the presence of developing seeds actively shortened the lifespan of the unfertilized ovules (Figure 8). In comparing the fate of unfertilized ovules in fertilized versus unfertilized flowers of the fruiting plants we may deduce that the degeneration of unfertilized ovules was accelerated by the fertilization and the subsequent development of the seed. This effect of the growing seed was more pronounced for those unfertilized ovules within the same ovary (Figure 9), but more importantly it was also observed in unfertilized flowers developing elsewhere on the same plant, particularly at 25 C where the developing seed grew faster (Table 1 and comparison of dashed lines at 25 C in Figures 7 and 8). Ample duration (10 days) of the EPP confirms that low fruit set in olive is not related to brief female fertility (Cuevas et al., 2009). On the contrary, the influence of fruit and seed set and development on ovule degeneration in other olive flowers could be interpreted as an early component of fruit competition which later results in intense fruitlet abscission (Rallo et al., 1981; Raporport and Rallo, 1991a). Furthermore, our finding that the effect of the growing seeds extends beyond the ovary barrier suggests that fertilized ovules may prevent syngamy and subsequent fruit set in neighboring flowers by shortening their EPP via reduced ovule longevity. In fact, embryo sac degeneration is frequent in the ovules of abscised pistils (Rapoport and Rallo, 1991a), although it is difficult to assert the extent to which ovule degeneration is a cause or a consequence of pistil abscission. Our observations that the functional ovule competitive effect is greatest over shorter distances (i.e. within the same ovary) are also in agreement with observations that fruit competition is most intense among nearby fruits (Rallo and Fernández-Escobar, 1985). Consistent with this interpretation, unpollinated flowers, when developing in the absence of fruit set and functional ovule influence, have notably longer periods of ovule viability (Figure 7) and pistil retention (Rapoport and Rallo, 1991b; this chapter). Strong competition among ovules of the same flower has been reported in other single-seeded species. Mogensen (1975) suggests that the first fertilized ovule of Quercus gambelii flowers arrests development of the other five ovules either by preventing their fertilization or by causing their abortion. In Cryptantha flava, postfertilization development of one ovule prevents seed set of the other three, often fertilized ovules (Casper and Wiens, 1981), while early ovule thinning increases the probability of seed set in the remaining ovules (Casper 1988, 1990). The same interaction seems to operate in Dalbergia sissoo, where Ganeshaiah and Uma Shaanker (1988) observed that dominant embryo situated at the tip of the pod usurps resources to supernumerary ovules, leading to starvation and death in sibling ovules. In olive, it is difficult to separate the influences of syngamy and seed growth on senescence of the other ovules, because syngamy is rapidly followed by seed growth (Cuevas, 1992; Table 1). However, we could separate their effects on ovule senescence by using differential temperatures to modify the seed growth rate. Partial correlations showed that, at 20 C, OS was significantly related with FOL (p=0.0007) and not with %F (p=0.12), suggesting that the latter parameter only contains redundant information about ovule senescence. The same trend was obtained at 25 C (p=0.09 versus p=0.40). The early timing of ovule abortion in olive and its coincidence with the establishment of size differences between ovules conforms with embryo abortion in surplus ovules of Crypthanta flava (Casper, 1990).

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The close relationship between abortive ovule degeneration and functional ovule growth suggests that competition among ovules within the same ovary in olive may be related to nutritional factors. The involvement of those factors is supported by observations of starch in developing ovules and ovary (Reale et al., 2009). However hormonal factors cannot be overruled because a minimal functional ovule development may be necessary for endosperm production of a sufficient phytohormone levels (see Lee, 1988 and Uma Shaanker et al., 1988 for a documented discussion regarding hormonal and nutritional control of ovule and seed abortion). The single seed pattern observed in olive fits within the relationship between seed pattern and dispersal structure for the Oleaceae (Taylor, 1945). A fixed pattern of ovule abortion in a one-seeded fruit, whether dry indehiscent or fleshy with a stony endocarp, may be the result of evolution against the intense sibling competition that would occur if more than one seed were dispersed together in a closed dispersal structure (Augspurger, 1986; Wiens et al., 1988). In fact, in a study using Hojiblanca‘, an olive cultivar prone to produce biseeded fruits, we found that seeds formed in one seeded drupes were significantly heavier than the seeds from biseeded drupes (Cuevas and Oller, 2002). Although their germination and early growth were not significantly reduced when those seeds were sown separately, the persistence of two seedlings in the same site is expected to be detrimental at the long run. Nevertheless, if dispersal efficiency, survival and/or seedling growth are enhanced by the abortion of the surplus ovules, an explanation for the production of these surplus ovules is needed. Uma Shaanker et al. (1988) stated that seed/ovule ratio reduction in species with one or few seeds per fruit represents an offspring strategy rather than one of the maternal plant. They argue that if the maternal parent's interest were the reduction of the seed/ovule ratio, selection would not favor the production of extra ovules. Uma Shaanker et al. propose rivalry between siblings of different strengths as a plausible explanation for seed/ovule ratio reduction and the ovule development and its subsequent abortion as an expression of parentoffspring conflict. However, the single seeded pattern in olive occurs because only one pollen tube reaches the ovary, so in this species seed/ovule ratio reduction is under maternal (or paternal via pollen tube competition) control, but not under offspring control. Difficulty in explaining the ―waste‖ of ovules from the point of view of the maternal plant underlies the parent-offspring conflict hypothesis. While, we agree with the occurrence of sibling rivalry in plants with postzygotic control of seed pattern, we would further suggest that the production of surplus ovules may provide advantages to the maternal plant. For example Sedgley (1981) proposed a fail-safe mechanism function for the two viable ovules of the single-seeded Macadamia fruit. Since immature and senescent ovules fail to attract pollen tubes to the micropyle (Herrero, 2001; Higashiyama et al., 2001), a species may also increase the time during which ovules at the optimum stage are present in the ovary by the sequential maturation of the ovules. In the olive tree, however, we suggest that the production of extra ovules could be a mechanism to achieve ovule selection. This hypothesis is, at present, merely speculative. However, female gametophyte selection correlates well with the achievement of strong selection of the progeny genotype achieved by intense pollen tube competition (male gametophyte selection) and by the selective abortion of the smaller fruitlets (offspring selection) (Cuevas et al., 1995), which were presumably fertilized later by slower pollen tubes. If pollen selection has gained acceptance as a powerful force in the evolution of flowering plants (Mulcahy, 1979), we do not find any reason to reject the possibility of ovule

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selection occurring in species where the ovules available for fertilization exceeds the number of pollen tubes entering the ovary. The contrary, fertilization at random, would be more troublesome to explain. An explanation of the mechanisms by which ovule competition could be possible follows. The arrival of just one pollen tube at the ovarian cavity of olive flowers results in a scenario in which ovule competition for gaining the favour of that male victor is possible. Since random fertilization is unlikely, the lack of motility of female gametes limits female competition to mechanisms based on pollen tube attractants. Such ovule selection could be carried out by means of the differential production of the factors implicated in pollen tube guidance by the synergid cells (Higashiyama et al., 2001). Short-range pollen tube attractants diffusing from the micropyle of virgin ovules have been identified in Tourneria fornieri (Higashiyama et al., 2001), a species in which the protruding egg apparatus facilitates its observation and manipulation, and in maize, where the strength of the signals guiding pollen tubes to the micropyle correlates well with the maturation of the egg apparatus during the female receptive period (Marton et al., 2005). Since the main source of the pollen tube attractants are the non-transcendent synergid cells, the assessment of ovule strength based on synergid capacity to attract winner pollen tubes must rely on the sisterhood among synergid, central and egg cells in Polygonum type embryo sacs, the type which occurs in olive (Altamura et al., 1982). This hypothesis needs confirmation. This proposed scenario of ovule competition does not annul previous and more intense male competition among several dozens of pollen tubes to gain access to the harem represented by the ovarian cavity. On the contrary, as it happens in the Animal Kingdom, a mechanism based on double (male and female) competition for sexual partners could likely occur as well in the Plant Kingdom. The occurrence of ovule competition deserves further studies, especially if the strength of the pollen tube guidance attractants can be manipulated. This ovule selection scenario is, to the best of our knowledge, the first documented case in which the usual roles of males and females (male competition, female choice) are inversed in flowering plants. Nonetheless, it could very likely be present in other species with supernumerary ovules, and successfully explains the evolutionary conservation of the apparently wasteful formation of equally viable and same-sized ovules occupying symmetrical positions in the ovary of an otherwise one-seeded fruit species. The profound implications that ovule abortion versus seed abortion strategies have on progeny genotype screening, resource savings, pollen-pistil interactions and control of the abortion decision certainly deserve further evaluation and analysis using varied approaches on model plants such as the olive tree.

REFERENCES Altamura, M., Pasqua, G. and Mazzolani, G. (1982). Embryogenesis in Olea europaea L. Annali di Botanica XL, 141-154. Augspurger, C. K. (1986). Double- and single-seeded indehiscent legumes of Platypodium elegans: consequences for wind dispersal and seedling growth and survival. Biotropica 18, 45-50.

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Augspurger, C. K. and Hogan, K. P. (1983). Wind dispersal of fruits with variable seed number in a tropical tree (Lonchocarpus pentaphyllus: Leguminosae). American Journal of Botany 70, 1031-1037. Casper, B. B. (1988). Evidence for selective embryo abortion in Cryptantha flava. American Naturalist 132, 318-326. Casper, B. B. (1990). Timing of embryo abortion and the effect of ovule thinning on nutlet mass in Cryptantha flava (Boraginaceae). Annals of Botany 65, 489-492. Casper, B. B. and Wiens, D. (1981). Fixed rates of random ovule abortion in Cryptantha flava (Boraginaceae) and its possible relation to seed dispersal. Ecology 62, 866-869. Cuevas, J. (1992). Incompatibilidad polen-pistilo, procesos gaméticos y fructificación de cultivares de olivo (Olea europaea L.). Ph.D. Thesis University of Córdoba, Spain. Cuevas, J. and Oller, R. (2002). Olive seed set and its impact on seed and fruit weight. Acta Horticulturae 586, 485-488. Cuevas, J., Pinillos, V. and Polito, V. (2009). Effective pollination period for ‗Manzanillo' and 'Picual' olive trees. Journal of Horticultural Science and Biotechnology 84, 370-374. Cuevas, J., Rallo, L. and Rapoport, H. F. (1994). Procedure to study ovule senescence in olive. Acta Horticulturae 356, 252-255. Cuevas, J., Rapoport, H. F. and Rallo, L. (1995). Relationships among reproductive processes and fruitlet abscission in Arbequina olive. Advances in Horticultural Sciences 9, 92-96. Eaton, G. W. (1959). A study of the megagametophyte in Prunus avium and its relation to fruit setting. Canadian Journal of Plant Science 39, 466-476. Ganeshaiah, K. H. and Uma Shaanker, R. (1988). Seed abortion in wind dispersed pods of Dulbergia sissoo: maternal regulation or sibling rivalry?. Oecologia 77, 135-139. Griggs, W. H., Hartmann, H. T., Bradley, M. V., Iwakiri, B. T. and Whisler, J. E. (1975). Olive pollination in California. California Agricultural Experiment Station Bulletin 869. Harrold, T. J. (1935). Comparative study of the developing and aborting fruits of Prunus persica. Botanical Gazette 96, 505-520. Herrero, M. (2001). Ovary signal for directional pollen tube growth. Sexual Plant Reproduction 14, 3-7. Higashiyama, T., Yabe, S., Sasaki, N., Nishimura, Y., Miyagishima, S-Y., Kuroiwa, H. and Kuroiwa, T. (2001). Pollen tube attraction by the synergid cell. Science 293, 1480-1483. King, J. R. (1938). Morphological development of the fruit of the olive. Hilgardia 11, 437453. Lange, J. H., Van der Walt, J. J. A. and Boucher, C. (1993). Autoecological studies on Audouinia capitata (Bruniaceae). 5. Seed development, abortion and pre-emergent reproductive success. South African Journal of Botany 59, 156-167. Lavee, S. Olive. In: Monselise, S. P., editor. Handbook of fruit set and development. Boca Raton: CRC Press Inc.; 1986; 261-276. Lee, T. D. Patterns of fruit and seed production. In: Lovett-Doust J. and Lovett-Doust L., editors. Plant Reproductive Ecology. Patterns and strategies. New York: Oxford Univ. Press; 1988; 179-202. Martínez-Téllez, J. and Crossa-Raynaud, P. (1982). Contribution à l'etude du processus de la fécondation chez trois espèces de Prunus: P. persica (L.) Batsch., P. cerasifera Ehrh., P. mahaleb L. grâce à l'utilisation de couples de variétés mâle-steriles et mâle-fertiles. Agronomie 2, 333-340.

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Márton, M. L., Cordts, S., Broadhvest, J. and Dresselhaus, T. (2005). Micropylar pollen tube guidance by egg apparatus 1 of maize. Science 307, 573-576. Mogensen, H. L. (1975). Ovule abortion in Quercus (Fagaceae). American Journal of Botany 62, 160-165. Mulcahy, D. L. (1979). The rise of the Angiosperms: A genecological factor. Science 206, 2024. Pimienta, E. and Polito, V. S. (1982). Ovule abortion in 'Nonpareil' almond (Prunus dulcis [Mill.] D.A. Webb). American Journal of Botany 69, 913-920. Pinillos, V. and Cuevas, J. (2008). Stardardization of the fluorochromatic reaction test to assess pollen viability. Biotechic and Histochemistry 83, 15-21. Rallo, L. and Fernández-Escobar, R. (1985). Influence of cultivar and flower thinning within the inflorescence on competition among olive fruit. Journal of the American Society for Horticultural Science 110, 303-308. Rallo, L., Martin, G. C. and Lavee, S. (1981). Relationship between abnormal embryo sac development and fruitfulness in olive. Journal of the American Society for Horticultural Science 106, 813-817. Rapoport, H. F. and Rallo, L. (1991a). Postanthesis flower and fruit abscission in 'Manzanillo' olive. Journal of the American Society for Horticultural Science116, 720-723. Rapoport, H. F. and Rallo L. (1991b). Fruit set and enlargement in fertilized and unfertilized olive ovaries. HortScience 26, 896-898. Reale, L., Sgromo, C., Ederli, L., Pasqualini, S., Orlandi, F., Fornaciari, M., Ferranti, F. and Romano, B. (2009). Morphological and cytological development and starch accumulation in hermaphrodite and staminate flowers of olive (Olea europaea L.). Sexual Plant Reproduction 22, 109-119. Sedgley, M. (1981). Early development of the macadamia ovary. Australian Journal of Botany 29, 185-193. Sedgley, M. (1983). Pollen tube growth in Macadamia. Scientia Horticulturae 18, 333-341. Stephenson, A. G. (1981). Flower and fruit abortion: proximate causes and ultimate functions. Annual Review of Ecology and Systematic 12, 253-79. Stösser, R. and Anvari, S. F. (1982). On the senescence of ovules in cherries. Scientia Horticulturae 16, 29-38. Taylor, H. (1945). Cyto-taxonomy and phylogeny of the Oleaceae. Brittonia 5, 337-367. Thompson, M. M. and Liu, L. J. (1973). Temperature, fruit set and embryo sac development in 'Italian' prune. Journal of American Society for Horticultural Science 98, 193-197. Tybirk, K. (1993). Pollination, breeding system and seed abortion in some African acacias. Botanical Journal of the Linnean Society 112, 107-137. Uma Shaanker, R., Ganeshaiah, K. H. and Bawa, K. S. (1988). Parent-Offspring conflict, sibling rivalry, and brood size patterns in plants. Annual Review of Ecology and Systematic 19, 177-205. Vishnyakova, M. A. (1991). Callose as an indicator of sterile ovules. Phytomorphology 41, 245-252. Wiens, D. (1984). Ovule survivorship, brood size, life history, breeding systems, and reproductive success in plants. Oecologia 64, 47-53. Wiens, D., Davern, C. I., and Calvin, C. L. (1988). Exceptionally low seed set in Dedeckera eurekensis: Is there a genetic component of extinction?. In: Hall C. A. and Doyle-Jones,

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V., editors. Plant Biology of Eastern California. Vol. 2. The Mary DeDecker Symposium, 19-29 Williams, R. R. (1965). The effect of summer applications on the quality of apple blossom. Journal of the Horticultural Science 40, 31-41.

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Chapter 4

APPLICATION OF AIRBORNE POLLEN DATA TO AGRONOMICAL RESEARCH H. García-Mozo Dpto. Botánica, Ecología y Fisiología Vegetal, Edif. Celestino Mutis (C4), Campus de Rabanales, Universidad de Córdoba, Córdoba, Spain

ABSTRACT

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Pollination is only one of the many events comprising the plant development cycle; however, it is extremely important for yield where seed is required. Although successful fertilization depends on a number of environmental and endogenous factors, including climate and plant nutritional status, a sufficient quantity of pollen must reach the receptive stigma in order to enhance fertilization potential. Aerobiological research focuses on the airborne dispersal of biological particles, including pollen grains from anemophilous plants. Airborne pollen data are currently used for various purposes in agricultural research. One major use is as a source of advance information concerning variations in the final fruit harvest of wind-pollinated species. This application, first introduced in the field of plant pathology in the 1940s, was further developed in the 1970s in French studies of vineyard yield; more recently, it has been successfully tested both in crops and in noncrop forest species such as oak or birch. Nowadays, aerobiological research into the influence of pollen emission on final fruit production takes into account a number of other variables, including weather-related factors and phytopathological data; it also uses new, computerized statistical tools to obtain more precise information on agricultural yield and phytopathological risks.



E-mail: [email protected].

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1. INTRODUCTION

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A plant constitutes a complex biological system, some of whose functional units (buds) undergo an annual, genetically-determined development cycle. The cycle comprises both vegetative and reproductive development; both forms involve a number of phenological phases characterized by specific morphological and/or physiological changes. Traditionally, the study of plant phenology relied almost solely on recording the timing of morphological changes; however, more recent research has shown that a deeper analysis of certain key phases (e.g. flowering or fruit ripening) provides a reliable biological evaluation that can usefully be applied to various crops. The phenological phases involved in flowering provide a macroscopic index of a key endogenous process influenced by external factors including soil, climate and crop husbandry. Aerobiology is a multidisciplinary science studying the release, dispersal and deposition of airborne living organisms; it deals with many different types of particles generated by natural or human activities, capable of producing biological effects (Edmonds, 1979). Aerobiological analysis enables the detection of airborne pollen and spores, thus providing information on plant phenology, potential crop production, plant distribution and the health of some species, allowing certain phytopathological risks to be identified. Airborne spore detection enables fungal diseases to be predicted and prevented; it provides valuable data which can be used to model the emission and deposition of phytopathogenic spores within crops, and to predict their transport from one crop to another (Frenguelli, 1998). The objective recording of pathogen spore levels provides the basis for Integrated Pest Management (IPM), a crop-husbandry strategy designed to overcome ecological problems (Figure 1).

Figure 1. Integrated Pest Management Scheme.

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IPM is a sustainable method of managing pests by combining biological, cultural, physical and chemical tools in a way that minimizes economic, health and environmental risks. These new methods must be implemented in three stages: prevention, observation, and intervention. The main goal is to eliminate or significantly reduce pesticide use whilst at the same time maintaining pest populations at acceptable levels. Recent studies using Aerobiology Modeling System (AMS) simulations in conjunction with meteorological information have provided the basis for communications and alerts from plant pathologists to farmers. Another major application of aerobiological data in agricultural research is the forecasting of crop production on the basis of airborne pollen data. Pollination is only one of the many events taking place in the plant development cycle; however, it is extremely important for yield where seed is required. Although successful fertilization depends on a number of environmental and endogenous factors, including climate and plant nutritional status, a sufficient quantity of pollen must reach the receptive stigma in order to enhance fertilization potential (Frenguelli, 1998). Moreover, in anemophilous plants a larger number of pollen grains are required to ensure pollination. Even at a distance of hundreds of kilometers, pollen incidence may be sufficient to effect at least some fertilization (Faegri and van der Pijl, 1979). Long-distance pollen dispersal is of great importance for pollination and seed-setting in isolated specimens, and also for the long-distance transport of genes (Faegri and van der Pijl, 1979). Wind pollination involves an indiscriminate, inefficient dispersal mechanism, and requires very large amounts of pollen in order to insure proper pollination in many crops (Frenguelli, 1998). If a stigmatic surface measures 1 mm2, then 1 million pollen grains distributed evenly over an area of 1 m2 are required for reasonable success in fertilizing a single ovule. The efficiency of wind pollination may be expressed by the equation: n = N x a/A, where n = effective pollen, N = total output of pollen produced, a = stigmatic surface, A = total area of the surroundings (Frankel and Galun, 1977). Pollen production, which is genetically and physiologically controlled, largely determines the pollination process (Allison, 1990; Tormo et al., 1996; Hidalgo-Fernández et al., 2000; Gómez-Casero et al., 2004; Prieto-Baena et al., 2003). Therefore, since wind pollination is a less controlled process than insect pollination, anemophilous plants have a very high ovule/pollen grain ratio averaging 1/500,000 (Tormo et al., 1996). The resulting elevated airborne pollen counts provide the basis for aerobiological crop-forecasting methods. Cour and Van Campo (1980) were the first to demonstrate the link between pollination levels in anemophilous species and subsequent yields; since then, a number of authors have used airborne pollen data as a tool for forecasting grape, olive and cereal crops (e.g. Besselat and Cour, 1990; Galán et al., 2005; Muñoz et al., 2000). Optimized production and reliable crop forecasting are essential for efficient product marketing: armed with an advance estimate of potential yields, producers can adopt the necessary strategies to offset year-on-year variations, and can also take informed decisions on harvest planning, pricing, insurance, and stock management (Gonzalez-Minero et al., 1998). This is especially necessary in the context of common international agricultural policies such as that operated in the European Union, whose farmers must meet production quotas in order to be eligible for subsidies. Over recent years, this application has been tested in non-crop forest species in order to account for variations in fruit production. Although this research is hindered by the absence of fruit production data of the sort available for agricultural crops, tentative results suggest that the considerable year-on-year annual variation in fruit production by anemophilous forest

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species (especially trees) is due largely to differences in pollen production and dispersal (Allison, 1990; Cecich and Sullivan, Litschauer, 2003; García-Mozo et al., 2007).

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2. PHENOLOGY Phenology, a term derived from the Greek phaino meaning ―to show‖ or ―to appear‖, is the study of periodical biological events in the animal and plant world as influenced by the environment (Schwartz, 2003). As soon as the first farmers began to settle, plant seeds, observe crop growth and obtain annual harvests, they became aware of the link between plant development and changes in the environment. The earliest phenological research naturally focussed on agricultural crops, in view of the economic importance of weather-induced effects (Pouget 1963, 1966; Richardson et al., 1974; Ashcroft et al., 1977; Swartz and Powell 1981). Airborne pollen monitoring provides an objective record of the various flowering phenophases in wind-pollinated plants. Phenological analysis enables the complex correlation between climate and floral productivity to be accurately charted in these species; plants are excellent indicators of climate change, since the onset of phenological events is closely governed by weather-related factors. As a result, plant phenology models are increasingly used for a wide range of purposes: predicting the impact of global warming on crops (Galán et al., 2005), improving primary productivity models (Lieth, 1970; Kramer and Mohren 1996), forecasting airborne pollen counts (Chuine et al., 1998; García-Mozo et al., 2009), and supporting foresters and farmers in management decisions such as the selection of reforestation sources in order to prevent frost damage (Cannell et al., 1985; García-Mozo et al., 2001). In general, phenological models are better termed ―pheno-meteorological‖ models, in that they use weather-related parameters to predict phenological events. In floral phenology, air temperature is the variable most influencing the flowering process (Chuine et al., 2003). Most of the variability in pollination onset is accounted for by heat accumulation over the preceding weeks, expressed as ―Growing-Degree-Days (GDDº)‖, especially in tree species. GDDº models must be defined by the start date for heat accumulation and by the threshold temperature above which the plant responds. These parameters may vary depending on the species and the study area. Other major variables in phenological studies include photoperiod and water availability, especially in herbaceous species (García-Mozo et al., 2009a; 2009b). Plant-phenology forecasting is becoming increasingly important in agriculture, since many crop practices – including the application of chemical, biological and hormonal treatments – must be carried out during specific phenological phases. Moreover, the combined monitoring of plant phenology and airborne pathogenic spore counts has been found to enhance the success of IPM strategies. Fungal spore germination occurs only under certain conditions and during specific phenological phases (Aira et al., 2009). Planning of chemical and biological treatments can thus be improved by taking into account not only spore thresholds but also favorable phenological phases. Aerobiological monitoring also has ecological applications. Analysis of airborne pollen data can provide an indication of species distribution, and can be used to monitor weed invasion. Aerobiological data thus serve as bioindicators of environmental change: in some areas of Central Europe, for example, the invasion of Ambrosia has been observed as a weed

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in summer crops. Ambrosia, Artemisia and other ruderal species are highly resistant to pollutants, and are seen as a sign of environmental decline; increased airborne pollen counts for these species, coupled with a decrease in tree-pollen counts, are thus indicative of the biodeterioration of vegetation.

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3. AGRICULTURAL PRODUCTIVITY Pollination is a key factor for crop yield. Although, theoretically, one pollen grain per ovule would be sufficient for fertilization, in several wind-pollinated plants the average number of pollen grains reaching the stigma ranges from 5 to 20 (Stefani, 1992). Seasonal pollen yields vary considerably and, though pollen output per plant also varies widely between species, most wind-pollinated species release relatively large amounts of pollen (Tormo et al., 1996). Pollen emission is the result of a long period of development, usually starting the year before, in late summer. The amount of pollen available for the next year is predetermined, since the cells designated to become pollen grains are already present. In anemophilous tree species flowering in early spring, such as Corylus and Betula, meiosis is observed in August or early September (Frenguelli et al., 1993; 1994). Therefore, for winterdormant trees the pollen yield depends on temperature and rainfall over the previous months. The stored resources of any plant are strained when both pollen and seeds are produced in large quantities. In many trees, variations in fruit production are due to the alternation of high-pollen-emission and low-pollen-emission years. Aerobiological data provide information not only regarding the timing and the trend of the phenophase, but also regarding its magnitude. Airborne pollen counts are an indicator of the amount of pollen actually produced by the plant. Numerous studies have reported a close link between the quantity and quality of emitted pollen and fruit production in windpollinated plants (Campbell and Halama, 1993; Frenguelli et al., 1998). Pollen data can provide information regarding the final fruit harvest several months in advance. This application, first developed in the 1970s in France by Cour (1974), has been successfully tested in both anemophilous crops and non-crop forest species (e.g. Galan et al., 2004; 2008; García-Mozo et al., 2007). Knowledge of the major biological and climate factors influencing the final harvest is becoming increasingly necessary in order to obtain reliable crop estimates and thus ensure optimized, effective crop management. This knowledge is also of great value to public agricultural institutions, for the planning of government subsidies (Sinclair and Seligman, 2000). Early and effective crop forecasting is proving essential in optimizing human and economic resources for harvesting, marketing strategies, and global commercial distribution. This is of particular importance for crops such as olive or grapes in Europe, which are major targets of European Union (EU) agricultural policy (Abassi, 2001). EU regulations establish production quotas, assign economic aid in cases of harvest loss due to weather-related disasters, encourage the planting or abandoning of certain crops, and establish channels of communication among producing countries to prevent market shortages and uncontrolled price rises in low-production years. Until now, the most widely-used forecasting methods were based on plot censuses, in which the observation of a limited number of plots provided an agronomic inventory from which the total production of a region could be

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extrapolated (Lletgos, 1987; Riera Mora, 1995). This forecasting method has certain drawbacks (Panigai and Moncomble, 1988; Besselat and Cour, 1990):

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a) Plot yield estimates are often affected by observer subjectivity. b) The method is costly because it requires numerous observation points. c) The earliest estimates often show an excessive margin of error, which can be corrected only in the period close to harvesting. As a result of these drawbacks, since the 1960s a number of authors have advocated forecasting methods based on the correlation between airborne pollen counts and fruit production in both cultivated and forest species (Sarvas, 1962; Hyde, 1963). The widely-used method developed by Cour and Van Campo (1980) has subsequently been applied to a range of crops, including olives, vines, cereals, citrus fruits and hazelnuts (Pinchon, 1983; Abid, 1984; Cour and Villemur, 1985; Lletjos et al., 1993; Gónzalez-Minero et al., 1998). The olive tree originated thousands of years ago in the eastern Mediterranean, and later spread westwards. The adult plant is estimated to have a million flowers that are either unisexual or hermaphrodite and are arranged in bunches (Abid, 1984). It is an amphiphilous species: primarily insect-pollinated, but with secondary wind-pollination. The fruit is a drupe from which olive oil is obtained. A large amount of farmland is devoted to olive production in the Mediterranean area (Bonazzi, 1997). Because of these floral, palynological and cultural characteristics, high airborne olive-pollen counts are recorded in many European Mediterranean regions. In southern Spain, Gónzalez-Minero et al. (1998) monitored olive pollen counts using a Cour trap; analysing their data in conjunction with agricultural yields and meteorological observations, they developed a forecasting method based on simple and multiple regression. They devised three sets of forecasting equations: for early July (the end of flowering, and six months before fruit picking); for late November (immediately before picking); and for late January (once fruit picking was over). Airborne pollen data have been used to determine optimum harvest dates in vineyards in France, Spain and Portugal (Gónzalez-Minero and Candau, 1995; Ciruelo et al. 1998; Jones and Davis, 2000; Ribeiro et al., 2005; Cunha et al., 2001): these studies generally note a trend towards earlier harvest dates. A correlation has also been detected between pollen counts and grape production, although the monitoring of fungal spores is essential in order to evaluate the impact of phytopathological diseases. Regression equations therefore take into account the effect of post-flowering growing conditions, and a minimum of 3-4 years are required to build reliable models. Analysis of results obtained in France indicate a strong correlation between estimated and real vine crops, with a mean R coefficient of 0.90 (Besselat and Cour, 1990). This method has proved effective in other anemophilous woody crops such as the hazelnut (Corylus avellana L.), for which Riera-Mora (1995) developed a forecasting equation capable of predicting fruit production up to 7-8 months prior to harvest. Over recent years, Hirst volumetric pollen traps (Hirst, 1952) have proved to be an accurate tool for crop forecasting, especially for olives – to which most research has been devoted (Galán et al., 2004; 2007; García-Mozo et al., 2008; Fornaciari et al., 2005; Orlandi et al., 2004). Most equations combine olive-tree phenology, airborne pollen counts, weather data and fruit production data to yield accurate results.

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Using a Hirst trap, Muñoz et al. (2000) evaluated the correlation between Poaceae pollen counts and cereal yields in Central Spain. The chief findings were a strong correlation between June pollen counts and dryland cereal yields (wheat, barley and triticale), and a lack of correlation between pollen counts and irrigated-crop yields (maize, rice and sorghum). A significant correlation was recorded between mean overall pollen counts in May and June and mean cereal yields, although this is likely to reflect the similar effect of environmental conditions on the wild flora producing most of the airborne pollen, and on cereal crops. Finally, attempts have been made to forecast fruit production in non-crop tree species, and especially in woody species such as Quercus (Cecich and Sullivan, 1999; García-Mozo et al., 2007), Taxus (Allison, 1990), and Betula (Litschauer, 2003), all of which are characterized by highly-variable fruit production. Various hypotheses have been put forward to account for the alternation between high and low production, although the variables involved remain unknown. In evergreen species such as Quercus, the resource-matching and seed-dispersal hypotheses have been scientifically ruled out by Koening et al. (1994). Other studies generally support the ―predator satiation‖ and ―wind pollination‖ hypotheses (Docousso et al., 1993; Koening et al., 1994; Vázquez, 1998); the results obtained applying the pollen-count method support the ―wind pollination‖ hypothesis. Combined use of aerobiological, field phenological and meteorological data could represent a major step forward in forest fruit production research. Apart from its ability to provide advance estimates, the pollen-count method has other advantages: deviations are lower than in the test-plot forecasting system; fewer collecting data points are needed; and it is more objective than other methods. However, the pollen-based forecasting method has certain limitations, due mainly to the lack of research programs and to the difficulty in calculating pollen-transport distances. Lack of knowledge of post-flowering factors is an additional major problem in Mediterranean areas. Improved definition of climate-related equations will help to overcome this difficulty and realize the full potential of this method. A further disadvantage is the neeed to establish the average distance over which pollen grains are transported in order to evaluate the fertilization potential in many plants.

4. AEROBIOLOGY AND PLANT PATHOLOGY Aerobiological data enable the distribution, ecology and concentration of fungal spores to be determined. Spores dispersed in the air can travel long distances. Airborne spore monitoring provides information on daily and hourly spore counts in a given crop. In 1946, Stakman and Christensen were the first researchers to apply aerobiological methods to plant pathology. A number of authors have since sought to correlate the extent of disease at a given time with airborne spore counts at the same time or previously (Jeger, 1984). Airborne spore counts are a bioindicator of the phenological cycle of pathogens. In the case of grapevine leaf attack by botrytis blight, a significant correlation was found between airborne conidia counts and lesions appearing one week later (Carise et al., 2008). In these cases, aerobiological data are more useful than weather data for detecting infections at an initial stage (inoculum), although the combined use of weather and spore-count data provides a valuable tool for the development of accurate, modern Integrated Pest Management (IPM) strategies. When the farmer knows the spore risk thresholds, spore counts can serve as a disease alert if weather

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conditions are favorable (Bugiani et al., 1995). The weather conditions favoring spore germination are usually humidity and dew temperature. The strategy most widely adopted by winegrowers to reduce the impact of fungal disease is the systematic application of chemical fungicides, generally following preset calendars based on the phenological growth stages of the grapevine (Bugiani et al., 1995). However, integrated control methods are associated with reduced application of chemical treatments, and with lower economic and ecological costs, e.g. 50%-80% saving of chemical sprays in the fight against Phytophora infestans (Bugiani et al., 1995). Reduction of chemical residues also leads to an improvement in wine quality; the value of wines produced under IPM conditions is thus greater (Albelda et al., 2005; Aira et al., 2009). Recently, several authors have combined aerobiological, phenological and meteorological data to produce equations for forecasting spore concentrations; in some cases, these equations account for up to 40% of spore-count variability when the variables with the highest correlation coefficients are included as estimators (Fernández-González et al., 2009). Over the last few years, certain dry areas of the Mediterranean area traditionally devoted to rainfed farming have been switched to irrigation. This may prompt an increase in the incidence of pathogenic fungi, which are more easily dispersed by irrigation than by rainsplash; since humid environments increase the active discharge of spores, heavy rain and irrigation favor the presence of certain airborne spore types (Gregory, 1973; Fitt et al., 1989).

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REFERENCES Abassi, F., 2001. Olive oil: a balanced world market. Olivae 89, 30-35. Abid, A. 1984. Contribution à l‘étude de la pollinisation de l‘olivier (Olea europaea). Université de Montpellier II. Thèse Doctorel. Montpellier. p. 91. Aira, M.J.; Fernández-González, M.; Rodríguez-Rajo, F.J. and Jato, V. (2009).- Modelo de predicción para Botrytis cinerea en un viñedo de Galicia (España). Boletín Micológico 24: 27-35 Albelda, Y.; Rodríguez-Rajo, F.J.; Jato, V. and Aira, M.J. (2005).- Concentraciones atmosféricas de propágulos fúngicos en viñedos del Ribeiro (Galicia. España). Boletin Micológico 20: 1-8. Allison, T.D., 1990. Pollen production and plant density affect pollination and seed production in Taxus canadensis. Ecology 71, 516-522. Ashcroft, G.L., E.A. Richardson and Seeley, S.D. A statistical method of determining chill unit and growing degree hour requirements for deciduous fruit trees, Hort Sci., 12, 347348, 1977. Besselat, B. and Cour, P. 1990. La prévision de la production viticole á l‘aide de la technique. Capture du pollen. Inf. Tech. CEMAGREF 78(3):1-4. Bonazzi, M. 1997. ―Les politiques Euro-Mediterraneennes et l'huile d' olive: Concurrence ou partage du travail?‖ MEDIT 3: 27–32 Bugiani, R., Govoni, P., Bottazi, R., Giannico, P., Montini, b., Pozza, M. 1995. Monitoring airborne concentrations of sporangia of Phytophora infestans in relation to tomato late blight in Emilis Romagna, Italy. Aerobiologia, 11:41-46.

