Social antagonism facilitates supergene expansion in ants.

Antagonistic selection has long been considered a major driver of the formation and expansion of sex chromosomes. For example, sexually antagonistic variation on an autosome can select for suppressed recombination between that autosome and the sex chromosome, leading to a neo-sex chromosome. Autosomal supergenes, chromosomal regions containing tightly linked variants affecting the same complex trait, share similarities with sex chromosomes, raising the possibility that sex chromosome evolution models can explain the evolution of genome structure and recombination in other contexts. We tested this premise in a Formica ant species, wherein we identified four supergene haplotypes on chromosome 3 underlying colony social organization and sex ratio. We discovered a novel rearranged supergene variant (9r) on chromosome 9 underlying queen miniaturization. The 9r is in strong linkage disequilibrium with one chromosome 3 haplotype (P2) found in multi-queen (polygyne) colonies. We suggest that queen miniaturization is strongly disfavored in the single-queen (monogyne) background and is thus socially antagonistic. As such, divergent selection experienced by ants living in alternative social "environments" (monogyne and polygyne) may have contributed to the emergence of a genetic polymorphism on chromosome 9 and associated queen-size dimorphism. Consequently, an ancestral polygyne-associated haplotype may have expanded to include the polymorphism on chromosome 9, resulting in a larger region of suppressed recombination spanning two chromosomes. This process is analogous to the formation of neo-sex chromosomes and consistent with models of expanding regions of suppressed recombination. We propose that miniaturized queens, 16%-20% smaller than queens without 9r, could be incipient intraspecific social parasites.

Unequal Reproduction Early in a Social Transition: Insights from Invasive Wasps.

AbstractIn eusocial insects, nestmate queens can differ in their reproductive output, causing asymmetries in the distribution of mutual benefits. However, little is known about how reproductive success is partitioned in incipiently polygynous species, which would provide clues about the evolutionary forces shaping the emergence of polygyny. Here, we leverage a recent transition from predominantly single-queen (monogyne) to multiple-queen (polygyne) colonies in an invasive yellowjacket species to investigate whether queens in incipiently polygyne colonies invest equally in reproductive effort or vary in their relative investment in each caste. We excavated nine polygyne Vespula pensylvanica colonies in Hawaii and used restriction site-associated DNA sequencing to infer the parentage of worker, male, and gyne (daughter queen) pupae from each nest comb. In four colonies with at least eight gyne pupae, a single queen produced most or all gynes. These queens had no male offspring and few worker offspring, suggesting that a subset of nestmate queens might exploit the collective benefits of newly polygyne societies. In contrast to most queens, gyne producers had offspring distributed nonrandomly across nest combs. Nestmate queens generally exhibited low relatedness levels. Our results suggest that rapid, ecologically driven transitions to polygyny among unrelated queens may, at their onset, be vulnerable to reproductive asymmetries that are likely evolutionarily unstable. More broadly, this study contributes to the understanding of social evolution by uncovering asymmetric partitioning of reproduction in a population with newly evolved polygyny and raises questions about the future trajectories of introduced populations.

Species recognition limits mating between hybridizing ant species.

Identifying mechanisms limiting hybridization is a central goal of speciation research. Here, we studied premating and postmating barriers to hybridization between two ant species, Formica selysi and Formica cinerea. These species hybridize in the Rhône valley in Switzerland, where they form a mosaic hybrid zone, with limited introgression from F. selysi into F. cinerea. There was no sign of temporal isolation between the two species in the production of queens and males. With choice experiments, we showed that queens and males strongly prefer to mate with conspecifics. Yet, we did not detect postmating barriers caused by genetic incompatibilities. Specifically, hybrids of all sexes and castes were found in the field and F1 hybrid workers did not show reduced viability compared to nonhybrid workers. To gain insights into the cues involved in species recognition, we analyzed the cuticular hydrocarbons (CHCs) of queens, males, and workers and staged dyadic encounters between workers. CHC profiles differed markedly between species, but were similar in F. cinerea and hybrids. Accordingly, workers also discriminated species, but they did not discriminate F. cinerea and hybrids. We discuss how the CHC-based recognition system of ants may facilitate the establishment of premating barriers to hybridization, independent of hybridization costs.

