Floral volatiles controlling ant behaviour

P>1. Ants show complex interactions with plants, both facultative and mutualistic, ranging from grazers through seed predators and dispersers to herders of some herbivores and guards against others. But ants are rarely pollinators, and their visits to flowers may be detrimental to plant fitness. 2. Plants therefore have various strategies to control ant distributions, and restrict them to foliage rather than flowers. These 'filters' may involve physical barriers on or around flowers, or 'decoys and bribes' sited on the foliage (usually extrafloral nectaries - EFNs). Alternatively, volatile organic compounds (VOCs) are used as signals to control ant behaviour, attracting ants to leaves and/or deterring them from functional flowers. Some of the past evidence that flowers repel ants by VOCs has been equivocal and we describe the shortcomings of some experimental approaches, which involve behavioural tests in artificial conditions. 3. We review our previous study of myrmecophytic acacias, which used in situ experiments to show that volatiles derived from pollen can specifically and transiently deter ants during dehiscence, the effects being stronger in ant-guarded species and more effective on resident ants, both in African and Neotropical species. In these plants, repellence involves at least some volatiles that are known components of ant alarm pheromones, but are not repellent to beneficial bee visitors. 4. We also present new evidence of ant repellence by VOCs in temperate flowers, which is usually pollen-based and active on common European ants. We use these data to indicate that across a wide range of plants there is an apparent trade-off in ant-controlling filter strategies between the use of defensive floral volatiles and the alternatives of decoying EFNs or physical barriers.

Spatio-temporal variation in seed production in three Euphorbia species and the role of animals on seed fate

Comparative studies on the reproductive biology of co-occurring related plant species have provided valuable information for the interpretation of ecological and evolutionary phenomena, with direct application in conservation management of plant populations. The aims of this thesis were to identify the causes of pre-dispersal reproductive losses in three Euphorbia species (the Mediterranean E. characias and the narrow endemics E. pedroi and E. welwitschii) and evaluate the variation of their effects in time, space and between individuals and species. Furthermore, we intended to study elaiosomes’ fatty acid profiles for the three Euphorbia and assess the role played by the elaiosome in ant attraction. Finally, we aimed to identify the major seed dispersal agents for each Euphorbia species in each site and study differences in short term seed fate due to differences in ant behaviour. The results indicated that intact seed production differed significantly between the three Euphorbia, mostly due to differences in cyathia production. Losses to pre-dispersal seed predators were proportionately larger for the endemic species which also suffered higher losses resulting in flower, fruit (in E. welwitschii) and seed abortion (in E. pedroi). The elaiosomes of E. pedroi are poor in fatty acids and for this reason seeds of this species were removed in lower proportion by mutualistic dispersers than those of their congeners, being more prone to seed predation. Two larger ant species – Aphaenogaster senilis and Formica subrufa – were responsible for a larger percentage of removals with seeds being transported at larger distances and being discarded in the vicinity of their nests following elaiosome removal. Our results highlight the role of insect-plant interactions as major determinants of seed survival for the three study plants and call for the need to include more information on insect-plant interactions in plant conservation programmes.

Size and lipid content of nonmyrmecochorous diaspores: Effects on the interaction with litter-foraging ants in the Atlantic rain forest of Brazil

Ants are often attracted to diaspores not adapted for dispersal by ants. These diaspores may occasionally benefit from this interaction. We selected six nonmyrmecochorous plant species (Virola oleifera, Eugenia stictosepala, Cabralea canjerana, Citharexylum myrianthum, Alchornea glandulosa and Hyeronima alchorneoides) whose diaspores differ in size and lipid content, and investigated how these features affect the outcome of ant-diaspore interactions on the floor of a lowland Atlantic forest of Southeast Brazil. A total of 23 ant species were seen interacting with diaspores on the forest floor. Ants were generally rapid at discovering and cleaning the diaspore pulp or aril. Recruitment rate and ant attendance were higher for lipid-rich diaspores than for lipid-poor ones. Removal rate and displacement distance were higher for small diaspores. The large ponerine ant Pachycondyla striata, one of the most frequent attendants to lipid-rich arillate diaspores, transported the latter into their nests and discarded clean intact seeds on refuse piles outside the nest. Germination tests with cleaned and uncleaned diaspores revealed that the removal of pulp or aril may increase germination success in Virola oleifera, Cabralea canjerana, Citharexylum myrianthum and Alchornea glandulosa. Gas chromatography analyses revealed a close similarity in the fatty acid composition of the arils of the lipid-rich diaspores and the elaiosome of a typical myrmecochorous seed (Ricinus communis), corroborating the suggestion that some arils and elaiosomes are chemically similar. Although ant-derived benefits to diaspores - secondary dispersal and/or increased germination - varied among the six plant species studied, the results enhanced the role of ant-diaspore interactions in the post-dispersal fates of nonmyrmecochorous seeds in tropical forests. The size and the lipid-content of the diaspores were shown to be major determinants of the outcome of such interactions.