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Campbell D.R., Halama K. J. 1993. Resource and pollen limitations to lifetime seed production in a natural plant population. Ecology, 74: 1043–1051 Cannell, M.G.R., Murray, M.B. and Sheppard, L.J. Frost avoidance by selection for late budburst in Picea sitchensis. J. Appl. Ecol., 22, 931-941, 1985. Carise, O., Savary, S., Willocquet, L. Spatiotemporal relationships between disease development and airborne inoculum in unmanaged and managed Botrytis leaf blight epidemics. Phytopathology 98(1):38-44. Cecich, R. A., Sullivan, N.H., 1999. Influence of weather at time of pollination on acorn production of Quercus alba and Quercus velutina. Canadian Journal of Forest Reseach. 29(12), 1817-1823. Chuine, I. Cour P. and Rousseau, D.D. Fitting models predicting dates of flowering of temperate-zone trees using simulated annealing. Plant, Cell and Env., 21, 455-466, 1998. Chuine, I., Kramer, K. and Hänninen, H. 2003. Plant Development Models. In: Schwartz, M.D., editor. Phenology: An Integrative Environmental Science. Dordrecht, Netherlands, 2003:217-237. Ciruelo, A., Pardo, C., Riera-Mora, S., Sotés, V. 1998. Consideraciones estadísticas referentes a la estimación precoz de producción de vino mediante el método aeropolínico. Viticultura y Enología Profesional 55: 5-18. Cour, P., Van Campo, M. 1980. Prèvisions de recortes a partir du contenu pollinique de l‘atmosphere. C.R. Acd. Sci. Paris 290, 1043-1046. Cour, P., Villemur, P. 1985. Fluctuations des émissions polliniques atmospheriques et previsions des recoltes des fruits. 5è Colloque sur les Recherches Frutieres. Bordeaux. Novembre 1985. Cour, P. 1974. Nouvelles techniques de détection des flux et retombées polliniques. Etude de la sédimentation des pollens et des spores à la surface du sol. Pollen et Spores 16: 103– 141. Docousso, A., Michaud, H. and Lumaret, R.1993. Reproduction and gene flow in the genus Quercus L. Ann. Sci. For. 50 (1): 91–106. Edmonds, R.L. 1979. Aerobiology: The Ecological System Approach. Stroudsburg, USA: Dowden Hutchinson and Ross. Faegri, K. and van der Pijl, L. 1979 The principles of pollination ecology. Pergamon Press, Oxford, New York Fernández-González, M., Rodríguez-Rajo, F.J. and Jato, V. and Aira, M.J. (2009).-Incidence of fungals in a vineyard of the denomination of origin Ribeiro (Ourense- NW Spain). Ann. Agric. Environ. Med. 16:263-271. Fitt, B.D.L., McCartney, H.A. Walklate, P.J. 1989. The role of rain in dispersal of pathogen inoculums. Annu. Rev. Phytopathol. 27:241-270. Frankel R, Galun E. 1977. Pollination mechanisms, reproduction, and plant breeding. Heidelberg: Springer-Verlag. Frenguelli, G, 1998., The contribution of Aerobiology to Agriculture. Aerobiol. 14, 95-100. Frenguelli, G., Ferranti, F., Fornaciari, M., Romano, B. 1993. Male flower development and pollination in hazel (Corylus avellana L.). In: XV International Botanical Congress, Yokohama, 1993:446. Frenguelli, G., Spieksma, F.Th.M., Ferranti, F., Fornaciari, M., Nikkels, H.A., Romano, B. Preliminary data about the growth of birch catkins in relation to pollen development. In: The Fifth International Conference on Aerobiology, Bangalore, India.

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Galán C., Vázquez L., García-Mozo H. and Domínguez E. 2004. Forecasting olive (Olea europaea L.) crop yield based on pollen emission. Field Crops Research, 86:43-51. Galán, C., García-Mozo, H., Vázquez, L., Ruiz, L., Díaz De La Guardia, C. and Domínguez, E. 2008. Modelling olive (Olea europaea L.) crop yield in Andalusia Region, Spain. Agronomy Journal.Vol 100(1); 98-104. Galán-Soldevilla, C; Garcia-Mozo, H.; Vazquez, L., Ruiz-Valenzuela, L; Díaz De La Guardia, C. and Trigo-Perez, M. 2005. Heat requirement for the onset of the Olea europaea L. Pollen season in several places of Andalusia region and the effect of the expected future climate change. International Journal of Biometeorology, Volume 49, Number 3, Pages: 184 – 188 Garcia-Mozo H, Orlandi F ,Galan C, Fornaciari M, Romano B, Ruiz L, Diaz de la Guardia C, Trigo MM. and Chuine I. 2009a. Olive flowering phenology variation between different cultivars in Spain and Italy: modelling analysis. Theoretical and Applied Climatology. 95:385:395 García-Mozo H., Galán, C., Belmonte, J., Bermejo. D., Candau, P., Díaz de la Guardia., C., Elvira, B., Gutiérrez, M., Jato, V., Silva, I., Trigo, M.M., Valencia, R. and Chuine, I. 2009b. Predicting the start and peak dates of the Poaceae pollen season in Spain using process-based models Agricultural and Forest Meteorology. Vol 149: 256-262. García-Mozo, H., Gómez-Casero, M.T., Dominguez, E. Galán, C. 2007. Influence of pollen emission and weather-related factors on variations in holm-oak (Quercus ilex subsp. ballota) acorn production. Environmental and Experimental Botany 61:35-40. García-Mozo, H., Hidalgo, P., Galan, C., Gómez-Casero, M.T. And Dominguez-Vilches E. 2001.Catkin frost damage in mediterranean cork-oak (Quercus suber L.). Israel Journal of Plant Science, 49:41-47. García-Mozo, H., Perez-Badía, R., Galán, C. 2008. Aerobiological and Meteorological factors‘ influence of olive (Olea europaea L.) crop. Aerobiologia, 24:13–18. Gómez-Casero, M.T., Hidalgo, P., García-Mozo, H., Domínguez, E., Galán, C., 2004. Pollen biology in four Mediterranean Quercus species. Grana, 43,1-9. González Minero, F. J., Candau, P. 1995. La aeropalinología como modelo de previsión de cultivos: los viñedos del condado de Huelva. Polen (7): 59-63 González- Minero, F.J., Candau, P., Morales, J., Tomas, C. 1998. Forecasting olive production based on ten consecutive years of monitoring airborne pollen in Andalusia (Southern Spain). Agr. Ecosyst. Environ. 69:201-215. Gónzalez-Minero, F.J., Candau, P., Morales, J. and Tomás, C. 1998. Forecasting olive crop production base don ten consecutive years of monitoring airborne pollen in Andalusia (southern Spain). Agriculture, Ecosystems and Environment 69:201-215. Gregory, P.H. 1973. The microbiology of the Atmosphere. 2nd ed. London. Leonard Hill. Hidalgo-Fernández, P., C. Galán Soldevilla, E. Domínguez-Vilches.; 2000. Pollen production of the genus Cupressus. Grana, 38,296-300. Hirst, J.M., 1952 An automatic volumetric spore Annals of Applied Biology, 39(2):257-265. Hyde, H.A. 1963. Pollen-fall as a means of sedes prediction in certain tres. Grana 4:217-230. Jeger, M.J. 1984. Relating disease progress to cumulative numbers of trapped spores: apple powdery mildew and scab epidemics in sprayed and unsprayed orchard plots. Plant Pathol. 33:517-523.

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Jones, G.V. and Davis, R.E. 2000. Climate Influences on Grapevine Phenology, Grape Composition, and Wine Production and Quality for Bordeaux, France Am. J. Enol. Vitic. 51:3:249-261 Koening, W.D., Mumme, R.L., Carmen, W.J. and Stanback M.T. 1994. Acorn production, by oaks in central coastal California, variation within and among years. Ecology 75: 99– 109. Kramer, K. and Mohren, G.M.J. 1996. Sensitivity of FORGRO to climatic change scenarios: a case study on Betula pubescens, Fagus sylvatica and Quercus robur in the Netherlands. Clim. Change, 34, 231-237, 1996. Lieth, H. Phenology in productivity studies, in Analysis of temperate forest ecosystems, 1, edited by D.E. Reichle, pp. 29-55, Springer Verlag, Heidelberg, 1970. Litschauer, R., 2003. Untersuchungen zum Reproduktionspotential im Bergwald. FBVA-130, 79-85. Lletgos, Ll. 1987. La previsión de cosechas. Revista de Fruticultura 2(3): 23-29. Lletgos, Ll., Bartroli, R., Esteban, A. and Riera, S. 1993. Forecasting hazelnut (Corylus avellana L.) crop production based on monitoring airborne pollen concentration 4th Int. Symp. On Fruit, Nut and Vegetables Production Engineering, Valencia-Zaragoza. Muñoz, A.F., Silva, I. and Tormo, R. 2000. The relationship between Poaceae pollination levels and cereal yields. Aerobiologia 16: 281-286. Orlandi, F., Romano, B., Fornaciari, M. 2005. Relationship between pollen emission and fruit production in olive (Olea europaea L.) Grana 44(2): 98–103. Panigai, L., Moncomble, D. 1988. La previsión de recoltes en champagne. Le Vigneron Champanois 6 : 359-367. Pinchon, O. 1983. Contribution al étude du pollen et de la pollinisation du pommier (Malus pimula Miller) et prévisions de recolte à partir de l‘analyse du contenu pollinique de l‘atmosphere. D.E.A. Agronomic. Ecol. Nat. Sup. Agron. De Montpellier. Pouget, R. 1963. Recherches physiologiques sur le repos végétatifs de la vigne (Vitis vinifera L.) : La dormance des bourgeons et le mécanisme de sa disparition, INRA, Paris. Pouget, R. 1966. Etude du rythme végétatif : caractères physiologiques liés à la précocité de débourrement chez la vigne. Annales de l‘amélioration des plantes, 16, 6-100. Prieto-Baena, J.C., P.J. Hidalgo, E. Domínguez, C. Galán., 2003. Pollen production in the Poaceae family. Grana, 42,153-160. Ribeiro, H., Abreu, I., Cunha M., Mota ,T. and Castro, R. 2005. Aeropalynological study of Vitis vinifera in the Braga region (1999–2003) Aerobiologia 21(2): 131-138. Richardson, E. A., S.D. Seeley, and D.R. Walker, a model for estimating the completion of rest for « Redhaven » and « Elberta» peach trees. Hort. Science, 9, 331-332,1974. Riera-Mora, 1995. Estimación de cosechas en cultivos leñosos a partir del contenido polínico de la atmósfera. Fruticultura Profesional 98: 17-29. Sarvas, R. 1962. Investigations on the flowering and seed crop of Pinus sylvestris. Comm. Inst. Forest Fenn. 53(3):1-198. Schwartz, M. D. 2003: Phenology: An Integrative. Environmental Science. Kluwer, Netherlands. Sinclair, Th., Seligman, N., 2000. Criteria for publishing papers on crop modelling. Field Crops Research 68, 165-172. Stakman, E.C., Christensen, C.M. 1946. Aerobiology in relation to plant disease. Bot. Rev. 12:205-253.

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Stefani, A., 1992; Pollination and Productivity. In: V Congreso Nazionale. Association Italiana de Aerobiologia, Montecatini, Italy. pp. 197-201. Swartz, H.J. and L.E. Powell. The effect of long chilling requirement on time of bud break in apple, Acta Horticulturae, 120, 173-177,1981. Tormo, R., Muñoz, A., Silva , I., Gallardo, F., 1996. Pollen production in anemophilous trees. Grana 35, 38-46.

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Chapter 5

2N POLLEN FORMATION: 40 CYTOLOGICAL MECHANISMS OF NUCLEAR MEIOTIC RESTITUTION

*

Nataliya V. Shamina Institute of Chemical Biology and Experimental Medicine, Siberian Branch of RAS, Novosibirsk, Russia, [email protected] The illustrated catalogue of meiotic division abnormalities, preferably cytoskeleton aberrations in karyo- and cytokinesis, leading to 2n gametes formation in plants; includes 40 meiotic restitution mechanisms in pollen mother cells.

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Keywords: division spindle, cell plate, pollen mother cells (PMCs), nuclear restitution, phragmoplast, cytokinesis, cytoskeleton, meiosis, 2n-gametes, plant cell division Gametes with diploid chromosome number play a considerable role in higher plant evolution and speciation and are an important instrument in breeding (Mendiburu, Peloquin, 1976; Peloquin et al., 1999; Jauhar, 2003; Cai and Xu, 2007). In angiosperms, about 30% 80% of species were estimated to be of polyploid origin. Polyploidisation is a key evolutionary process in higher plants that lead to the formation of new species. The majority of higher plant species evolutionised this way, or by means of hybridization with more or less widely related species and further polyploidisation. Such important phenomenon as gametophytic apomixis is also associated with polyploidy (Estrada-Luna et al., 2002; d‘Erfurth et al., 2009). Apomixis is a specific reproduction method that allows to obtain absolute genetic copies of mother plants and is related with the process of 2n gametes formation. Despite impressive advancements of genetic engineering in development of transgenic plants, wide hybridization remains the most important and, so far, indispensible method to obtain breeding material. It is explained by the thing that most of the traits important for breeding have polygenic control and cannot be transferred by transformation. Overcoming hybrids F1 sterility is the key problem of plant wide hybridization (Udall and Wendel, 2006). Under complete or partial absence of homologs conjugation, regular chromosome distribution To Munikote Ramanna, who encouraged me to study nuclear restitution mechanisms in higher plants Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

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in two subsequent meiotic divisions becomes impossible. Developing microspores have their aneuploid chromosome number and are non-viable. Rapid development of fertile wide hybrids is possible due to sexual polyploidisation realized by 2n gametes. Such gametes form in parents or in hybrids (allohaploids) as a result of meiotic restitution process (Consiglio et al., 2004). Restitution is understood as absence of segregation of daughter or parent genomes during gametes formation. These aberrations may occur in each of two meiotic divisions, also in preand postmeiotic mitoses. If, as a consequence of such abnormalities, the results of the first meiotic division are exterminated, the first division restitution (FDR) with the formation of 2n nonreduced gametes proceeds. As for the second division restitution (SDR) – 2n reduced gametes develop (see review Hermsen, 1984). Non-reduced 2n gametes from taxonomically distant parents fuse and initiate fertile progenies, and integration of different species genomes is of hybrid advantage for the organism. Allopolyploid species are very viable and widely spread. Many of them are important agricultural crops. Wide hybridization combined with sexual polyploidization is the most important stage in the evolution for the majority of cultivated plants (Hutchinson, 1953). The first such example is the Raphanobrassica hybrid obtained at the beginning of the 20th century crossing 2n gamet producents (Karpechenko, 1927). Though it had a cabbage root and radish leaves, it laid ground for such wonderful synthetic agronomic forms as triticale (wheat-rye amphidiploid) and others. It is well known that, in certain plant cross variants, wide F1 hybrids set seeds both in backcrosses and self-pollination (Love, Craig, 1919; Muntzing, 1939; Mitsuoka, 1953; Tanaka, 1959). When studying chromosome behavior at meiosis in fertile wide F1 hybrids of monocotyledonous species, it was detected that certain aberrations of chromosome segregation in the first, second or both meiotic divisions is the condition of viable gametes formation in them. Frequent are the reports on viable gametes formation as a combination result of low level of homeological chromosome conjugation, univalents equational division type at anaphase I and the subsequent chromosome non-segregation in the second meiotic division (Маап, Sasakuma, 1977; Sasakuma, Kihara, 1981; Xu and Joppa, 2000). A number of phenomena that lead to the restitution nuclei formation in the first meiotic division with the following equational division in the second meiosis and development of viable dyads of microspores were described. In these cases the reasons for restitution nucleus formation may be drastically unequal univalents distribution between the poles (Sears, 1953; Avers, 1954; Stefani, 1986, Jauhar, 2003), absence of their movement in anaphase I (Rhoades, Dempsey, 1966; Wagenaar, 1968; Fukuda, Sakamoto,1992) and even the movement of segregated univalents from the poles to the cell centre again (Riley, Chapman, 1957). It was also reported on the formation of restitution nuclei as a result of cytokinesis absence (Bielig et al., 2003; Barba-Gonzalez et al., 2005; Risso-Pascotto et al., 2006). It is obvious that these meiotic restitution processes are the result of some aberrations in the division spindle cytoskeleton apparatus and, possibly, phragmoplast, but there was no cytological analysis carried out with the structures visualization in these contributions. Meiosis in dicotyledonous species is characterized by the so-called simultaneous cytokinesis which proceeds at telophase II and immediately autonomises 4 haploid nuclei located in the common cytoplasm by means of 6 phragmoplasts activities (Tiezzi et al., 1992). Nuclei are the products of the first meiotic division also located in the common cytoplasm, as cytokinesis does not proceed at telophase I. These peculiarities provide additional possibilities for the reunion of segregated chromosome groups and condition original meiotic restitution

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mechanisms in this taxonomic group (Carputo et al., 2000; Andreuzza and Siddiqi, 2008). At the same time, some restitution mechanisms observed monocots are typical of dicots. It is known that meiotic restitution in dicotyledonous plant species can be realized by means of 1) chromosome non-segregation at anaphase I or II (Lam, 1974; Gill et al., 1985); 2) premature cytokinesis after the first meiotic division and the absence of chromosome segregation in the second one (Mok, Peloquin, 1975; Gill et al., 1985; Watanabe, Peloquin, 1993); 3) univalents equational division at anaphase I in asynaptic meiosis and absence of the second meiotic division (Gustafsson, 1935; Ramanna, 1983; Jongedijk et al., 1991; Vorsa, Ortiz, 1992); 4) absence of the second meiotic division (Conicella et al., 1991; Werner, Peloquin, 1991); 5) disorientation and shift of second division spindles (parallel and ―tripolar‖ spindles configurations) (Mok, Peloquin, 1975; Ramanna, 1979; Genualdo et al., 1998; El Mokadem et al., 2002; Nelson et al., 2009); 6) fused spindles at metaphase II (Ramanna, 1979; Veilleux et al., 1982; Gill et al., 1985); 7) undetected division spindle abnormalities (Qu and Vorsa, 1999); 8) cytokinetic aberrations (Ramanna, 1974; Werner and Peloquin, 1991), 9) chromosome non-segregation in the first and second post-meiotic mitoses (Prakken, Swaminatham, 1952; Bastiaanssen et al., 1998); 10) aberrations of pre-meiotic mitosis (Prakken, Swaminatham, 1952). 2n-gametes, in most cases, form as a result of restitution nucleus formation in the first or the second meiotic division. Caryo- and cytokinetic abnormalities are the base for the restitution nucleus formation process. Separation of chromosome groups in the cell space and their further autonomisation are realized by transient cytoskeleton structures: division spindle and phragmoplast (see rev. Mathur, 2004; Wasteneys and Yang, 2004). Unfortunately, despite the key role which cytoskeleton structures play during meiotic restitution, investigations in this field are not numerous (Alfano et al., 1997; Genualdo et al., 1998). It is explained with several reasons. First, it is because the lack of knowledge about the cytoskeleton rearrangements cycle during plant cell division. The centrosome and polar spindle organizers have not been identified as morphological structures in plant cell; thus organelles that control the cytoskeleton cycle are not known for it (Mineyuki, 2007). Transient stages of cytoskeleton cycle in plant cell division have also not been sufficiently studied. Second, slow progress in studying cytoskeleton mechanisms of meiotic restitution is explained by the known methodic difficulties: immunocytochemical methods of cytoskeleton structure visualization have their own restrictions (sampling problem), whereas the percent of meiocytes in which restitution proceeds is not always sufficiently high. In the combination, these two circumstances considerably complicate investigations. Third, there exists the clear notion of interdisciplinary isolation of experts in cell biology specialized in cytoskeleton studies and those involved in breeding programs and mostly dealing with the analysis of chromosome behavior during restitution (Ramanna, 1974, 1979, 1983; Mok, Peloqun, 1975; Werner, Peloquin, 1991). However, there are efficient classical, but unfortunately forgotten methods for cytoskeleton structure visualization which are very suitable for the above-described tasks. They consist in the use of acetoformol fixation (according to Bouin, Chamberlain, Navashin) that preserves the cytoskeleton with further staining using common stains for protein (acetocarmine, acetoorseine, etc.). The value of these methods consists in, first, their simplicity, cheap price and low labour costs; it allows to analyse comparatively big amounts of material and to trace the division even in a low percent of abnormal cells. Second, they are not different in their expressed sampling problem, - loss of information during making up the

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preparation; it provides the analysis of synchronized PMCs of only one anther or its separate chamber for the correct identification of abnormal meiotic stages. And, finally, it is very important that alongside with the cytoskeleton and chromosomes, classical visualization methods allow us also to observe the nuclear envelope, nuclear zone, cell membranes, cell plate, conglomerates of membrane vesicules which are indispensible for the correct definition of abnormal cell division stage. Application of these methods in studying higher plant meiocytes is most effective due to these big cell sizes. To analyse abnormal meiosis, whose results are presented lower, I used the method of Navashin modified fixation (Wada, Kusunoki, 1964). Under meiosis, buds were being fixed for 24 hours at room temperature using the modified Navashin fixative on the following protocol:  

Solution A: 1,1 g CdCl2, 10 ml glacial acetic acid, 65 ml distilled water; Solution B: 40 ml of 40% (or 37%) formol, 35 ml of distilled water.

The solutions are prepared separately and mixed in equal volumes. The material can be preserved in the fixative at room temperature during a year, and up to 3 years at +4oC without quality loss. Before making up the preparations, it is necessary to rinse buds with running water. Anthers are stained on slide under heating in a drop of 3% acetocarmine; then they are squashed with cover glass. The images were made in white light at magnitude 100 x 10.

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1. NORMAL CYTOSKELETON DYNAMICS DURING POLLEN MOTHER CELLS MEIOTIC DIVISION Before the description of abnormal phenotypes, for better orientation in the cytoskeleton cycle course, I present an illustration of trivial meiotic division with successive cytokinesis in PMCs of wheat x wheatgrass (WWG) hybrid F1 (Shamina, 2005a, b; Shamina et al., 2007). Trivial is the phenotype in which other abnormalities are not observed, but the absence of homological chromosome synapsis. The meiotic process with simultaneous cytokinesis of wild type tomato PMCs is also illustrated. The cytoskeleton cycle and meiotic chromosome cycle do not differ from each other in mono- and dicoteledonous plant species till the telophase stage.

1.1. Cytoskeleton Cycle during First Meiosis with Successive Cytokinesis in Monocotyledonous Species At early prophase, the zygothene stage, chromosomes are in the local nuclear zone attaching with their telomeres to the limited nuclear envelope region forming the ‗‘bouquet‘‘ configuration. At this stage, the cytoskeleton fibres radiate from the nuclear envelope surface to the cytoplasm periphery (Fig. 1.1, a). During pachythene, chromosomes (in WWG F1 hybrids they are univalent) realize from the bouquet and are distributed all over the nuclear volume. The cytoskeleton begins to consolidate around the nuclear surface (Fig 1.1, b). In diplothene-diakinesis, a cytoskeleton ring forms around the nucleus in the meridional plane

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(Fig 1.1, c). Further on, simultaineously with the nuclear envelope breakdown (NEB) and the onset of prometaphase, the perinuclear ring also decays into constitutive elements – microtubule bundles (Fig 1.1, d), which then straighten, turn and enter the former nuclear area. As a result, a chaotic net of cytoskeleton elements (MT bundles) (Fig 1.1, e) which attach to chromosome kinetochores, developing spindle kinetochore fibers, also with each other, developing bipolar central spindle fibers is forming. At late prometaphase (Fig. 1.1, f), the developed kinetochore fibers with attached univalents and bipolar central fibers orient along the division axis, interact with each other, converge on the poles and form the division spindle (Fig. 1.1, g). Univalents, as a rule, are of reductional orientation, i.e. each carries one kinetochore and is aligned with only one of the poles. Therefore, the metaphase plate is absent; though at early metaphase, univalents randomly distributing over between the poles consolidate on the spindle equator. At anaphase, univalents are scattered over the spindle body and gradually reach the poles (Fig. 1.1, h). At early telophase, the spindle consists of a bundle of central fibers (early phragmoplast), which are the base of the forming phragmoplast, and aneuploid telophase chromosome groups. Then the cell plate forms on the telophase spindle equator (Fig 1.1, i); the central spindle fibers surround it having a hollow cylinder configuration (mid phragmoplast). Then its fibers progressively curve and lengthen. As a result, their central points centrifugally move to the cell periphery (late phragmoplast) (Fig. 1.1, j, k). Having reached the mother cell membrane, cell plate membrane vesicules (plastosomes) fuse forming daughter cell membranes, and cytokinesis is completed. A dyad with aneuploid members is the division product (Fig 1.1, l). The second meiotic division in the trivial phenotype proceeds normally, a tetrad of aneuploid non-viable microspores is the meiotic product.

а

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Figure 1.1. Process of the first meiotic division in PMCs of wheat x wheatgrass hybrid F1 of trivial phenotype: a) zygothene, chromosomes are in bouquet configuration; cytoskeleton represents radial MT bundles; b) pachythene, chromosomes realize from the bouquet; c) diakinesis, a perinuclear cytoskeleton ring is forming; d) nuclear envelope breakdown and perinuclear ring disintegration at the prometaphase onset; e) chaotic prometaphase stage (mid-prometaphase); f) late prometaphase I, univalents assemble on the spindle equator imitating the metaphase plate; g) conventional metaphase I – anaphase I; h) late anaphase I – early telophase I; i – k) successive stages of telophase I with phragmoplast/ cell plate centrifugal movement; l) dyad.

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1.2. Cytoskeleton Cycle in Male Meiosis in Dicotyledonous Species with Simultaneous Cytokinesis Cytoskeleton and chromosome behavior at meiotic stages till telophase I in PMCs with simultaineous cytokinesis is not different from those having in PMCs with successive cytokinesis (Fig. 1.2, a – h). The images of cytoskeleton structures at prophase of PMC of wild type tomato, placed here to illustrate meiosis of dicots, is slightly different from that described for WWG F1 hybrids. Thus, there is no visible cytoskeleton perinuclear ring and also its intermediate formation stages at prophase and its disassembly at early prometaphase found. It is explained by the dual cytoskeleton behavior type at prophase stage: radial fibers, moving in the cytoplasm during perinuclear system formation can preserve their morphology and be found during rearrangements, or can depolymerise. It occurs as shortening towards the nucleus. Such short bundles undergo the same movements to the tangential position to the nuclear surface and form a very thin perinuclear ring invisible with classical methods and revealed using the immunostaining (Shamina, 2005a). This thin ring exists up to the nuclear envelope breakdown and the beginning of prometaphase; then the ring disintegrates, its short MT bundles get longer and enter the nuclear zone forming a chaotic prometaphase figure. Complete microtubule cytoskeleton depolymerisation and complete disappearance of ring perinuclear antitubuline staining at prophase I was not observed by us in any of the species analysed using immunostaining method (tomato, potato, tobacco, maize, wheat, rye). Our observations showed that this or that cytoskeleton behavior type at prophase (rearrangements of long or shortened fibers) is encountered with this or that frequency in PMCs of all investigated species regarding both mono- and dicotyledonous plants. Sometimes, cells with developed perinuclear rings, just as without them, can be observed among PMCs within one anther at light microscopic level. The cytoskeleton cycle without depolymerisation at prophase of 100% PMCs is realized in the wild type meiosis of Elytrigia elongatum and common wheat Triticum aestivum for cultivar Albidum 114, and also in the half of the wide cereal hybrids (WWG and wheat x rye WR F1 hybrids) we analysed. Out of 105 cross variants we analysed for F1 of WWG F1 hybrids, in 53 of them, developed perinuclear rings were frequently encountered at prophase I, i.e. their either prevailed or appeared in 100% of PMCs. In the rest of cross variants, PMCs with developed perinuclear rings were constantly present as an admixture in all the anthers. Apparently the same picture was observed in the wheat-rye F1 hybrids we analysed, also in maize and rice haploids. After the division spindle formation and anaphase process, telophase chromosome groups at early telophase I become separated by the central spindle fibers (Fig. 1.2, h). At mid telophase, on the poles, mass polymerization of new MT bundles directed to the equator with their (+) ends begins. At late telophase, this system of developing interzonal fibers often fills up the space between daughter nuclei (Fig. 1.2, i, j). It consists of the central spindle fibers and newly-formed opposite polar MT bundles conjuncted with their (+) ends. During the interzonal cytoskeleton system formation, the nuclear envelope forms around telophase chromosome groups, and microtube bundles diverge from its surface. Such configuration of interzonal system connecting daughter nuclei and filling in practically the whole cytoplasm has the cytoskeleton in meiotic interkinesis of all the dicotyledonous species we investigated. To the onset of prophase II, the interzonal cytoskeleton system gets depolymerized (Fig. 1.2, k, l). MTs (+)- ends disjunct, MTs shorten becoming closer to daughter nuclei. Cytoskeleton rings develop at the end of prophase II around daughter nuclei.

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Spindles at metaphase II of the dicots PMCs are mutually perpendicular or located at the angle of 600C; so, their polar regions are maximally distant from each other (Fig. 1.2, m, n). After the formation of 4 daughter nuclei, the formation of the so-called secondary spindles begins in the common cytoplasm at telophase II, i.e. radial MT bundles connecting non-sister daughter nuclei (VanLammeren et al., 1985; Traas et al., 1989). Setting of cell plates proceeds simultaineously in the system of six phragmoplasts (Fig. 1.2, o). Daughter cell membranes form also simultaineously, realizing the so-called simultaineous cytokinesis and developing four microspores, disposed tetrahedrically (Fig. 1.2, p).

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Figure 1.2. Cytoskeleton dynamics in the meiotic division of tomato (Lycopersicon esculentum) PMCs: a) diakinesis; b) early prometaphase I; c, d) mid prometaphase I (chaotic stage); e) late prometaphase; f) metaphase I; g, h) anaphase I; i, j) interzonal cytoskeleton system at telophase I – interkinesis; k, l) interzonal cytoskeleton depolymerisation at prophase II; m) prometaphase II; n) metaphase II; o) late telophase II; the 4th nucleus is out of focus; o) tetrad.

1.3. Simultaneous and Successive Cytokinesis Compared During evolution, plants have elaborated the way of cytokinesis which is modified exocytocis. Membrane vesicules transport from Golgi apparatus to the equatorial cytoplasmic zone, form a monolayer there (cell plate), then fuse and develop daughter cell membranes (Staehelin and Hepler, 1996). Transport of membrane vesicules to the equator is realized by the temporal cytoskeleton structure – phragmoplast. In meiosis, cytokinesis can be successive, after each division (as in most monocots) or only at the telophase of the second meiosis, simultaneously automising four daughter nuclei

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by six phragmoplast activities (as in most dicots). Phragmoplasts that form in successive and simultaneous cytokinesis do not principally differ in their structure: it is a system of MT bundles diverging from the telophase chromosome groups and overlapping with their (+)ends on the equator. However, phragmoplasts in successive and simultaneous cytokinesis differ in their fibers architecture, also in the way of cell plate formation. The phragmoplast in the successive cytokinesis, in monocots, is a system of long fibers encircling the growing cell plate edge as a palisade and centrifugally moving. In simultaneous cytokinesis of dicots, phragmoplasts are a multitude of long fibers that cut through the whole cytoplasm in the interzone between nuclei. This phragmoplast has no hollow cylinder configuration and does not make centrifugal movement. It is a solid structure similar with a metaphase spindle, but more voluminous (Shamina and Dorogova, 2006). The phragmoplast function in dicots and monocots does not principally differ: transport of membrane vesicules into the region of MTs overlap for cell plate formation. However, cell plate formation is realized in different ways. In monocots, the phragmoplast expands centrifugally together with a growing cell plate, but it is immobile in dicots. The points into which plastosomes are to move during the monolayer formation (cell plate) are determined by overlapping points of phragmoplast MT (+) ends. Overlapping points of MTs in mobile phragmoplast fiber ‗draw‘ the cell plate plane during their centrifugal movement. In case of an immobile phragmoplast, the whole multitude of points into which plastosomes are to move, is set by a multitude of MT (+)-ends overlap that densely cut through the whole cytoplasm and are located on the equatorial plane. All these distinctions allow us to point out two different phragmoplast types in higher plant meiosis: mobile and immobile – in successive and simultaneous cytokinesis, respectively. To form up these two phragmoplast types, central division spindle fibers are utilized. It is not surprising, as central spindle fibers are in fact a phragmoplast – a system of MT bundles oriented to the equator with their (+)-ends and overlapping in this region. In the immobile phragmoplast formation, polar MT s play a considerable role, they complete the central spindle fibers in phragmoplasts between sister nuclei and completely make up phragmoplasts between nonsister nuclei. The role of polar MTs in the mobile phragmoplast formation and function was hypothesized (Shamina et al., 2007a). The most important distinction of simultaneous cytokinesis from that of successive consists in the absence of cytoplasm division after the first meiotic division. An immobile phragmoplast develops between daughter nuclei at telophase I in dicot PMCs with simultaneous cytokinesis, but it does not build a cell plate. Probably, this cytokinetic arrest is realized by means of a switch off of one of the basic cytokinetic processes – production of cell plate membrane vesicules (plastosomes). If it takes place, just as in transgenic tobacco line Res91, then cytokinesis is realized at telophase I of dicot meiosis by means of cell plate development and formation of daughter cell membranes (Shamina et al., 2000b).

2. PROPHASE ABNORMALITIES AS THE REASON FOR MEIOTIC RESTITUTION In the prophase of a dividing eukaryotic cell, the cytoskeleton begins the process of transfer from the interphase system to the division spindle. In PMCs it is to proceed in the way that microtubule bundles orientation would change for the opposite in a cell. MTs (+)-

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ends of radial interphase system are on the cell periphery, and (-)-ends are by the nucleus, as plant cell MTOCs are concentrated on the nuclear envelope surface (Vantard et al., 1990; Stoppin et al., 1994; Lambert, Lloyd, 1994; Azimzadeh et al., 2001). It is on the contrary in the division spindle: (+) MT ends are in the cell centre, on the spindle equator and kinetochores, and (-) ends are located on the spindle poles, on the cell periphery (Euteneuer et al., 1982). In animal cells such elements reorientation of microtubule cytoskeleton performs due to the doubling of polar organizers – centrioles – and interaction of MT bundles diverging from them. Plant cell is deprived of centrioles and any other morphologically identified spindle polar organizers. Therefore, mechanisms of cytoskeleton reorientation during interphase-metaphase transition fall away from observation. Studying the cytoskeleton reorientation process at prophase is especially complicated by its depolymerization or drastic shortening of fibers for this period (Сhan, Cande, 1998; Peirson et al., 1997; Zee, Ye, 2000). But cytoskeleton rearrangement can be traced at these stages in the PMCs where cytoskeleton depolymerization does not proceed. In plant cell prophase, the cytoskeleton undergoes considerable rearrangements, the result of which is the perinuclear cytoskeleton system formation (DeMey et al., 1982; Vos et al., 2008). It is necessary to note that, in plant cell mitosis, perinuclear cytoskeleton systems were described, though their function and formation mechanisms remain obscure (Ambrose and Cyr, 2007). They are a loose cage of straight chaotic MT bundles converging with their (-)ends in several points (Schmit et al., 1983; Wick, Duniec, 1984). During prophase, such a ‗‘loose cage‘‘ can have a higher degree of structural organization, making the so-called prophase spindle or polar caps as a result of MTs bipolar orientation and even their convergence on the poles. During the nuclear envelope breakdown, the prophase spindle disappears (Wang et al., 1991), and its relation with prometaphase cytoskeleton has not been shown. At meiotic prophase, a cytoskeleton transfer from the interphase radial system to the perinuclear ring (Shamina, 2005a) is realised. As a result of our study of wild-type phenotypes with and without cytoskeleton depolymerisation at transient stages and of abnormal meiotic divisions, the cytoskeleton cycle undergoes the following stages at prophase (Shamina, 2005a): 1) transition from the reticular cytoskeleton to straight radial MT bundles located in the meridional plane, whose proximal ends are in the narrow ring zone (plane reorientation); 2) MT bundles reorientation at 90 oC from the radial orientation to that of tangential to the nucleus; 3) prophase perinuclear cytoskeleton system formation – the meridional microtubular ring; the last process includes an MT bundles curvature and their coorientation in the ring body. According to our data. variations of MT depolymerization/ repolymerisation do not principally affect the cytoskeleton cycle at this stage. Due to the formation of perinuclear system at late prophase, the cytoskeleton fibers become closer to the nucleus (VanLammeren et al., 1985; Staiger, Cande, 1990; 1991; Suzuki, Tanaka, 1999; Ambrose et al., 2007) that, further on, provides – after the nuclear envelope breakdown - their contact with chromosome kinetochores. Aberrations of this important stage lead to drastic abnormalities in division spindle formation, corresponding disturbances of chromosome segregation and nuclear meiotic restitution.

