Jürgen Liebig

Genomic comparison of the ants Camponotus floridanus and Harpegnathos saltator.

Ant Variation Ants of the same genotype can exhibit numerous phenotypic forms and develop multiple functional castes within a colony. Bonasio et al. (p. 1068) sequenced the genomes of two ant species exhibiting differences in caste development—Camponotus floridanus and Harpegnathos saltator—and used the sequences to compare gene expression and identify differences in epigenetic gene regulation that lead to the phenotypic differences. Ants may offer a model system for studying the role of epigenetics in behavior and development. Comparison reveals the epigenetic controls on caste development in ants. The organized societies of ants include short-lived worker castes displaying specialized behavior and morphology and long-lived queens dedicated to reproduction. We sequenced and compared the genomes of two socially divergent ant species: Camponotus floridanus and Harpegnathos saltator. Both genomes contained high amounts of CpG, despite the presence of DNA methylation, which in non-Hymenoptera correlates with CpG depletion. Comparison of gene expression in different castes identified up-regulation of telomerase and sirtuin deacetylases in longer-lived H. saltator reproductives, caste-specific expression of microRNAs and SMYD histone methyltransferases, and differential regulation of genes implicated in neuronal function and chemical communication. Our findings provide clues on the molecular differences between castes in these two ants and establish a new experimental model to study epigenetics in aging and behavior.

Genome-wide and caste-specific DNA methylomes of the ants Camponotus floridanus and Harpegnathos saltator.

BACKGROUND Ant societies comprise individuals belonging to different castes characterized by specialized morphologies and behaviors. Because ant embryos can follow different developmental trajectories, epigenetic mechanisms must play a role in caste determination. Ants have a full set of DNA methyltransferases and their genomes contain methylcytosine. To determine the relationship between DNA methylation and phenotypic plasticity in ants, we obtained and compared the genome-wide methylomes of different castes and developmental stages of Camponotus floridanus and Harpegnathos saltator. RESULTS In the ant genomes, methylcytosines are found both in symmetric CG dinucleotides (CpG) and non-CpG contexts and are strongly enriched at exons of active genes. Changes in exonic DNA methylation correlate with alternative splicing events such as exon skipping and alternative splice site selection. Several genes exhibit caste-specific and developmental changes in DNA methylation that are conserved between the two species, including genes involved in reproduction, telomere maintenance, and noncoding RNA metabolism. Several loci are methylated and expressed monoallelically, and in some cases, the choice of methylated allele depends on the caste. CONCLUSIONS These first ant methylomes and their intra- and interspecies comparison reveal an exonic methylation pattern that points to a connection between DNA methylation and splicing. The presence of monoallelic DNA methylation and the methylation of non-CpG sites in all samples suggest roles in genome regulation in these social insects, including the intriguing possibility of parental or caste-specific genomic imprinting.

Cuticular hydrocarbons mediate discrimination of reproductives and nonreproductives in the ant Myrmecia gulosa

In many species of social insects, the cuticular hydrocarbons of adults vary with both colony identity and individual physiology (oogenesis). Such variations have been shown in some ants and social wasps to function in nestmate recognition, but as yet there is no demonstration of their use by workers to recognize egg layers. We report that in the ant Myrmecia gulosa, workers can discriminate queens and fertile workers from infertile individuals based on distinctive blends of long-chained hydrocarbons present both on the cuticle and in the postpharyngeal gland. The purified hydrocarbon fraction of cuticular extracts from queens elicited high interest in workers, unlike the nonhydrocarbon fraction. However, both fractions were necessary to trigger a response of maximal intensity. In contrast, extracts of mandibular and Dufour glands from queens or infertile workers were not treated differentially by workers. We suggest that cuticular hydrocarbons function as pheromones allowing for recognition of the queen as well as egg-laying workers.