Muscle Protein Synthesis Responses Following Aerobic-Based Exercise or High-Intensity Interval Training with or Without Protein Ingestion: A Systematic Review.

BACKGROUND: Systematic investigation of muscle protein synthesis (MPS) responses with or without protein ingestion has been largely limited to resistance training. OBJECTIVE: This systematic review determined the capacity for aerobic-based exercise or high-intensity interval training (HIIT) to stimulate post-exercise rates of MPS and whether protein ingestion further significantly increases MPS compared with placebo. METHODS: Three separate models analysed rates of either mixed, myofibrillar, sarcoplasmic, or mitochondrial protein synthesis (PS) following aerobic-based exercise or HIIT: Model 1 (n = 9 studies), no protein ingestion; Model 2 (n = 7 studies), peri-exercise protein ingestion with no placebo comparison; Model 3 (n = 14 studies), peri-exercise protein ingestion with placebo comparison. RESULTS: Eight of nine studies and all seven studies in Models 1 and 2, respectively, demonstrated significant post-exercise increases in either mixed or a specific muscle protein pool. Model 3 observed significantly greater MPS responses with protein compared with placebo in either mixed or a specific muscle fraction in 7 of 14 studies. Seven studies showed no difference in MPS between protein and placebo, while three studies reported no significant increases in mitochondrial PS with protein compared with placebo. CONCLUSION: Most studies reporting significant increases in MPS were confined to mixed and myofibrillar PS that may facilitate power generating capacity of working skeletal muscle with aerobic-based exercise and HIIT. Only three of eight studies demonstrated significant increases in mitochondrial PS post-exercise, with no further benefits of protein ingestion. This lack of change may be explained by the acute analysis window in most studies and apparent latency in exercise-induced stimulation of mitochondrial PS.

A socially polymorphic Formica ant species exhibits a novel distribution of social supergene genotypes.

Most supergenes discovered so far are young, occurring in one species or a few closely related species. An ancient supergene in the ant genus Formica presents an unusual opportunity to compare supergene-associated phenotypes and the factors that influence the persistence of polymorphism in different species. We investigate the genetic architecture of social organization in Formica francoeuri, an ant species native to low- and mid-elevation semiarid regions of southern California, and describe an efficient technique for estimating mode of social organization using population genomic data. Using this technique, we show that F. francoeuri exhibits polymorphism in colony social organization and that the phenotypic polymorphism is strongly associated with genotypes within the Formica social supergene region. The distribution of supergene haplotypes in F. francoeuri differs from that of related species Formica selysi in that colonies with multiple queens contain almost exclusively workers that are heterozygous for alternative supergene haplotypes. Moreover, heterozygous workers exhibit allele-specific expression of the polygyne-associated haplotype at the candidate gene Knockout, which is thought to influence social organization. We also report geographic population structure and variation in worker size across a large fraction of the species range. Our results suggest that, although the Formica supergene is conserved within the genus, the mechanisms that maintain the supergene and its associated polymorphisms differ among species.

Effects of social organization and elevation on spatial genetic structure in a montane ant.

Studying patterns of population structure across the landscape sheds light on dispersal and demographic processes, which helps to inform conservation decisions. Here, we study how social organization and landscape factors affect spatial patterns of genetic differentiation in an ant species living in mountainous regions. Using genome-wide SNP markers, we assess population structure in the Alpine silver ant, Formica selysi. This species has two social forms controlled by a supergene. The monogyne form has one queen per colony, while the polygyne form has multiple queens per colony. The two social forms co-occur in the same populations. For both social forms, we found a strong pattern of isolation-by-distance across the Alps. Within regions, genetic differentiation between populations was weaker for the monogyne form than for the polygyne form. We suggest that this pattern is due to higher dispersal and effective population sizes in the monogyne form. In addition, we found stronger isolation-by-distance and lower genetic diversity in high elevation populations, compared to lowland populations, suggesting that gene flow between F. selysi populations in the Alps occurs mostly through riparian corridors along lowland valleys. Overall, this survey highlights the need to consider intraspecific polymorphisms when assessing population connectivity and calls for special attention to the conservation of lowland habitats in mountain regions.