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2.1. Cytoskeleton Conservation in the Interphase Radial Configuration

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The cytoskeleton block in the radial position was observed by us during meiosis of 10% of PMCs of WWG F1 hybrids № 9-00 (Triticum aestivum L. SP-718 x Agropyron glaucum L.). It is rather a rare abnormality of cytoskeleton cycle. If, in wild type meiosis and the trivial phenotype of WWG F1 hybrids, MT bundles become closer to the nuclear envelope at late prophase, all the processes of cytoskeleton reorganization are arrested in this abnormal phenotype. Cytoskeleton fibers, up to telophase, preserve their interphase orientation; perinuclear ring, division spindle, and the phragmoplast are not developing. Accordingly, processes of caryo- and cytokinesis do not proceed. At the same time nuclear envelope breaks down at the beginning of conventional prometaphase and its reformation at the end of conventional telophase proceeds normally. Between these events chromosomes remain immobile in the former nuclear area. The product of the first meiotic division is a monad with a restitution nucleus. In the second meiotic division, a common spindle is developing in such a cell. After the regular division in the second meiosis, such a cell forms a dyad with nonreduced 2n members. The reason for meiotic restitution in this phenotype is the impossibility of MT (+)-ends attachment to chromosome kinetochores due to their spatial distance: in the radial central configuration, MT (-)-ends are located in the central cytoplasm region, and (+) ends are on the periphery (Shamina et al., 2003а).

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Figure 2.1. Conservation of radial configuration of MT bundles during the first meiotic division in PMCs of WWG F1 №9-00: a) PMC before nuclear envelope breakdown; b, c) radial cytoskeleton at non-nuclear stages from prometaphase I to telophase I (nuclear envelope is absent, the former nuclear area is not different from the cytoplasm; d) monad with restitution nucleus at prophase II.

2.2. Fused Spindle. Approachment of Nuclei at Prophase II of Meiosis with Simultaineous Cytokinesis in Dicotyledonous Species As a mechanism of meiotic restitution, the phenomenon of fused spindles has been known for a long time in dicotyledons (Jorgensen, 1928). It consists in the formation of common division spindle at metaphase II after normal first meiotic division. We observed five reasons initiating this process: approachment of prophase nuclei, fusion of perinuclear rings (p. 2.3 of the present review), underformation of interzonal cytoskeleton system in interkinesis (p. 7.9), aberration of interzonal cytoskeleton system in interkinesis (p. 7. 10), approachment of chaotic prometaphase figures in common cytoplasm (p. 4.4). In this part, behavioral aberrations of daughter nuclei are described at the prophase of the second meiotic division.

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In wild type meiosis with simultaineous cytokinesis in dicotyledons, daughter nuclei of the first meiotic division keep at the distance from each other in the common cytoplasm due to the developed system of interzonal cytoskeleton (VanLammeren et al., 1985; Hogan, 1987; Traas et al., 1989). At prophase II this system depolymerises (Peirson et al., 1997), interzonal MT cytoskeleton is replaced for the perinuclear MT system shaped as rings (Shamina, 2005a), but nuclei do not approach each other. After the nuclear envelope breakdown, the second meiotic division spindles form out of microtubules of perinuclear systems. Mutually perpendicular orientation of these spindles is achieved at late prometaphase II, after the chaotic stage. Spindles are located in the common cytoplasm, six phragmoplasts develop at telophase II connecting all the nuclei, and simultaineous cytokinesis autonomising four haploid nuclei proceeds. During male meiosis in two potato (Solanum tuberosum) clone groups 1) CD1015, CD1050 and 2) RH95-237-06, RH95-237-14; RH95-237-03; RH96-2013-03 abnormal behaviour of daughter nuclei at prophase II is observed. In 50% PMCs of the first clones group and in 90-100% PMCs of the second clones group, at this stage, migration of daughter nuclei and their abnormal approachment occur. After the nuclear envelope breakdown, a common spindle possessing chromosomes of both daughter nuclei is developing in the cell centre. After chromosomes segregation by this spindle, two unreduced 2n nuclei form in the common cytoplasm, and then a dyad forms out of such cells. Studies of MT cytoskeleton behavior at meiotic stages preceding the appearance of fused spindles (at telophase I – interkinesis – prophase II) do not reveal any deviations in all enumerated clones, both under cytoskeleton visualization using the classical method (Navashin fixation) and immunostaining. Approachment of nuclei begins after the transition from the interzonal cytoskeleton in interkinesis to the perinuclear one at prophase II. After chromosome segregation within the common spindle, two daughter nuclei with diploid chromosome number develop in the common spindle at anaphase II. At telophase II, an immobile phragmoplast develops between them, and cytokinesis with the formation of a dyad of unreduced microspores proceeds (Shamina et al., 2004).

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Figure 2.2. Approachment of daughter nuclei in PMCs of potato (Solanum tuberosum) clone with fused spindle phenotype at prophase II: a) normal position of daughter nuclei in interkinesis; b) approachment of nuclei in the common cytoplasm at prophase II; c) common spindle at metaphase II.

2.3. Fused Spindle. Fusion of Cytoskeleton Perinuclear Rings at Prophase II Potato (Solanum tuberosum) clone СЕ10 is the producent of nonreduced gametes with fused spindle phenotype. In PMCs of this clone, the above-described (p. 2.2) approachment of

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daughter nuclei at prophase II is accompanied by the reorganization of perinuclear cytoskeleton system. This clone is characterized by complete asynapsis and curved C-shaped spindle formation at metaphase I. Due to the spindle curvature, its poles and, as a consequence, daughter nuclei become close at telophase I (see p. 3.2). However, their position corrects itself and becomes normal as a result of development of cytoskeletal interzonal system at late telphase I. Interzonal cytoskeleton system elongates and separates daughter nuclei,.so they become maximally distant from each other, just as in normal meiosis. At prophase II, the nuclei surrounded by perinuclear cytoskeleton rings move to the cell centre and approach each other. Then perinuclear rings disjunct, fuse and form one common ring encircling both nuclei. After nuclear envelope breakdown, a common division spindle forms out of this common ring. Chromosome segregation proceeds normally, the phragmoplast develops between two daughter cells, and cytokinesis occurs. The product of meiosis is a dyad with unreduced 2n members (Conicella et al.,2003). Approachment of nuclei and fusion of perinuclear rings at prophase II occurs in 50% of PMCs of clone CЕ10. This phenomenon indicates the thing that MTs of perinuclear rings have their own active dynamics.

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Figure 2.3. Fusion of perinuclear rings (marked in arrows) of daughter nuclei at prophase II in potato clone CE10 PMCs a) disjunction of rings of approached nuclei b) rings fusion, c) common perinuclear ring around daughter chromosome groups, d) common chaotic cytoskeleton figure at prometaphase II

2.4. Cortical Cytoskeleton Ring and Meiotic Restitution In the meiotic division of PMCs of maize (Zea mays) haploid №1498, multiple spindle formation in the mononuclear PMCs in the first meiosis, and the deviation in the structure of cytoskeleton configuration – the perinuclear ring at prophase II - was found in 50% of cells. After nuclear envelope breakdown, in the first meiotic prometaphase, a normally appearing chaotic cytoskeleton figure was developing, out of which several spindles were developing instead of one; chromosomes were randomly distributed among them (‗‘multiple spindles‘‘ phenotype). In interkinesis, multinuclear monads were forming out of PMCs with multiple spindles. Usually, in such cells, at the second meiotic division, several spindles develop (due to multiple nuclei), and the meiotic division product of monocotyledons PMCs with multiple spindles phenotype is a multinuclear monad at tetrad stage. But in haploid №1498, unlike them, at prophase II, a cortical cytoskeleton ring was forming encircling all micronuclei. At prometaphase II, in such cells, there developed a common chaotic cytoskeleton figure out of which one common biplolar spindle was forming. Chromosome segregation and cytokinesis in the second meiosis occurs normally, and a dyad was the meiotic division product. The cells – members of such a dyad – are predecessors of unreduced 2n-gametes (Shamina et al., 2007c).

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Figure 2.4. Cortical cytoskeleton ring at prophase II in multinuclear PMCs of maize haploid leads to chromosome integration into a common spindle at metaphase II a) multiple spindles at metaphase I; b, c) common cortical ring at prophase II (arrows), d) common division spindle at metaphase II.

2.5. Autonomous Cytoskeleton Ring

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In WWG hybrid №14-2 (T. aestivum, cv. Novosibirskaya 67 x A. glaucum 13-1), at prophase I, PMCs are characterized by a drastically excentric position of nuclei in 7-10% of cells. Radial cytoskeleton is also asymmetrical: MT bundles radiate from the nucleus to the periphery are of different length. At late prophase, in the free cytoplasm region, where radial MT bundles had the biggest length, a ring figure of curved MT bundles is gradually forming. Under normal meiosis, an MT ring closely encircles the nucleus, but in PMCs of this hybrid, the ring is fully autonomous to the nucleus lying in the free cytoplasmic region. It is orientated also in the meridional plane within flattened tablet-shaped cereal meiocyte, just as a normal perinuclear ring. Later, a bipolar spindle without chromosomes forms out of this ring close to the nucleus.

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Figure 2.5 Formation of autonomous perinuclear ring and autonomous division spindle as a consequence of excentric position of the nucleus in PMCs WWG F1 hybrids: a) excentric nucleus at prophase I, radial cytoskeleton is asymmetrical, b, c) autonomous cytoskeleton ring at late prophase I, d) autonomous spindle in the cell with a restitution nucleus at interkinesis.

Chromosomes cannot contact spindle fibers and form a restitution nucleus. At telophase, in the equatorial zone of the autonomous spindle, the phragmoplast/cell plate are developing which expand centrifugally. As the spindle and, respectively, phragmoplast are on the periphery, the cytoplasm incompletely divides, and daughter cell membranes form like incisions on the mother cell membrane. Sometimes there occurs cytokinesis with the separation of unnuclear cytoplast; in part of cells, cytokinesis may be arrested (Shamina et al., 2003a).

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2.6. Chromosome Arrest in the Zygothene „‟Bouquet” Configuration: Monopolar Chromosome Migration in a Bipolar Spindle

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At prophase, chromosome behavioral abnormalities can be the direct reason for the restitution nucleus formation at telophase on the background of normally proceeding cytoskeleton cycle. In male meiosis in WWG F1 hybrid №1-2-1, rice (Oryza sativum) № C45 and maize (Zea mays) №4479-4 haploids, we observed chromosome arrest in the ‗‘bouquet‘‘ configuration up to NEB. Occurrence frequency of PMCs with chromosome arrest in the bouquet varied from 30 to 80% in these genotypes. Chromosome compactisation was normal. Stages of cytoskeleton cycle: perinuclear ring at late prophase and bipolar division spindle formation – were also normal. At prometaphase, obviously due to the preserving univalents orientation, a monopolar figure consisting of reductionally oriented univalents and their kinetochore fibers, was developing. Then this monopole interacted with central spindle fibers and, as a result, a bipolar spindle was forming with all chromosomes (univalents) were attached to one pole. At anaphase, all univalents migrated to this pole forming a restitution nucleus. At telophase, on the spindle equator, a phragmoplast and a cell plate were forming, cytokinesis proceeded normally; the division result was a dyad. One cell had a 2n nucleus (or n in haploids), the other was unnuclear (Shamina et al., 2007b). Similar meiotic products – dyads with one anucleate member and the other with restitution nucleus - were described in meiosis in a wide lily hybrid – a producent of 2n gametes (Barba-Gonzalez et al., 2005).

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Figure 2.6. Consequence of chromosome arrest in the ‗‘bouquet‘‘: monopolar segregation in the bipolar spindle: a) chromosomes in the ‗‘bouquet‘‘ configuration at diakinesis; b) monopole at prometaphase I; c) monopolar segregation of univalents at anaphase I in bipolar spindle; d) telophase I with drastically unequal telophase chromosome groups.

3. EARLY PROMETAPHASE ABNORMALITIES LEADING TO NUCLEAR RESTITUTION As a result of perinuclear cytoskeleton ring formation at prophase, (+) and (-) MT ends become to be at the equal distance from the nuclear area. It can be determined as the first stage of cytoskeleton reorientation during the interphase – metaphase transition. Prometaphase is separated from prophase by such an important morphological event as nuclear envelope breakdown. As a result, cytoskeleton can contact with chromosomes, and prometaphase - the process of direct division spindle formation – begins. Early prometaphase is determined by us as a period from the perinuclear ring breakdown (during NEB) to chaotic cytoskeleton figure formation (Shamina, 2005a). The main outcome of early prometaphase is

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entering the former nuclear area by microtubular bundles to contact chromosomes. (+) Mt ends become turned to the cell center having finished their turn at 180° at this stage compared to their orientation at interphase. The stages of MTs rearrangement during early prometaphase, include 1) perinuclear ring disassembly into separate fibers (MT bundles), 2) straightening of these bundles, 3) their invasion the former nuclear area. The result of these processes is the formation of the chaotic prometaphase cytoskeleton figure and a transfer to mid prometaphase. As for the process of MT bundles entering the former nuclear area after NEB, A. Bajer supposed it in his research of mitotic ultrastructure in the endosperm of African lily Haemanthus (Bajer, 1968; 1987; Bajer, Mole-Bajer, 1972). One can say that the meiotic perinuclear ring is an analog of the mitotic prophase spindle. It is interesting to compare their behavior during prophaseprometaphase transition. At mitosis, the prophase spindle develops from a perinuclear cytoskeletal cage by means of bipolarization of MT convergence centers (DeMey et al., 1982; Smirnova, Bajer, 1994). After NEB, MT bundles are chaotically oriented, i.e. the prophase spindle loses its clear organization and structure (Lambert, Bajer, 1975; DeMey et al., 1982; Wang et al., 1991). Unfortunately, the relationship of prophase – metaphase spindles is not shown because of methodic problems (low cell number in the analysis does not allow us to trace intermediate stages of the process). The meiotic perinuclear ring does not have convergence points and the traits of bipolar orientation of its constitutive bundles. Besides, it is not a loose cage, but a well organized and oriented ring structure. Probably, it is explained by differences in MT perinuclear system at mitotic and meiotic prophase. Phenotypes with abnormal cycles at meiosis allow us to demonstrate the direct relationship between the perinuclear ring at prophase, chaotic prometaphase figure and the metaphase spindle in the sense that spindle formation proceeds by means of perinuclear ring MTs utilization.

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3.1. Cytoskeleton Conservation in the Perinuclear Ring Configuration In WWG F1 hybrids № 9-3 (Triticum. aestivum L., cv. Novosibirskaya 67 x Agropyron glaucum L.), № 2-2 и 2-5 T. durum сv. Altaiskaya Niva х E. elongatum the perinuclear ring disintegration is arrested at the earlierst stage of this process in 5 -7% of PMCs. As a result, after NEB, chromosomes remain scattered in the former nuclear area and do not contact with the microtubular skeleton. The perinuclear ring with a completely preserved structure encircles the former nuclear area at the stages from prometaphase I till telophase I or interkinesis. The division spindle does not develop. No cytokinetic traits are found in such cells at the light level. At late (conventional) telophase I, a nuclear envelope re-forms around the chromosome group. In the second meiotic division, the formed mononuclear monad divides with a dyad formation having potentially viable predecessors of unreduced 2n gametes (Shamina, 2005b). Perinuclear ring conservation is also observed in WWG F1 hybrids № 2-2 и 2-5 T. durum cv. Altaiskaya Niva х E. elongatum. Abnormality frequency – about 10%. Abnormal conservation of perinuclear cytoskeleton system during the first meiotic division was described in PMCs of mutant mel1 in rice (Oryza sativum) (Nonomura et al., 2007).

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Figure 3.1. Conservation of cytoskeleton ring configuration at prometaphase-telophase I in PMCs of WWG F1 hybrids: a) late prophase I: perinuclear ring is formed, the nuclear area is distinct, b, c) conventional metaphase-telophase I: nuclear envelope broke down, the perinuclear ring is preserved in the cytoplasm, the former nuclear area is indistinguished from the cytoplasm; d) monad with a restitution nucleus.

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3.2 Aberration in Straightening of Microtubules of Perinuclear Ring: C-Spindle According to our observations, this abnormality is the most widely spread and a mass spindle aberration in plants and is encountered in the meiosis of 80% WWG F1 hybrids and in all WR F1 hybrids we studied with the frequency 30-100%. It is found only in the phenotypes with univalents reductional orientation (each univalent is attached only to one spindle pole, i.e. carries one kinetochore fiber). Such spindles also develop in 100% of PMCs of synaptic tomato mutant as6, Brassica juncea haploids and potato clones СЕ10, ВЕ62, ВЕ1050, characterized by complete asynapsis. C-shaped spindles are also typical of the phenotype of meiotic mutation ms28 in maize with normal synapsis of homological chromosomes. The spindle curvature in M1 was described for haploids (Sadasivaiah, Kasha, 1971), synaptic higher plant mutants (Iwanaga, 1984; Chan, Cande, 1998), wide hybrids (Darlington, 1965). In this phenotype, after nuclear envelope breakdown, the perinuclear ring disintegrates into individual MT bundles and disappears as a consolidated structure. The further spindle formation process proceeds the following way. At early prometaphase, univalents with attached MT bundles are chaotically scattered in the former nuclear area. One can often observe the thing that MT bundles are curved at the chaotic prometaphase stage. Later on, they develop a spindle of abnormal shape and location. The shape abnormality is caused by the fiber curvature, so the metaphase spindle is crest-shaped. The abnormal spindle location is expressed in the thing that its equatorial zone has a shift from the center to the cell periphery. Reductionally oriented univalents roughly congress on the spindle equator and imitate the metaphase plate. We defined this stage as metaphase I. Our observations show that the location of reductionally oriented univalents on the spindle equator is an obligatory stage of asynaptic meiosis. We constantly observed the formation of such a ‗‘pseudometaphase plate‘‘ in all the phenotypes with asynaptic meiosis we analysed – both in straight and curved spindles. At anaphase I univalents have their uncoordinated migration to the poles and turn scattered along the whole spindle body. This stage is very prolonged compared to the ―metaphase‖, it is more often encountered in preparations and, probably, due to this reason, it

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is mistakenly determined as abnormal metaphase. Finally, univalents reach the poles, and aneuploid chromosome groups develop on the poles of a curved spindle. On the curved spindle equator, a phragmoplast/cell plate develops and cytokinesis occurs. The cytokinetic process is normal, but, when the cell plate reaches the closest mother cell membrane, the plastosomes fusion and development of daughter cell membranes, which incompletely divide the cytoplasm as an incision, proceeds. A premature stop of cytokinetic processes takes place. An incised binucleate monad is the division product in this case. Due to the spindle bending, its poles are close, and telophase chromosome groups can separate from the poles and migrate to the cell center. Based on the close daughter nuclei in the common cytoplasm in the second meiosis, a common division spindle develops most often. A dyad with unreduced members is the product of meiosis (Shamina et al., 1999). Such a phenotype is observed in a number of WWG F1, WR F1 hybrids, maize haploids, in meiotic maize mutant ms28 .

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Figure 3.2. С-spindle and meiotic restitution of PMCs of maize haploid № 2906. a) curved spindle at conventional metaphase – anaphase I, b) assymetrical phragmoplast position (arrows) due to the spindle curvature; c) binucleate monad with an incision (arrow) at interkinesis.

However, telophase figure correction occurs at this stage in many of genotypes with Cspindles we analysed. MT bundles polymerise from the curved spindle poles towards each other, connect on the equator and form a broad phragmoplast which crosses most part of the cytoplasm. In such phenotypes the cytokinesis proceeds normally. In additionl, there may be a correction of phragmoplast/cell plate centrifugal movement, which proceeds on the polarized cytokinesis type (Cutler and Ehrhardt, 2002; Cook, 2004), i.e. centrifugal movement continues also after the system reaches the closest mother cell membrane. The spindle bending is also typical for the meiosis in haploids (Sadasivaiah, Kasha, 1971) and synaptic higher plant mutants (Iwanaga, 1984; Chan, Cande, 1998). It proves our supposition on this abnormality reasons at asynaptic meiosis. Besided, the curved spindle abnormality is almost exclusively typical of the first meiotic division. The spindle bending in the second metaphase was traced by us only in two phenotypes out of many dozens of forms with C-spindles we analysed. C-shaped spindles were described earlier in wide hybrid meiosis (Darlington, 1965), but their formation mechanism was not investigated.

3.3 Arrest of Cytoskeleton Invading the Former Nuclear Area In WWG F1 hybrid № 5-5 (T. aestivum сv. ANK9 x A. glaucum 52-3), the process of cytoskeleton cycle, at early prometaphase, is arrested at later stages. The perinuclear ring loses its integrity and becomes loose, microtubules straighten in its composition, but do not

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move. They do not leave the perinuclear zone, do not enter the former nuclear area and do not contact with chromosomes and each other. From prometaphase to conventional telophase, MTs encircle the former nuclear area – but not as a consolidated ring, as it is in the abovedescribed phenotype (see. p. 3.1), but they encircle it as a set of tangentially oriented straight microtubular bundles. As an outcome, the division spindle is not forming and chromosomes remain scattered in the center of the cell without attachment to the cytoskeleton. This cytoskeleton configuration preserves till interkinesis. As an outcome of this, chromosome segregation and cytokinesis are arrested in the first meiotic division. At late telophase, the nuclear envelope reforms around the whole chromosome set, and a restitution nucleus is developing. The PMC share is – 5-7% with this abnormality. The analogous phenotype is also demonstrated by WWG F1 hybrid № 59-8 (T. durum cv. Altaika х E. elongatum) at frequency 10%. (Shamina et al., 2003c). Our results show that the cytoskeleton dynamics at meiotic prophase is similar with that of mitotic plant cell division (see review Baskin, Cande, 1990). Actually, the meiotic perinuclear ring could be interpreted as an analogue of the mitotic prophase spindle.

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Figure 3.3. Cytoskeleton configuration at conventional metaphase I - telophase I under the arrest of fibers penetration into the former nuclear area of PMCs in WWG F1 hybrids a - c) cytoskeleton conservation in the configuration of a disorganized perinuclear ring during metaphase I – telophase I, d) monad with restitution nucleus at interkinesis.

4. MID PROMETAPHASE ABNORMALITIES AND MEIOTIC RESTITUTION The period of cytoskeleton chaotic configuration that forms after the penetration of MT bundles into the former nuclear area is indicated as mid prometaphase. This period makes us refer it to a separate substage due to its temporal prolongation compared to other prometaphase substages. First, it is possible to assume that, at this time, MTs attach to chromosome kinetochores, and our observations prove it. Kinetochore spindle fibers formation can normally be observed at the chaotic stage in objects with quite big PMCs, also in forms characterized by the prolonged prometaphase stage. It is a most important stage of division spindle formation, during which the bipolar spindle fiber development occurs (Shamina, 2005a). MT bundles attach to chromosome kinetochores forming bipolar kinetochore spindle fibers. If chromosomes are bivalent or they are a pair of chromatids with oppositely oriented kinetochores, then the system of ‗‘kinetochore MT bundle – chromosome – kinetochore MT bundle‖ – is a bipolar fiber, which is the bipolar spindle element. If chromosomes are univalent, and each of them has one (unsplitted) kinetochore, each element

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carries one kinetochore MT bundle. and, further on, it attaches only to one of the spindle poles. Free MT bundles, at the chaotic prometaphase stage attach each other with their (+) ends and form another important spindle element – bipolar central fibers. Later they serve as the base of a developing phragmoplast. Analysis of abnormal phenotype of WWG F1 hybrid № 27-1 (T. aestivum cv. Saratovskaya 29 x A. glaucum) and WWG F1 № 5-98 (T. durum cv. Altaika x E. elongatum) with a very prolonged chaotic stage and a full arrest of kinetochore spindle fiber formation allowes to reveal the process. The main process of mid-prometaphase or the chaotic stage is the formation of the basic elements in bipolar spindle development, its bipolar fibers: central and oppositely oriented kinetochore fibers (Shamina, 2005 a). Aberrations of these processes lead to those of division spindle structure and phragmoplast, abnormalities in caryo- and cytokinesis and to the meiotic restitution.

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4.1. Monopolar Spindle One of the key stages of plant spindle bipolarity setting is the formation of bipolar fibers out of antiparallel MT bundles. Under the aberration of this stage, and the rest prometaphase processes preservation (MT co-orientation in the spindle body, their convergence on the poles), monopolar spindles develop. As two types of bipolar fibers – central and the opposite kinetochore ones – participate in the plant spindle formation, the absence of these both elements leads to the monopolar spindle development. Bipolarity of central spindle fibers is provided as a result of MT bundles (+) ends junction. Double kinetochore fiber bipolarity is provided by the opposite orientation of chromosome kinetochores. Therefore, for monopolar plant spindle formation, a combination of two abnormalities is necessary: absence of mutual attachment of free MTs (+) ends, and absence, due to some reason, of oppositely oriented kinetochore fibers. A combination of two such abnormalities is rather a rare event. In the meiosis of WWG F1 hybrid № 30-2, chromosomes are presented by univalents with an unsplitted kinetochore (reductional univalents). Thus double kinetochore fiber formation, in this case, is impossible; only one MT bundle is attached to each univalent. During central spindle fibers formation, interaction of free MT (+)-ends is aberrated, probably by the effect of the corresponding mutation which is manifested in conditions the allohaploid F1 hybrid genotype. Instead of bipolar fibers, free MT bundles and univalents with a single kinetochore MT bundle, participate in the spindle formation. Such monopolar spindle elements interact; their (-)-ends converge with the formation of a monopolar figure. Such a monopolar cone is compact, there is no anaphase chromosome movement in it, as a rule, phragmoplast is absent. At telophase, a monad with a restitution nucleus is developing (Shamina et al., 2003b). Such a phenotype was also observed by us in WR F1 hybrid and haploid of Brassica juncea. In a number of wide cereal hybrid phenotypes, the monopolar spindle has a looser structure and the shape of a non-compact cone, but that of a monoaster. At telophase, a monad developes out of such cells with many micronuclei. However, sometimes, in such spindles, anaphase univalent migration is realized to the single pole, and a restitution nucleus is forming (Shamina, 2005b). The phragmoplast does not form in such monopoles, and cytokinesis does not proceed.

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Figure 4.1. Monopolar spindles in PMCs of WWG F1 hybrids at metaphase I: a) compact monopolar spindle at conventional metaphase I – telophase I; b) one of not numerous chromosome anaphase movements in a compact monopolar spindle; c) loose radial monopolar spindle at metaphase I; d) telophase I in PMCs with a loose monopole.

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Monopolar spindles are a well known and most frequently encountered abnormality of centriolar spindle in animal cells. This abnormality leads to the arrest of chromosome segregation and was described in the phenotype of homozygotes on mutations polo (Sunkel and Glover, 1988), mgr (Gonzalez et al., 1988), mast in Drosophila melanogaster (Lemos et al., 2000), in cell culture ot the Chinese hamster (Wang et al., 1983). Monopolar spindles also form as a result of the influence on dividing animal cells by specific inhibitors (Mazia et al., 1981). Besides, monopolar spindles function in some types of modified cell division, e.g. in the meiosis of Sciara (Abbot et al., 1981; Fuge, 1994). The reason for monopolar spindle formation instead of a normal bipolar one in animal cells is segregation arrest of polar spindle organizers (centrosomal structures) (Sawin et al., 1992; Heck et al., 1993; Blangy et al., 1995). In acentriolar plant cell, the monopolar spindle has not been described in natural conditions for a long time; it was reported on its formation under the effect of a number of chemical agents (Tiwari et al., 1984; Binarova et al., 1998; Smirnova et al., 2002).

4.2. Autonomous Spindle Among abnormal meiotic divisions, there are those in which spindle kinetochore fibers formation is completely arrested. Chromosomes remain a disoriented group in the former nuclear area, and MT bundles, with lesser of bigger success, undergo the division spindle formation process. Under the aberration of MT bundles attachment to kinetochores, the socalled autonomous spindle - bipolar cytoskeleton system deprived of chromosomes and consisting of only central spindle fibers – is developing. Immobile chromosomes lying separately form restitution nucleus. A phragmoplast and a cell plate can develop on the autonomous spindle equator, which are indicative of such spindle fibers bipolarity. A chaotic configuration is also encountered. Under simultaineous aberration of MT (+)-ends to kinetochores and their conjuction with each other, an autonomous monopolar spindle is forming. During male meiosis in WWG F1 № 27-1 (T. aestivum cv.Saratovskaya 29 х A. glaucum) and TA F1 № 5-98 (T. durum сv. Altaika x E. elongatum), the cytoskeleton cycle is of normal procedure till the chaotic prometaphase stage. Free MTs conjunct with each other by their (+)-ends forming central spindle fibers, whereas kinetochore ones are absent. Chromosomes

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are chaotically scattered in the former nuclear area in the cell center, close to the formed bipolar spindle, or they form a compact group close to it, but do not connect to the spindle fibers. The division in the second meiosis of obtained monad with a restitution nucleus results in the formation of a dyad with 2n members at tetrad stage.

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Figure 2. Autonomous spindles at metaphase I im PMCs of WWG F1 hybrid: a, b) bipolar autonomous spindle (arrows) at metaphase I; c) chaotic autonomous ―spindle‖ at metaphase I (arrows); d) common spindle in the monad with restitution nucleus at metaphase II.

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4.3. Chromosomes Monopolar Migration in a Bipolar Spindle “Comet” Phenotype This abnormality was observed by us in the meiosis of wide cereal F1 hybrids: WR F1 (T. aestivum cv. Saratovskaya 29 х S. cereale cv. Onokhoiskaya, WWG F1 № 27-1, (T. aestivum cv. Saratovskaya 29 х A. glaucum), № 21-1 (T. durum cv. Altaiskaya Niva х E. elongatum), also in wheat-rye alloplasmic line CYANK9 and in monosomic line 2А of wheat T. aestivum cv. Milturum 533. In this phenotype, the spindle kinetochore fibers formation is aberrated in 15% of PMCs. The metaphase spindle is developing bipolar, but all chromosomes move to one of the poles, whereas the opposite pole remains empty. An autonomous bipolar spindle is forming in such cells, and it is composed of exclusively central spindle fibers. Chromosomes contact it with their arms and slide along its surface to one of the poles as a unified group (Seryukova et al., 2003). The telophase chromosome group on one pole and spindle fibers diverging from it looks very originally, reminding of a comet. A cell plate develops at telophase on the spindle equator (in the middle of a ―comet tail‖), and the spindle becomes barrel-shaped which is typical of the phragmoplast. Centrifugal movement of phragmoplast/cell plate and daughter cell membrane formation occur normally, and an anucleate cytoplast splits from the cell. The other cell contains a restitution nucleus. Rather often cytokinesis may be arrested. Chromatin movement along the autonomous spindle have been described in mouse oocytes (Deng et al., 2009). Migration of all chromosomes to one pole was described as a reason for restitution nucleus formation (Sears, 1953; Avers, 1954; Stefani, 1986, Jauhar, 2003). This phenomenon was also reported in animal meiosis of meiotic mutants fusolo и solofuso (Drosophyla) (Bucciarelli et al., 2003). However, the mechanisms of this phenomenon remained unknown, as there was no division spindle visualization as a cytoskeleton structure. We managed to reveal two reasons for chromosome monopolar segregation in a bipolar spindle (see also p. 2.6).

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Figure 4.3. Result of chromosome monopolar migration along the autonomous spindle (arrows) in PMCs of WWG F1 hybrid a, b) late anaphase I; c) telophase I; d) autonomous phragmoplast (arrow).

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4.4. Fused Spindle. Approachment and Fusion of Cytoskeleton Chaotic Figures at Mid-Prometaphase II in PMCs with Simultaneous Cytokinesis in Dicots During normal meiosis chaotic prometaphase II figures representing randomly oriented MT bundles in a complex with chromosomes are located at the maximal distance from each other in the common cytoplasm. Then spindle fibers orient along the axes of the future division, co-orient, converge on the poles, and two mutually perpendicular division spindles develop in the common cytoplasm (Traas et al., 1989). In potato clones – producents of 2n gametes ВЕ1050; ВЕ62 in 50% PMCs, and in tomato (Lycopersicon esculentum) meiotic mutant as6 in 100% of PMCs, prometaphase chaotic figure migration proceeds to the cell center and their fusion into a common chaotic figure takes place. All the enumerated genotypes are characterized by a complete asynapsis, curved spindles at metaphase I, approachment of daughter nuclei on the poles of a curved spindle at telophase I and their further correction by the developing cytoskeleton interzonal system. These nuclei enter prometaphase II and are located at a normal distance from each other. But, after NEB and the onset of prometaphase, chaotic figures approach each other and fuse in the cell center. Then their fibers co-orient, converge on the poles and form a common spindle. We have never observed such a fusion of cytoskeleton figures at late prometaphase II (set of roughly bipolarly oriented spindle fibers non-converged on the poles without a developed metaphase plate) or developed spindles at M II (Shamina et al., 2004). Results of our observations using the method of immunostaining showed that the MT cytoskeleton per se does not play any role in chaotic figures approachment.

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Figure 4.4. Approachment of chaotic cytoskeleton figures at prometaphase II in PMCs of tomato (Lycopersicon esculentum) meiotic mutant as6. a, b) successive migration of chaotic prometaphase figures to the cell center; c) formation of common chaotic prometaphase figure; d) common spindle at metaphase II. Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

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5. ABNORMALITIES OF LATE PROMETAPHASE AS A MEIOTIC RESTITUTION MECHANISM

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The span between the chaotic stage and metaphase is indicated as late prometaphase. The basic cytoskeleton dynamic process at this period is cytoskeleton realization from the chaotic configuration and bipolar spindle formation, i.e.spindle fibers orientation along the future division axis. Research of late prometaphase with the help of abnormalities typical of this stage allows to reveal some of its constitutive morphological processes and to determine their interrelation. At late wild type meiotic prometaphase, the formed bipolar spindle elements (chromosomes with oppositely directed spindle kinetochore fibers and bipolar central spindle fibers) orient along the future division axis and form a system of bipolar fibers – the division spindle. At the same stage, the final spindle formation processes – fibers (-)-ends convergence and spindle poles development, also development of metaphase plate (chromosome congression) - take place. During normal meiosis, these processes occur rapidly, so that many their details are indiscernible. Multiple observations of this stage in normal and abnormal meiosis allow us to assert that bipolar spindle fibers orientation proceeds as a result of their turn and migration in the cytoplasm; each fiber is independent from the rest (Shamina, 2005a). These peculiarities can be traced directly under the formation of (relatively) normal bipolar spindles in the meiosis of wide F1 hybrids. Due to a number of reasons (probably univalent chromosomes) their prometaphase stage is considerably prolonged compared to the norm, and its distinct events and intermediate stages become accessible for direct observation. The main result of late prometaphase is the completion of bipolar spindle formation – the pledge and condition of mother cell genome division into two parts. Aberrations of this process can lead to the formation of restitution nuclei.