Molecular traces of alternative social organization in a termite genome

Although eusociality evolved independently within several orders of insects, research into the molecular underpinnings of the transition towards social complexity has been confined primarily to Hymenoptera (for example, ants and bees). Here we sequence the genome and stage-specific transcriptomes of the dampwood termite Zootermopsis nevadensis (Blattodea) and compare them with similar data for eusocial Hymenoptera, to better identify commonalities and differences in achieving this significant transition. We show an expansion of genes related to male fertility, with upregulated gene expression in male reproductive individuals reflecting the profound differences in mating biology relative to the Hymenoptera. For several chemoreceptor families, we show divergent numbers of genes, which may correspond to the more claustral lifestyle of these termites. We also show similarities in the number and expression of genes related to caste determination mechanisms. Finally, patterns of DNA methylation and alternative splicing support a hypothesized epigenetic regulation of caste differentiation.

Worker policing limits the number of reproductives in a ponerine ant

Reproductive division of labour is an essential feature of insect sociality, but the regulation of sterility among colony members remains incompletely understood. Ant workers and queens are morphologically divergent and workers are only capable of producing males in a colony, although they usually do not do so. Worker policing is one mechanism proposed for their infertility and it can be expressed as either aggressive inhibition of ovarian activity among workers or destruction of worker–laid eggs. A few studies have shown that workers with developed ovaries are preferentially attacked by nest–mates, but adequate demonstration of worker policing also requires evidence that these attacks result in the suppression of ovarian activity or death. We investigated worker policing in the ponerine ant Harpegnathos saltator in which workers are able to mate and replace the founding queen. Five colonies were each divided into two groups, one of which consisted exclusively of infertile workers. Some individuals in the orphaned groups began laying eggs during the three–week separation and upon reunification these were vigorously attacked by infertile workers of the other groups. The ovarian activity of these new egg layers became inhibited, as revealed by subsequent dissection of marked individuals. Worker policing in H. saltator appears to function primarily in preventing an excess of reproductive workers.

Cuticular Hydrocarbons Reliably Identify Cheaters and Allow Enforcement of Altruism in a Social Insect

Cheaters are a threat to every society and therefore societies have established rules to punish these individuals in order to stabilize their social system. Recent models and observations suggest that enforcement of reproductive altruism (policing) in hymenopteran insect societies is a major force in maintaining high levels of cooperation. In order to be able to enforce altruism, reproductive cheaters need to be reliably identified. Strong correlational evidence indicates that cuticular hydrocarbons are the means of identifying cheaters, but direct proof is still missing. In the ant Aphaenogaster cockerelli, we mimicked reproductive cheaters by applying a synthetic compound typical of fertile individuals on nonreproductive workers. This treatment induced nestmate aggression in colonies where a queen was present. As expected, it failed to do so in colonies without a queen where workers had begun to reproduce. This provides the first direct evidence that cuticular hydrocarbons are the informational basis of policing behaviors, serving a major function in the regulation of reproduction in social insects. We suggest that even though cheaters would gain from suppressing these profiles, they are prevented from doing so through the mechanisms of hydrocarbon biosynthesis and its relation to reproductive physiology. Cheaters are identified through information that is inherently reliable.

Phylogenetic and Transcriptomic Analysis of Chemosensory Receptors in a Pair of Divergent Ant Species Reveals Sex-Specific Signatures of Odor Coding

Ants are a highly successful family of insects that thrive in a variety of habitats across the world. Perhaps their best-known features are complex social organization and strict division of labor, separating reproduction from the day-to-day maintenance and care of the colony, as well as strict discrimination against foreign individuals. Since these social characteristics in ants are thought to be mediated by semiochemicals, a thorough analysis of these signals, and the receptors that detect them, is critical in revealing mechanisms that lead to stereotypic behaviors. To address these questions, we have defined and characterized the major chemoreceptor families in a pair of behaviorally and evolutionarily distinct ant species, Camponotus floridanus and Harpegnathos saltator. Through comprehensive re-annotation, we show that these ant species harbor some of the largest yet known repertoires of odorant receptors (Ors) among insects, as well as a more modest number of gustatory receptors (Grs) and variant ionotropic glutamate receptors (Irs). Our phylogenetic analyses further demonstrate remarkably rapid gains and losses of ant Ors, while Grs and Irs have also experienced birth-and-death evolution to different degrees. In addition, comparisons of antennal transcriptomes between sexes identify many chemoreceptors that are differentially expressed between males and females and between species. We have also revealed an agonist for a worker-enriched OR from C. floridanus, representing the first case of a heterologously characterized ant tuning Or. Collectively, our analysis reveals a large number of ant chemoreceptors exhibiting patterns of differential expression and evolution consistent with sex/species-specific functions. These differentially expressed genes are likely associated with sex-based differences, as well as the radically different social lifestyles observed between C. floridanus and H. saltator, and thus are targets for further functional characterization. Our findings represent an important advance toward understanding the molecular basis of social interactions and the differential chemical ecologies among ant species.