A novel distribution of supergene genotypes is present in the socially polymorphic ant Formica neoclara.

BACKGROUND: Supergenes are chromosomal regions with tightly linked clusters of alleles that control compound phenotypic traits. Supergenes have been demonstrated to contribute to the maintenance of polymorphisms within populations in traits as diverse as mimetic wing coloration in butterflies, mating strategies in birds, and malarial susceptibility in mosquitoes. A large supergene also underlies variation in social organization in Formica ants. Alternative supergene haplotypes are associated with the presence of either a single queen (monogyny) or multiple queens (polygyny) within colonies. Here, we assess the social structure and supergene status of the North American species Formica neoclara. RESULTS: We sequenced a subset of the genome in 280 individuals sampled in populations from California to northern British Columbia using ddRADseq. We determined that F. neoclara is socially polymorphic in queen number, and we show that the social polymorphism is associated with alternative haplotypes at the social supergene. Intriguingly, polygyne colonies can harbor workers that are homozygous for both haplotypes as well as heterozygotes. CONCLUSIONS: This colony genetic composition contrasts with other Formica species, in which almost all individuals in polygyne colonies have the polygyne-associated haplotype. The social polymorphism is present in widely distributed and genetically subdivided populations of F. neoclara. In studying this system in F. neoclara, we expand our understanding of the functional evolution of supergene haplotypes as they diverge in different lineages.

Early queen joining and long-term queen associations in polygyne colonies of an invasive wasp revealed by longitudinal genetic analysis.

Invasive social insects rank among the most damaging of terrestrial species. They are responsible for extensive damage and severely threaten the biodiversity of environments where they are introduced. Variation in colony social form commonly occurs in introduced populations of yellowjacket wasps (genus Vespula). In particular, invasive colonies may contain multiple queens (i.e., polygyne) and persist several years, while in the native range, the colonies are usually annual and harbor a single queen (i.e., monogyne). In this study, we used genome-wide loci obtained by double digest restriction site-associated DNA sequencing (RADseq) to investigate the genetic structure and queen turnover in colonies of the western yellowjacket, Vespula pensylvanica, in their introduced range in Hawaii. Of the 27 colonies monitored over four months (October-January), 19 were polygyne and already contained multiple queens on the first day of sampling. Contrary to previous speculation, this finding suggests that polygyny often arises early in the annual colony cycle, before the production of new queens in the fall. Furthermore, polygyne colonies exhibited a prolonged average lifespan relative to those headed by a single queen. As a result, there is no clear window during which colony eradication efforts would be more effective than upon first discovery. The relatedness among nestmate queens was slightly above zero, indicating that these colonies are generally composed of nonrelatives. The queen turnover within each colony was low, and we detected some full-sibling workers sampled up to four months apart. Finally, we did not detect any population structure among colonies, suggesting that queens disperse up to several kilometers. Taken together, our results provide the first insights into the requeening dynamics in this invasive and incipiently polygyne population and illuminate the early establishment of multiple long-lasting queens in these damaging colonies.

Ant nest architecture is shaped by local adaptation and plastic response to temperature.

Social insects are among the most abundant arthropods in terrestrial ecosystems, where they provide ecosystem services. The effect of subterranean activity of ants on soil is well-studied, yet little is known about nest architecture due to the difficulty of observing belowground patterns. Furthermore, many species' ranges span environmental gradients, and their nest architecture is likely shaped by the climatic and landscape features of their specific habitats. We investigated the effects of two temperature treatments on the shape and size of nests built by Formica podzolica ants collected from high and low elevations in the Colorado Rocky Mountains in a full factorial experiment. Ants nested in experimental chambers with soil surface temperatures matching the local temperatures of sample sites. We observed a plastic response of nest architecture to conditions experienced during excavation; workers experiencing a high temperature excavated deeper nests than those experiencing a cooler temperature. Further, we found evidence of local adaptation to temperature, with a significant interaction effect of natal elevation and temperature treatment on nest size and complexity. Specifically, workers from high elevation sites built larger nests with more tunnels when placed in the cool surface temperature treatment, and workers from low elevation sites exhibited the opposite pattern. Our results suggest that subterranean ant nest architecture is shaped by a combination of plastic and locally adapted building behaviors; we suggest that the flexibility of this 'extended phenotype' likely contributes to the widespread success of ants.