5.1. Chaotic Spindle Prometaphase I of meiosis in WWG F1 hybrid № 27-1 (T. aestivum cv. Saratovskaya 29 x A. glaucum) is drastically abnormal. First, it is expressed in the unusually longstage of ‗‘chaotic bundles net‘‘ at which spindle fibers are disorderly scattered over the cytoplasm. At this stage, the course of prometaphase is arrested in part of the cells, and the division spindle is not developing. This figure (spindle fibers chaotic net and a group of disoriented chromosomes in the center) is preserved in the metaphase, anaphase and telophase of the first meiotic division. At the end of telophase I, chromosomes are encircled by the nuclear envelope, and a restitution nucleus or a set of micronuclei is forming. Chromosomes carry MT bundles on kinetochores, i.e. spindle kinetochore fibers are developing. But there is no chromosome anaphase movement and chromosomes remain in the cell centre. It is possible to distinguish long central spindle fibers oriented chaotically. This abnormality is the result of abrerration of bipolar orientation process of the developed spindle fibers at late prometaphase. Cytokinesis does not proceed due to the absence of a developed bipolar phragmoplast (it can also be called chaotic here). In the second meiotic division, a common spindle and a dyad with 2n members (Seriukova et al., 2003) develop. The abnormalities frequency – up to 10%.

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Figure 5.1. Chaotic spindle and its consequences in PMCs of WWG F1 hybrid. a, b) chaotic spindle at metaphase I; c) monad with a restitution nucleus in interkinesis; d) common spindle at metaphase II.

The similar picture is observed in the abnormal meiosis in dicots, at the prometaphase of both first and second meiotic division. In PMCs of pea (Pisum sativum) meiotic mutant ms3, at metaphase I, abnormal cytoskeleton figures in a form of a network of chaotically oriented MT bundles, built instead of a bipolar spindle. Chromosomes with attached kinetochore fibers are disorderly scattered among free MT bundles; individual spindle fibers do not interact with each other, do not obtain bipolar orientation, do not converge with their (-)-ends. Chromosome movement is arrested at anaphase, a monad with micronuclei develops in the interkinesis. As a result of these aberrations, at telophase II, the cell divides into many fragments forming a polyad at the stage of tetrads (Shamina et al., 2000a). A net of chaotically scattered fibers in the cytoplasm at metaphase II, after the normal first meiotic division is a typical phenotypic feature of sugar beet mutant line SOAN-112. The first male meiotic division proceeds without deviations. Drastic abnormalities of cytoskeleton dynamics appear at prometaphase II. After NEB, the MT cytoskeleton concentrated in the perinuclear region, begins to rearrange for the spindle formation of the second meiotic division. However, instead of two bipolar mutually perpendicular spindles, in 30% of the mutant PMCs, a chaotic net of disoriented criss-crossed MT bundles with disorderly scattered chromosomes with kinetochore fibers, is developing. A bipolar system of mutually parallel MT bundles, typical of spindles, does not develop. MTs do not converge on pole ends and it can be well seen when operating a microscope vintage. At anaphase, chromosomes segregate into chromatides, but anaphase movement is arrested or, either, chromosomes move at a small distant. Abnormal daughter cell membranes sometimes divide the cell incompletely and look like incisions on the mother cell membrane. This is the result of multiple short cell plates formation on the chaotic fibers. Herewith, an incised monad with micronuclei is forming

5.2. Spindle Disorientation at Metaphase II in the Meiosis with Simultaineous Cytokinesis in Dicots Orientation of developed bipolar spindle fibers, just as that of spindles proper, along the cell division axis is realized at late prometaphase. In the abnormal phenotype ‗‘disoriented spindle‘‘, bipolar spindles develop, but orientation of division axes changes in the cell. Herein, in the second meiotic division of dicots PMCs, poles of spindles located in the common cytoplasm, can converge, and it leads to the formation of a nucleus with a doubled chromosome set (Mok, Peloquin, 1975; Dorogova and Shamina, 2000; Taschetto and Pagliarini, 2003; Camadro et al., 2008).

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The AtPS1 gene and a corresponding set of mutants that produce pollen grains which are up to 65% diploid and give rise to numerous triploid plants in the next generation was described in Arabidopsis (d‘Erfurth et al., 2008).

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Figure 5.2. Spindle poles convergence under the spindles disorientation in the second meiotic division in tobacco (Nicotiana tabacum) PMCs. a) normal spindle position at MII in PMCs of wildtype tobacco; left spindle is seen from equator, right – from the pole, b - d) spindle disorientation with poles convergence in the second meiotic division of tobacco PMCs, line Res91.

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6. ANAPHASE ABNORMALITIES THAT LEAD TO MEIOTIC RESTITUTION It is surprising, but chromosome segregation abnormalities per se (chromosome anaphase movement arrest in the normal spindle) are not so often encountered among forms with abnormal meiosis as one would expect There are literary reports on chromosome anaphase movement aberrations (Rhoades, Dempsey, 1966; Wagenaar, 1968; Fukuda, Sakamoto,1992). But, as these observations were made without spindle visualization, it is impossible to exclude the thing that these abnormalities are caused by spindle structure aberrations, but not the mechanism of anaphase movement proper. Chromosome movement can be decelerated in a normal spindle, there may be chromosome laggards. As our wide observations of hundreds of different plant forms with abnormal meiosis show, if the anaphase began and the division spindle is normally developed, then chromosomes reach the poles, sooner or later. We manage to trace univalents that do not reach the poles and remain in the body of a normal spindle till the end of telophase only in several genotypes of haploids and allohaploids. Cytokinesis is not aberrated, as a rule, and a dyad with mironuclei is the product of such a division in most cases. If cytokinesis does not proceed, monads with micronuclei or a restitution nucleus develop. In the case of micronuclei formation, univalents move at a bigger distance, and the spindle may be longer. Restitution nucleus forms when chromosomes migrate at a comparatively short distance at anaphase and remain close to the equator (p. 6.1).

6.1. Aberration of Anaphase Chromosome Movement This phenotype is observed in PMCs of WWG F1 hybrid № 519 in 20% of cells. At metaphase I, univalents, just as in the trivial phenotype, group on the spindle equator imitating the metaphase plate. At anaphase I, univalents begin to move, but do not diverge far from the equator. The phragmoplast can develop independently from the chromosomes

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position, or its development or function may be arrested. Cytokinesis can be realized (incomplete, so daughter cell membranes are incision-shaped) or not. In any case, nondiverged chromosomes form a restitution nucleus out of which a common division spindle is developing at metaphase II.

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Figure 6.1. Arrest of chromosomes at early anaphase in PMCs of WWG F1 hybrid a) PMCs at early telophase I; b, c) attemption of phragmoplast (arrows) centrifugal movement of PMCs with chromosome arrest in the spindle body; d) monad with a restitution nucleus.

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6.2. Spindle Shortening During Anaphase The process of chromosome segregation is provided by two processes: kinetochore fiber shortening (anaphase A) and central spindle fiber lengthening (anaphase B) (Bruss-Mascher et al., 2004; Bouck and Bloom, 2005). By means of anaphase A, chromosomes become spatially distant from each other; by means of anaphase B – spindle poles. In dividing higher plant cells, the process of anaphase B is realized alongside with anaphase A (Yu and Russell, 1993). In PMCs of maize dihaploid №4611 we observed a drastic shortening of normal division spindle at anaphase I in about 35% of cells. Chromosome synapsis is of normal procedure in the dihaploid, the division spindle has normal size and shape at metaphase I. Chromosomes start anaphase movement coordinatedly, as anaphase groups. After chromosomes pass the distance, which is about one half of half-spindles length, the spindle begins to shorten, and chromosomes move backward. Finally, they are on the equator, there where they began their movement from. At telophase, chromosomes are encircled by the common nuclear envelope forming a restitution nucleus. Two closely approached daughter nuclei may develop. Cytokinesis is arrested: phragmoplast fibers are also abnormally shortened, centrifugal movement of phragmoplast/cell plate does not proceed. A common division spindle is forming in the monad at metaphase II. As a result of the second meiotic division, dyads with 2n members develop at the stage of tetrads. One can hypothesise that shortening of a normal metaphase spindle, under the onset of anaphase, is caused by the aberration of activity balance of motor cytoskeleton MAPs, kinesins (Lee and Liu, 2007). Some of them provide transport towards (+)-MT ends separating the spindle poles, others – towards (-)-ends closing the poles (Hamada, 2007; Bannigan et al., 2007). The first ones activity prevails at normal anaphase. Abnormally shortened spindles are described in the phenotype of Arabidopsis mor-1 and clasp-1 mutants (Kawamura et al., 2006; Ambrose et al., 2007). The coordination of spindle length is important for regular cell division, and it is a conserved feature of eukaryotic cell (McNally et al., 2006; Yang et al., 2003; 2005).

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Figure 6.2. Spindle shortening during anaphase I in PMCs of maize dihaploid a) normal chromosome segregation at the onset of anaphase I; b) spindle shortening, disappearance of its polar regions; c) dumb-bell restitution nucleus; a reduced phragmoplast is seen on the nuclear sides, d) common spindle in the monad in metaphase II.

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7. CYTOSKELETON ABNORMALITIES AT TELOPHASE AND MEIOTIC RESTITUTION Cytokinesis is the fundamental cell division process whose mechanisms remain mostly unclear in plant cell (Jurgens, 2005; Segui-Simarro et al., 2007; VanDamme et al., 2008). Cytoskeleton abnormalities and daughter genomes autonomisation failure also lead to the meiotic restitution. Daughter nuclei, being in the common cytoplasm, can approach each other, and it leads to the congression of their chromosomes in the common spindle in the further cell division – both meiotic and post-meiotic. Due to the analysis of meiotic division abnormalities, we managed to describe the process of mobile phragmoplast functioning and formation in PMCs of monocots and also to divide the telophase into substages (Shamina et al., 2007). Bipolar phragmoplast fibers (central spindle fibers) develop at mid prometaphase, and the phragmoplast proper (central spindle, bipolar system of bipolar fibers, early phragmoplast) is developing at late prometaphase. After chromosome segregation to the opposite poles at anaphase, the spindle cytoskeleton composition changes. Kinetochore fibers depolymerise and shorten and, at early telophase, the spindle consists of only central fibers. At late anaphase – early telophase, the central spindle looks like a solid column between telophase chromosome groups. We determined this structure as an early phragmoplast. At early telophase, on the spindle equator, a cell plate begins to develop as a monolayer of membrane vesicules (plastosomes). These vesicules are Golgi-derived, and are transported here by the central spindle fibers. After the cell plate crosses the whole spindle, its central fibers redistribute so that they surround the cell plate growing edge and obtain a form of a hollow cylinder – mid phragmoplast. The fibers encircle the growing cell plate as a palisade. Probably, at this stage, central spindle fibers lose their lateral connections with each other, which they had in the anaphase spindle. The phragmoplast in PMCs is not a set of short fibers around the cell plate, but a system of long fibers linking polar regions with the equator and surrounding the growing cell plate edge. At mid telophase, phragmoplast fibers become longer, curve and, due to this, their central points make centrifugal movement (Shamina et al., 2007a). We called this expanding barrel-shaped structure with progressively curving fibers late phragmoplast. As the phragmoplast moves to the periphery, its fibers become more and more curved. The cell plate expands by means of new membrane vesicules attachment to its growing edge, and phragmoplast MT bundles continue to encircle its growing edge providing

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vesicules transport to this region. During centrifugal movement the number of phragmoplast fibers increases, probably by means of polar synthesis of new MTs (Shamina et al., 2007a). When the cell plate reaches the mother cell membrane, plastosomes fuse in its composition with the formation of daughter cell membranes. After this, the formation of daughter nuclei envelopes proceeds. The developed membranes cut the phragmoplast fibers on the equator in the region of MT (+)-ends overlapping. Separated phragmoplast fibers, after this, become part of radial interphase cytoskeleton of daughter cells. At late telophase, when the phragmoplast/cell plate practically completely cross the mother cell cytoplasm, MT bundles begin to polymerise from telophase chromosome groups to the direction opposite to the equator. They do not participate in cytokinesis, but they are part of the radial cytoskeleton typical of interkinesis. The cytoskeleton cycle returns to the initial point here. It is shown, that the process of phragmoplast formation and operation during mitosis in meristematic cells can also be divided into phases: phragmoplast initials, solid phragmoplast, transitional phragmoplast, and ring-shaped phragmoplast (Seguí-Simarro et al., 2004). Phragmoplast abnormalities were described in a number of phenotypes: embryonic mutants pilz and hinkel (Arabidopsis thaliana) (Mayer et al., 1999; Strompen et al., 2002), in the meiosis of mutant aph (Zea mays) (Staiger, Cande, 1993), in PMCs in wild populations of the grass Paspalum (Pagliarini et al., 1999) and wide cereal hybrids F1 (Shamina et al., 2007), also under physical and chemical influences on mitotic cells (Mole-Bajer, 1969; Smirnova and Bajer, 1998). A number of embryonic mutations that disturb Arabidopsis cytokinesis (Sollner et al., 2002) is known, including those arresting membrane vesicules fusion and daughter cell membrane formation. Mutants with cytokinetic aberrations in male meiosis were described in Arabidopsis (Chen and McCormick, 1996; Hulskamp et al., 1997; Spielman et al., 1997; Hauser et al., 2000; Magnard et al., 2001).

7.1. Arrest of Basic Telophase Processes In male meiosis in WWG F1 hybrid № 9-02, about 20% dyads form at the stage of tetrads. It is explained by a complete arrest of the basic processes of telophase I: phragmoplast centrifugal movement, cell plate formation and nuclear envelope restoration around daughter nuclei. Only the amplification of phragmoplast fibers occurs, and it becomes wider, but it does not make any centrifugal movement and does not approach the mother cell membrane. At conventional interkinesis, the radial cytoskeleton is developing in the cell existing alongside with the telophase spindle (early phragmoplast). At prophase II, a cytoskeleton ring that encircles both chromosome telophase groups develops out of radial cytoskeleton elements. A common division spindle is developing at prometaphase II out of this common ring, and a dyad forms at the stage of tetrads. Cells – the dyad members – have unreduced 2n chromosome number. Aberration of regulation of the corresponding cell cycle stages is, probably, the reason for this phenotype‘s abnormalities. Experiments with the expression of a nondegradable cyclin in plant mitotic cells resulted in a phenocopy of the described abnormality (Weingartner et al., 2004). Arrest of daughter nuclei envelopes reformation and arrest of cytokinesis at telophase were described in experiments on kinase CDK1 inhibition (Wheatley et al., 1997), in dividing rat kidney cells. In S. cerevisiae (budding yeast), high levels of nondegradable cyclin CLB2

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arrest cells late in mitosis, with segregated chromosomes and the presence of an elongated mitotic spindle (Surana et al., 1993). Indestructible cyclin Cdc13 arrests Schizosaccharomyces pombe cells in anaphase with separated and condensed chromosomes and no septa (Yamano et al., 1996).

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Figure 7.1. Meiotic process under the complete arrest of telophase I processes in PMCs of WWG F1 hybrid: a) telophase I, b) interkinesis, c) prophase II with a common perinuclear ring (arrows), d) common spindle at metaphase II.

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7.2. Complete Arrest of Cytokinetic Processes at Early Phragmoplast Stage This phenotype observed in a rice haploid № 207 and WWG F1 hybrid № 738 differes from the pervious one in the thing that not all telophase processes are arrested in it, but only cytokinetic processes. At telophase I – interkinesis, the cytoskeleton preserves its early phragmoplast configuration, i.e. it is the central spindle with telophase chromosome groups (then with the interphase nucleus) on the poles. Mid-phragmoplast formation as a hollow cylinder, also barrel-shaped late phragmoplast with curved fibers expanding centrifugally, does not occur. Phragmoplast fibers do not amplify. The cell plate does not develop. Unlike the previous abnormal phenotype, nuclear envelope formation proceeds here around daughter nuclei, so they co-exist with the cytoskeleton which remains in the anaphase spindle configuration. As a result, a binucleate monade is forming. During interkinesis, daughter nuclei often migrated from telophase spindle poles and approach in the common cytoplasm. At the second meiotic division, in monads with approached daughter nuclei, a common division spindle is forming, and then – a dyad of unreduced microspores at tetrad stage. Abnormality frequency – to 40%. Arrest on the solid phragmoplast stage is described in postmeiotic mitosis in double kinesin mutant in Arabidopsis (Lee et al., 2007).

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Figure 7.2. Arrest of all cytokinetic processes at telophase I in PMCs of rice haploid. a) metaphase I; b) late anaphase I; c) monad with an anaphase spindle and interphase nuclei at interkinesis; d) common spindle at metaphase II. Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

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7.3. Arrest at Early Phragmoplast Stage with Element of Centrifugal Movement: Gamma-Phenotype In a number of phenotypes of WWG hybrids F1 (Triticum aestivum cv Novosibirskaya 67 x Agropyron glaucum) and WR (Triticum aestivum cv Yubileinaya x Secale cereale v Hariuchiban) F1 hybrids, after chromosomes reach the poles, a weakly curved or a straight spindle curves more and more during telophase, so that the polar regions approach. Sometimes, the spindle curves so strongly that telophase chromosome groups interchange their positions, and the whole figure looks like the Greek letter gamma. That is why we called this abnormality gamma-phenotype. The cell plate is not forming; but in many PMCs, it is possible to observe the appearance of big vacuole-like structures (not shown) as we interpreted as plastosomes conglomerates. Phragmoplast fibers lengthening and curvature are realized, but they do not amplificate. Approached chromosome telophase groups then either encircled by the common nuclear envelope and form the restitution nucleus, or they are encircled by the nuclear envelope individually. Mono- and binucleate monads enter their second meiotic division. The restitution nucleus or approached daughter nuclei form the common division spindle at prometaphase II (Shamina et al., 2009). In this case, the meiotic product is dyads with nonreduced members at the stage of tetrads. The number of cells with this abnormality in the anther may be high (up to 80%). Unlike the previous abnormality (p. 7.2), where not only mid-phragmoplast formation is arrested, but also further cytokinetic processes, in this phenotype, the early phragmoplast structure is affected by the processes of phragmoplast centrifugal movement (fiber curvature and their elongation) at mid-late telophase. Normal mid-phragmoplast (hollow cylinder) fibers perform their autonomous centrifugal movement. Normally, the central point of each fiber (MT (+)-ends overlap) moves centrifugally, indicating its own line along the radius from the center to the periphery. Movements of all fibers accompanied by plastosome transport to their central points lead to the centrifugal cell plate growth. Probably, in the gammaphenotype, anaphase spindle central fibers do not lose their lateral connections, cannot autonomies, and the early phragmoplast behaves then like a single fiber. It is necessary to point out the principle distinction of curved spindles in the gammaphenotype and C-shaped spindles that appear because of aberrations in straightening of perinuclear ring MTs at early prometaphase (p. 3.2). The reasons for the formation of meiotic restitution nuclei and its mechanisms are completely different in these two cases.

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Figure 7.3. Progressive spindle curvature during telophase I in PMCs of WWG F1 hybrid a) anaphase I; b - d) telophase chromosome groups approacment caused during telophase I.

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7.4. Consolidation of Laggard Chromosomes into the Restitution Nucleus

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The following process of restitution nucleus formation was described in a number of contributions on cytological research of meiosis in allohaploids: univalents do not reach the poles at anaphase being scattered in the spindle body, and then consolidate into a restitution nucleus at telophase. As a rule, cytokinesis does not occur. This consolidation mechanism is unknown. A mononuclear monad, but not a monad with micronuclei, is developing; it would not occur without this consolidation. Restitution nuclei have a very original torus shape with a cytoplasm piece in the center (Chistyakova, 1974; Xu and Joppa, 2000, Fig. 1a). We believe that this variant of gamma-phenotype is the mechanism for restitution nucleus formation: an arrest at the stage of hollow cylinder formation and a progressive curvature of earlier phragmoplast on the background of univalents anaphase movement arrest. Chromosomes do not reach the poles, remain scattered in the spindle body and, as it curves, consolidate into a spiral or ring figure. The cell plate is absent, cytokinesis is completely arrested. At the end of the conventional telophase, chromosomes, which are located as a ring (or even spiral), are encircled by the nuclear envelope. A restitution nucleus is forming often ring-shaped: as a torus with a cytoplasmic ‗‘cork‘‘ in the center. Nevertheless, at metaphase II, in such cells, a common division spindle is forming, cytokinesis occurs normally, and a dyad of nonreduced microspores is the product of meiosis.

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Figure 7.4. Consolidation of laggard chromosomes scattering within the spindle body into the restitution nucleus at telophase I in PMCs WWG F1 hybrid by means of progressive curvature of telophase spindle; a) late anaphase I, b) spindle curvature with scattered chromosomes in it at midtelophase I; c) ring spindle at late telophase I, d) spiral spindle at late T1.

7.5. Arrest of Phragmoplast Development at the Stage of Hollow Cylinder In this abnormal phenotype, the stage of transition from mid-phragmoplast (hollow cylinder) to configuration of late phragmoplast and centrifugal movement is arrested: no progressive fiber elongation, curvature and amplification. The cell plate is not forming. The cytoskeleton is at ‗‘hollow cylinder‘‘ stage till late telophase, when a nuclear envelope develops around telophase chromosome groups. Cytokinesis does not occur. Daughter nuclei approach each other in the common cytoplasm, often penetrating the cylinder. As a result, binucleate monads with closely approached daughter nuclei are developing. In the second meiotic division, a common spindle at metaphase II and a dyad with 2n cells at the stage of tetrads are forming. Abnormality frequency is up to 35%.

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Figure 7.5. Arrest of phragmoplast development at the stage of hollow cylinder in PMCs of WWG F1 hybrid № 30-546. a, b) phragmoplast conservation at hollow cylinder stage in interkinesis; daughter nuclei migrate inside the phragmoplast and approach each other c) binucleate monads in interkinesis; d) common division spindle at metaphase II.

7.6. Excessive Curvature of Phragmoplast Fibers during Centrifugal Movement

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One of the models of phragmoplast centrifugal movement at meiosis presupposes the thing that this process is a modification of B-anaphase (Shamina et al., 2007a). Phragmoplast fibers not only elongate, as in B-anaphase, but also curve; due to this their central points move centrifugally, and the poles remain fixed. Aberrations in phragmoplast fibers curvature contribute to the restitution process.

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Figure 7.6. Excessive curvature of phragmoplast fibers as a reason for telophase chromosome groups approachment and their congression into the restitution nucleu in PMCs of WWG F1 hybrid: a) midtelophase I: cell plate formation is aberrated; b, c) abnormally drastic phragmoplast fibers curvature at late telophase I; telophase chromosome groups approach the equator following fibers polar ends; d) telophase chromosome groups congression into a restitution nucleus.

In PMCs of WWG F1 hybrid № 328, at mid – late telophase I, the phragmoplast morphology considerably changes. Telophase chromosome groups move to the cell equator and closely approach each other. This shift takes place because of an excessive phragmoplast fibers bending whose polar ends abnormally approach each other on the equator. Because of the thing that the cell plate does not develop in 30% of PMCs, approached chromosome telophase groups congress forming a restitution nucleus. The mononuclear monad, in the second meiotic division, results in a dyad with non-reduced 2n members. The percentage of cells with abnormal phragmoplast fibers curvature is up to 50%.

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7.7. Aberration of Phragmoplast Centrifugal Movement

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In some phenotypes, aberration of cytokinesis occurs because of abnormalities in centrifugal phragmoplast movement and consequent secondary aberrations of cell plate formation. It is expressed in the thing that phragmoplast expansion decelerates, or stops halfway and does not reach the mother cell membrane. The cell plate, in such a phragmoplast, is always abnormal: it is not a monolayer of vesicules, but cloud-shaped or is strongly meandered. Sometimes, in such case, the phragmoplast/cell plate has assymetrical expansion, i.e. one edge irregurally moves and the other does not. As a result, the system reaches the mother cell membrane and contact it with its only one part; the cell plate incompletely divides the cytoplasm, and daughter cell membranes are incision-shaped on the mother cell membrane or invaginations inside the mother cytoplasm. If the phragmoplast/cell plate does not reach the mother cell membrane, then membrane vesicules do not fuse, and there is no daughter cell membranes formation. In such cases, the cell plate then disappears dispersing into its constitutive membrane vesicules, and the daughter nuclei approach each other. In such phenotypes, at late telophase, the cytoplasm turns to be completely filled with plastosome conglomerates whose formation also continues at interkinesis. This cytokinetic abnormality was observed by us in the phenotype of meiotic mutant ms43 (Shamina and Dorogova, 1995) in maize, also in WWG F1 hybrid № 13-2 and № 13-4 (T. aestivum ANK9 x A. glaucum 52-3). The abnormality frequency – from 15 to 60%. It is shown that failure of kinesins function leads to aberration of phragmoplast centrifugal movement, and cell plates formed by these phragmoplasts did not reach the mother cell membrane (Hiwatshi et al., 2008).

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Figure 7.7. Phenotypic manifestation of aberration in phragmoplast centrifugal movement of PMCs of WWG F1 hybrid; a - c) abnormal cell plate (marked in arrow) does not reach the mother cell membrane, its shape is drastically aberrated; d) daughter chromosome groups from previously approached nuclei encircled by the common perinuclear cytoskeleton ring at prophase-early prometaphase II; such common ring is a predecessor for common spindle in M II.

7.8. Arrest of Radial Cytoskeleton System Formation at TII (in „‟Parallel Spindles‟‟ Phenotype) in the Simultaineous Cytokinesis in the Dicot PMCs This mechanism of nuclear meiotic restitution is one of the most important in potato breeding (Alfano et al., 1999; Carputo et al., 2003). Simultaineous cytokinesis at telophase II of wild-type dicot male meiosis is realized due to the formation of six immobile phragmoplasts that conjunct four nuclei lying on the tetrahedron tops. Such location is

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achieved by means of the corresponding spindles orientation at metaphase II. In this case, the tetrad microspores also have their tetrahedral location. Four phragmoplasts form at metaphase II under parallel spindle co-orientation, and a coplanar microspores tetrad is the division product. Binucleate dyads are the meiotic product at telophase II under parallel spindles location and simultaineous arrest of rafial MT (additional spindles, phragmoplsts) formation. This leads to the meiotic restitution, i.e. reunion of chromosomes of these nuclei in the first postmeiotic mitosis. This mechanism was described in the phenotype of potato mutant clone with parallel spindles and aberration of radial MT bundles (Alfano et al, 1997; Genualdo et al., 1998).

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Figure 7.8. Consequences of arrest of radial MT systems at TII in sugar beet (Beta vulgaris) PMCs. a) metaphase I; b) prophase II; c) metaphase II with parallel spindles; b) dyad with binucleate members at tetrad stage.

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7.9. Fused Spindles. Underformation of Cytoskeleton Interzonal System at Telophase I Approachment of daughter nuclei is observed at telophase I in sugar beet (Beta vulgaris) mutant lines В-24 and SOAN 5-10, which demonstrate high percentage of nonreduced gametes. Analysing MT cytoskeleton dynamics in the meiosis of this mutant line, we found that cytoskeleton aberrations at telophase I – interkinesis lead to the backward movement of daughter nuclei to the cell center and the restitution process. Normally, at telophase I, the system of central spindle fibers separating daughter chromosome groups and, later, interphase nuclei, symmetrically widen due to new polar MTs polymerization, and interzonal fibers amplificate because of this process. Thus, the so-called immobile phragmoplast develops between daughter nuclei. It disappears only at prophase II Unlike this, the system of interzonal MTs is aberrated in mutant meiocytes at telophase I. Its fibers do not amplify, most part of MT bundles disappears and, as a result, the system looks abnormal: daughter nuclei not supported by MTs move to the center and abnormally approach each other. Approachment of daughter nuclei is preserved at interkinesis and prophase II, which has the major influence on the process of the second meiotic division. At prometaphase II and metaphase II, a common spindle is developing in such cells. At telophase II, on the common spindle equator, a phragmoplast/cell plate is forming, and cytokinesis takes place. Instead of a tetrad a dyad forms; it initiates the formation of nonreduced gametes (Dorogova et al.,1999). Abnormality frequency – about 60%.

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Figure 7.9. Approachment of daughter nuclei at interkinesis at underformation of intezonal cytoskeleton system in PMCs of sugar beet (Beta vulgaris) line B-24: a) underformation of interzonal cytoskeleton system, b, c) approachment of nuclei at interkinesis; d) common spindle at MII.

Approachment of daughter nuclei, as a result of abnormal MT dynamics in the mutant, reveals the framework function of interzonal fibers at TI of dicots. This abnormality differs in its mechanism from that described in p. 3.2., which also causes fused spindles formation. In phenotype 3.2, approachment of daughter nuclei occurs later, at prophase II, because of an unknown mechanism on the background of normal MT cytoskeleton behavior. The interzonal system of polar MT bundles forming de novo can perform an unexpected function in the correction of daughter nuclear position. If a division spindle forms as a curvature (C-shaped) – it practically always happens under asynaptic meiosis, - telophase chromosome groups and daughter nuclei become abnormally approached. Approachment of daughter nuclei would inevitably lead to their restitution with the absence of cytokinesis after the first meiotic division in dicots. Such a mechanism of restitution nuclei formation was described by us in the meiosis of monocots: development of C-spindle at metaphase I and arrest of cytokinesis at telophase I lead to the approachment of daughter nuclei and their restitution (p. 3.2). However, approachment of nuclei on the poles of a curved spindle in the common cytoplasm at telophase I has never led to restitution in any of the numerous cases of asynaptic meiosis with C-spindle of different dicot species we analysed. The thing is that the system of straight interzonal microtubules forming at telophase from the spindle poles inevitably separted abnormally closed telophase nuclei. As a result, at interkinesis, nuclei were always diametrically opposite to each other in the cortical cytoplasm region, just as in wild type meiosis, i.e. the position of nuclei was completely corrected. We observed this picture under the spindle curvature in the asynaptic meiosis of potato clones (Solanum tuberosum L.) CE10, BE1050, ВЕ62, tomato mutant as6 (Lycopersicon esculentum L.), haploids of Brassica juncea Rajat.

7.10. Fused Spindles. Aberration of Fibers of Interzonal Cytoskeleton System Brassica juncea is a natural tetraploid. It is believed that it originated from the cross of diploid species Brassica rapa and Brassica nigra (Prakash, 1980). The tetraploid progeny from such a cross could appear only under the formation of nonreduced gametes by both parents. In this case, it is possible to expect this trait manifestation in the meiosis of haploids B. juncea. True, the haploids meiotic products at the stage of tetrads have an admixture of dyads up to 20%. Analysis of meiotic process in PMCs with cytoskeleton visualization revealed two mechanisms of meiotic restitution that lead to dyads formation.

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1st mechanism: aberration of interzonal cytoskeleton system formation at telophase Iinterkinesis. This abnormality represents a failure of MTs (+)-ends overlap of interzonal system on the equator. Under normal polymerization of polar MTs, but with aberration of their (+)-ends overlapping on the equator, the structure of interzonal cytoskeleton system becomes aberrated. Instead of an immobile phragmoplast, it is radial MT bundles diverging from daughter nuclei arise. Distal ends of these MTs criss-cross with each other in the cell equatorial region, and this is a good diagnostic trait of this abnormality. As a result, the interzonal MT system is not consolidated, and leads to a shift of daughter nuclei, their approachment and to the common spindle formation in the second meiotic division. 2nd mechanism – monopolar spindle formation at metaphase I in about 7% of cells. This abnormality was described above (p. 4.1). It leads to a complete arrest of chromosome segregation at anaphase I and the restitution nucleus formation at telophase I. At metaphase II, a common division spindle is forming and a dyad instead a tetrad at telophase II. Monopolar spindle formation was described by us also in the meiosis of monocot plant forms: WWG and WR F1 hybrids, where they are also the reason for meiotic restitution. The both mehanisms of nuclear restitution in the meiosis of Brassica juncea are based on one abnormality of cytoskeleton dynamics: aberration of connection of (+)-ends of antiparallel polar MTs that proceeds either at prometaphase I which, in its turn, leads to the monopolar spindle formation or at telophase I, and it aberrates the structure of interzonal cytoskeleton system.

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Figure 7.10. Meiosis in PMCs of Brassica juncea haploid №6. Aberration of MTs (+)-ends overlap of interzonal cytoskeleton system and the shift of daughter nuclei caused by it (disconnected fibers are indicated by arrows); a - c) disconnection of interzonal MT bundles at telophase I (arrows); c) displacement of daughter nuclei at interkinesis; d) dyad at tetrad stage.

8. CELL PLATE ABNORMALITIES LEADING TO MEIOTIC RESTITUTION The cytoskeleton play a key role in the cell plate formation: membrane vesicules transport occurs along phragmoplast fibers from Golgi apparatus into the cell equatorial zone (Lee et al., 2001). Nevertheless, many aspects of cell plate and daughter cell membranes formation are realized independently from the cytoskeleton function (Otegui and Staehelin, 2000). Aberration of these processes leads to that in daughter genomes autonomisation and, as a consequence, it can be the reason for meiotic restitution. Formation of daughter cell membranes is a very important terminal cytokinetic stage. It occurs when the cell plate

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reaches the mother cell membrane and contact it (Ehrhardt and Cutler, 2002). After this, plastosomes fuse and daughter cell membranes develop (Esseling-Ozdoba et al., 2008).

8.1. Absence of Cell Plate

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This aberration is quite widely spread in abnormal cereal meiosis. We observed it at the frequency from 3 to 30% in several dozens of haploid and allohaploid genotypes of maize, rice, wheat, rye, wheat-grass and their wide hybrids. Under the presence of a developed phragmoplast, no membrane vesicules observed, - either as a cell plate (monolayer) or other associations. The phragmoplast fibers move centrifugally and can reach the mother cell membrane, but the formation of daughter cell membranes is impossible. Daughter nuclei, in rather a high cell percentage, approach in the common cytoplasm, and their chromosomes can congress into the common division spindle at metaphase II (Shamina et al., 1999). A dyad of unreduced microspores is the meiotic product in PMCs with such a phenotype. Investigations carried out at the light level, cannot reveal the thing if the cell plate absence is the consequence of arrest in the synthesis of membrane vesicules by Golgi apparatus or an aberration of phragmoplast transport. Indirect data (absence of plastosome conglomerates in the cytoplasm) are in favour of the first supposition. Abnormal cytokinesis without cell plate in mobile phragmoplast was also described in meiosis of Magnolia (Brown and Lemmon, 1992; Dinis and Mesquita, 1993) and Conocephalum (Bryophyta) (Brown and Lemmon, 1998).

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Figure 8.1. Approachment of telophase chromosome groups under the absence of cell plate on the background of the realization of phragmoplast (marked in arrows) centrifugal movement in PMCs of WWG F1 hybrids: a, b) telophase process with the absence of cell plate, c) binucleate monade at prophase II, nucleiare encircled by the common perinuclear cytoskeleton ring (arrows), d) common spindle at metaphase II.