Biology and behaviour of the neotropical ant-mimicking spider Aphantochilus rogersi (Araneae: Aphantochilidae): nesting, maternal care and ontogeny of ant-hunting techniques

Aphantochilus rogersi is an ant-mimicking spider that preys exclusively on cephalotine ants. The spiders oviposit in close proximity to nests of the model ant Zacryptocerus pusillus, and emergent spiderlings tend to remain in the vicinity of natal egg sacs. Females of A. rogersi actively defend their egg sacs against approaching workers of Z. pusillus, but the latter may sometimes destroy the eggs. Feeding specialization on these ants is confirmed by more than 300 observations of young and adult A. rogersi carrying ant corpses in the held. Although A. rogersi possesses several behavioural traits which may reduce the risk of being injured by ants during subjugation, field and laboratory observations showed that social defence by Z. pusillus may cause mutilation to the spiders. Tests in captivity revealed an ontogenetic change in the prey-capture techniques employed by A. rogersi. Early-instar spiderlings can apparently only seize the ant's petiole tightly if they are able to approach the ant from the front. As the ant is paralysed, the spiderling positions itself vertically in relation to the substratum. Larger spiders, on the other hand, attack ants most frequently from behind, and seem better equipped to seize the ant's petiole firmly with their larger chelicerae. Owing to their greater strength, late-instar spiders are able to Lift the struggling ant aloft. The selection of a suitable oviposition site, the mother's ability to defend herself and the eggs from nearby ants, and the capacity to capture and subdue ants safely from emergence to maturity, are regarded as crucial traits inherent in the mimetic and feeding specialization by A. rogersi.

Visual Navigation In The Neotropical Ant Odontomachus Hastatus (formicidae, Ponerinae), A Predominantly Nocturnal, Canopy-dwelling Predator Of The Atlantic Rainforest.

The arboreal ant Odontomachus hastatus nests among roots of epiphytic bromeliads in the sandy forest at Cardoso Island (Brazil). Crepuscular and nocturnal foragers travel up to 8m to search for arthropod prey in the canopy, where silhouettes of leaves and branches potentially provide directional information. We investigated the relevance of visual cues (canopy, horizon patterns) during navigation in O. hastatus. Laboratory experiments using a captive ant colony and a round foraging arena revealed that an artificial canopy pattern above the ants and horizon visual marks are effective orientation cues for homing O. hastatus. On the other hand, foragers that were only given a tridimensional landmark (cylinder) or chemical marks were unable to home correctly. Navigation by visual cues in O. hastatus is in accordance with other diurnal arboreal ants. Nocturnal luminosity (moon, stars) is apparently sufficient to produce contrasting silhouettes from the canopy and surrounding vegetation, thus providing orientation cues. Contrary to the plain floor of the round arena, chemical cues may be important for marking bifurcated arboreal routes. This experimental demonstration of the use of visual cues by a predominantly nocturnal arboreal ant provides important information for comparative studies on the evolution of spatial orientation behavior in ants. This article is part of a Special Issue entitled: Neotropical Behaviour.

A simple genetic basis for complex social behaviour mediates widespread gene expression differences.

A remarkable social polymorphism is controlled by a single Mendelian factor in the fire ant Solenopsis invicta. A genomic element marked by the gene Gp-9 determines whether workers tolerate one or many fertile queens in their colony. Gp-9 was recently shown to be part of a supergene with two nonrecombining variants, SB and Sb. SB/SB and SB/Sb queens differ in how they initiate new colonies, and in many physiological traits, for example odour and maturation rate. To understand how a single genetic element can affect all these traits, we used a microarray to compare gene expression patterns between SB/SB and SB/Sb queens of three different age classes: 1-day-old unmated queens, 11-day-old unmated queens and mated, fully reproductive queens collected from mature field colonies. The number of genes that were differentially expressed between SB/SB and SB/Sb queens of the same age class was smallest in 1-day-old queens, maximal in 11-day-old queens and intermediate in reproductive queens. Gene ontology analysis showed that SB/SB queens upregulate reproductive genes faster than SB/Sb queens. For all age classes, genes inside the supergene were overrepresented among the differentially expressed genes. Consistent with the hypothesized greater number of transposons in the Sb supergene, 13 transposon genes were upregulated in SB/Sb queens. Viral genes were also upregulated in SB/Sb mature queens, consistent with the known greater parasite load in colonies headed by SB/Sb queens compared with colonies headed by SB/SB queens. Eighteen differentially expressed genes between reproductive queens were involved in chemical signalling. Our results suggest that many genes in the supergene are involved in regulating social organization and queen phenotypes in fire ants.