Linked supergenes underlie split sex ratio and social organization in an ant.

Sexually reproducing organisms usually invest equally in male and female offspring. Deviations from this pattern have led researchers to new discoveries in the study of parent-offspring conflict, genomic conflict, and cooperative breeding. Some social insect species exhibit the unusual population-level pattern of split sex ratio, wherein some colonies specialize in the production of future queens and others specialize in the production of males. Theoretical work predicted that worker control of sex ratio and variation in relatedness asymmetry among colonies would cause each colony to specialize in the production of one sex. While some empirical tests supported theoretical predictions, others deviated from them, leaving many questions about how split sex ratio emerges. One factor yet to be investigated is whether colony sex ratio may be influenced by the genotypes of queens or workers. Here, we sequence the genomes of 138 Formica glacialis workers from 34 male-producing and 34 gyne-producing colonies to determine whether split sex ratio is under genetic control. We identify a supergene spanning 5.5 Mbp that is closely associated with sex allocation in this system. Strikingly, this supergene is adjacent to another supergene spanning 5 Mbp that is associated with variation in colony queen number. We identify a similar pattern in a second related species, Formica podzolica. The discovery that split sex ratio is determined, at least in part, by a supergene in two species opens future research on the evolutionary drivers of split sex ratio.

The maintenance of polymorphism in an ancient social supergene.

Supergenes, regions of the genome with suppressed recombination between sets of functional mutations, contribute to the evolution of complex phenotypes in diverse systems. Excluding sex chromosomes, most supergenes discovered so far appear to be young, being found in one species or a few closely related species. Here, we investigate how a chromosome harbouring an ancient supergene has evolved over about 30 million years (Ma). The Formica supergene underlies variation in colony queen number in at least five species. We expand previous analyses of sequence divergence on this chromosome to encompass about 90 species spanning the Formica phylogeny. Within the nonrecombining region, the gene knockout contains 22 single nucleotide polymorphisms (SNPs) that are consistently differentiated between two alternative supergene haplotypes in divergent European Formica species, and we show that these same SNPs are present in most Formica clades. In these clades, including an early diverging Nearctic Formica clade, individuals with alternative genotypes at knockout also have higher differentiation in other portions of this chromosome. We identify hotspots of SNPs along this chromosome that are present in multiple Formica clades to detect genes that may have contributed to the emergence and maintenance of the genetic polymorphism. Finally, we infer three gene duplications on one haplotype, based on apparent heterozygosity within these genes in the genomes of haploid males. This study strengthens the evidence that this supergene originated early in the evolution of Formica and that just a few loci in this large region of suppressed recombination retain strongly differentiated alleles across contemporary Formica lineages.

Evolution of specialization in a plant-microbial mutualism is explained by the oscillation theory of speciation.

Specialization in mutualisms is thought to be a major driver of diversification, but few studies have explored how novel specialization evolves, or its relation to the evolution of other niche axes. A fundamental question is whether generalist interactions evolve to become more specialized (i.e., oscillation hypothesis) or if partner switches evolve without any change in niche breadth (i.e., musical chairs hypothesis). We examined alternative models for the evolution of specialization by estimating the mutualistic, climatic, and edaphic niche breadths of sister plant species, combining phylogenetic, environmental, and experimental data on Acmispon strigosus and Acmispon wrangelianus genotypes across their overlapping ranges in California. We found that specialization along all three niche axes was asymmetric across species, such that the species with broader climatic and edaphic niches, Acmispon strigosus, was also able to gain benefit from and invest in associating with a broader set of microbial mutualists. Our data are consistent with the oscillation model of specialization, and a parallel narrowing of the edaphic, climatic, and mutualistic dimensions of the host species niche. Our findings provide novel evidence that the evolution of specialization in mutualism is accompanied by specialization in other niche dimensions.

High-Quality Assemblies for Three Invasive Social Wasps from the Vespula Genus.