8.2. Rotation of Phragmoplast on the Background of Cell Plate Absence A variant of the previous phenotype is continuation of ‗‘empty‘‘ phragmoplast centrifugal movement also after it reaches the mother cell membrane. At early - mid telophase, the cell plate is absent, and then an ‗‘empty‘‘ phragmoplast – as a hollow cylinder, and, later, as the hollow barrel (as its fibers become curved), begins the centrifugal movement to the mother cell membrane. Having reached it, the phragmoplast does not stop moving and continues to expand. Daughter nuclei are formed in interphase, but centrifugal movement of the phragmoplast still continue. As a result, it turns within the PMC - which is tablet-shaped

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in cereals – in complex with telophase chromosome groups. The telophase spindle axis becomes to be located along the short axis of a tablet-shaped PMC as a consequence of this turn. Daughter nuclei or chromosome groups approach each other in the common cytoplasm, which is the reason for the restitution process. The percentage of cells with a common spindle is much higher at the phragmoplast turn than without it at metaphase II. Such excessive phragmoplast movement without a cell plate is observed in most of WWG F1 and WR F1 hybrids with the ―cell plate absence‘‘ phenotype. In the hybrid wheatgrass x wheat F1 № 1-99 (Е. еlongatum x T. aestivum cv. Lutescence 132), this abnormality was mass (in 30% PMCs).

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Figure 8.2. Approachment and abnormal co-orientation of daughter nuclei within the cell under the cell plate absence in PMCs of WWG F1 hybrid: a) late telophase without cell plate; b, c) two optical sections of the same PMC with displaced daughter nuclei; c) abnormal position of nuclei at telophaseinterkinesis.

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8.3. Aberration of Cell Plate Shape and Structure In male meiosis of several wide cereal hybrids, the cell plate abnormalities are observed, whereas the phragmoplast centrifugal movement proceeds regularly. The abnormal cell plate is either fragmental or it looks like an amorphous congression of vesicules or cisternaea on the cell equator, or it has an abnormal wide diffusional shape. Concomitantly, the daughter cell membranes do not form or form abnormal. In such cases, the daughter cell membranes do not divide the cytoplasm completely and are an incision or a cave on the mother cell membrane. As a result of cell division with such a phenotype, binucleate monads are developing. In the second meiotic division, chromosomes of both nuclei sometimes congress in the common spindle, and a dyad at tetrad stage is forming. The percentage of such abnormalities is, as a rule, not so high. Aberrant cell plate formation is described in the phenotype of Arabidopsis mutant mor1 (Eleftheriou et al., 2005).

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Figure 8.3. Abnormalities of the cell plate formation during the first meiotic division in cereal PMCs: a) phragmental cell plate as a chain of lacoons; b) amorphous congression of vesicules on the cell equator; c) wide diffuse cell plate not forming daughter cell membranes; d) binucleate monad.

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8.4. Arrest of Plastosomes (Cell Plate Membrane Vesicules) Fusion In the phenotype of meiotic mutant pam1 (Zea mays), at the stage of dyads (interkinesis), the appearance of some amount – about 10% - of binucleate monads with drastically changed morphology is observed. Daughter nuclei are located one above the other in them. Nevertheless, all telophase PMCs of the mutant have no deviations from the norm; in particular, the phragmoplast and cell plate are normally developed. Telophase chromosome groups in such PMCs are of normal position, i.e. they are always in the plane parallel to the slide. It is necessary to note that in normal maize dyads and tetrads the nuclei are also always in one plane parallel to the slide. Normally, in cereals, the division axis of PMCs on squashed preparations is also oriented this way, as cells have ratios 3:3:1 counting from the division axis. Thus, in the mutant, after telophase I, during interkinesis, the cells – precursors of deformed monads – considerable change their shape, turn flattened from the poles and, as a consequence of this, they have an abnomal position on the preparation. Such cells ratios become 1:3:3: counting from the division axis. The analysis of cytokinesis in them showed that it proceeds normally, but the final stage – fusion of membrane vesicules and daughter cell membranes formation - do not occur in such cells. Having reached the mother cell membrane, the phragmoplast/cell plate do not stop and continue their centrifugal movement. Herein, the cells expand laterally and, hence, they flatten from the poles. As a result of the phragmoplast/cell plate excessive expansion, the cell deforms and changes its positon on the squashed preparation: division axis is perpendicular to the slide. Telophase chromosome groups turn into interphase daughter nuclei, so that cytokinesis continues in the conventional interkinesis. Simultaineous presence of formed interphase nuclei and cell plate (but not daughter cell membranes) in cereal PMC is not encountered in wild type meiosis and is a typical phenotypic trait of mutant pam1. During ‗‘excessive‘‘ cytokinetic process, the formed cell plate gradually disperses. Plastosome dispersion begins from the center: edges of semidispersed cell plate sometimes turn into incomplete daughter cell membranes as ring incisions around the mother cell membrane. At metaphase II, in common cytoplasm, a common division spindle often develops, as daughter nuclei close approach each other. 2n dyads at the stage of tetrads are the meiotic product (Dorogova and Shamina, 2001). Such a phenotype is also observed in number of maize haploids at a high PMC percentage (25%). In those cases the phragmoplast turn during excessive centrifugal movement is also observed (see also p.8.2).

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Figure 8.4. Aberration of daughter cel membranes formation in PMCs of meiotic mutant pam1 (Zea mays). a) interphase nuclei and cell plate at early interkinesis; daughter cell membranes do not form, b) turn of telophase figure inside PMCs, as a result of continuation of phragmoplast/cell plate centrifugal movement after they reach the mother cell membrane; c) approachment of daughter nuclei in the common cytoplasm; the cell plate is dispersed; d) common division spindle at MII.

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Gu and Verma (1997) showed that in BY-2 cells, under arrest of membrane vesicules fusion cased by the phragmoplastin overexpression, the cell plate gradually becomes abnormally S-shaped and even shifts into the diagonal or longtitudinal (along the long cell axis) position. We believe that it is consequence of the cell plate excessive growth under nonstop cytokinesis. As BY-2 cells are enveloped by a rigid cell wall, the cell plate excessive growth leads not to the cell expansion in the equatorial region, but to the deformation and shift of the cell plate per se. This shift is quite possible: rotation of the phragmoplast/cell plate was described in some plant cell types under their differentiation (Palevitz, Hepler, 1974). Peculiarities of knolle mutant phenotype with failure of plastosomes fusion (Lukowitz, Mayer, 1996; Lauber et al., 1997; Batoko and Moore, 2001) also indicate the processes of ―non-stop cytokinesis‖. First, it is the disorientation of daughter cell wall stubs that indicates their shift mentioned by the authors. Second, it is excessive thickness of these stubs which is indicative of excessive amount of membrane vesicules that moved into this region. Phenotypes of mutants tangled and discordia characterized with division plane disorientation (Walker et al., 2007; Wright et al., 2009) could also be interpreted as examples of ―non-stop‖ cytokinesis. Vivid traits of excessive cytokinesis are observed in dividing plant cells under caffeine action. This agent specificaly inhibits the fusion of plastosomes and daughter cell membranes formation. In studying the ultrastructure of dividing cell of stamen hairs in spiderwort under the action of caffeine, Helper and Bonsignore (1990) pointed out considerable plastosome conglomerates on the cell plate edges adjoining the mother cell membrane. In our opinion, this is the result of excessive, non-stop cytokinesis when the phragmoplast/cell plate are deprived of their ability to continue centrifugal movement within the walled cell. In the present review, the pragmoplast non-stop movement is described in p. 8.2. for the phenotype with cell plate absence. In this case – because of the impossibility of daughter cell membranes formation – the ‗‘empty‘‘ phragmoplast continues its centrifugal movement after it has reached the mother cell membrane and, as a result of this, it turns inside it.

8.5. Longtitudinal Orientation of Cell Plate and Daughter Cell Membranes According to Division Axis In PMCs of WWG F1 hybrid № 27 and № 734, at frequency 7-10%, a considerable aberration in cell plate formation, which proceeds because of aberrations in cell plate membrane vesicules (plastosomes) transport to the equator and, to be more precise, their distribution over the phragmoplast. Membrane vesicules are not transported along all the phragmoplast fibers towards the cell equator, but are distributed along some closely located fibers and fill in the space among them – from pole to pole as a stretched conglomerate. If this plastosome congression adjoins the mother cell membrane, daughter cell membranes develop as a narrow cistern, the incision. Abnormal cytokinesis, thus, occurs not perpendicularly to the division spindle axis, but along to it. An unnuclear cytoplast separates from the cell, or an incision is developing on the mother cell membrane, and daughter nuclei turn to be in the common cytoplasm. Their chromosomes can congress in the common spindle in the following division which is, as a rule, of normal procedure.

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Figure 8.5. Abnormal cell plate formation under aberration of phragmoplast fibers transport function in PMCs of WWG F1 hybrid: a - c) longtitudinal location of a mass of cell plate membrane vesicules (arrows) among the phragmoplast fibers in local part of the phragmoplast, d) abnormal daughter cell membranes located along the cell division axis.

This phenotype adds to the group of plant cell division abnormalities which are characterized by an incomplete arrest of some process that refers to the whole multitude of cytoskeleton elements (fibers, MT bundles) in a cell. In these abnormalities, part of fibers population continues to follow the cytoskeleton cycle, and part of it stops at the previous stage. Such are i) the ―combined spindle‖ phenotypes that consist of a mixture of straight and curved fibers (Shamina, 2005 b), ii) part of centrifugally moving phragmoplast fibers (p. 9.5), iii) triple phragmoplast in a cell with tripolar spindle in which, this or that cytokinetic process sometimes ‗‘switches off‘‘ in one or two of its three consituents Thus, the cell plate formation or phragmoplast expansion may occur in only one or two of the three phragmoplasts (Shamina, paper in preparation).

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9. INCOMPLETE CYTOKINESIS AS INCISIONS ON THE MOTHER CELL MEMBRANE AND ITS CONSEQUENCES In all cases, when the phragmoplast/cell plate contact the mother cell membrane – due to this or that reason – not simultaneously over its whole circumpherence, daughter cell membranes separate the cytoplasm incompletely, and look like more or less deep incisions on the mother cell membrane. It happens as a consequence of the thing that an assymetrically located cell plate, at the moment of contacting the mother cell membrane, do not completely cross the cytoplasm. A binucleate incised monad formed as a result of such a division is often the reason for 2n cells formation in the following division. Approachment of daughter nuclei in the common cytoplasm can often be the reason for the development of 2n gametes out of incised binucleate monads. In many cases, two division spindles form in the common cytoplasm at metaphase II of such a monad; the meiotic product is a triad with two haploid or aneuploid members and one binucleate cell. This last one can congress chromosomes of two nuclei in the common spindle at postmeiotic mitosis and result in a cell – the precursory of a 2n gamete. Sometimes, at metaphase II, spindles are not in parallel, but located at an angle to each other. As a result, the polar regions of two spindles approach, often in one point, and a restitution nucleus forms here.

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Figure 9. Restituion process in binucleate incised monads formed as a result of incomplete cytokinesis in cereal PMCs: a) binucleate monad with abnormal daughter cell membranes as an incision on the mother cell membrane, b) parallel spindles at metaphase II, c) displaced spindles at metaphase II with a converged pair of poles (arrow), d) triad with a 2n member.

According to our observations of simultaneous cytokinesis in PMCs of monocots, the reasons for asymmetrical cell plate position and incisions development can be the following:

9.1. Assymmetrical Position of the Division Spindle and Phragmoplast

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For regular cytokinesis, a spindle should be positioned right in the cell center. If it becomes approached to some mother cell membrane area, the first cell plate contact with mother cell membrane leads to fusion of vesicules and the stop of cytokinesis. We observed this phenomenon in the meiosis of WWG F1 № 86-2 (T. aestivum ANK26А х A. glaucum), № 97-13 (T. aestivum AHK9 x A. glaucum) in multinucleate meiocyte division, where some spindles were located on the cytoplasm periphery (also the second meiotic division in PMCs on mutant ms43 in maize), in the abnormal phenotype of a multiple spindle in WR F1.

9.2. Spindle Curvature When the spindle is curved, its equatorial region, as a consequence, is displaced to the mother cell membrane. Incomplete cytokinesis is observed in the phenotype of maize meiotic mutant ms28, also in curved spindles under asynaptic meiosis (wide cereal hybrids F1, haploids, synaptic mutants) (see p. 3.2).

9.3. Cytoskeleton Disorganization Under spindle disorganization, when it represents a chaotic network of fibers, multiple cell plates may develop on random MT bundles. Only those cell plates that met the mother cell membrane turn into abnormal daughter cell membranes in a form of short incisions. Cell plates, located deep in the cytoplasm, are unable to reach the mother cell membrane, disperse, because the fusion of vesicules and formation of daughter cell membranes do not proceed (p. 5.1).

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9.4. Abnormal Phragmoplast/Cell Plate Expansion Under the pragmoplast aberrated centrifugal movement, the cell plate expands asymmetrically and often reaches the mother cell membrane not simultaneously over the whole circumpherence. After the contact, there is a fusion of plastosomes, and daughter cell membranes look like incisions. This abnormality is typical of the phenotype of maize meiotic mutant ms43 (p. 7.7).

9.5. Under Asymmetrical Phragmoplast Movement in the Lateral Direction In PMCs of wheat– wheatgrass hybrid F1 no. 625 (T. aestivum cv. Novosibirskaya 67 × A. glaucum) and wheat–rye hybrid F1 no. 8245 (T. aestivum cv. Lutescens x Secale cereale cv. Onokhoiskaya), a curious phenomenon is observed at TI with 15% frequency.

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Figure 9.5. Fragmentary phragmoplast function and incomplete asymmetrical cytokinesis in WR F1: a, b) centrifugal movement of lateral phragmoplast part (arrow) and arrest of its rest parts movement at telophase I, c) result of cell division with this phenotype: binucleate incised monade, d) approachment of spindle poles at metaphase II

The phragmoplast, normal and completely developed out of the straight spindle, moves centrifugally and forms the cell plate only with its some part, and the rest parts remain immobile. As a result, the cell plate incompletely crosses the cytoplasm and, having contacted the local part of mother cell membrane, performs into abnormal daughter cell membranes in a form of incision.

9.6. Cell Plate Dispersion Incomplete daughter cell membranes in a shape of an incision arises under the break of a normal cell plate as a consequence of plastosomes dispersion with the aberration of daughter cell membranes formation (maize mutant phenotype pam1). Here, in part of cells, a full ring incision that encircles the cell equatorial area arise (p. 8.4).

9.7. In Phenotypes with Cell Plate Structure Aberrations (P. 8.3), Incomplete Cytokinesis as Incisions is a Common Phenomenon Such a mechanism of restitution nuclei formation was described in microsporogenesis in Brachiaria (Gallo et al., 2007).

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9.8. Cytokinesis Correction in the Gamma-Phenotype In many of WWG F1 and WR f1 hybrids with an arrest of middle phragmoplast (hollow cylinder) formation and its progressive curvature during telophase (p. 7.3), an addditonal phragmoplast that consists of polar MT bundles, overlapping on the equator, forms berween approached telophase groups (Shamina et al., 2009). This phragmoplast forms a cell plate and carries out cytokinesis, which is successful in many cases, but it is incomplete in some PMC percentage with daughter cell membranes as an incision.

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Figure 9.8. Correction of cytokinesis in gamma-phenotype WR F1 with the formation of binucleate incised monads: a) approachment of telophase chromosome groups at late telophase I as a result of an early phragmoplast progressive curvature, b, c) additional phragmoplast (arrow) formation between approached telophase chromosome groups, d) binucleate incised monad at prophase II – the result of incomplete cytokinetic correction.

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10. GENERAL MEIOTIC ABNORMALITIES LEADING TO RESTITUTION Most of meiotic restitution mechansms consists in the aberration of chromosome spatial separation during karyokinesis or/and avtonomisation of diverged chromosome groups (or daughter nuclei) during cytokinesis. Intracellular distant transport, including chromosome segregation and cell plate formation, is realized by the cytoskeleton. However, in this part of the catalog, meiotic abnormalities are considered, that are not related to cytoskeleton cycle abnormalities, but conditioned by the aberration of the whole set of meiotic division processes.

10.1. Omission of one or Both of Meiotic Divisions Haploid microspores are the result of two successive meiotic divisions. If meiosis is omitted (apomeiosis), 2n gametes genetically identical to parental genotype are formed (Tavoletti, 1994). If only one of the meiotic divisions proceeds, then microspores are also diploid (Carputo et al., 2003; Hayashi et al., 2009). It was reported on the equational division of univalents (into chromatids) at anaphase I in the asynaptic meiosis and the absence of the second meiotic division (Gustafsson, 1935; Ramanna, 1983; Jongedijk et al., 1991; Vorsa, Ortiz, 1992; Ramanna et al., 2003; Barrel and Grossniklaus, 2005), Our observations of male gametogenesis in the natural apomict Arabis holboelli and maize haploid №4607 showed that, under complete asynapsis, univalents

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equational orientation, normal chromatids segregation and cytokinesis, pollen grains develop out of the only dyad members formed as a result of this process. The cytoskeleton rearrangements were not aberrated. We believe that this phenotype, just as those described in the above cited contributions, is an ‗‘omission‘‘ of not the second one, but the first meiotic division, as the processes are exclusively typical of the second meiotic division. Such phenotype represents first division restitution mechanism. In literary sources, it was also reported on the absence of the second meiotic division in the synaptic forms (Conicella et al., 1991; Werner, Peloquin, 1991; Park et al., 2002). Omission of the second meiotic division in forms with normal chromosome synapsis and bivalent formation is genetically equivalent to the second division restitution mechanism.

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Figure 10.1. First meiotic division in maize haploid with univalents equational division and the absence of the second meiotic division: a) PMC at diakinesis, b) MI with the metaphase plate organized by equationally oriented univalents, c) anaphase I, d) dyad of haploid microspores.

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10.2. Equational Division of Univalents in the First Meiotic Division and Failure of Chromosomes Segregation at the Second Meiosis This abnormality leads to the restitution nucleus formation at the first meiotic division. In most cases, further random segregation of chromatides at the second meiosis leads to the formation of nonviable aneuploid gametes. However, in a number of wide F1 cereal hybrid genotypes, undetected aberrations of division spindle formation at prometaphase II lead to chromosome nondivergence, formation of restitution nuclei and potentially viable 2n gametes. Such phenotypes were described in literature (Маап, Sasakuma, 1977; Sasakuma, Kihara, 1981).

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Figure 10.2. Equational division of univalents at MI with a furthet arrest of division spindle formation at MII: a) metaphase plate in the division spindle at MI consisting of equationally oriented univalents, b) dyad with nonreduced members, c) dyad at MII without traits of division spindle formation: cytoskeleton fibers are not revealed, d) dyad at the stage of tetrads: nuclear envelope encircled the nondiverged chromosomes.

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10.3. Cytomixis The known phenomenon of migration of nuclei from one PMC into the other through cytomictic channels (Caetano-Pereira and Pagliarini, 1977; DeSousa and Pagliarini, 1977; Utsunomiya et al., 2004) can be the reason for 2n gametes formation. This process was described as mass in the wild type meiosis of Dactylis glomerata, where it causes gametes formation with a doubled chromosome number out of recipient cells (Falistocco et al., 1995). In a number of forms with abnormal meiosis we analysed, cytomixis was observed. We found out that the transfer of nuclear material is realized together with the nuclear envelope (Shamina et al., 2000b), and the common perinuclear cytoskeleton ring develops at late prophase around the basic and additional nuclei. This ring functions then as a base for common division spindle formation and congression of chromosomes of both nuclei. In some transgenic tobacco lines, 10% of nucleus cells transferred completely to the neighbouring meiocyte at early prophase I (Shamina et al., 2000b), and that was the base for meiotic restitution. It is necessary to note viable 2n gametes, by means of cytomixis can develop only in diploids with a normal homological chromosome synapsis. Formation of 2n gametes as a result of meiocytes fusion was described in cereal meiosis (Risso-Pascotto et al., 2006).

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Figure 10.3. Congression of neighbouring PMCs chromosomes as a result of cytomixis in WWG F1 hybrid: a) donor cell at initial stages of cytomixis: the nucleus stretches towards cytomictic channels (contact point of two PMCs is marked with arrow), b) recipient cell with an additional nucleus that migrated into it as a result of cytomixis, c) double division spindle in the recipient cell at MI, d) common cytoskeleton ring in the binucleate recipient cell at prophase I, the condition for further spindle formation with a doubled chromosome number.

10.4. Cytokinesis Aberrations in the Last Premeiotic Mitosis The phenotype of meiotic mutant pam1 and haploid № 2905 in maize is characterized by cytokinetic arrest in the meiosis and part of cells of the last premeiotic mitosis. It leads to the thing that some cells turn binucleate at prophase I. At late prophase, a common perinuclear ring initiating the common diision spindle at MI is forming round these nuclei. As this spindle includes a double chromosome number, diploid microspores will be the product of such cell (Dorogova and Shamina, 2001). Such phenotype is also encountered in WWG F1hybrids with frequency of 10-15%. 2n gametes formation as a result of failure of the last premeiotic cytokinesis is reported for some forms of oat Avena vaviloviana (Katsiotis and Forsberg, 1995).

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Figure 10.4. Meiotic restitution as a result of the absence of cytokinesis in the last premeiotic mitosis. Binucleate PMCs at prophase I, common cytoskeleton perinuclear ring (marked with arrows) in binucleate PMCs at prophase I of meiotic mutant pam1, (Zea mays) and WWG F1 hybrid: a) binucleate PMC at prophase I in the maize haploid, b) common perinuclear ring around two nuclei at prophase I in PMCs of mutant pam1 c) common perinuclear ring around two nuclei at prophase I in PMCs of WWG F1 hybrid, d) common spindle with a double chromosome set at metaphase I.

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10.5. Backward Movement of Daughter Chromosome Groups from the Spindle Poles into the Cell Center This phenomenon was earlier described under the formation of restitution nuclei (Sears, 1953; Avers, 1954; Stefani, 1986, Jauhar, 2003). However, as there was no cytoskeleton visualization made in these contributions, its mechanisms remained unknown. Abovedescribed are some cytoskeleton abnormalities that lead to the congression of chromosome groups diverged to the poles: anaphase spindle shortening (p. 6.2), earlier phragmoplast curvature or gamma-phenotype (p. 7.3), excessive fibers curvature of late phragmoplast (p. 6.6), absence of cell plate (pp. 8.1, 8.2). An original process of telophase chromosome groups congression in the cell center is observed in WWG F1 hybrid № 27-645 (T.aestivum c Алтайская Нива x E. elongatum), Univalents that constitute aneuploid daughter chromosome groups move at telophase I from poles to the equator sliding with their arms along the telophase spindle surface. They can move both in a group or individually. Congressed on the equator, chromosomes form a unified group which is surrounded by the nuclear envelope and forms a restitution nucleus. Cytokinesis is arrested here: there is no phragmoplast centrifugal movement and cell plate formation. In the second meiotic division, a common division spindle and a dyad with nonreduced members are developing.

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Figure 10.5. Backward movement of telophase chromosomes and telophase chromosome groups to the spindle equator in PMCs of WWG F1 hybrid № 27-645: a) backward movement of telophase chromosome groups to the cell center, b) formation of restitution nucleus out of chromosomes approached on the equator, c, d) return of telophase chromosomes to the equator from the poles.

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One can hypothesise that this abnormality is a result of untimely activity of chromokinesines. These transport proteins are localized on chromosome arms and transport them to MT (+)-ends in the process of metaphase plate formation during late prometaphase (Mazumdar and Misteli, 2005). It is possible, that this process additionally acts at telophase I in WWG №.27-645. Such a phenomenon of untimely onset of intracell process is known in cytokinesis in Schizosaccharomyces pombe, when the cell forms several septs one by one (Fankhausher and Simanis, 1994; Song et al., 1996). The return of chromosomes completely diverged to the poles back to the cell equator was reported on in animal cells under division with the expression of nondestructible form of cyclin B (Wheatley et al., 1997).

DISCUSSION Cell meiotic division is realized by a set of many complicated multistage processes aimed at one result: obtaining haploid spores. Thus, it is not surprising that, in detailed research of abnormal meiosis, there are so many different ways leading to meiotic restitution and 2n gametes formation. Besides this, abnormal processes of pre- and postmeiotic divisions can also affect the ploidy of gametes.

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Meiotic Restituion Mechanisms Typical of only Species with Successive Cytokinesis Formation of restitution nuclei in meiosis with successive cytokinesis has its peculiarities. They consist both in the fact of successive autonomisation of two meiotic division products and in the aberrations of specific mechanisms of mobile phragmoplast formation and operation. Thus, the central position of the division spindle is very important in the cell. If it is abnormal, the cytokinetic abnormalities lead to the formation of incomplete cell membranes as incisions which, in its turn, often results in meiotic restitution. On the contrary, in PMCs of wild type dicots with simultaneous cytokinesis, the division spindle is very often peripheric or excentric in the first meiosis; it does not cause any abnormalities, as cytokinesis is not realized at telophase I. Incomplete cytokinesis after the first meiotic division is one of the restitution mechanisms typical of only monocots with simultaneous cytokinesis. Out of p. 9 it is seen how various the reasons for incomplete cytokinesis are in male meiosis in monocots. Incomplete daughter cell membranes can be a consequence of cytoskeleton and other abnormalities during a long period from prometaphase to telophase completion. It is obvious that abnormal daughter cell membranes as an incision on the mother cell membrane in walless cell division are stubs analogs in the mitosis of walled cells (Nacry et al., 2000; Assaad et al., 2001; Muller et al., 2002). Thus, when analyzing abnormal mitosis, it is also necessary to keep in mind that the appearance of stubs proper in meiotic division products of walled cells does not say anything about the reasons of their development and requires a detailed analysis of the whole mitotic process. Approachment of daughter nuclei on the poles of a curved spindle with parallel cytokinetic aberrations (Shamina et al., 1999) refers to monocots meiotic restitution

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mechanisms. Restitution process can occur under cytokinesis which, here, looks like a not deep incision on the mother cell membrane. An important feature of this restitution mechanism in monocots is the absence of the process that corrects abnormal approachment of daughter nuclei or chromosome groups. On the contrary, in the male meiosis with simultaneous cytokinesis of dicots, such a mechanism functions, and it is based on the thing that, unlike monocots, formation of daughter nuclei proceeds in them also at early telophase. Further on, from daughter nuclear envelopes to the equator, polar MTs polymerise, and they form a developed system of interzonal fibers (immobile phragmoplast). These fibers separate daughter nuclei if they are abnormally approached due to some reason. This correction mechanism is absent in PMCs of monocots with simultaneous cytokinesis, as their phragmoplast is a derivative of the central spindle, and the role of polar MTs is secondary. It is obvious that the described correction mechanism will function in dicots under daughter chromosome pretelophase approachment caused not only by the spindle curvature, but other reasons. Thus, the approachment of chromosome groups on a curved spindle poles (p. 3.2), as a result of spindle shortening at anaphase (p. 6.2), bachward movement of telophase groups to the equator (p. 10.5), are the restitution mechanisms typical of meiosis with successive cytokinesis. They correct in dicots meiosis with simultaneous cytokinesis. Cytokinesis abnormalities in the first meiosis are a typical mechanism of meiotic restitution in monocot plant species. In this case, daughter nuclei easily approach in the cytoplasm, and it leads to the congression of their chromosomes in the common spindle of the following cell division. The original aberrations of successive cytokinesis that are a reason for meiotic restitution, are numerous and diverse (points 7-1 – 7.7). They are a valuable source of information to study temporal and spatial regulation of plant cytokinesis, also the mechanisms of centrifugal movement of phragmoplast/cell plate at normal meiosis (Shamina et al., 2007a).

Meiotic Restitution Mechanisms Typical of only Species with Simultaneous Cytokinesis As in male meiosis of the majority of dicots, cytokinesis is absent after the first meiotic division, they often have occasions in genetic material congression in the common cytoplasm before and during the second meiotic division and common division spindle formation at metaphase II. This abnormality is called ―fused spindles‖ and it is the basic source of nonreduced gametes formation in dicots (see review Ramanna, 1979). In the present review, there are 5 mechanisms that lead to fused spindles and they are described in more or less detail. Two of these mechanisms are abnormalities of interkinetic interzonal cytoskeleton system that corrects the position of daughter nuclei in the common cytoplasm, the gist of rest 3 is so far unclear (Dorogova et al., 1999; Shamina et al., 2001). 1)

mechanism fs: aberration of polar MT polymerization from the polar regions of telophase spindles and from the envelope of daughter nuclei in sugar beet line phenotype В-24 (p .7.9), 2) mechanism fs: aberration of opposite polar MT (+)- ends attachment under the formation of interzonal cytoskeleton system, as in haploid Brassica juncea (p. 7-10),

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3) mechanism fs: approachment of daughter nuclei at prophase II unaccompanied by visible abnormalities of cytoskeleton structural behaviour (p. 2.2), 4) mechanism fs: fusion of perinuclear cytoskeleton rings of approached prophase nuclei and the common perinuclear ring formation, out of which a common fused spindle forms (p. 2.3). Possibly, it is a necessary condition for a fused spindle formation at the approachment of daughter nuclei. 5) mechanism fs: approachment, due to an unknown reason, and fusion of chaotic prometaphase figures (mid-prometaphase) in the second meiotic division (p. 4.4). Partial or complete arrest of polar MTs polymerization at telophase I that leads to fused spindles, is interesting to be compared to the similar cytoskeleton abnormality that proceeds in the second meiotic division on the background of parallel spindles (ps) (p. 7.8). This abnormality also leads to the formation of unreduced 2n gametes and consists in polar MTs polymerization arrest and, thus, the impossibility of phragmoplasts formation between nonsister daughter nuclei. Despite the identity of aberrations proper, the restitution mechanisms turn different in these two cases (fused spindles and parallel spindles). Approachment of daughter nuclei at prophase II is the reason for fused spindles phenotype, typical of many different dicot forms producing 2n gametes. Normally, at this stage, the interzonal cytoskeleton depolymerises, nuclei become deprived of its support, but do not approach each other in wild type meiosis. According to our data obtained both with the classical method and immunostaining, approachment of prophase nuclei is not accompanied by any aberrations in the microtubule skeleton behaviour (Conicella et al.,2003). Thus, the reason for the approachment of nuclei at prophase II remains unknown in these phenotypes. An interesting phenomenon is the fusuion of perinuclear cytoskeleton rings in approached prophase nuclei. It is a necessary stage of daughter cell genomes congression at prometaphase II and the development of common division spindle. It is unknown whether this phenomenon is cytoskeleton behavioural abnormalities or it is part of a special mechanism that provides genome consolidation in the division and‘‘switches on‘‘ untimely. The importance of common cortical cytoskeleton ring for genome consolidation by means of a common spindle formation also demonstrates the restitution mechanism described in p. 2.4 of maize haploids. In this case, many micronuclei in a monad – the first meiotic abnormality product – are encircled by a cortical cytoskeleton ring at prophase II. As a result, a common division spindle at metaphase II and a dyad of normal microspores at the stae of tetrads are forming. Approachment and fusion of mid-prometaphase figures are not accompanied by any visible aberrations in MT cytoskeleton structures, and the reason for this phenomenon is not yet known. Processes that occur in the second meiotic division in PMCs with ‗‘fused spindles‘‘ phenotype can be compared to the process of genetic material congression at prophaseprometaphase of the first meiotic division in multinucleate PMCs of both mono - and dicot plant species. PMCs can be multinucleate when entering meiosis as a result of cytoskeleton abnormalities of the last premeiotic mitosis, also as a resulu of cytomixis. We observed the first meiotic division in binucleate PMCs of maize meiotic mutant pam1 (Dorogova, Shamina, 2001), and also phenotypes with cytomixis in different mono- and dicot species (Shamina et al., 2000). In all these numerous cases, the first meiotic division had the same procedure in multinucleate cells: all nuclei approached in the center, were encircled by the

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common perinuclear MT ring and formed their unified division spindle at metaphase I. If multinucleate cells formed as a result of chromosome segregation abnormalities and/or cytokinesis in the first meiotic division, in the second one several division spindles developed out of multiple nuclei (Shamina et al., 1981). In other words, in the second meiotic division, the process of genetic material consolidation is not observed, which is obligatory in the first meiotic division. Obviously, nuclear cytoplasmic interactions proceed in a different way in dicots: active genetic material consolidation is realized at the first meiosis, its autonomisation – at meiosis II. One can hypothesise that consolidation of genetic material of daughter nuclei in the second meiotic division of dicots with ‗‘fused spindles‘‘ phenotype (points 2.2. 2.3. 4.4), and ‗‘common cortical ring‘‘ (p. 2.4), there is some aberration of signal mechanism that makes the cell divide on the ‗‘scenario‘‘ of the first meiotic division. Spindle disorientation at metaphase II in the common cytoplasm refers to specific restitution mechanisms of dicot plant species: parallel (p. 7.8) and the so-called ‗‘tripolar‘‘ configurations (p. 5.2) (Mok, Peloquin, 1975). As it was noted above, parallel spindle coorientation leads to the restitution only in combination with an additional abnormality: arrest in polar MT bundles formation at telophase II (Genualdo et al., 1998). Tripolar configurations are disoriented spindles whose two poles are in one point, i.e. considerably approached (p. 5.2) (Mok, Peloquin, 1975; Ramanna, 1979).

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Meiotic Restitution Mechanisms Common for Monoand Dicotyledonous Species Cytoskeleton reorganization from prophase till the onset of telophase occurs in the meiosis of mono- and dicotyledonous plants the same way. Thus, cytoskeleton abnormalities observed at these stages and leading to the formation of restitution nuclei can be considered universal. And some of such abnormalities were really found by us both mono- and dicot plant species, and we are convinced that further wide investigations will find out the same for the rest of abnormalities. Aberration in chromosome segregation that leads to the formation of restitution nuclei in meiosis of monocots was described also in the meiosis of dicots (without division spindle visualization) (Lam, 1974). Out of the abnormalities described in this chapter, it is possible to consider the following common restitution mechanisms for monoand dicotyledonous plant species: 1) chromosome arrest in the ‗‘bouquette‘‘ and drastically unequal chromosome segregation to the poles related with this thing (p. 2.6), 2) arrest of cytoskeleton cycle at the stage of radial bundles (p. 2.1), 3) autonomous cytoskeleton ring formation under accentric nucleus position (p. 2.5), 4) cortical cytoskeleton ring formation at prophase of the second meiosis (p. 2.4), 5) arrest of perinuclear cytoskeleton ring disintegration at early prometaphase (p. 3.1), 6) arrest of cytoskeleton penetrating the former nuclear area at early prometaphase (p. 3.3) (found in dicot lines Res91, Res79 transgenic tobacco Nicotiana tabacum (Sidorchuk, personal communication); 7) monopolar spindle (p. 4.1) (in dicots - haploid Brassica juncea), 8) chromosome monopolar movement in a bipolar spindle under the absence of kinetochore fibers (p. 4.3 ‗‘comet‘‘ phenotype), 9) arrest of kinetochore fibers formation at mid-prometaphase (p. 4.2), 10) arrest of cytoskeleton realising from the chaotic stage at late prometaphase (p. 5.1, chaotic spindle) (it dicots it is observed in the phenotype of pea meimutant ms3) 11) abnormalities of anaphase chromosome movement (p. 6.1), 12) spindle shortening at anaphase (p. 6.2), 13) cell plate absence (p. 8.1) (in dicots –

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meimutant tetraspore in Arabidopsis) (Spielman et al., 1997), 14) cell plate abnormalities (p. 8.3), 15) arrest of daughter cell membranes formation (p. 8.4) and 16) the so-called general abnormalities (p. 10), but p. 10.5. Absence of one of meiotic divisions (Ramanna, 1983; Gill et al., 1985; Conicella et al., 1991) and ‗‘premature cytokinesis‘‘ – when the cytoplasm divides at telophase I and the second meiotic division does not occurs (p. 10.1) (Watanabe, Peloquin, 1993) was described in dicots among them.