Queen fertility, egg marking and colony size in the ant Camponotus floridanus

In ant societies, workers do not usually reproduce but gain indirect fitness benefits from raising related offspring produced by the queen. One of the preconditions of this worker self-restraint is sufficient fertility of the queen. The queen is, therefore, expected to signal her fertility. In Camponotus floridanus, workers can recognize the presence of a highly fertile queen via her eggs, which are marked with the queens specific hydrocarbon profile. If information on fertility is encoded in the hydrocarbon profile of eggs, we expect workers to be able to differentiate between eggs from highly and weakly fertile queens. We found that workers discriminate between these eggs solely on the basis of their hydrocarbon profiles which differ both qualitatively and quantitatively. This pattern is further supported by the similarity of the egg profiles of workers and weakly fertile queens and the similar treatment of both kinds of eggs. Profiles of queen eggs correspond to the cuticular hydrocarbon profiles of the respective queens. Changes in the cuticular profiles are associated with the size of the colony the queen originates from and her current egg-laying rate. However, partial correlation analysis indicates that only colony size predicts the cuticular profile. Colony size is a buffered indicator of queen fertility as it is a consequence of queen productivity within a certain period of time, whereas daily egg-laying rate varies due to cyclical oviposition. We conclude that surface hydrocarbons of eggs and the cuticular profiles of queens both signal queen fertility, suggesting a major role of fertility signals in the regulation of reproduction in social insects.

Eusocial insects as emerging models for behavioural epigenetics

Understanding the molecular basis of how behavioural states are established, maintained and altered by environmental cues is an area of considerable and growing interest. Epigenetic processes, including methylation of DNA and post-translational modification of histones, dynamically modulate activity-dependent gene expression in neurons and can therefore have important regulatory roles in shaping behavioural responses to environmental cues. Several eusocial insect species — with their unique displays of behavioural plasticity due to age, morphology and social context — have emerged as models to investigate the genetic and epigenetic underpinnings of animal social behaviour. This Review summarizes recent studies in the epigenetics of social behaviour and offers perspectives on emerging trends and prospects for establishing genetic tools in eusocial insects.

Direct assessment of queen quality and lack of worker suppression in a paper wasp

Assessing a conspecifics potential is often crucial to increase ones fitness, e.g. in female choice, contests with rivals or reproductive conflicts in animal societies. In the latter, helpers benefit from accurately assessing the fertility of the breeder as an indication of inclusive fitness. There is evidence that this can be achieved using chemical correlates of reproductive activity. Here, we show that queen quality can be assessed by directly monitoring her reproductive output. In the paper wasp Polistes dominulus, we mimicked a decrease in queen fertility by regularly removing brood. This triggered ovarian development and egg-laying by many workers, which strongly suggests that brood abundance is a reliable cue of queen quality. Brood abundance can be monitored when workers perform regular brood care in small size societies where each brood element is kept in a separate cell. Our results also show that although the queen was not manipulated, and thus remained healthy and fully fertile, she did not control worker egg-laying. Nevertheless, when workers laid eggs, the queen secured a near reproductive monopoly by selectively destroying these eggs, a mechanism known as ‘queen policing’. By contrast, workers destroyed comparatively few queen-laid eggs, but did destroy each others eggs.