Social wasps of the genus Vespula have spread to nearly all landmasses worldwide and have become significant pests in their introduced ranges, affecting economies and biodiversity. Comprehensive genome assemblies and annotations for these species are required to develop the next generation of control strategies and monitor existing chemical control. We sequenced and annotated the genomes of the common wasp (Vespula vulgaris), German wasp (Vespula germanica), and the western yellowjacket (Vespula pensylvanica). Our chromosome-level Vespula assemblies each contain 176-179 Mb of total sequence assembled into 25 scaffolds, with 10-200 unanchored scaffolds, and 16,566-18,948 genes. We annotated gene sets relevant to the applied management of invasive wasp populations, including genes associated with spermatogenesis and development, pesticide resistance, olfactory receptors, immunity and venom. These genomes provide evidence for active DNA methylation in Vespidae and tandem duplications of venom genes. Our genomic resources will contribute to the development of next-generation control strategies, and monitoring potential resistance to chemical control.

Cas13b-dependent and Cas13b-independent RNA knockdown of viral sequences in mosquito cells following guide RNA expression.

Aedes aegypti and Aedes albopictus mosquitoes are vectors of the RNA viruses chikungunya (CHIKV) and dengue that currently have no specific therapeutic treatments. The development of new methods to generate virus-refractory mosquitoes would be beneficial. Cas13b is an enzyme that uses RNA guides to target and cleave RNA molecules and has been reported to suppress RNA viruses in mammalian and plant cells. We investigated the potential use of the Prevotella sp. P5-125 Cas13b system to provide viral refractoriness in mosquito cells, using a virus-derived reporter and a CHIKV split replication system. Cas13b in combination with suitable guide RNAs could induce strong suppression of virus-derived reporter RNAs in insect cells. Surprisingly, the RNA guides alone (without Cas13b) also gave substantial suppression. Our study provides support for the potential use of Cas13b in mosquitoes, but also caution in interpreting CRISPR/Cas data as we show that guide RNAs can have Cas-independent effects.

Maternal effect killing by a supergene controlling ant social organization.

Supergenes underlie striking polymorphisms in nature, yet the evolutionary mechanisms by which they arise and persist remain enigmatic. These clusters of linked loci can spread in populations because they captured coadapted alleles or by selfishly distorting the laws of Mendelian inheritance. Here, we show that the supergene haplotype associated with multiple-queen colonies in Alpine silver ants is a maternal effect killer. All eggs from heterozygous queens failed to hatch when they did not inherit this haplotype. Hence, the haplotype specific to multiple-queen colonies is a selfish genetic element that enhances its own transmission by causing developmental arrest of progeny that do not carry it. At the population level, such transmission ratio distortion favors the spread of multiple-queen colonies, to the detriment of the alternative haplotype associated with single-queen colonies. Hence, selfish gene drive by one haplotype will impact the evolutionary dynamics of alternative forms of colony social organization. This killer hidden in a social supergene shows that large nonrecombining genomic regions are prone to cause multifarious effects across levels of biological organization.

Expanding the CRISPR Toolbox in Culicine Mosquitoes: In Vitro Validation of Pol III Promoters.

CRISPR-Cas9-based "gene drive" technologies have been proposed as a novel and effective means of controlling human diseases vectored by mosquitoes. However, more complex designs than those demonstrated to date-and an expanded molecular toolbox with which to build them-will be required to overcome the issues of resistance formation/evolution and drive spatial/temporal limitation. Foreseeing this need, we assessed the sgRNA transcriptional activities of 33 phylogenetically diverse insect Polymerase III promoters using three disease-relevant Culicine mosquito cell lines (Aedes aegypti, Aedes albopictus, and Culex quinquefasciatus). We show that U6 promoters work across species with a range of transcriptional activity levels and find 7SK promoters to be especially promising because of their broad phylogenetic activity. We further show that U6 promoters can be substantially truncated without affecting transcriptional levels. These results will be of great utility to researchers involved in developing the next generation of gene drives.

An Ancient and Eroded Social Supergene Is Widespread across Formica Ants.