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General Review of Meiotic Restitution Cytoskeleton Mechanisms According to their mechanisms, the abnormalities we described, that lead to meiotic restitution, are divided into three groups;1) formation of restitution nuclei, 2) congression of daughter genomes, 3) consolidation of cytoskeleton structures, 4) regulatory aberrations. Formation of Restitution Nuclei is a Result of a Complete Arrest in Spatial Separation of Daughter Genomes. We detected several cytoskeleton abnormalities that lead to the formation of restitution nuclei. Theses are the above-described arrest of radial cytoskeleton reorientation, perinuclear ring conservation, autonomous ring, ring spindle, arrest of MT entering the former nuclear area, arrest of kinetochore MT formation, arrest in bipolar spindle fibers development, arrest in bipolar spindle formation (chaotic nonpolar spindles). Thus development of restitution nuclei occurs due to drastical aberrations of segregation apparatus, - the division spindle. Congression of daughter genomes, as a restitution process, occurs after chromosomes segregation, and the formation of the nuclear envelope around them. After this, as a result of cytoskeleton abnormalities, a recurrent approachment of daughter nuclei in the common cytoplasm can proceed. The basic reason for this is aberrations in structures that realize daughter genomes autonomisation, i.e. cytokinetic structures. As a consequence, daughter nuclei approach in the common cytoplasm and, in the next division, after NEBs, both daughter genomes become involved in one common division spindle. As a result of segregation, nuclei with a doubled chromosome number are developing. We have never observed the fusion of envelopes of approached daughter nuclei. According to our data, approachment of nuclei in the common cytoplasm can occur as a consequence of abnormalities of cytoskeleton-maintaining structures: cenral spindle fibers and interzonal MT system, also due to the absence of cell plate or daughter cell membranes. Approachment of daughter nuclei can be spontaneous in the common cytoplasm, just as in phenotype fs in dicots. Consolidation of cytoskeleton structures, as our data testify, also lead to the congression of segregated daughter genomes and, hence, to the meiotic restitution. Spontaneous approachment of daughter nuclei with developed perinuclear cytoskeleton rings occurs in 50% of PMC potato clone CE10 at prophase II. After complete approachment of nuclei, rings disjunct and fuse forming one common ring around both nuclei. After NEB, a common division spindle forms out of a common perinuclear cytoskeleton system. In 100% of PMCs of tomato meiotic mutant, as6, at mid prometaphase II, approachment and fusion of cytoskeleton chaotic figures proceed. No other MT cytoskeleton aberrations are observed in this phenotype. Regulatory aberrations make up a considerably big group of meiotic restitution mechanisms. It is the omission of one or both meiotic divisions (pp. 10.1, 10.2) and also the

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manifestation of processes typical of other studies at certain meiotic stages (p. 10.5, fused spindle phenotype, pp. 2.2, 2.3, 4.4).

ACKNOWLEDGMENTS The author would like to express her gratitude to G.M Seriukov and E.G. Seriukova for their kindly provided material of wheat-wheat-grass F1, L.F. Dudka and V.Ya. Kovtunenko for their wheat-rye F1 hybrids, O.A. Shatskaya for maize haploids, Zh.M Mukhina for rice haploids and Alexander V. Zhuravlev for the English version of this big contribution.

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REFERENCES Abbot AG, Hess JE, Gerbi SA. 1981. Spermatogenesis in Sciara coprophila. 1. Chromosome orientation on the monopolar spindle of meiosis 1. Chromosoma 83: 1-18. Alfano F, Cammareri M, Errico A, Frusciante L, Conicella C. 1999. 2n Gametes in Solanum tuberosum dihaploids. Amer J Potato Res 76: 281-285. Alfano F, Errico A, Conicella C. 1997. Potato meiotic mutants and importance of changes in the cytoskeleton. Cell Biol Internat. 21: 3-5. Amanda J. Wright,1 Kimberly Gallagher,2 and Laurie G. Smith. 2009. Discordia1 and alternative discordia1 function redundantly at the cortical division site to promote preprophase band formation and orient division planes in maize. Plant Cell 21: 234-247 Ambrose JC, Cyr R. 2007. The kinesin ATK5 functions in early spindle assembly in Arabidopsis . Plant Cell 19: 226–236. Ambrose JC, Shoji T, Kotzer AM, Pighin JA, Wasteneys GO. 2007. The Arabidopsis CLASP Gene Encodes a Microtubule-Associated Protein Involved in Cell Expansion and Division. Plant Cell 19: 2763–2775. Andreuzza S, Siddiqi I. 2008. Spindle positioning, meiotic nonreduction, and polyploidy in plants. PLoS Genet 4: e1000272. Assaad FF, Huet Y, Mayer U, Jürgens G. 2001.The cytokinesis gene KEULE encodes a Sec1 protein that binds the syntaxin KNOLLE. J Cell Biol 152:531–544. Avers CJ. 1954. Chromosome behaviour in fertile triploid aster hybrids. Genetics 39 : 117— 126. Azimzadeh J, Traas J, Pastuglia M. 2001. Molecular aspects of microtubule dynamics in plants. Curr Opin Plant Biol 4:513–519. Bajer A. 1968. Behavior and fine structure of spindle fibers during mitosis in endosperm. Chromosoma 25: 249-281. Bajer A. 1987. Substructure of the kinetochore and reorganization of kinetochore microtubules during early prometaphase. Eur J Cell Biol 43: 23-34. Bajer A, Mole-Bajer J. 1972. Spindle dynamics and chromosome movements. Int Rev Cytol, Suppl 3: 1-217. Banningan A, Scheible W-R, Lukowitz W, Fagerstrom C, Wadsworth P, Somerville C, Baskin T. 2007. A conserved role for kinesin-5 in plant mitosis. J Cell Sci 120: 28192827.

Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

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

138

Nataliya V. Shamina

Barba-Gonzalez R, Lim K-B, Ramanna MS, Visser RGF, Van Tuyl JM. 2005. Occurrence of 2n gametes in the F1 hybrids of Oriental X Asiatic lilies (Lilium): relevance to intergenomic recombination and backcrossing. Euphytica 143: 67-73. Baskin TI, Cande WZ. 1990. The structure and function of the mitotic spindle in flowering plants. Annu Rev Plant Physiol Plant Mol Biol 41: 27-315. Bastiaanssen HJM, Van den Berg PMM, Lindhout P, Jacobsen E, Ramanna MS. 1998. Postmeiotic restitution in 2n-egg formation of diploid potato. Heredity 81: 20-27. Batoko H, Moore I. 2001. Plant cytokinesis: KNOLLE joins the club. Curr Biol 11: 423-426. Bielig LM, Mariani A, Berding N. 2003. Cytological studies of 2n male gamete formation in sugarcane, Saccharum, L. Euphytica 133: 117-124. Binarova P., Dolezel J, Draber P, Heberle-Bors E, Strnad M, Bogre L. 1998. Treatment of Vicia faba root tip cells with specific inhibitors to cyclin-dependent kinases leads to abnormal spindle formation. Plant J 16: 697-707. Blangy A, Lane HA, d‘Hefin P, Harper M, Kress M, Nigg EA. 1995. Phosphorilation by p34cdc2 regulates spindle association of human Eg5, a kinesin-related motor essential for bipolar spindle formation in vivo. Cell 83: 1159-69. Bouck DC, Bloom KS. 2005. The kinetochore protein Ndc10p is required for spindle stability and cytokinesis in yeast. Proc Natl Acad Sci USA 102: 5408-5413. Brown RC, Lemmon BE. 1992. Control of division plane in normal and griseofulvin-treated microsporocytes of Magnolia. J Cell Sci 103: 1031-1038. Brown RC, Lemmon BE. 1998. Cytokinesis occurs at boundaries of domains delimited by nuclear-based microtubules in sporocytes of Conocephalum conicum (Bryophyta). Cell Motil Cytoskeleton 11: 139-146. Brust-Mascher I, Civelekoglu-Scholey M, Kwon M, Mogilner A, Scholey JM. 2004. Model for anaphase B: Role of three mitotic motors in a switch from poleward flux to spindle elongation. Proc Natl Acad Sci USA 101: 15938-15943. Bucciarelli E., Giansanti M.G., Bonaccorsi S., Gatti M. 2003. Spindle assembly and cytokinesis in the absence of chromosomes during Drosophila male meiosis. J Cell Biol 160: 993-999. Caetano-Pereira MC, Paliarini MS. 1997. Cytomixic in maize microsporocytes. Cytologia 62: 351-355. Cai X, Xu SS. 2007. Meiosis-Driven Genome Variation in Plants. Curr Genomics 8: 151– 161. Camadro EL, Saffarano SK, Espinillo JC, Castro M, Simon PW. 2008. Cytologica mechanisms of 2n pollen formation in the wild potato Solanum okadae and pollen-pistil relations with the cultivated potato Solanum tuberosum. Genet Resour Crop Evol 55: 471-477. Carputo D, Barone A, Frusciante L. 2000. 2n Gametes in the potato: essential ingredients for breeding and germplasm transfer. Theor Appl Genet 101: 805-813. Carputo D, Frusciante L, Peloquin SJ. 2003. The role of 2n gametes and endosperm balance number in the origin and evolution of polyploids in the tuber-bearing Solanum. Genetics 163: 287-294. Chan A, Cande WZ.1998. Maize meiotic spindles assemble around chromatin and do not require paired chromosomes. J Cell Sci 111: 3507-3515.

Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

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2n Pollen Formation: 40 Cytological Mechanisms of Nuclear Meiotic Restitution

139

Chen YC, McCormick S. 1996. Sidecar pollen, an Arabidopsis thaliana male gametophytic mutant with aberrant cell divisions during pollen development. Development 122: 3243-3253. Chistyakova VN. 1974. Cytogenetic studies of polyhaploids of incomplete Triticum x Agropyron amphidiploids (2n = 56). II. Formation of microspores. Genetika (Russ.) 10: 5-16. Conicella C, Barone A, Del Guidice A, Frusciante L, Monti LM. 1991. Cytological evidences of SDR-FDR mixture in the formation of 2n eggs in a potato diploid clone. Theor Appl Genet 81: 59-63. Conicella C, Capo A, Cammareri M, Errico A, Shamina N, Monti L. 2003. Elucidation of meiotic nuclear restitution mechanisms in potato through analysis of microtubular cytoskeleton. Euphytica 133: 107-115. Consiglio F, Carputo D, Monti L, Conicella C. 2004. Exploitation of genes affectin meiotic non-reduction: Arabidopsis as a model? Sex Plant Reprod 17: 97-105. Cook M. 2004. Cytokinesis in Coleochaete orbicularis (Charophyceae): an ancestral mechanism inherited by plants. Am J Bot 91: 313-320. Cutler SR, Ehrhardt DW. 2002. Polarized cytokinesis in vacuolated cells of Arabidopsis. Proc Natl Acad Sci USA 99: 2812-2817. d'Erfurth I, Jolivet S, Froger N, Catrice O, Novatchkova M, Mercier R. 2009. Turning Meiosis into Mitosis. PLoS Biol 7: e1000124. d'Erfurth I, Jolivet S, Froger N, Catrice O, Novatchkova M, Simon M, Jenczewski E, Mercier R. 2008. Mutations in AtPS1 (Arabidopsis thaliana Parallel Spindle 1) lead to the production of diploid pollen grains. PLoS Genet 4: e1000274. Darlington CD. 1965. Cytology. London, J & A Churchill LTD. DeMey J, Lambert AM, Bajer AS, Moeremans M, DeBrabander M. 1982. Visualization of microtubules in interphase and mitotic plant cells of Haemanthus endosperm with the immunogold staining method. Proc Natl Acad Sci USA 79: 1898-1902. DeSouza AM, Pagliarini MS. 1997. Cytomixis in Brassica napus var. oleifera and Brassica campestris var. oleifera (Brassicaceae). Cytologia 62: 25-29. Deng M, Gao J, Suraneni P, Li R.. 2009. Kinetochore-independent chromosome poleward movement during anaphase of meiosis II in mouse eggs. PLoS ONE 4(4): e5249 Dinis AM, Mesquita JF.1993. The F-actin distribution during microsporogenesis in Magnolia soulangeana Soul. Sex Plant Reprod 6: 57-63. Dorogova NV, Shamina NV. 2000. Mutation acd disturbing microtubule rearrangements during male meiosisin shugar beet. Tsitologiya (Russ.) 42: 372-377. Dorogova NV, Shamina NV. 2001. Phenomenon of excessive cytokinesis in maize meiotic mutant pam1 phenotype. Tsytologiya (Russ).43: 471-476. Dorogova NV, Shamina NV, Maletskii SI. 1999. The mutational variation of a cytoskeleton and forming of the unreduced male gametes in sugar beet. Shugar Tech 1: 89-92. Dujardin M, Hanna WW. 1988. Cytology and breeding behavior of a partially fertile triploid pearl millet. J Hered 79:216–218. El Mokadem H, Crespel L, Meynet J, Gudin S. 2002. The occurrence of 2n pollen and the origin of sexual polyploids in dihaploid roses (Rosa hybrida L.). Euphytica 125: 169177. Eleftheriou EP, Baskin TI, Hepler PK. 2005. Aberrant cell plate formation in the Arabidopsis thaliana microtubule organization 1 mutant. Plant Cell Physiol 46: 671-675.

Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

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

140

Nataliya V. Shamina

Esseling-Ozdoba A, Vos JW, van Lammeren AAM, Emons AMC. 2008. Synthetic lipid (DOPG) vesicles accumulate in the cell plate region but do not fuse. Plant Physiol 147: 1699–1709. Estrada-Luna AA, Huanga-Mamani W, Acosta-Garsia G, Leon-Martinez G, Becerra-Flora A, Perez-Ruiz R, Vielle-Caizada JP. 2002. Beyond promiscuity: From sexuality to apomixis in flowering plants. In Vitro Cell Devel Biol – Plant 38: 146-151. Euteneuer U, Jackson WT, McIntosh JR. 1982. Polarity of spindle microtubules in Haemanthus endosperm. J Cell Biol 91: 644-653. Falistocco E, Tosti N, Falcinelli M. 1995. Cytomixis in pollen mother cells of diploid Dactylis, one of the origins of 2n gametes. J Hered 86: 448-453. Fankhauster C, Simanis V. 1994. Cold fission: splitting the pombe cell at room temperature. Trend Cell Biol 4:96-101. Fuge H. 1994. Unorthodox male meiosis in Trichosia pubescence (Sciaridae). J Cell Sci 107: 299-312. Fukuda K, Sakamoto S. 1992. Studies on unreduced gamete formation in hybrids between tetraploid wheats and Aegilops squarrosa L. Hereditas 116: 253-255. Gallo PH, Micheletti PL, Boldrini KR, Risso-Pascotto C, Pagliarini MS, do Valle CB. 2007. 2n gametes formation in the genus Brachiaria (Poaceae: Paniceae). Euphytica 154: 255-260. Genualdo G., Errico A., Tiezzi A., Conicella C. 1998. -Tubulin and F-actin distribution during microsporogenesis in a 2n pollen producer of Solanum. Genome 41: 636-641. Gill BS, Kam-Morgan LWW, Shepard JF. 1985. An apparent meiotic mutation in a mesophyll cell protoclone of the “Russel Burbank” potato. J Hered 76: 17-20. Goddard RH, Wick SM, Silflow CD, Snustad DP. 1994. Microtubule components of the plant cell cytoskeleton. Plant Physiol 104: 1-6. Gonzalez C, Saunders RD, Casal J, Molina I, Carmena M, Ripoll P. 1990. Mutation in the asp locus of Drosophila lead to multiple free centrosomes in syncitial embrios, but restrict centtrosome duplication in larval neuroblasts. J Cell Sci 96: 605-616. Gu X., Verma DPS. 1997. Dynamics of phragmoplastin in living cells during cell plate formation and uncoupling of cell division. Plant Cell 9: 157-169. Gustafsson A. 1935. Studies on mechanisms of parthenogenesis. Hereditas 21: 1-111. Hamada T. 2007. Microtubule-associated proteins in higher plants. J Plant Res 120: 79-98. Harper L, Golubovskaya IN, Cande WZ. 2004. A bouquet of chromosomes. J Cell Sci 117: 4025-4032. Hauser BA, He JQ, Park SO, Gasser CS. 2000. TSO1 is a novel protein that modulates cytokinesis and cell expansion in Arabidopsis. Development 127: 2219-2226. Hayashi M, Kato J, Ohashi H, Mii M. 2009. Unreduced 3x gametes formation of allotriploid hybrid derived from the cross of Primula denticulata (4x) x P. rosea (2x) as a causal factor for producing pentaploid hybrids in the backcross with pollen of tetraploid P. denticulata. Euphytica 169: 123-131. Heck MMS, Pereira A, Pesavento P, Yannoni Y, Spradling AC. 1993. The kinesin-like protein KLP61F is essential for mitosis in Drosophila. J Cell Biol 123: 665-679. Hepler PK, Bonsignore CL. 1990. Caffeine inhibition of cytokinesis: ultrastructure of cell plate formation/degeneration. Protoplasma 157: 182-192. Hermsen JG. 1984. Mechanisms and genetic implications of 2n-gamete formation. Iowa State J Research 58: 421-434.

Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

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

2n Pollen Formation: 40 Cytological Mechanisms of Nuclear Meiotic Restitution

141

Hiwatashi Y, Obara M, Sato Y, Fujita T, Murata T, Hasebe M. 2008. Kinesins are indispensable for interdigitation of phragmoplast microtubules in the moss Physcomitrella patens. Plant Cell 20: 3094-3106. Hogan C. 1987. Microtubule patterns during meiosis in two higher plant species. Protoplasma 138: 126-136. Hulskamp M, Parekh NS, Grini P, SchneitzK, Zimmermann I, Lolle SJ, Pruitt RE. 1997. The STUD gene is required for male-specific cytokinesis after telophase II of meiosis in Arabidopsis thaliana. Dev Biol 187: 114-124. Hutchinson GE. 1953. Marginalia. Amer Sci 41: 630. Iwanaga M. 1984. Discovery of a synaptic mutant in potato haploids and its usefulness for potato breeding. Theor Appl Genet 68: 87-93. Jauhar PP. 2003. Formation of 2n gametes in durum wheat haploids: sexual polyploidization. Euphytica 133: 81-94. Jongedijk E, Ramanna MS, Sawor Z, Hermsen JGTh. 1991. Formation of first division restitution (FDR) 2n-megaspores through pseudohomotypic division in ds-1 (desynapsis) mutants of diploid potato: routine production of tetraploid progeny from 2xFDR x 2xFDR crosses. Theor Appl Genet 82: 645-656. Jorgensen CA. 1928. The experimental formation of heteroploid plants in the genus Solanum. J Genet 19: 133-210. Jurgens G. 2005. Cytokinesis in higher plants. Ann Rev Plant Biol 56: 281-299. Karpechenko GD. 1927. The production of polyploidy gametes in hybrids. Hereditas 9: 349368. Katsiotis A, Forsberg RA. 1995. Discovery of 2n gametes in tetraploid oat Avena vaviloviana. Euphytica 81: 1-6. Kawamura E, Himmelspach R, Rashbrooke MC, Whittington AT, Gale KR, Collings DA, Wasteneys GO. 2006. MICROTUBULE ORGANIZATION 1 regulates structure and function of microtubule arrays during mitosis and cytokinesis in the Arabidopsis Root. Plant Physiol 140: 102–114. Lam S. L. 1974. Origin and formation of unreduced gametes in the potato J. Heredity. 65: 175-178. Lambert AM, Bajer A. 1975. Fine sttructure dynamics of the prometaphase spindle. J Microscope Biol Cell 23: 181-194. Lambert AM, Lloyd CW. 1994. The higher plant microtubule cycle. In: Microtubules, Wiley-Liss, Inc. pp. 325-341. Lauber MN, Waithenegger I, Steinmann T, Schwarz H, Mayer U. 1997. The Arabidopsis KNOLLE protein is a cytokinesis specific syntaxin. J Cell Biol 139: 1485-1493. Lee Y-RJ, Giang HM, Liu B. 2001. A novel plant kinesin-related protein specifically associates with the phragmoplast organelles. Plant Cell 13: 2427–2440. Lee Y-RJ, Li Y, Liu B. 2007. Two Arabidopsis phragmoplast-associated kinesins play a critical role in cytokinesis during male gametogenesis. Plant Cell 19: 2595–2605 Lee YRJ, Liu B. 2007. Cytoskeletal motor proteins in plant cell division. Plant Cell Monogr 9: 169-193. Lemos CL, Sampaio P, Maiato H, Costa M, Omel‘yanchuk L, Liberal V, Sunkel K. 2000. Mast, a conserved microtubule-associated protein required for bipolar mitotic spindle organization. EMBO J 19: 3668-3682. Love H, Craig WJ. 1919. Fertile wheat-rye hybrids. J Hered 10: 195—207.

Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

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

142

Nataliya V. Shamina

Lukowitz W, Mayer U, Jurgens G. 1996. Cytokinesis in the Arabidopsis embrio involves the syntaxin-related KNOLLE gene product. Cell 84: 61-71. Maan SS, Sasakuma T. 1977. Fertility of amphihaploids in Triticinae. J Hered 68: 87— 94. Magnard JL, Yang M, Chen YC, Leary M, McCormick S. 2001. The Arabidopsis gene tardy asynchronous meiosis is required for the normal pace and synchrony of cell division during male meiosis. Plant Physiol 127: 1157-1166. Mathur J. 2004. Plant cytoskeleton: reinforcing lines of division in plant cells. Curr Biol. 14(7):R287-9. Mayer U, Herzog U, Berger F, Inze D, Jurgens G. 1999. Mutation in the pilz group genes disrupt the microtubule cytoskeleton and uncouple cell cycle progression from cell division in Arabidopsis embryo and endosperm. Eur J Cell Biol 78: 100-108. Mazumdar M., Misteli T. 2005. Chromokinesins: multitalented players in mitosis. Trends Cell Biol 15: 349-355. McNally K, Audhya A, Oegema K, McNally FJ. 2006. Katanin controls mitotic and meiotic spindle length. J Cell Biol 175: 881-891. Mendiburu AO, Peloquin SJ. 1976. Sexual polyploidization and depolyploidization: some terminology and definitions. Theor Appl Genet 48: 137-143. Mineyuki Y. 2007. Plant microtubule studies: past and present. J Plant Res 120: 45-51. Mitsuoka S. 1953. Morphology and fertility of the Fl hybrids between certain varietes of Triticum dicoccum and Aegilops squarrosa. Seiken Ziho 6: 91—96. Mok DWS, Peloquin SJ. 1975. Three mechansms of 2n pollen formaton in diploid potatoes. Can J Genet Cytol 17: 217-225. Mole-Bajer J. 1969. Fine structural studies of apolar mitosis. Chromosoma 26: 427-448. Muntzing A. 1939. Studies on the properties and the ways of production of rye-wheat amphidiploids. Hereditas 25: 387—430. Müller S, Fuchs E, Ovecka M, Wysocka-Diller J, Benfey PN, Hauser M-T. 2002. Two new loci, PLEIADE and HYADE, implicate organ-specific regulation of cytokinesis in Arabidopsis. Plant Physiol 130: 312–324. Nacry P, Mayer U, Jurgens G. 2000. Genetic dissection of cytokinesis. Plant Mol Biol 43: 719-733. Nelson MN, Mason AS, Castello MC, Thomson L, Yan G, Cowling WA. 2009. Microspore culture preferentially selects unreduced (2n) gametes from an interspecific hybrid of Brassica napus L X Brassica carinata Braun. Theor Appl Genet 119: 497-505. Nonomura K-I, Morohoshi A, Nakano M, Eiguchi M, Miyao A, Hirochika H, Kurata N. 2007. A germ cell–specific gene of the ARGONAUTE family is essential for the progression of premeiotic mitosis and meiosis during sporogenesis in rice. Plant Cell 19: 2583–2594. Otegui M, Staehelin LA. 2000. Syncytial-type cell plates. A novel kind of cell plate involved in endosperm cellularization of Arabidopsis. Plant Cell 12: 933–948. Pagliarini MS, Takayama SY, de Freitas PM, et al. 1999. Failure of cytokinesis and 2n gametes formation in Brasilian accessions of Paspalum. Euphytica 108: 129-135. Palevitz BA, Hepler PK. 1974. The control of the plane of division during stomatal differentiation in Allium. I. Spindle reorientation. Chromosoma 46: 297-326. Park SM, Wakana A, Hiramatsu M, Uresino K. 2002. A tetraploid hybrid plant form 2x x 4x crosses in Vitis and its origin. Euphytica 126: 345-353.

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Peirson IH, Bowling SE, Makaroff CA. 1997. A defect in synapsis causes male sterility in a T-DNA-tagged Arabidopsis thaliana mutant. Plant J 11: 659-669 Peloquin SJ, Boteux LS, Carputo D. 1999. Meotic mutants n potato: valuable variants. Genetics 153: 1493-1499. Prakken R, Swaminatham MS. 1952. Cytological behaviour of some interspecific hybrids in the genus Solanum sect. Tuberarium. Genetica 26: 77-101. Qu L, Vorsa N. 1999. Desynapsis and spindle abnormalities leading to 2n pollen formation in Vaccinium darrowi. Genome 42: 35-40. Ramanna MS. 1974. The origin of unreduced microspores due to aberrant cytokinesis in the meiocytes of potato and its genetic signifcance. Euphytica 23: 20-30. Ramanna MS. 1979. A re-examination of the mechanisms of 2n gamete formation in potato and its implications for breeding. Euphytica 28: 537-561. Ramanna MS. 1983. First division restitution gametes through fertile desynaptic mutants of potato. Euphytica 32: 337-350. Ramanna MS, Kuipers AGJ, Jacobsen E. 2003. Occurrence of numerically unreduced (2n) gametes in Alstroemeria interspecific hybrids and their significance for sexual polyploidisation. Euphytica 133: 95-106. Rhoades MM, Dempsey E. 1966. Induction of chromosome doubling at meiosis by the elongate gene in maize. Genetics 54: 505-522. Riley R, Chapman V. 1957. Haploids and polyhaploids in Aegilops and Triticum. Heredity 11: 195—207. Risso-Pascotto C, Pagliarini MS, Valle CB. 2006. Microsporogenesis in Brachiaria dictyoneura (Fig. & De Not.) Stapf (Poaceae; Paniceae). Getet Mol Res 5(4): 837-845. Sadasivaiah RS, Kasha KJ. 1971. Meiosis in haploid barley - an interpretation of nonhomologous chromosome association. Chromosoma 35: 247-263. Sasakuma Т, Kihara H. 1981. A synthesized common wheat obtained from a triploid hybrid, Aegilops squarrosa var. Strangulata x Triticum durum . Wheat Inf. Serv 52: 14—18. Sawin KE, LeGuellec K, Philippe M, Mitchison TJ. 1992. Mitotic spindle organization by a plus end-directed microtubule motor. Nature 359: 540-543. Schmit AC, Vantard M, de Mey J, Lambert AM. 1983. Aster-like microtubule centers establish spindle polarity during interphase – mitosis transition in higher plant cells. Plant Cell Reports 2: 285-288. Sears ER. 1953. Addition of the genome of Haynaldia villosa to Triticum aestivum. Amer J Bot 40: 168—174. Seguí-Simarro JM, Austin II JR, White EA, Staehelin LA. 2004. Electron tomographic analysis of somatic cell plate formation in meristematic cells of Arabidopsis preserved by high-pressure freezing. Plant Cell 16: 836-856. Segui-Simarro JM, Otegui MS, Austin II JR, Staehelin LA. 2007. Plant cytokinesis – insights gained from electron tomography studies. Plant Cell Monogr 9: 251-287. Seryukova EG, Dorogova NV, Zharkov NA, Shamina NV. 2003. Aberrations in prometaphase leading to the formation of restitution nuclei. Tsitologiya (Russ.) 45: 243-248. Shamina N.V.. 2005a. Formation of division spindle in higher plant meiosis. Cell Biology International 29: 309-318.

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Nataliya V. Shamina

Shamina N.V.. 2005b. Catalogue of division spindle abnormalities in higher plants. Cell Biology International 29: 384-391. Shamina NV, Dorogova NV. 1995. Ultrastructural study of abnormal meiosis in mutant ms43 in Zea mays. Tsitologiya (Russ.) 37: 561-566. Shamina NV, Dorogova NV. 2006. Dynamics of microtubular cytoskeleton in higher plant meiosis. VII. Mechanisms of simultaneous cytokinesis. Tsitologiya (Russ.) 48: 127-132. Shamina NV, Dorogova NV, Cherednichenko AE. 2004. Consolidation of cytoskeleton during plant spindle formation. II. Fused spindles. Tsitologiya 46: 685-689. Shamina NV,. Dorogova NV, Orlova AG,. Trunova SA. Gontcharov NP. 1999. Abnormalities of spindle and cytokinesis behavior leading to meiotic restitution in cereals. Cell Biol Internatl 23: 863-870. Shamina NV, Dorogova NV, Perelman PL. 2000a. Disruption of male meiosis in Pisum sativum L. induced by mutation ms3. Tsitologiya (Russ.) 42: 404-411. Shamina NV, Dorogova NV, Seriukova EG. 2003a. Dynamics of microtubular cytoskeleton in higher plant meiosis. II. The stage of prophase perinuclear band formation. Tsitologiya (Russ.) 45: 655-660. Shamina NV, Dorogova NV, Seriukova EG, Silkova OG. 2003c. Dynamics of microtubular cytoskeleton in higher plant meiosis. III. Abnormalities of the early prometaphase. Tsitologiya (Russ.) 45: 661-667. Shamina NV,. Dorogova NV, Sidorchuk Yu V, Zagorskaya AA, Deineko EV, Shumny VK. 2000b. Abnormalities of meiotic division caused by T-DNA tagged mutation in tobacco (Nicotiana tabacum L.). Cell Biol Internatl 25: 367-369. Shamina NV, Dudka LF, Kovtunenko VY, Bolobolova EU. 2009. Additional phragmoplast corrects abnormal cytokinesis in wheat x rye hybrid PMC. Tsitologiya (Russ.) 54: 297301. Shamina NV, Golubovskaya IN, Gruzdev AD. 1981. Spindle disturbances in meiotic mutants in maize. Tsitologiya (Russ.) 23: 275-282. Shamina NV, IA Zaporozhchenko, Yu R Maksiutova, ОА Shatskaya. 2007c. Cortical cytoskeletal ring at prophase II leads to correction of abnormalities of the first meiotic division and to meiotic restitution. Tsitologiya (Russ.) 49: 865-869 Shamina NV, Mukhina Zh M, Gordeeva EI, Orlov PS. 2007b. Bouquet arrest and its consequences for asynaptic meiosis in cereals – haploids and wide hybrids F1. Tsytologiya (Russ.) 49: 690-695. Shamina N.V., Silkova O.G., Seriukova E.G. 2003b. Monopolar spindles in meiosis of intergeneric cereal hybrids. Cell Biol Internatl 27: 657-664. Shamina N.V., Gordeeva E.I., Kovaleva N.M., Seriukova E.G., Dorogova N.V. 2007a. Formation and function of phragmoplast during successive cytokinesis stages in higher plant meiosis. Cell Biol Intern 31: 626-635. Smirnova EA, Bajer AS. 1994. Microtubule converging centers and the reorganization of interphase cytoskeleton and the mitotic spindle in higher plant Haemanthus. Cell Motil Cytoskel 27: 219-233. Smirnova ES, Bajer AS. 1998. Early stages of spindle formation and independence of chromosome and microtubule cycles in Haemanthus endosperm. Cell Motil Cytoskeleton 40: 22-37.

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2n Pollen Formation: 40 Cytological Mechanisms of Nuclear Meiotic Restitution

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Smirnova EA, Svetlitskaya OM, Chentsov YS. 2002. Disruption of mitotic microrubule organization in root meristem cells of Allium cepa induced by chloralhydrate. Tsitologiya (Russ.) 44: 120-130. Sollner R, Glasser G, Wanner G, Somerville CR, Jurgens G, Assaad FF. 2002. Cytokinesisdefective mutants of Arabidopsis. Plant Physiol 129: 678-690. Song K, Mach KE, Chen CY, Reynolds T, Albright CF. 1996. A novel suppressor of ras1 in fission yeast, byr4, is a dosage-dependent inhibitor of cytokinesis. J Cell Biol; 133:1307-19. Spielman M, Preuss D, Feng-Lan Li, Brown WE, Scott RJ. 1997. TETRASPORE is required for male meiotic cytokinesis in Arabidopsis tthaliana. Development 124: 2645-2657. Staiger CH, Cande WZ. 1990. Microtubule distribution in dv, a maize meiotic mutant defective in the prophase to metaphase transition. Developmental Biology 138: 231242. Staiger CH, Cande WZ. 1991. Microfilament distribution in maize meiotic mutants correlates with microtubule organization. Plant Cell 3: 637-644. Staiger CH, Cande WZ. 1993. Cytoskeletal analysis of maize meiotic mutants. In: Molecular and Cell Biology of the Plant Cell Cycle. JC Omrod and D. Francis (eds), Kluver Academic Press, pp. 157-171. Stefani A. 1986. Unreduced gametes in the Fl hybrid of Triticum durum Desf. x Haynaldia villosa Schur. Z. Pflanzenzuchtg 96: 8—14. Stoppin V, Vantard M, Schmit AM, Lambert AM. 1994. Isolated plant nuclei nucleate microtubule assembly: the nuclear surface in higher plants has centrosome-like activity. Plant Cell 6: 1099-1106. Strompen G, El Kasmi F, Richter S, Lukowitz W, Assaad FF, Jurgens G, Mayer U. 2002. The Arabidopsis HINKEL gene encodes a kinesin-related protein involved in cytokinesis and is expressed in a cell-cycle dependent manner. Curr Biol 12: 153-158. Sunkel CE, Glover DM. 1988. Polo, a mitotic mutant of Drosophila displaying abnormal spindle poles. J Cell Sci 89: 25-38. Surana U, Amon A, Dowzer C, McGrew J, Byers B, Nasmyth K. 1993. Destruction of the CDC28/CLB mitotic kinase is not required for the metaphase to anaphase transition in budding yeast. EMBO J 12: 1969–1978. Suzuki T, Tanaka I. 1999. Attachment of kinetochores to spindle microtubules during meiosis 1 of Lilium microsporocytes. Chromosome Res 7: 121-129. Tanaka M. 1959. Newly synthesized amphidiploid from the hybrids, Emmer wheat x Aegilops squarrosa varieties. Wheat Inf Serv 8: 8. Taschetto OM, Pagliarini MS, 2003. Occurrence of 2n and jumbo pollen in the Brasilian ginseng (Pfaffia glomerata and P. tuberosa). Euphytica 133: 139 – 145. Tavoletti S. 1994. Cytological mechanisms of 2n eggs formation in a diploid genotype of Medicago sativa sbbsp falcata. Euphytica 75: 1-8. Tiezzi A, Bednara J, Del Casino C, Bartalesi A, Cai G, Moscatelli A. 1992. The microtubular cytoskeleton during pollen development and pollen tube growth in Nicotiana tabacum. In: Angiosperm pollen and ovules. New York: Spronger – Verlag, pp 224-232. Tiwari SC, Wick SM, Williamson RE, Gunning BES. 1984. Cytoskeleton and integration of cellular function in cells of higher plants. J Cell Biol 99: 63S-69S.