Kin structure and queen execution in the Argentine ant Linepithema humile

Every spring, workers of the Argentine Ant Linepithema humile kill a large proportion of queens within their nests, Although this behaviour inflicts a high energetic cost oil the colonies, its biological significance has remained elusive so far. An earlier study showed that the probability of a queen being executed is not related to her weight, fecundity, or age. Here we test the hypothesis that workers collectively eliminate queens to which they are less related, thereby increasing their inclusive fitness. We found no evidence for this hypothesis. Workers of a nest were on average not significantly less related to executed queens than to surviving ones. Moreover, a population genetic analysis revealed that workers were not genetically differentiated between nests. This means that workers of a given nest are equally related to any queen in the population and that there can be no increase in average worker-queen relatedness by selective elimination of queens. Finally, our genetic analyses also showed that, in contrast to workers, queens were significantly genetically differentiated between nests and that there was significant isolation by distance for queens.

Pupal cocoons affect sanitary brood care and limit fungal infections in ant colonies.

BACKGROUND: The brood of ants and other social insects is highly susceptible to pathogens, particularly those that penetrate the soft larval and pupal cuticle. We here test whether the presence of a pupal cocoon, which occurs in some ant species but not in others, affects the sanitary brood care and fungal infection patterns after exposure to the entomopathogenic fungus Metarhizium brunneum. We use a) a comparative approach analysing four species with either naked or cocooned pupae and b) a within-species analysis of a single ant species, in which both pupal types co-exist in the same colony. RESULTS: We found that the presence of a cocoon did not compromise fungal pathogen detection by the ants and that species with cocooned pupae increased brood grooming after pathogen exposure. All tested ant species further removed brood from their nests, which was predominantly expressed towards larvae and naked pupae treated with the live fungal pathogen. In contrast, cocooned pupae exposed to live fungus were not removed at higher rates than cocooned pupae exposed to dead fungus or a sham control. Consistent with this, exposure to the live fungus caused high numbers of infections and fungal outgrowth in larvae and naked pupae, but not in cocooned pupae. Moreover, the ants consistently removed the brood prior to fungal outgrowth, ensuring a clean brood chamber. CONCLUSION: Our study suggests that the pupal cocoon has a protective effect against fungal infection, causing an adaptive change in sanitary behaviours by the ants. It further demonstrates that brood removal-originally described for honeybees as "hygienic behaviour"-is a widespread sanitary behaviour in ants, which likely has important implications on disease dynamics in social insect colonies.

Stay or drift? Queen acceptance in the ant Formica paralugubris

The acceptance of new queens in ant colonies has profound effects on colony kin structure and inclusive fitness of workers. Therefore, it is important to study the recognition and discrimination behaviour of workers towards reproductive individuals entering established colonies. We examined the acceptance rate of queens in populations of the highly polygynous ant F. paralugubris, where the genetic differentiation among nests and discrimination ability among workers suggest that workers might reject foreign queens. We experimentally introduced young queens in their natal nest and in foreign nests. Surprisingly, the survival rate of mated queens did not differ significantly when introduced in a foreign male-producing nest, a foreign female-producing nest, or the natal nest. Moreover, the survival of virgin queens in their natal nest was twice the one of mated queens, suggesting that mating status plays an important role for acceptance. The results indicate that other factors than queen discrimination by workers are implicated in the limited long-distance gene flow between nests in these populations.

Effects of ploidy and sex-locus genotype on gene expression patterns in the fire ant Solenopsis invicta.

Males in many animal species differ greatly from females in morphology, physiology and behaviour. Ants, bees and wasps have a haplodiploid mechanism of sex determination whereby unfertilized eggs become males while fertilized eggs become females. However, many species also have a low frequency of diploid males, which are thought to develop from diploid eggs when individuals are homozygous at one or more sex determination loci. Diploid males are morphologically similar to haploids, though often larger and typically sterile. To determine how ploidy level and sex-locus genotype affect gene expression during development, we compared expression patterns between diploid males, haploid males and females (queens) at three developmental timepoints in Solenopsis invicta. In pupae, gene expression profiles of diploid males were very different from those of haploid males but nearly identical to those of queens. An unexpected shift in expression patterns emerged soon after adult eclosion, with diploid male patterns diverging from those of queens to resemble those of haploid males, a pattern retained in older adults. The finding that ploidy level effects on early gene expression override sex effects (including genes implicated in sperm production and pheromone production/perception) may explain diploid male sterility and lack of worker discrimination against them during development.