Supergenes, clusters of tightly linked genes, play a key role in the evolution of complex adaptive variation [1, 2]. Although supergenes have been identified in many species, we lack an understanding of their origin, evolution, and persistence [3]. Here, we uncover 20-40 Ma of evolutionary history of a supergene associated with polymorphic social organization in Formica ants [4]. We show that five Formica species exhibit homologous divergent haplotypes spanning 11 Mbp on chromosome 3. Despite the supergene's size, only 142 single nucleotide polymorphisms (SNPs) consistently distinguish alternative supergene haplotypes across all five species. These conserved trans-species SNPs are localized in a small number of disjunct clusters distributed across the supergene. This unexpected pattern of divergence indicates that the Formica supergene does not follow standard models of sex chromosome evolution, in which distinct evolutionary strata reflect an expanding region of suppressed recombination [5]. We propose an alternative "eroded strata model" in which clusters of conserved trans-species SNPs represent functionally important areas maintained by selection in the face of rare recombination between ancestral haplotypes. The comparison of whole-genome sequences across 10 additional Formica species reveals that the most conserved region of the supergene contains a transcription factor essential for motor neuron development in Drosophila [6]. The discovery that a very small portion of this large and ancient supergene harbors conserved trans-species SNPs linked to colony social organization suggests that the ancestral haplotypes have been eroded by recombination, with selection preserving differentiation at one or a few genes generating alternative social organization.

Are personalities genetically determined? Inferences from subsocial spiders.

BACKGROUND: Recent research has revealed that polymorphic behavioral strategies shape intra-and interspecific interactions and contribute to fitness in many animal species. A better understanding of the proximate mechanisms underlying these behavioral syndromes will enhance our grasp this phenomenon. Spiders in the genus Anelosimus exhibit inter-individual behavioral variation on several axes: individuals have consistent responses to stimuli (e.g. bold vs. shy individuals) and they are subsocial (exhibiting extended maternal care and sibling cooperation) across most of their range, but they sometimes form permanent social groups in northern temperate regions. Here, we seek genetic variants associated with boldness and with social structure in a socially polymorphic population of the spider Anelosimus studiosus. We also develop preliminary genomic resources, including a genome assembly and linkage map, that support this and future genomic research on this group. RESULTS: Remarkably, we identify a small genomic scaffold (~ 1200 bp) that harbors seven single nucleotide polymorphisms (SNPs) associated with boldness. Moreover, heterozygotes are less common than expected based on Hardy-Weinberg equilibrium, suggesting that either assortative mating or selection against heterozygotes may be occurring in this system. We find no loci significantly associated with social organization. Our draft genome assembly allows us to localize SNPs of interest in this study and to carry out genetic comparisons with other published genomes, although it remains highly fragmented. CONCLUSIONS: By identifying a locus associated with a well-studied animal personality trait, this study opens up avenues for future research to link behavioral studies of animal personality with genotype and fitness.

Asymmetric assortative mating and queen polyandry are linked to a supergene controlling ant social organization.

Nonrecombining genomic variants underlie spectacular social polymorphisms, from bird mating systems to ant social organization. Because these "social supergenes" affect multiple phenotypic traits linked to survival and reproduction, explaining their persistence remains a substantial challenge. Here, we investigate how large nonrecombining genomic variants relate to colony social organization, mating system and dispersal in the Alpine silver ant, Formica selysi. The species has colonies headed by a single queen (monogynous) and colonies headed by multiple queens (polygynous). We confirmed that a supergene with alternate haplotypes-Sm and Sp-underlies this polymorphism in social structure: Females from mature monogynous colonies had the Sm/Sm genotype, while those from polygynous colonies were Sm/Sp and Sp/Sp. Queens heading monogynous colonies were exclusively mated with Sm males. In contrast, queens heading polygynous colonies were mated with Sp males and Sm males. Sm males, which are only produced by monogynous colonies, accounted for 22.9% of the matings with queens from mature polygynous colonies. This asymmetry between social forms in the degree of assortative mating generates unidirectional male-mediated gene flow from the monogynous to the polygynous social form. Biased gene flow was confirmed by a significantly higher number of private alleles in the polygynous social form. Moreover, heterozygous queens were three times as likely as homozygous queens to be multiply mated. This study reveals that the supergene variants jointly affect social organization and multiple components of the mating system that alter the transmission of the variants and thus influence the dynamics of the system.