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Traas JA, Doonan JH, Rawlins DJ, Shaw PJ, Watts J, Lloyd CW. 1987. An actin network is present in cytoplasm throughout the cell cycle of carrot cells and associates with the dividing nucleus. J Cell Biol 105: 387-395. Udall JA, Wendel JF. 2006. Polyploidy and crop improvement. The Plant Genome 1: S3– S14. Utsunomiya KS, Pagliarini MS and do Valle CB (2004). Chromosome transfer among meiocytes in Brachiaria nigropedata (Ficalho & Hiern) Stapf (Gramineae). Cytologia 69: 395-398. VanDamme D, Inze D, Russinova E. 2008. Vesicle trafficking during somatic cytokinesis. Plant Physiol 147: 1544-1552 Van Lammeren AAM, Keijzer CJ, Willemse MTM, Kieft H. 1985. Structure and function of the microtubular cytoskeleton during pollen development in Gasteria verrucosa (Mill.) H. Duval. Planta 165: 1-11. Vantard M, Levilliers N, Hill AM, Adoutte A, Lambert AM. 1990. Incorporation of Paramecium tubulin into higher plant cells reveals functional sites of microtubule assembly. Proc Natl Acad Sci USA 87: 8825-8829. Veilleux RE, McHale NA, Lauer FI. 1982. 2n gametes in diploid Solanum: frequency and types of spindle abnormalities. Can J Genet Cytol 24: 301-314. Vorsa N, Ortiz R. 1992. Cytology of 2n pollen formation in a blueberry aneuploid (2n= 4x+9=57). J Hered 83: 346-349. Vos JW, Pieuchot L, Evrard J-L, Janski N, Bergdoll M,et al. 2008. The plant TPX2 protein regulates prospindle assembly before nuclear envelope breakdown. Plant Cell 20: 2783–2797 Wada B, Kusunoki F. 1964. Spindle membrane in meiosis of pollen mother cells of Tradescantia and in mitosis of endosperm cells of Zephyranthes. Cytologia (Tokyo) 29: 109-111. Wagenaar EB. 1968. Meiotic restitution and the origin of polyploidy. II. Prolonged duration of metaphase I as causal factor of restitution induction. Can J Genet Cytol 10: 844-852. Walker KL, Müller S, Moss D, Ehrhardt D W, Smith LG. 2007. Arabidopsis tangled identifies the division plane throughout mitosis and cytokinesis. Curr Biol 17: 18271836 Wang H, Cutler AJ, Fowke LC. 1991. Microtubule organization in cultured soybean and black spruce cells: interphase – mitosis transition and spindle morphology. Protoplasma 162: 46-54. Wang RJ, Wissinger W, King EJ, Wang G. 1983. Studies on cell division in mammalian cells. VII. A temperature sensitive cell line abnormal in centriole separation and chromosome movement. J Cell Biol 96: 301-306. Wasteneys GO, Yang Z. 2004. New views on the plant cytoskeleton. Plant Physiol. 136: 3884–3891 Watanabe K, Peloquin SJ. 1993. Cytological basis of 2n pollen formation in a wide range of 2x, 4x, and 6x taxa from tuber-bearing Solanum species. Genome 36: 8-13. Weingartner M, Criqui MC, Mészáros T. et al. 2004. Expression of a nondegradable cyclin B1 affects plant development and leads to endomitosis by inhibiting the formation of a phragmoplast. Plant Cell 16: 643-657.

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Werner JE, Peloquin SJ. 1991. Occurrence and mechanisms of 2n egg formation in 2x potato. Genome 34: 975-982. Wheatley SP, Hinchcliffe EH, Glotzer M, Hyman AA, Sluder G, Yu-li Wang. 1997. CDK1 inactivation regulates anaphase spindle dynamics and cytokinesis in vivo. J Cell Biol 138: 385-393. Wick SM, Duniec J. 1984. Immunofluorescence microscopy of tubulin and microtubule arrays in plant cells. II. Transition between the pre-prophase band and the mitotic spindle. Protoplasma 122: 45-55. Xu SJ, Dong Y. 1992. Fertility and meiotic mechanisms of hybrids between chromosome autoduplication tetraploid wheats and Aegilops species. Genome 35:379–384. Xu SJ, Joppa LR. 1995. Mechanisms and inheritance of first division restitution in hybrids of wheat, rye, and Aegilops squarrosa. Genome 38: 607-615. Yamano H, Gannon J, and Hunt T. 1996. The role of proteolysis in cell cycle progression in Schizosaccharomyces pombe. EMBO J 15: 5268–5279. Yang HY, Mains PE, McNally FJ. 2005. Kinesin-1 mediates translocation of the meiotic spindle to the oocyte cortex through KCA-1, a novel cargo adapter. J. Cell Biol. 169:447–457. Yang HY, McNally K, McNally FJ. 2003. MEI-1/katanin is required for translocation of the meiosis I spindle to the oocyte cortex in C. elegans. Dev. Biol. 260:245–259 Yu Hong-Shi, Russell SD. 1993. Three-dimentional ultrastructure of generative cell mitosis in the pollen tube of Nicotiana tabacum. Eur J Cell Biol 61: 338-348. Zee SY, Ye XL. 2000. Microtubule reorganization during pollen development of rice (Oryza sativa L.). Protoplasma 210: 188-201.

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Chapter 6

ORBICULES IN RELATION TO THE POLLINATION MODES B. G. Galati1, M. M Gotelli1,3, S. Rosenfeldt2, J. P. Torretta1,3 and G. Zarlavsky1 1

Cátedra de Botánica. Facultad de Agronomía. Universidad de Buenos Aires. Argentina 2 DBBE. Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires, Argentina 3 Conicet

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ABSTRACT Orbicules or Ubisch bodies are corpuscles of sporopollenin that appear in the anther locule during pollen grain development. Their size ranges from 0.14 µm to 20 µm. They present different shapes with a smooth or ornamented surface. Orbicules often form aggregates and sometimes have a plaque-like appearance. Ultrastructurally, they may present a central core with different degree of transparency to electrons. Those that do not have a central core are observed completely solid. Orbicules are resistant to acetolisis, autofluorescent when irradiated with ultraviolet light and have the same reaction to colorants that the exine of pollen grains. Their presence is generally associated with a tapetal membrane in species with secretor type tapetum and with a peritapetal membrane in species with intermediate or plasmodial type tapetum. Although the shed of orbicules out of the anther along with the pollen grains is cited, they are usually attached to the inner surface of the locule when the anther opens. Investigations suggest that orbicules appear in approximately 80 families of Angiosperms and Gimnosperms. It is not certain whether orbicules are not developed in the rests of the families or are just not informed. Researches on ontogeny and ultrastructure of orbicules are rare. However, their tapetal origin and their simultaneous formation with the pollen grain wall are well established. The systematic value of orbicules is known and considered in a few families, such as Loganiaceae, Gentianaceae, Apocynaceae, Rubiaceae and Oxalidaceae. Evolutionary studies on these bodies or on its relationship with the different modes of pollination are lacking. Even though orbicules are so common among angiosperms, their function is unknown and only speculations are made. On this report a review on orbicules is made and an analysis of their presence,

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B. G. Galati, M. M Gotelli, S. Rosenfeldt et al… ontogeny and morphology is presented. Our aim is to supply information that will help understand orbicules function. Therefore, the orbicules morphology in relation with the pollination mode is studied.

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INTRODUCTION During the second halve of the XIX century and the beginnings of the XX, small granular bodies in the anther locule of species of Solanaceae, Boraginaceae and Liliaceae, draw the attention of scientists (Rosanoff, 1865; Mascré, 1922; Schnarf, 1923). These are the first references of the presence of orbicules in Angiospermae. Years later, Ubisch (1927) observed particles that resembled perforated plaques on the inner tangential wall of the tapetal cells of Oxalis rosea. This author found surprising to be the first to describe such striking bodies. On the other hand, she makes reference to the similar response to colorants of these bodies and the exine of the pollen grains. Since Ubisch (op cit.) was the first to give a detailed description of these corpuscles, in a first instance, they were named after her as Ubisch bodies (Rowley, 1962). Simultaneously, Kosmath (1927) investigated the presence of these bodies in 69 taxa of 35 families. Heslop-Harrison (1968a) denominated these corpuscles as orbicules. According to Madjd & Roland-Heydacker (1978) and Abadie & Hideux (1979) there are differences between Ubisch bodies and orbicules. These authors define Ubisch bodies as hollow bodies with a surface comparable to the exine of pollen grains and orbicules as spherical structures that can be deformed. Nowadays, ―Ubisch bodies‖ and ―orbicules‖ are used as synonyms. Orbicules can be described as small bodies which size varies between 0.14 and 20 µm. They present the same reaction to colorants, auto-fluorescence and resistance to acetolysis than the exine of pollen grains. Huysmans et al. (1998) studied orbicules morphology, function, distribution and relation to the different types of tapeta in Angiosperms. According to these authors, orbicules are present in 314 species of 72 families of Angiosperms. Studies in a large number of families should be made in order to confirm the presence or absence of orbicules. Since these bodies may be extremely small, it is probable that in some families considered to lack orbicules, these were not seen. The taxonomic importance of orbicules was considered for different taxa, between them, Euphorbiaceae (El-Ghazaly & Chaudhary, 1993), Rubiaceae (Huysmans et al., 1997; Vinckier et al., 2000), Gentianales (Vinckier & Smets, 2002c, 2003), Loganiaceae (Vinckier & Smets, 2002a), Apocynaceae (Vinckier & Smets, 2002b) and Oxalidaceae (Rosenfeldt & Galati, 2008). According to these studies, orbicule morphology can be used to confirm, justify or reject taxonomic conclusions. Most authors consider that orbicules are exclusive of secretor type tapeta (Pacini et al., 1985; Huysmans et al., 1998) and that never appear on ameboid or plasmodial tapeta (Vinckier et al., 2000). However, further studies confirmed their presence in species of Asteraceae (Gotelli et al., 2008) and Malvaceae (Strittmatter et al., 2000; Galati et al., 2007) with plasmodial or invasive non-syncytial tapeta. Therefore, the presence of orbicules seems to be a fact much more generalized in Angiosperms as previously believed. Orbicules are disposed coating the interior of the anther locule. They may be in direct contact with the inner tangential face of the endothecium cells or on a special membrane, also

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resistant to acetolysis. This is a tapetal type membrane in the secretory tapeta, that is formed from the inner tangential face of these cells (Bhandari & Kishori, 1973; Lovisolo & Galati, 1999; Galati & Strittmatter, 1999a, b; Strittmatter & Galati, 2001), and peritapetal in the plasmodial (Strittmatter et al., 2000) and invasive non-syncytial tapeta (; Galati et al., 2007), since it develops from the outer tangential face of these cells. Several authors studied the ultrastructure of the tapetal membrane of Poaceae (Banerjee, 1967; Christensen et al., 1972; El-Ghazaly & Jensen, 1987). This membrane is composed by two layers or strata and is observed fenestrate. Different origins were adjudicated to orbicules. On the beginning of the XX century, Krjatchenko (1925) considered the mitochondria of tapetal cells to be the centre of origin of orbicules. This idea was supported by Heslop-Harrison (1962). Numerous researches on the ontogeny of these bodies in different Angiosperm species confirmed their tapetal origin and its simultaneous formation with the pollen grain wall. The increased development of the endoplasmic reticulum of rough type coincides with the appearing of the pro-orbicules. Therefore, the participation of this organelle in the synthesis of orbicules may be confirmed (Galati & Strittmatter, 1999a; Strittmatter et al., 2000; Amela et al., 2002; Rosenfeldt & Galati, 2005; Galati et al., 2007). Orbicule morphology is varied and can only be interpreted using transmission and scanning electron microscope techniques together. Unfortunately, there are very few papers where both techniques are applied, and, therefore, the morphology may not be completely understood. An approach to orbicules classification was made by Galati (2003). Externally, they are spherical to sub-spherical o piriform. On the first case, they may present a smooth surface, invaginations or spicules. Orbicules of some species have perforations, and when it is a single perforation, they have the aspect of a doughnut or a croissant. Sometimes orbicules group into two or more units forming aggregates that present a vermiform or plaque aspect. Internally they can be observed as completely solid masses or with a central core, more or less transparent to electrons at maturity. The external wall resembles the exine of the pollen grain in coloration and electron-density. According to Clement & Audran (1993a, b) the orbicular core is surrounded by a cytomembrane supporting a glycocalyx on which the sporopollenin orbicular wall is built bordered by a peripheral sheet. Many speculations were made trying to understand the role of orbicules. However, none of them could be confirmed. One of the first hypotheses was that orbicules contributed with the development of the exine (Maheshwari, 1950). Orbicules would represent a mechanism to transport sporopollenin from the tapetum towards the developing microspores. Studies on Poaceae showed strands of sporopollenin that could be indicating a possible passage or transference of substrates from tapetal cells towards the exine (El-Ghazaly & Jensen, 1986; Banerjee & Barghoorn, 1971). However, this theory was discarded since it is now known that orbicules and pollen grain exine are formed simultaneously (Christensen et al., 1972; ElGhazaly & Jensen, 1986, 1987; Galati & Strittmatter, 1999a; Amela et al., 2002; Strittmatter et al., 2000; Galati et al., 2007). A second hypothesis was proposed by Christensen et al. (1972) who mentioned that the orbicular wall could be considered as a vestigial capacity of the tapetum. According to Hesse (1986) there would be an homology between the tapetum and the sporogenous tissue. A third hypothesis was suggested by Rowley & Erdtman (1967) who thought orbicules could play an important role in the lisis and degeneration of tapetal cells. Herich & Lux

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(1985) observed that the central core of Ubisch bodies in Lilium henryi is connected to the plasma membrane of tapetal cells through a channel. Therefore, these authors support the litic activity theory. El-Ghazaly & Jensen (1986) and El-Ghazaly & Nilsson (1991) suggest that the function of orbicules is to retain in their surface the excess of sporopollenin that could interfere with the normal development of the microspores. Heslop-Harrison (1968a, b) was the first to consider that the orbicules could be associated to pollen dispersal forming a not-wettable surface from which pollen can easily be freed. Later, Keijzer (1984) supported this theory. Pacini & Franchi (1993) wondered if pollen grains and orbicules would repulse each other, since they are formed by the same substance and, therefore, charged in the same way. Vaknin et al. (1999) analyzed the role of electrostatic forces. Although the hypothesis of orbicules implicated on pollen dispersal was announced more than four decades ago, the relation between orbicule morphology and pollination syndromes was not considered. This line of investigation could confirm this last hypothesis. In this research, orbicule morphology of representative species of diverse families of Angiosperms with different modes of pollen dispersal (anemophily, melittophily, ornithophily and psicophily) is studied. Observations are made with transmission and scanning electron microscope, and are compared with previous knowledge.

MATERIALS AND METHODS

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Species Studied Fourteen species belonging to eight families of Angiosperms with different modes of pollination, were analyzed (Table I).

Methods For transmission electron microscopy (TEM) studies, the anthers were pre-fixed in 1 % glutaraldehyde in phosphate buffer (pH 7.2) overnight and then post-fixed in 1 % OsO4 at 2ºC in the same buffer for 3 h. Following dehydration in acetone series, the material was embedded in Spurr‘s resin. Ultrathin sections (750 to 900 nm) were made on a Reichert-Jung ultramicrotome and then stained with uranyl acetate and lead citrate (O‘Brien & McCully, 1981). The sections were observed and photographed with a Philips EM 301 at 60.0 kV and a JEOL 100c TEM at 80.0 kV. For scanning electron microscopy (SEM), anthers were fixed in FAA (formalin-aceticalcohol mixture) for 48 h and stored in 70% ethanol. Then, they were dehydrated in an ascendant ethanol series and air dried. The material was then sputter-coated with gold and examined using a Philips XL 30 and a Zeiss supra 40 microscope.

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Table I. Species studied.

Species Juglans australis Gris. Plantago tomentosa Lam. Piper nigrum L. Eleusine tristachya (Lam.) Lam. Briza subaristata Lam. Distichlis spicata (L.) Greene Solanum granulosum-leprosum Dun. Pyrostegia venusta (Ker Gawl.) Miers Ceiba insignis (Kunth) Gibbs & Semir. Sida rombifolia L. Modiolastrum malvifolium (Griseb.) K. Schum. Jacaranda mimosifolia D. Don. Eryngium coronatum Hook. & Arn.

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Eryngium pandanifolium Cham. & Schtdl.

Family

Order

Pollination Mode

Juglandaceae

Fagales

anemophily (Culley et al., 2002)

Plantaginaceae

Lamiales

Piperaceae

Piperales

Poaceae

Poales

Poaceae

Poales

Poaceae

Poales

Solanaceae

Solanales

buzz-pollination (Endress, 1998)

Bignoniaceae

Lamiales

ornithophily (Galetto et al., 1994; Rivera, 2000)

Malvaceae

Malvales

psichophily (Gibbs & Semir, 2003)

Malvaceae

Malvales

melittophily (according to floral morphology)

Malvaceae

Malvales

melittophily (according to floral morphology)

Bignoniaceae

Lamiales

melittophily (Rivera, 2000)

Apiaceae

Apiales

generalist (Molano-Flores, 2001)

Apiaceae

Apiales

anemophily (Proctor et al., 1996; Nitiu, 2006) anemophily (according to floral morphology) anemophily (Proctor et al., 1996; Culley et al., 2002) anemophily (Proctor et al., 1996; Culley et al., 2002) anemophily (Proctor et al,. 1996; Culley et al., 2002)

generalist (Molano-Flores, 2001)

RESULTS Orbicule and Pollen Grain Exine Morphology a) Anemophilous species

Juglans Australis Gris Spherical to sub-spherical orbicules, with a media diameter of 0.9 µm, and a central core transparent to electrons placed towards the base of the orbicule, in direct contact with the anther wall. The orbicule wall presents superficial spicules (Fig. 1 A-B). Pollen grain exine microechinate (Fig. 3 A).

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Figure 1. Orbicules morphology. A-B. J. australis. A. SEM. Scale bar: 2 µm. B. TEM. Scale bar: 0.5 µm. C-D. P. tomentosa. C. SEM. Scale bar: 1 µm. D. TEM. Scale bar: 0.5 µm. E-F. P. nigrum. E. SEM. Scale bar: 0.2 µm. F. TEM. Scale bar: 0.1 µm.

Figure 3. Pollen exine morphology. SEM. A. J. australis. Scale bar: 10 µm. B. P. tomentosa. Scale bar: 2 µm. C. P. nigrum. Scale bar: 0.3 µm. D. S. granulosum-leprosum. Scale bar: 1 µm. Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

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Plantago Tomentosa Lam Orbicules with a plaque aspect, with a media diameter of 0.7 µm, and one to several central cores. The upper surface, which is in direct contact with the anther locule, is speculate (Fig. 1 C-D). Pollen grain exine verrucate, microechinate (Fig. 3 B). Piper Nigrum L Orbicules almost spherical, with a media diameter of 0.2 µm, a central core with moderate electron-density, and an orbicular wall covered by spicules with a wide base (Fig. 1 E-F). Pollen grain exine verrucate, microechinate (Fig. 3 C).

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Eleusine Tristachya (Lam.) Lam Spherical to sub-spherical orbicules, with a media diameter of 0.6 µm. Most of them present the face that contacts the anther wall straight. They show a central core transparent to electrons and an orbicular wall with micro-channels and superficial spicules (Fig. 2 A-B). Pollen grain exine microechinate.

Figure 2. Orbicules morphology. A-B. E. tristachya. A. SEM. Scale bar: 2 µm. B. TEM. Scale bar: 2 µm. C. B. subaristata. SEM. Scale bar: 2 µm. D. D. spicata. SEM. Scale bar: 2 µm.

Briza Subaristata Lam Spherical orbicules, with a media diameter of 0.9 µm and a central core that is transparent to electrons. The orbicular wall has micro-channels and superficial spicules (Fig. 2 C). Pollen grain exine microechinate.

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Distichlis Spicata (L.) Greene Spherical orbicules, with a media diameter of 0.6 µm and a central core that is transparent to electrons. The orbicular wall has micro-channels and superficial spicules (Fig. 2 D). Pollen grain exine microechinate. b) Buzz-pollination species

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Solanum Granulosum-Leprosum Dun Spherical orbicules, with a media diameter of 0.7 µm, solid, without a central core and with irregularly disposed superficial spicules of variable length. Orbicules are found immersed in a continuous orbicular wall, formed by the union of micro-orbicules (Fig. 4 AB). Pollen grain exine microechinate (Fig. 3 D).

Figure 4. Orbicules morphology. A-B. S. granulosum. A. SEM. Scale bar: 1 µm. B. TEM. Scale bar: 1 µm. C-D. P. venusta. C. SEM. Scale bar: 1 µm. D. TEM. Scale bar: 1.5 µm. E-F. C. insignis. E. SEM. Scale bar: 2 µm. F. TEM. Scale bar: 1 µm.

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c) Ornitophilous species

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Pyrostegia Venusta (Ker Gawl.) Miers Orbicules resembling croissants, with a media diameter of 0.9 µm, a central core transparent to electrons, and a conspicuous perforation that goes through its middle part. The orbicular wall presents a smooth surface (Fig. 4 C-D). Pollen grain exine foveolate in the mesocolpium, pertectatum in the apocolpium (Fig. 6 A-B).

Figure 6. Pollen exine morphology. SEM. A-B. P. venusta. A. Pollen grain in polar view. Scale bar: 10 µm. B. Detail of the exine. Scale bar: 1 µm. C. C. insignis. Scale bar: 3.5 µm. D. S. rombifolia. Scale bar: 20 µm. E. J. mimosifolia. Scale bar: 10 µm. F. E. pandanifolium. Scale bar: 12 µm.

d) Psicophilous species

Ceiba Insignis (Kunth) Gibbs & Semir Spherical orbicules, with a media diameter of 1 µm, a central core of moderate electrondensity, and an orbicular wall, that has invaginations. Some orbicules present a central perforation (Fig. 4 E-F). Pollen grain exine reticulate (Fig. 6 C). e) Melittophilous species

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Sida Rombifolia L

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Modiolastrum Malvifolium (Griseb.) K. Schum Piriform orbicules, with a smooth surface and a media longitud of 3.5 µm. They are solid, without a central core (Fig. 5 A-B). Pollen grain exine echinate (Fig. 6 D).

Figure 5. Orbicules morphology. A. S. rombifolia. SEM. Scale bar: 5 µm. B. M. malvifolium. TEM. Scale bar: 0.5 µm. C-D. J. mimosifolia. C. SEM. Scale bar: 2 µm. D. TEM. Scale bar: 0.5 µm. E. E. coronatum. Scale bar: 1 µm. SEM. F. E. pandanifolium. TEM. Scale bar: 0.5 µm.

Jacaranda Mimosifolia D. Don Spherical orbicules, with a smooth surface and a media diameter of 0.7 µm. They present a central core transparent to electrons (Fig. 5 C-D). Pollen grain exine psilate (Fig. 6 E). f)

Generalist species

Eryngium Coronatum Hook. Et Arn Spherical orbicules with a smooth surface, a central perforation, a media diameter of 0.6 µm and a central core of moderate electron-density (Fig. 5 E). Pollen grain exine psilate. Eryngium Pandanifolium Cham. & Schtdl Spherical orbicules with a smooth surface, a media diameter of 0.5 µm and a central core of moderate electron-density (Fig. 5 F). Pollen grain exine psilate (Fig. 6 F).

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CONCLUSION Juglans australis, Plantago tomentosa and Piper nigrum are anemophilous species (Culley et al, 2002; Nitiu, 2006; De Figueiredo & Sazima, 2000). Although they belong to distant angiosperm families, they present spiculate orbicules. They have one or more central cores, they are spherical or in plaque form, but their surfaces are always microespiculate. Species of the Poaceae family are anemophilous and present the same orbicule morphology. Orbicules have a central core transparent to electrons at maturity and an orbicular wall trespassed by micro-channels, with superficial spicules (Christensen et al., 1972; El-Ghazaly & Jensen, 1987; Lovisolo & Galati, 1999). Several Gymnosperms, Ephedra americana, Welwitschia mirabilis (Hesse, 1984); Pinus sylvestris (Rowley & Walles, 1987); Juniperus chinensis, Thuja orientalis, Cunnighamia konishii (Chen, 1988) and Ephedra americana (Doores et al., 2007) present orbicules with spiculate surface. In this research, Solanum granulosum-leprosum was studied as an example of buzzpollination. The orbicular surface is very dense in the interior of the locule. Many small orbicules are connected to bigger ones, constituting a continuous orbicular membrane. Bigger orbicules present small spicules on their surface. Exacum oldenlandioides (Gentianaceae) is pollinated in the same way (Endress, 1998; Proctor et al., 1996; Buchmann, 1983) and has spiculate orbicules (Vinckier & Smets, 2002c). According to their floral morphology, Sida rombifolia and Modiolastrum malvifolium are melittophylous. In these species, orbicules may be spherical, sub-spherical or piriform, but they always have a smooth surface. Rauvolfia mattfeldiana, Allamanda cathartica (Apocynaceae); Geniostoma borbonicum, G. rupestre (Geniostomaceae); Mandevilla atroviolaceae (Apocynaceae) and Jacaranda mimosifolia (Bignoniaceae) are all examples of species that present the same pollination mode (Humeau et al., 2003; Castro & Robertson, 1997; Ehrenfeld, 1979; Löhne et al., 2004; Torres & Galeto, 1998; Rivera 2000) and orbicule morphology (Vinckier & Smets, 2002a, b; El-Ghazaly & Chaudhary, 1993; Hess, 1986; Huysmans et al., 1997; Galati & Strittmatter, 1999a, b). Pyrostegia venusta is an ornithophilous species (Galleto et al., 1994; Rivera 2000). Its pollen is dispersed by hummingbirds. Orbicules have a conspicuous eccentric perforation that gives them the aspect of croissants. Its surface is completely smooth. There are species with a similar orbicular morphology whose pollination syndromes are unknown, as Logania nuda (Loganiaceae) (Vinckier & Smets, 2002a). Ceiba insignis is a psichophilous species, visited by nocturnal lepidopterous (Gibbs & Semir, 2003). Its orbicules are characterized by an irregular surface with invaginations, which are perfectly observed with MET. Lilium sp. presents the same pollination syndrome and the Ubisch bodies have an orbicular wall with invaginations, which gives them an undulated aspect in section (Clément & Audran, 1993a, b). Alstonia congensis (Apocynaceae), Tabernaemontana coronaria (Apocynaceae), Acokanthera oblongifolia (Apocynaceae), Vinca rosea (Apocynaceae), Fagraea racemosa (Gentianaceae), Acokanthera oblongifolia (Apocynaceae) y Beaumontia grandiflora (Apocynaceae) are examples of probable psychophilous species whose orbicules present an irregular surface, not smooth, and may also have one or more perforations (Vinckier & Smets, 2002a, b, c; Cousin, 1979).

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Eryngium pandanifolium and E. coronatum have orbicules with a smooth surface. Some of these orbicules present a perforation. Several characteristics of these Ubisch bodies are coincident with that of melittophilous, ornithophilous and psychophilous species. This could be in relation with the fact that these species are generalist (Molano-Flores 2001). The exine of the pollen grain in anemophilous species is predominantly microechinate. This coincides basically with the morphology of the orbicules. Species with other pollination syndromes possess variable exine morphology. We can conclude that families, which may be taxonomically distant but share the mode of pollen dispersal present similar orbicule morphology. According to our observations and the data available in other published works, we could consider that spiculate orbicules, with or without a central core, are characteristic of anemophilous species. The only studied species with buzz-pollination also presents Ubisch bodies with superficial spicules. Since pollen repulsion in both pollination modes is similar, the same orbicule morphology was expected. Unfortunately, the orbicule morphology and effective pollinator are known in a few species. Therefore, we can only make some speculations. More studies are needed to confirm this theory. Apparently, orbicules with a smooth surface and without perforations would be characteristic of melitophilous species. Orbicules with perforations and/or an irregular surface seem to be common among species pollinated by hummingbirds or long- tongued insects. The coincidences observed between orbicule morphology and pollination syndrome allow us to adjudicate a possible function to orbicules. We support the theory proposed by Heslop-Harrison (1968a, b), who suggested that orbicules could be related to pollen dispersal by a non-wetable surface from which pollen can be easily freed. According to Pacini & Franchi (1993), since orbicules and pollen exine are constituted by sporopollenin, they repeal each other. This would allow the expulsion of pollen grains. We can now add that the different orbicule morphology would facilitate each pollination mode. This would also be related to exine morphology. According to Hess (2000), several studies show possible correlations between pollen structure and pollen vectors, while others report the absence of such association. Here, we can only establish a relation between pollen and orbicule morphology in anemophilous species. Exine morphology of species with other pollination syndromes is very variable. Thus, this is an open road to continue this research.

REFERENCES Abadie, M. & Hideux, M (1979). L’anthère de Saxifraga cymbalaria L. ssp. huetiana (Boiss.) Engl. et Iemsch en microscopie électronique (M.E.B. et M.E.T). 1. Généralités. Ontogenèse des orbicules. Ann. Sci. Nat. Bot., 13 (1), 199-233. Amela García, M. T.; Galati, B. G. &. Anton, A. M. (2002). Microsporogenesis, microgametogenesis and pollen morphology of Passiflora spp. (Passifloraceae). Botanical Journal of the Linnean Society, 139, 383-394. Banerjee, U.C. (1967). Ultrastructure of the tapetal membranes in grasses. Grana Palynologica, 7, 2-3.

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Orbicules in Relation to the Pollination Modes

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Banerjee, U. C. & Barghoorn, E. S. (1971). The tapetal membranes in grasses and Ubisch body control of mature exine pattern. In: J. Heslop-Harrison (ed.) Pollen development and physiology. (1971, pp 1-338). Butterworths, London Bhandari, N. N. & Kishori, R. (1973). Development of tapetal membrane and Ubisch-granuls in Nigella damascene- A histochemical approach. Beitr. Biol. Pflanzen, 49, 59-72. Buchmann, S. L. (1983). Buzz pollination in Angiosperm. In: Jones, C. E., Little, R. J. (eds.), Handbook of experimental pollination biology. (1983, pp 1-558). Van Nostrand Reinhold, New York Castro, I. & Robertson, A. (1997). Honeyeaters and the New Zealand forest flora: the utilization and profitability of small flowers. New Zealand Journal of Ecology, 21 (2), 169-179. Chen, Su-Hwa. (1988). A scanning electron microscope survey of common airborne pollen grains in Tapel, Taiwan. Taiwania 33, 75-108. Christensen, J. E.; Horner, H. T. Jr. & Lersten, N. R. (1972). Pollen wall and tapetal orbicular wall development in Sorghum bicolor (Gramineae). Amer. J. Bot., 59 (1), 4358. Clément, C. & Audran, J. C. (1993a). Cytochemical and ultrastructural evolution of orbicules in Lilium. Pl. Syst. Evol. (Suppl.) 7, 63-74. (1993b). Orbicule wall surface characteristics in Lilium (Liliaceae). Grana, 32, 348-353. Cousin, M-T. (1979). Tapetum and pollen grains of Vinca rosea (Apocynaceae). Grana 18, 115-128. Culley, T. M.; Weller, S. G. & Sakai, A. K. (2002). The evolution of wind pollination in angiosperms. Trends in Ecology & Evolution, 17, 361-369. De Figueiredo, R. A. & Sazina, M. (2000). Pollination Biology of Piperaceae species in southeastern Brazil. Annals of Botany, 85, 455-460. Doores, A. S.; Osborn, J. M. & El-Ghazaly, G. (2007). Pollen ontogeny in Ephedra americana (Gnetales). Int. J. Plant Sci., 168 (7), 985-997. Ehrenfeld, J. G. (1979). Pollination of Three Species of Euphorbia Subgenus Chamaesyce, with Special Reference to Bees. American Midland Naturalist, 101 (1), 87-98 El-Ghazaly, G. & Jensen, W. A. (1986). Studies of the development of wheat (Triticum aestivum) pollen. I. Formation of the pollen wall and Ubisch bodies. Grana, 25, 1-29. 1987. Development of wheat (Triticum aestivum) pollen. II. Histochemical differentiation of wall and Ubisch bodies during development. Amer. J. Bot., 74 (9), 1396-1418. El-Ghazaly, G. & Nilsson, S. (1991). Development of tapetum and orbicules of Catharanthus roseus (Apocynaceae). In: S. Blackmore & S. H. Barnes (eds.) Pollen and spores. Systematics Association Special, 44. Clarendon Press, Oxford. El-Ghazaly, G. & Chaudhary, R. (1993). Morphology and taxonomic application of orbicules (Ubisch bodies) in the genus Euphorbia. Grana suppl., 2, 26-32. Endress, P. K. (1998). Diversity and evolutionary biology of tropical flowers. Cambridge University, Cambridge, England. Galati, B. G. (2003). Ubisch bodies in Angiosperms. In: A.K. Pandey & M.R. Dhakal (eds.) Advances in Plant Reproductive Biology. (2003, pp.1-21). Narendra Publishing House, Delhi. Galati, B. G. & Strittmatter, L. I.. (1999a). Correlation between pollen development and Ubisch bodies ontogeny in Jacaranda mimosifolia (Bignoniaceae). Beitr. Biol. Pflanzen,71, 1-12.

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162

B. G. Galati, M. M Gotelli, S. Rosenfeldt et al…

(1999b). Microsporogenesis and microgametogenesis in Jacaranda mimosifolia (Bignoniaceae). Phytomorphology, 49(2), 147-156. Galati, B. G.; Monacci, F.; Gotelli, M. M. & Rosenfeldt, S. (2007). Pollen, Tapetum and Orbicule Development in Modiolastrum malvifolium (Malvaceae). Annals of Botany, 99(4), 755-763. Galetto, L., Bernardello, L. M. & Juliani, H. R. (1994). Characteristics of secretion of nectar in Pyrostegia ventusa (Ker-Gawl.) Miers (Bignoniaceae). New Phytologists, 127, 465471. Gibbs, P. & Semir, J. (2003). A taxonomic revision of the genus Ceiba Mill. (Bombacaceae). Annales Jardin Botanico de Madrid, 60 (2). Gotelli, M. M.; Galati, B. G. & Medan, D. (2008). Embryology of Helianthus annuus (Asteraceae). Ann. Bot. Fennici, 45(2), 81-96. Herich, R. & Lux, A. (1985). Lytic activity of Ubisch bodies (orbicles). Cytologia, 50, 563569. Heslop-Harrison, J. (1962). Origin of exine. Nature, 195, 1069-1071. (1968a). Pollen wall development. Science, 161, 230-237. (1968b). Tapetal origin of pollen-coat substances in Lilium. New Phytol., 67, 779-786. Hesse, M. (1984). Pollenkitt is lacking in Gnetatae: Ephedra and Welwitschia; further proof for its restriction to the Angiosperms. Pl. Syst. Evol., 144, 9-16. (1986). Orbicules and the ektexine are homologous sporopollenin concretions in Spermatophyta. Pl. Syst. Evol., 153, 37-48. (2000). Pollen wall stratification and pollination. Plant Syst. Evol., 222, 1-17. Humeau, L.; Strasberg, D. & Pailer, T. (2003). Dioécie cryptique chez Geniostoma borbonica, espèce pionnière endémique de La Réunion. Can. J. Bot., 81, 897-904. Huysmans, S.; El-Ghazaly, G.; Nilsson, S.& Smets, E. (1997). Systematic value of tapetal orbicules: A preliminary survey of the Cinchonoideae (Rubiaceae). Canad. J. Bot., 75, 815-826. Huysmans, S.; El-Ghazaly, G. & Smets, E. (1998). Orbicules in Angiosperms: Morphology, function, distribution, and relation with tapetum types. The Botanical Review, 64 (3), 240-272. Keijzer, C.J. (1984). Pollen dispersal and the function of the orbicules. Acta Bot. Neerl., 33, 244. Kosmath, L. von. (1927). Studien tiber das antherentapetum. Osterr. Bot. Z., 76, 235-241. Krjatchenko, D. (1925). De l’activité des chondriosomes pendant le développement des grains de pollen et des cellules nourricières du pollen dans Lilium croceum chaix. Rev. Gén. Bot., 37, 193-211. Löhne, C.; MacHado, I. C.; Porembski, S.; Erbar, C. & Leins, P. (2004). Pollination biology of a Mandevilla species (Apocynaceae), characteristic of NE-Brazilian inselberg vegetation. Botanische Jahrbücher, 125 (2), 229-243(15) Lovisolo, M. & Galati, B.G. (1999). Pollen development in Deyeuxia viridiflavescens (Poir) Kunth (Gramineae: Poaceae). Phytomorphology, 49(4), 411-418 Madjd, A. & Roland-Heydacker, F. (1978). Sécrétions et dégénérescence des celles du tapis dans l’anthère du Soja hispida Moench, Papilionaceae. Grana, 17, 167-174. Maheshwari, P. (1950). Introduction to the embryology of angiosperms. McGraw-Hill, New York. Mascré, M. (1922). Sur l’étamine des Vorágines. C.R. Acad. Sci. Paris, 175, 987-988.