Influence of the number of queens on nestmate recognition and attractiveness of queens to workers in the Argentine ant, Iridomyrmex humilis (Mayr)

To investigate the influence of the number of queens per colony on nestmate recognition in Iridomyrmex humilis, comparative assays were performed to study the attraction of workers to queens. These assays demonstrated that a phenomenon of recognition is superimposed on the attraction of workers to queens. Workers could discriminate non-nestmate queens from their nestmate queen to which they were significantly more attracted. This discrimination is probably based on the learning by workers of queen and colony odour. The level of attraction of workers to non-nestmate queens was similar in monogynous and polygynous colonies, whereas the level of attraction of workers to nestmate queens was significantly lower in polygynous colonies. This difference in the level of attraction of workers to nestmate queens almost certainly resulted from a lower efficiency in nestmate recognition in polygynous colonies. It is hypothesized that the mixture of several pheromonal sources produced by less related individuals in polygynous colonies may result in a less distinct colony odour than in monogynous colonies. The results are discussed with regard to some implications of polygyny and particularly to the loss of intercolonial aggression in I. humilis as well as in other polygynous ant species

Queen execution in the Argentine ant, Iridomyrmex humilis

Field censuses and laboratory experiments show that in the Argentine ant, Iridomyrmex humilis (Mayr), c. 90% of the queens are executed by workers in May, at the beginning of the reproductive season. The reduction in the number of queens probably decreases the inhibition exerted by queens on the differentiation of sexuals and thus allows the production of new queens and males shortly thereafter. In the laboratory, there was no correlation between the percentage of queens executed and their weight or fecundity. At the time of execution of queens, nearly all queens were of the same age; less than 1 year. Therefore it is not likely that the age of queens plays any role in the choice that workers make in the queens they executed. Execution of these queens results in a heavy energetic cost for the colony which amounts c. 8% of the total biomass. This behaviour of workers executing nestmate queens is discussed with regard to possible evolutionary significance at the queen and worker level.

Species- and nestmate brood discrimination in the sibling wood ant species Formica paralugubris and Formica lugubris

Formica lugubris and E paralugubris are sympatric sibling species of wood ants, both of which are widely distributed in Switzerland. Until 1996 they were considered the same species, E lugubris. To investigate whether the two species can be distinguished based on discrimination cues used by the workers we used the pupa-carrying test first introduced by Rainer Rosengren. In this test workers of discriminator colonies are faced with two kinds of pupae and their preferences for one of the types are recorded based on differential retrieval. Interspecific comparisons showed that ants preferred conspecific worker pupae to those of the sibling species regardless whether the pupae were con-colonial or hetero-colonial. Hence, this test can be used as a taxonomic tool to identify wood ants hardly distinguishable by morphological characters. In intraspecific comparisons the highly polygynous (many queens per colony) E paralugubris, the polygynous form of E lugubris and the monogynous (single queen per nest) to weakly polygynous form of E lugubris expressed different trends in their preference behaviour (with nestmate recognition in 14%, 20% and 31% of replicates, respectively). Only F paralugubris presented no significant nestmate recognition.

Expression of foraging and Gp-9 are associated with social organization in the fire ant Solenopsis invicta.

The aim of this study was to investigate levels of expression of two major genes, the odorant binding protein Gp-9 (general protein-9) and foraging, that have been shown to be associated with behavioural polymorphisms in ants. We analysed workers and young nonreproductive queens collected from nests of the monogyne (single reproductive queen per nest) and polygyne (multiple reproductive queens) social forms of Solenopsis invicta. In workers but not young queens, the level of foraging expression was significantly associated with social form and the task performed (ie localization in the nest or foraging area). The level of expression of Gp-9 was also associated with social form and worker localization. In addition there was a higher level of expression of the Gp-9(b) allele compared with the Gp-9(B) allele in the heterozygote workers and the young nonreproductive queens. Finally, in the polygyne colonies the level of expression of foraging was not significantly associated with the Gp-9 genotype for either workers or young nonreproductive queens, suggesting that both genes have independent non-epistatic effects on behaviour in S. invicta.

Queen acceptance in a socially polymorphic ant

A central question in social evolution is what processes regulate the number of breeders in each social group. Here, we tested whether differences in the rate of acceptance of new queens by resident workers could be a proximate cause explaining the coexistence of single- and multiple-queen colonies in an ant population. We found that Formica selysi workers discriminated against foreign (non-nestmate) queens, which contributes to maintaining the genetic integrity of the social group essential to kin selection. All the young and newly mated foreign queens introduced into experimental groups of workers died within 48 h. In contrast, workers frequently accepted young newly mated nestmate queens. The survival of nestmate queens was not significantly lower in groups of workers originating from single- queen colonies than in groups of workers originating from multiple-queen colonies. Finally, virgin queens had significantly higher survival than mated queens. Together, these results show that the maintenance of single-queen and multiple-queen colonies in the same population is unlikely to be caused by strong differences between the two types of colonies in their rate of acceptance of new queens by workers. They also suggest that the discrimination of queens by resident workers restricts the dispersal of foreign queens among colonies, but not the acceptance of additional nestmate queens.