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Orbicules in Relation to the Pollination Modes

163

Molano-Flores, B. (2001). Reproductive biology Eryngium yuccifolium (Apiaceae), a prairie species. Journal of the Torrey Botanical Society, 128 (1), 1-6. Nitiu, D. S. (2006). Aeropalynologic analysis of La Plata City (Argentina) during a 3-year period. Aerobiologia, 22, 79-87. O'Brien, T. P., McCully, M. E. (1981). The study of plant structure. Principles and selected methods. Termarcarphi Pty. Ltd.: Melbourne, Australia. Pacini, E. & Franchi, G. G. (1993). Role of the tapetum in pollen and spore dispersal. Pl. Syst. Evol., Suppl. 7, 1-11. Pacini, E.; Franchi, G. G. & Hesse, M. (1985). The tapetum: its form, function, and possible phylogeny in Embryophyta. Plant Syst. Evol., 149, 155-185. Proctor, M.; Yeo, P. & Lack, A. (1996). The Natural History of Pollination. Timber Press, Portland, OR. Rivera, G. L. (2000). Nuptial nectary structure of Bignoniaceae from Argentina. Darwiniana, 38(3-4), 227-239 Rosanoff, S. (1865). Zur kenntnniss des baues und der entwickelungsgeschichte des pollens der Mimosaceae. Jahrb. Wiss. Bot., 4, 441-450. Rosenfeldt, S. & Galati, B. G. (2005). Ubisch bodies and pollen ontogeny in Oxalis articulata Savigny. Biocell, 29(3), 271-278. (2008). Orbicules diversity in Oxalis species from Buenos Aires province (Argentina). Biocell, 32(1), 41-47. Rowley, J. R. (1962). Nonhomogeneous sporopollenin in microspores of Poa annua L. Grana Palynol., 3, 3-19. Rowley, J. R. & Erdtman, G. (1967). Sporoderm in Populus and Salix. Grana Palynol., 7, 518-567. Rowley, J. R. & Walles, B. 1987. Origin and structure of Ubisch bodies in Pinus sylvestris. Acta Societatis Botanicorum Poloniae, 56 (2), 215-227. Schnarf, K. (1923). Kleine beiträge zur entwicklungschichte der angiospermen. IV. Über das verhalten des antherentapetums einiger pflanzen. Österr. Bot. Z., 72, 242-245. Strittmatter, L. I.; Galati, B. G. & Monacci, F. (2000). Übisch bodies in the peritapetal membrane of Abutilon pictum Gill (Malvaceae). Beitr. Biol. Pflanzen, 71, 1-10. Strittmatter, L. I. & Galati, B. G. (2001). Development of anthers, microsporogenesis and microgametogenesis of Myosotis azorica and M.laxa (Boraginaceae). Phytomorphology, 51 (1), 1-10. Torres, C. & Galetto, L. (1998). Patterns and implications of floral nectar secretion, chemical composition, removal effects, and standing crop in Mandevilla pentlandia (Apocynaceae). Botanical journal of the Linnean Society, 172, 207-223. Ubisch, G. von. (1927). Kurze mitteilungen zur entwicklungsgeschiehte der antheren. Planta, 3, 490-495. Vaknin, Y.; Gan-Mor, S.; Bechar, A.; Ronen, B. & Eisikowitch, D. (1999). The role of electrostatic forces in pollination. In: Dafni, Hesse & Pacini (eds.). Pollen and Pollination. Springer-Verlag Wien New York. Vinckier, S. & Smets, E. (2002a). Morphology, ultrastructure and typology of orbicules in family Loganiaceae s.l. and related genera, in relation to systematics. Rev. Palaeobot. Palynol., 119 (3-4), 161-189. (2002b). Morphological and ultrastructural diversity of orbicules in relation to evolutionary tendencies in Apocynaceae s.l. Ann. Bot., 90, 647-662.

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(2002c). Systematic importance in orbicule diversity in Gentianales. Grana, 4, 158-182. (2003). Morphological and ultrastructural diversity of orbicules in Gentianaceae. Ann. Bot., 92, 657-672. Vinckier, S.; Huysmans, S. & Smets, E. (2000). Morphology and ultrastructure of orbicules in the subfamily Ixoroideae (Rubiaceae). Rev. Palaeobot. Palynol., 108, 151-174.

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INDEX # 20th century, 86

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A access, 38, 43, 44, 45, 46, 66, 69 accessions, 142 acetic acid, 88 acetone, 152 acid, 9, 11, 14, 16, 17, 29 activity theory, 152 adaptation, 2, 15, 21, 47 adaptations, 21, 28 adhesion, 12, 59, 62 advancements, 85 Africa, 27 age, 61 agricultural ecosystems, vii, 1 agriculture, 76 air temperature, 18, 76 alcohols, 10, 11 aldehydes, 10 ambient air, 18 amines, 16, 31 amino, vii, 1 amino acid, vii, 1 amino acids, vii, 1 anatomy, 25, 51 androgynophore, vii, 33, 34, 35, 38, 44, 45, 46 aneuploid, 86, 89, 101, 125, 129, 131, 146 angiosperm, 2, 12, 19, 28, 29, 31, 159 Angiosperms, ix, 38, 71, 149, 150, 152, 161, 162 aniline, viii, 57, 59, 60, 63, 64, 65 annealing, 81 anther, ix, 21, 26, 34, 35, 37, 39, 46, 49, 50, 88, 90, 114, 149, 150, 153, 155 anther locule, ix, 149, 150, 155

anthocyanin, 15, 28 antibiotic, 18 antioxidant, 9 apex, 38, 41, 62, 63 Apocynaceae, ix, 8, 16, 25, 26, 149, 150, 159, 161, 162, 163 Arabidopsis thaliana, 112, 139, 141, 143 Argentina, 22, 33, 37, 47, 51, 53, 55, 163 aromatic compounds, 15 arrest, 92, 98, 102, 103, 104, 109, 110, 112, 113, 115, 118, 119, 120, 121, 124, 125, 127, 128, 129, 130, 134, 135, 136, 144 arrests, 67, 113 assessment, 69 atmosphere, 9, 81, 83 attachment, 94, 102, 103, 104, 111, 133 attractant, 7, 11, 12, 16, 20, 23 avoidance, 54, 55, 81

B backcross, 140 bacteria, 17 base, 8, 10, 19, 37, 42, 62, 63, 82, 87, 89, 103, 130, 153, 155 beetles, 3, 14, 15, 16 bending, 35, 36, 39, 47, 48, 49, 51, 52, 53, 101, 116 benefits, 2, 8, 22, 24 biochemical processes, 15 biochemistry, 29 bioindicators, 76 biological roles, 31 biosynthesis, 5, 29, 31 biosynthetic pathways, 14 biotic, 2 birds, 2, 3, 7, 18, 45, 47, 53 botrytis, 79 Brazil, 54, 55, 161

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breakdown, 89, 90, 93, 94, 95, 96, 98, 100, 146 breeding, 55, 58, 71, 81, 85, 87, 117, 138, 139, 141, 143 budding, 112, 145

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C cabbage, 25, 86 caffeine, 124 capillary, 16 carbon, 9 carbon atoms, 9 carotenoids, 4, 9, 27 case study, 32, 83 Catharanthus roseus, 161 cation, 4 cell biology, 87 cell culture, 104 cell cycle, 112, 142, 146, 147 cell division, 85, 87, 88, 102, 104, 108, 110, 111, 122, 125, 127, 132, 133, 139, 140, 141, 142, 146 cell line, 146 cell membranes, 88, 89, 91, 92, 97, 101, 108, 110, 112, 117, 120, 121, 122, 123, 124, 125, 126, 127, 128, 132, 136 cell size, 88 Central Europe, 76 centriole, 146 centrosome, 87, 145 chemical, vii, 1, 2, 9, 12, 13, 17, 21, 22, 23, 26, 29, 75, 76, 80, 104, 112, 163 chemicals, 10, 11, 13, 14, 20 chromatography, 32 chromosome, 85, 86, 87, 88, 89, 90, 92, 93, 94, 95, 96, 97, 98, 99, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 128, 129, 130, 131, 132, 133, 135, 136, 137, 139, 143, 144, 146, 147 circadian rhythm, 9, 13 City, 163 clarity, 63 classes, 4, 9 classical methods, 90 classification, 37, 45, 151 climate, viii, 3, 73, 74, 75, 76, 77, 79, 82 climate change, 76, 82 climates, 18 clone, 95, 96, 118, 136, 139 clusters, 8 color, vii, 1, 3, 4, 5, 7, 8, 9, 14, 19, 22, 24, 27, 28, 32 combined effect, 27 commercial, 77 communication, 29, 30, 77

communities, 7, 8 community, 5, 9, 54 competition, viii, 52, 58, 66, 67, 68, 69, 71 compilation, 36 complex interactions, 2 composition, 10, 22, 24, 26, 31, 101, 111, 112, 163 compounds, 9, 10, 12, 14, 15, 16, 23, 25, 29, 30, 31 conditioning, 25 configuration, 88, 89, 90, 92, 94, 96, 98, 100, 102, 104, 107, 113, 115 conflict, 68, 71 congress, 100, 116, 121, 122, 124, 125 Congress, 81 conifer, 29 conjugation, 85, 86 conservation, 69, 99, 102, 116, 136 consolidation, 115, 134, 135, 136 constituents, 15, 18, 21 construction, 17 consumption, 11 contamination, 12 convergence, 32, 93, 99, 103, 107, 109 coordination, 110 corolla, 2, 5, 15, 19, 31, 39 correlation, 5, 8, 51, 62, 66, 76, 78, 79, 80 correlation coefficient, 66, 80 correlations, 13, 66, 67, 160 cortex, 147 cost, 7, 19, 52 courtship, 16 crop, vii, ix, 1, 27, 58, 73, 74, 75, 76, 77, 78, 79, 82, 83, 146, 163 crop production, vii, 1, 74, 75, 82, 83 crops, ix, 54, 73, 74, 75, 76, 77, 78, 79, 86 cross-fertilization, 18 cues, 5, 9, 10, 13, 14, 15, 16, 22, 30, 32 cultivars, 13, 82 culture, 52, 142 cycles, vii, 33, 34, 38, 41, 99, 144 cytokinesis, ix, 85, 86, 88, 89, 90, 91, 92, 94, 95, 96, 97, 98, 101, 102, 103, 105, 109, 112, 115, 117, 118, 119, 121, 123, 124, 126, 127, 128, 129, 130, 131, 132, 133, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147 cytoplasm, 86, 88, 90, 91, 92, 94, 95, 97, 100, 101, 106, 107, 108, 111, 112, 113, 115, 117, 119, 121, 122, 123, 124, 125, 126, 127, 133, 135, 136, 146 cytoskeleton, ix, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 104, 105, 106, 107, 108, 110, 111, 112, 113, 115, 117, 118, 119, 120, 121, 125, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 139, 140, 142, 144, 145, 146

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D damages, 7, 8, 18, 38 database, 20 deformation, 124 degradation, 27 dehiscence, viii, 33, 49, 50 dehydrate, 49 dehydration, 152 Denmark, 53 depolymerization, 93 deposition, vii, 33, 34, 59, 74 depth, 29, 38 derivatives, 4, 9, 11, 16 desorption, 12 detectable, 12 detection, 11, 14, 29, 74 deviation, 96 dew, 80 diakinesis, 88, 89, 91, 98, 129 diet, 20 diffusion, 18 dimorphism, 6, 54 diploid, 85, 95, 109, 119, 128, 130, 138, 139, 140, 141, 142, 145, 146 direct observation, 107 discrimination, 4, 26 diseases, 74, 78 dispersion, 3, 123, 127 displacement, 120 disposition, 43 distilled water, 88 distribution, 4, 15, 25, 46, 51, 74, 76, 77, 79, 85, 86, 124, 139, 140, 145, 150, 162 divergence, 22 diversification, 22 diversity, 6, 25, 31, 32, 163, 164 DNA, 143, 144 donors, viii, 34 dosage, 145 Drosophila, 104, 138, 140, 145 dusts, 19

E ecology, vii, 1, 3, 5, 21, 23, 25, 26, 27, 28, 29, 32, 53, 54, 79, 81 economic resources, 77 editors, 70, 72 Effective Pollination Period, viii, 57 egg, 69, 71, 138, 147 electromagnetic, 3, 4

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electron, 143, 151, 152, 155, 157, 158, 161 electrons, ix, 149, 151, 153, 155, 156, 157, 158, 159 elongation, 18, 114, 115, 138 embryo sac, 58, 67, 69, 71 embryology, 162 emission, ix, 9, 10, 11, 12, 13, 14, 15, 20, 24, 27, 30, 73, 74, 77, 82, 83 endosperm, 68, 99, 137, 138, 139, 140, 142, 144, 146 enemies, 23 energy, 3, 5, 18, 19 engineering, 85 England, 161 enlargement, 71 environment, 2, 9, 14, 76 environmental change, 76 environmental conditions, 79 enzymes, 8, 9, 10, 14 equational division, 86, 87, 128, 129 equipment, 13 ester, 19 ethanol, 152 EU, 77, 144 eukaryotic, 92, 110 eukaryotic cell, 92, 110 Europe, 77 European Union, 75, 77 evidence, 12, 21, 28, 49 evolution, 21, 22, 23, 25, 26, 27, 28, 29, 31, 32, 53, 54, 55, 68, 85, 86, 91, 138, 161 experimental condition, 53 expulsion, 160 extinction, 71 extraction, 12, 24 extracts, 24 exudate, 8, 18

F FAA, 152 families, ix, 2, 7, 8, 16, 18, 19, 149, 150, 152, 159, 160 farmers, 75, 76 farmland, 78 fatty acids, 18 feces, 16 fertility, 67, 142 fertilization, viii, 1, 2, 5, 39, 61, 62, 64, 65, 67, 69, 73, 75, 77, 79 fiber, 92, 100, 102, 103, 107, 110, 114, 115 fibers, 89, 90, 92, 93, 94, 97, 98, 99, 102, 103, 104, 105, 106, 107, 108, 110, 111, 112, 113, 114, 116, 118, 119, 120, 121, 124, 125, 126, 129, 131, 133, 135, 136, 137

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filament, 35, 39, 50 fission, 140, 145 fitness, 12 fixation, 87, 88, 95 flavonoids, 3, 4, 5, 25, 30 flexibility, 51 flight, 7, 18, 19 flora, 79, 161 floral scents, vii, 1, 11 flowering period, 46, 52 flowers, vii, viii, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 29, 30, 31, 32, 33, 34, 35, 36, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 57, 58, 59, 61, 62, 63, 64, 65, 66, 67, 69, 71, 78,뫰161 fluorescence, viii, 31, 57, 59, 60, 61, 63, 64, 65, 150 food, vii, 1, 3, 5, 7, 10, 12, 14, 16, 23, 24 food industry, vii, 1 force, 68 forecasting, 75, 76, 77, 78, 79, 80 forest ecosystem, 83 formation, ix, 4, 9, 29, 69, 85, 86, 87, 90, 91, 92, 93, 94, 95, 96, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 149, 151 fragments, 108 France, 77, 78, 83 freezing, 143 frost, 76, 82 fruits, 4, 13, 14, 24, 39, 48, 50, 52, 58, 67, 68, 70, 78, 81 fungi, 9, 28, 29, 80 fungus, 8, 29 fusion, 94, 96, 101, 106, 112, 123, 124, 126, 127, 130, 134, 136

G gamete, 125, 138, 140, 143 gametogenesis, 128, 141 gametophyte, 68 genes, 15, 23, 75, 139, 142 genetics, 32 genome, 107, 134, 143 genotype, 68, 69, 103, 128, 145 Gentianaceae, ix, 149, 159, 164 genus, viii, 7, 9, 13, 16, 17, 19, 23, 34, 41, 42, 43, 45, 46, 47, 52, 81, 82, 140, 141, 143, 161, 162 germination, 2, 59, 62, 68, 76, 80

Gimnosperms, ix, 149 ginseng, 145 global warming, 76 glue, 17 glycoside, 4 Gori, 5, 24 grass, 112, 121, 137 grasses, 160, 161 gravitational orientation, 41 grouping, 6 growth, vii, viii, 57, 58, 59, 61, 62, 63, 66, 67, 68, 69, 70, 71, 76, 80, 81, 114, 124, 145 growth rate, 62, 67 guidance, 69, 71

H habitats, 18 hair, 59 haploid, 86, 95, 96, 97, 101, 103, 113, 120, 121, 125, 128, 129, 130, 131, 132, 133, 135, 143 harvesting, 77, 78 Hawaii, 53 health, 74, 75 height, 5 hermaphrodite, 7, 34, 35, 39, 47, 48, 49, 71, 78 heterogeneity, 25 history, 30, 54, 58, 71 hives, 14 honey bees, 3, 14 host, 8, 29, 54 House, 161 hue, 3, 4 human, 3, 5, 10, 12, 74, 77, 138 humidity, 49, 50, 59, 80 husbandry, 74 hybrid, 86, 88, 89, 97, 98, 101, 103, 105, 106, 107, 108, 109, 110, 112, 113, 114, 115, 116, 117, 122, 124, 125, 127, 129, 130, 131, 140, 142, 143, 144, 145 hybridization, 85, 86 hypothesis, 10, 19, 68, 69, 79, 151, 152

I ideal, 11, 19 identification, 12, 88 identity, 134 image, 45, 63 images, 88, 90 imitation, 22 improvements, 12

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Index in vivo, 24, 138, 147 incidence, viii, 19, 34, 75, 80 incompatibility, 6, 31 independence, 144 India, 81 inducer, 29 induction, 146 infancy, 10 infection, 8 inferences, 28 infundibulum, 8, 26 ingredients, 138 inheritance, 147 inhibition, 11, 112, 140 inhibitor, 145 inoculum, 79, 81 insects, vii, 1, 2, 3, 4, 5, 6, 8, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 22, 31, 42, 45, 160 insertion, 38 institutions, 77 integration, 86, 97, 145 integrity, 101 interference, 4, 41, 54, 55 interphase, 92, 93, 94, 98, 112, 113, 118, 121, 123, 139, 143, 144, 146 intervention, 75 ions, 32 Iowa, 140 irrigation, 80 islands, 7, 20 isolation, 7, 30, 87 isoprene, 9, 28 Israel, 23, 26, 82 Italy, 1, 14, 15, 24, 27, 80, 82, 84

J Jordan, 6, 25, 26

K ketones, 11 kidney, 112 kinetochore, 89, 98, 100, 102, 103, 104, 105, 107, 108, 110, 135, 136, 137, 138 Korea, 22

L landscapes, 29 larvae, 14, 18

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lead, 3, 14, 85, 86, 93, 103, 107, 111, 114, 118, 119, 129, 131, 132, 133, 136, 139, 140, 152 learning, 26 legs, 16, 18, 45, 48 Lepidoptera, 15 lesions, 79 lifetime, 81 light, ix, 2, 4, 13, 15, 25, 59, 88, 90, 99, 121, 149 light conditions, 13 lignin, 9 lipids, 22, 31 loci, 142 locus, 23, 140 Loganiaceae, ix, 149, 150, 159, 163 longevity, viii, 31, 57, 61, 63, 67 LTD, 139 lying, 97, 104, 117

M magnet, 25 magnets, 22 magnitude, 77, 88 majority, vii, 1, 41, 47, 85, 86, 133 mammalian cells, 146 mammals, 20, 22, 24 man, 4 management, 75, 76, 77 manipulation, 69 marketing, 77 mass, 25, 32, 70, 90, 100, 122, 125, 130 mass spectrometry, 32 measurement, 59 media, 153, 155, 156, 157, 158 Mediterranean, 28, 58, 78, 79, 80, 82 megagametophyte, 70 meiosis, 77, 85, 86, 87, 88, 90, 91, 92, 94, 95, 96, 97, 98, 99, 100, 101, 103, 104, 105, 106, 107, 108, 109, 112, 115, 116, 117, 118, 119, 120, 121, 122, 123, 126, 128, 129, 130, 132, 133, 134, 135, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147 meiotic restitution, ix, 85, 86, 87, 93, 94, 101, 103, 111, 114, 117, 118, 119, 120, 128, 130, 132, 133, 136, 144 membranes, 112, 117, 120, 122, 123, 126, 127, 132, 160, 161 mercury, 59 meristem, 145 mesophyll, 140 meta-analysis, 11 metabolism, 22, 27, 29, 31 metabolites, vii, 1, 13, 26 metal ion, 4

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metal ions, 4 metaphase, 87, 89, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 104, 105, 106, 107, 108, 109, 110, 111, 113, 115, 116, 118, 119, 120, 121, 122, 123, 125, 126, 127, 129, 131, 132, 133, 134, 135, 145, 146 metaphase plate, 89, 100, 106, 107, 109, 129, 132 methodology, 37 Mexico, 20 micrometer, 62 microorganisms, 18 micropyle, 68, 69 microscope, 59, 62, 108, 151, 152, 161 microscopy, 147 migration, 95, 100, 103, 106, 107, 130 mildew, 82 mimicry, 8, 21, 22, 24, 25, 26, 27, 30 Missouri, 21 mitochondria, 151 mitosis, 87, 93, 99, 112, 113, 118, 125, 130, 131, 132, 134, 137, 140, 141, 142, 143, 146, 147 mixing, 4 modelling, 82, 83 models, 13, 76, 78, 81, 82, 116 modifications, 9 molecular biology, 29 molecules, vii, 1, 4 monolayer, 91, 92, 111, 117, 121 Montenegro, 7, 28 morphology, ix, 7, 8, 19, 22, 30, 36, 42, 51, 90, 116, 123, 146, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160 mother cell, ix, 85, 89, 97, 101, 107, 108, 112, 117, 121, 122, 123, 124, 125, 126, 127, 132, 133, 140, 146 multiple regression, 78 muscles, 18 mutant, 99, 100, 101, 106, 108, 112, 113, 117, 118, 119, 122, 123, 124, 126, 127, 130, 131, 134, 136, 139, 141, 143, 144, 145 mutation, 100, 103, 140, 144 mutations, 104, 112

N NEB, 89, 98, 99, 106, 108, 136 Netherlands, 81, 83 neuroblasts, 140 New Zealand, 54, 55, 161 next generation, 109 nicotine, 11 nitrogen, 9, 11, 16 NMR, 30

normal development, 152 nucellus, 59 nuclear surface, 88, 90, 145 nuclei, 86, 90, 91, 92, 94, 95, 96, 97, 101, 106, 107, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 143, 145 nucleus, 86, 87, 88, 90, 91, 93, 94, 97, 98, 100, 102, 103, 104, 105, 107, 108, 109, 110, 111, 113, 114, 115, 116, 120, 125, 129, 130, 131, 135, 146 nutrients, vii, 1 nutritional status, viii, 73, 75

O oil, vii, 1, 17, 18, 21, 23, 24, 58, 80 olfaction, 13, 19 olive flowers, viii, 14, 57, 62, 67, 69 olive oil, 78 olive tree, viii, 57, 58, 68, 69, 70, 78 omission, 129, 136 oocyte, 147 opportunities, 46 orchid, 8, 16, 17, 20, 23, 25, 28 organ, 16, 142 organelles, 87, 141 organic compounds, 9 organism, 86 organs, 5, 13, 16, 17, 24, 27 ovaries, 7, 51, 59, 71 overlap, 92, 114, 120 ovule, vii, viii, 2, 52, 57, 58, 59, 60, 61, 62, 65, 66, 67, 68, 69, 70, 75, 77 ovules, viii, 1, 36, 57, 58, 59, 61, 62, 63, 64, 65, 67, 68, 69, 71, 145 ox, 1, 2 Oxalidaceae, ix, 149, 150

P parallel, 5, 34, 38, 39, 44, 45, 48, 87, 108, 118, 123, 125, 126, 132, 134, 135 parasite, 27 parents, 86, 119 parthenogenesis, 140 Passiflora species, vii, 33, 34, 41, 47 Passionvines, vii, 33 pathogens, 9, 10, 12, 13, 17, 79 pathology, viii, 73, 79 pathways, 15 PCA, 13 pedicel, 38, 41

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Index perforation, 151, 157, 158, 159, 160 perianth, 7, 8, 38, 45 permit, 5, 6, 9, 20 personal communication, 135 pest populations, 75 pesticide, 75 pests, 2, 75 pH, 4, 152 phenocopy, 112 phenotype, 88, 89, 94, 95, 96, 100, 101, 102, 103, 104, 105, 108, 109, 110, 112, 113, 114, 115, 117, 118, 119, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 133, 134, 135, 136, 137, 139 phenotypes, 88, 93, 100, 101, 103, 112, 114, 117, 125, 129, 134 phenylalanine, 9 phosphate, 152 photographs, 36, 37 physiology, 161 pigmentation, 4, 15, 23, 27 pistil, 58, 59, 63, 67, 69, 138 plant-animal relationships, vii, 1 plants, vii, viii, ix, 1, 2, 3, 5, 6, 7, 8, 9, 11, 12, 13, 14, 16, 17, 18, 19, 20, 21, 22, 25, 26, 27, 28, 29, 30, 31, 41, 47, 48, 49, 50, 54, 57, 58, 59, 61, 62, 63, 64, 65, 66, 67, 68, 69, 71, 73, 75, 76, 77, 79, 85, 86, 90, 91, 100, 109, 135, 137, 138, 139, 140, 141, 144, 145 plaque, ix, 149, 151, 155, 159 plasma membrane, 152 plasmodial type tapetum, ix, 149 platform, 43 playing, 18 ploidy, 132 PM, 142 polar, 87, 90, 91, 92, 93, 104, 111, 114, 116, 118, 119, 120, 125, 128, 133, 134, 135, 157 polarity, 143 policy, 77 pollen, vii, viii, ix, 1, 2, 3, 5, 6, 7, 8, 10, 11, 12, 14, 16, 17, 18, 19, 20, 21, 22, 23, 24, 27, 28, 30, 32, 33, 34, 35, 39, 40, 41, 48, 49, 50, 52, 53, 54, 55, 57, 59, 61, 62, 63, 66, 68, 69, 70, 71, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 85, 109, 129, 138, 139, 140, 142, 143, 145, 146, 147, 149, 150, 151, 152, 159, 160, 161, 162, 163 pollen tube, vii, viii, 57, 59, 61, 62, 63, 66, 68, 69, 70, 71, 145, 147 pollination, vii, ix, 1, 2, 3, 5, 6, 7, 8, 9, 10, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 36, 38, 39, 40, 41, 42, 44, 46, 47, 48, 49, 52, 53, 54, 55, 58, 59, 61,

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70, 75, 76, 78, 79, 80, 81, 83, 86, 149, 152, 153, 156, 159, 160, 161, 162, 163 pollination ecology, vii, 1, 3, 5, 29, 32, 54, 81 pollinators, vii, viii, 1, 3, 4, 5, 6, 7, 8, 9, 10, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 25, 26, 28, 30, 32, 33, 34, 35, 38, 39, 40, 41, 44, 45, 46, 47, 49, 52, 53, 54 Pollinators, vii, 1 pollutants, 77 polymerization, 90, 118, 120, 133, 134 polymorphism, 7 polymorphisms, 31 polyploid, 85 polyploidy, 85, 137, 141, 146 pools, 11 population, 7, 31, 32, 48, 49, 51, 52, 81, 125 Portugal, 78 potato, 90, 95, 96, 100, 106, 117, 118, 119, 136, 138, 139, 140, 141, 143, 147 predation, 6 preparation, 88, 123, 125 preservation, 52, 103 prevention, 75 principles, 54, 81 probability, 2, 11, 46, 49, 67 probe, 46 producers, 75 product market, 75 production quota, 75, 77 profit, 18 profitability, 161 prophase, 88, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 102, 112, 113, 117, 118, 119, 121, 128, 130, 131, 134, 135, 136, 144, 145, 147 protection, 7, 29, 31 proteins, 3, 16, 132, 140, 141 proteolysis, 147 publishing, 83 pulp, 52

R radiation, 26 radius, 38, 114 rainfall, 77 reactions, 18 reception, 5, 52 recognition, 5, 13, 14 recombination, 138 reflectance spectra, 20 reforms, 102 regulations, 77 relative size, vii, 33, 34

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relatives, 58 relevance, 138 repellent, 11 reproduction, 12, 28, 30, 81, 85 reproductive organs, 6, 45 reptile, 20 reptile species, 20 repulsion, 160 requirements, 5, 13, 22, 80 researchers, 79 residues, 80 resins, 17 resistance, 23, 150 resolution, 5 resource availability, 46 resources, 6, 11, 21, 46, 47, 67, 77 response, 9, 29, 150 restitution, ix, 85, 86, 87, 93, 94, 97, 98, 100, 101, 102, 103, 104, 105, 107, 108, 109, 110, 111, 114, 115, 116, 117, 118, 119, 120, 122, 125, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 138, 139, 141, 143, 144, 146, 147 restoration, 112 restrictions, 87 reticulum, 151 rewards, 3, 5, 6, 7, 8, 10, 13, 16, 17, 22, 23, 27 rhythm, 27 rhythmicity, 20 rings, 90, 94, 95, 96, 134, 136 risk, 6, 79 risks, ix, 73, 74, 75 rodents, 14 room temperature, 88, 140 root, 86, 138, 145 Royal Society, 20, 21, 26, 30 Rubiaceae, ix, 149, 150, 162, 164 runoff, 7 Russia, 85

S saturation, 3 savings, 69 scanning electron microscopy, 152 scattering, 115 scent, vii, 1, 3, 8, 9, 10, 14, 16, 18, 19, 21, 23, 26, 27, 29, 30, 32 science, 74 scope, 2, 19 seasonal component, 49 secrete, 17 secretion, vii, 1, 17, 25, 162, 163

seed, viii, 3, 4, 23, 24, 27, 28, 31, 55, 57, 58, 61, 62, 64, 65, 66, 67, 68, 69, 70, 71, 73, 75, 79, 80, 81, 83 seedlings, 46, 68 segregation, 86, 87, 93, 95, 96, 98, 102, 104, 105, 109, 110, 111, 120, 128, 129, 135, 136 selectivity, 20 self-fertilization, 6, 21 semicircle, 45 senescence, viii, 57, 59, 61, 62, 63, 65, 67, 70, 71 senses, 13 services, vii, 1, 17, 19 sex, 5, 16, 17, 36 sexual behavior, 17 sexual reproduction, 1, 9, 20 sexuality, 1, 36, 140 shade, 46, 49 shape, 2, 4, 5, 7, 15, 22, 28, 44, 100, 103, 110, 115, 117, 122, 123, 127 shoot, 47 short supply, 3 showing, 35 sibling, 67, 68, 70, 71 siblings, 68 signals, 2, 3, 5, 6, 7, 8, 10, 13, 17, 20, 21, 27, 69 signs, 7 simulation, 21 simulations, 75 skeleton, 4, 9, 18, 99, 134 solution, 12, 30, 59 solvents, 12 somatic cell, 143 South Africa, 19, 27, 70 South America, 22 Spain, 57, 70, 73, 78, 79, 81, 82 specialization, 23 speciation, 85 species, vii, viii, ix, 2, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 20, 22, 25, 26, 31, 32, 33, 34, 35, 37, 38, 39, 40, 41, 42, 45, 46, 47, 48, 49, 50, 51, 52, 53, 55, 58, 67, 68, 69, 73, 74, 75, 76, 77, 78, 79, 82, 85, 86, 88, 90, 119, 133, 134, 135, 141, 146, 147, 149, 150, 151, 152, 153, 156, 157, 158, 159, 160, 161, 162, 163 spending, 3 spin, 34, 38, 39, 50 spindle, 85, 86, 87, 89, 90, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 140, 141, 142, 143, 144, 145, 146, 147 spore, 74, 76, 79, 80, 82, 163

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Index sporopollenin, ix, 149, 151, 152, 160, 162, 163 stability, 138 stamens, 3, 6, 7, 10, 17, 19, 27, 34, 40, 50 starch, 68, 71 starvation, 67 state, 2, 14, 39, 46 stereospecificity, 9 sterile, 7, 16, 19, 34, 48, 71 sternum, 17 stigma, viii, 1, 2, 17, 18, 19, 20, 34, 35, 39, 41, 46, 48, 50, 51, 58, 59, 61, 62, 65, 66, 73, 75, 77 storage, 9 stratification, 162 stress, 9, 31 stress factors, 31 structure, 5, 15, 58, 68, 87, 91, 92, 96, 99, 100, 103, 105, 109, 111, 114, 120, 137, 138, 141, 160, 163 style, viii, 33, 34, 35, 50, 51, 58, 62, 63, 66 Styles, 40, 47, 50, 51 subjectivity, 78 substrates, 151 sucrose, 8 sugar beet, 108, 118, 119, 133, 139 sugarcane, 138 sulfur, 9, 14, 16 sulphur, 30 Sun, 8, 31 surplus, 58, 67, 68 survival, 13, 68, 69 symptoms, 59 syndrome, 3, 5, 10, 17, 34, 38, 42, 45, 58, 159, 160 synergid, 69, 70 synthesis, 112, 121, 151

T Taiwan, 161 taxa, 15, 26, 46, 146, 150 taxonomy, 71 techniques, 39, 81, 151 telophase, 86, 88, 89, 90, 91, 92, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 127, 128, 131, 132, 133, 134, 135, 141 TEM, 152, 154, 155, 156, 158 temperature, 16, 18, 31, 49, 50, 61, 63, 76, 77, 80, 146 tension, 9, 19 teratology, 50 terpenes, 9 terrestrial ecosystems, 15 tetrad, 89, 91, 96, 105, 113, 118, 120, 122

173

Thailand, 26 theft, 19 thinning, 67, 70, 71 thorax, 34, 41 tibia, 16 tissue, 51, 62, 66, 151 tobacco, 90, 92, 109, 130, 135, 144 torus, 115 total product, 77 toxic effect, 17 trafficking, 146 traits, viii, 3, 5, 10, 34, 35, 45, 85, 99, 124, 129 transcription, 28 transference, 151 transformation, 62, 85 translocation, 147 transmission, 151, 152 transmission electron microscopy, 152 transparency, ix, 149 transport, 12, 74, 75, 79, 91, 92, 110, 112, 114, 120, 121, 124, 125, 128, 132, 151 trial, 61 triploid, 109, 137, 139, 143 two-dimensional space, 2 tyrosine, 4

U Ubisch bodies, ix, 149, 150, 152, 159, 160, 161, 162, 163 ultrastructure, ix, 99, 124, 140, 147, 149, 151, 163, 164 ultraviolet irradiation, 31 unfertilized flowers, viii, 57, 61, 62, 63, 64, 65, 67 uniform, 5 United, 25 United States, 25 USA, 31, 81, 138, 139, 146 UV, 3, 4, 5, 7, 8, 9, 29, 30, 60 UV light, 7, 60

V vacuole, 114 Valencia, 82, 83 vapor, 9 variables, ix, 66, 73, 76, 79, 80 variations, viii, 7, 59, 73, 75, 77, 82, 93 varieties, 145 vascular bundle, 60, 61 vector, 2 vegetation, 19, 54, 58, 77, 162

Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and

Index

174 vehicles, 2 vertebrates, 2, 3, 14, 18 viscosity, 19 vision, 3, 4, 26 visualization, 86, 87, 95, 105, 109, 119, 131, 135

W

Y yeast, 112, 138, 145 yield, viii, ix, 73, 75, 77, 78, 82

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walking, 46 waste, 3, 6, 68 water, 2, 18, 19, 24, 76, 88

wild type, 88, 90, 94, 95, 107, 119, 123, 130, 132, 134 wind-pollinated species, viii, 73, 77 wood, 17 workers, 11

Pollination: Mechanisms, Ecology and Agricultural Advances : Mechanisms, Ecology and Agricultural Advances, edited by Nichole D. Raskin, and