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Our understanding of the gene regulatory network that constitutes the circadian clock has greatly increased in recent decades, notably due to the use of Drosophila as a model system. In contrast, the analysis of natural genetic variation that enables the robust function of the clock under a broad range of environments has developed more slowly. In the current study, we analyzed comprehensive genome sequencing data from wild European populations of Drosophila, which were densely sampled through time and space. We identified hundreds of single nucleotide polymorphisms (SNPs) in nine genes associated with the clock, 276 of which exhibited a latitudinal cline in their allele frequencies. While the effect sizes of these clinal patterns were small, indicating subtle adaptations driven by natural selection, they provided important insights into the genetic dynamics of circadian rhythms in natural populations. We selected nine SNPs in different genes and assessed their impact on circadian and seasonal phenotypes by reconstructing outbred populations fixed for either of the SNP alleles, from inbred DGRP strains. The circadian free-running period of the locomotor activity rhythm was affected by an SNP in doubletime (dbt) and eyes absent (Eya). The SNPs in Clock (Clk), Shaggy (Sgg), period (per), and timeless (tim) affected the acrophase. The alleles of the SNP in Eya conferred different levels of diapause and the chill coma recovery response.

Since the pioneering work of Dobzhansky in the 1930s and 1940s, many chromosomal inversions have been identified but how they contribute to adaptation remains poorly understood. InDrosophila melanogaster, the widespread inversion polymorphismIn(3R)Payneunderpins latitudinal clines in fitness traits on multiple continents. Here, we use single-individual whole-genome sequencing, transcriptomics and published sequencing data to study the population genomics of this inversion on four continents: in its ancestral African range and in derived populations in Europe, North America, and Australia. Our results confirm that this inversion originated in sub-Saharan Africa and subsequently became cosmopolitan; we observe marked monophyletic divergence of inverted and non-inverted karyotypes, with some substructure among inverted chromosomes between continents. Despite divergent evolution of this inversion since its out-of-Africa migration, derived non-African populations exhibit similar patterns of long-range linkage disequilibrium between the inversion breakpoints and major peaks of divergence in its center, consistent with balancing selection and suggesting that the inversion harbors alleles that are maintained by selection on several continents. Using RNA-seq we identify overlap between inversion-linked SNPs and loci that are differentially expressed between inverted and non-inverted chromosomes. Expression levels are higher for inverted chromosomes at low temperature, suggesting loss of buffering or compensatory plasticity and consistent with higher inversion frequency in warm climates. Our results suggest that this ancestrally tropical balanced polymorphism spread around the world and became latitudinally assorted along similar but independent climatic gradients, always being frequent in subtropical/tropical areas but rare or absent in temperate climates.

Temperature fluctuations are challenging for ectotherms which are not able to regulate body temperature by physiological means and thus have to adjust their thermal environment via behavior. However, little is yet known about whether microbial symbionts influence thermal preference (Tp) in ectotherms by modulating their physiology. Several recent studies have demonstrated substantial effects ofWolbachiainfections on hostTpin differentDrosophilaspecies. These data indicate that the direction and strength of thermal preference variation is strongly depend on host and symbiont genotypes and highly variable among studies. By employing highly controlled experiments, we investigated the impact of several environmental factors including humidity, food quality, light exposure, and experimental setup that may influenceTpmeasurements in adult flies. Additionally, we assessed the effects ofWolbachiainfection onTpofDrosophilaat different developmental stages, which has not been done before. We find no effects ofWolbachiaon hostTpat any life stage. Instead, we show that environmental variation has a substantial influence onTpmeasurements which demonstrates the necessity of careful experimental design and cautious interpretations ofTpmeasurements together with a thorough description of the methods and equipment used to conduct behavioral studies.

Wolbachia bacteria are common endosymbionts of many arthropods found in gonads and various somatic tissues. They manipulate host reproduction to enhance their transmission and confer complex effects on fitness-related traits. Some of these effects can serve to increase the survival and transmission efficiency of Wolbachia in the host population. The Wolbachia-Drosophila melanogaster system represents a powerful model to study the evolutionary dynamics of host-microbe interactions and infections. Over the past decades, there has been a replacement of the ancestral wMelCS Wolbachia variant by the more recent wMel variant in worldwide D. melanogaster populations, but the reasons remain unknown. To investigate how environmental change and genetic variation of the symbiont affect host developmental and adult life-history traits, we compared effects of both Wolbachia variants and uninfected controls in wild-caught D. melanogaster strains at three developmental temperatures. While Wolbachia did not influence any developmental life-history traits, we found that both lifespan and fecundity of host females were increased without apparent fitness trade-offs. Interestingly, wMelCS-infected flies were more fecund than uninfected and wMel-infected flies. By contrast, males infected with wMel died sooner, indicating sex-specific effects of infection that are specific to the Wolbachia variant. Our study uncovered complex temperature-specific effects of Wolbachia infections, which suggests that symbiont-host interactions in nature are strongly dependent on the genotypes of both partners and the thermal environment.

The insulin insulin-like growth factor signaling pathway has been hypothesized as a major determinant of life history profiles that vary adaptively in natural populations. InDrosophila melanogaster, multiple components of this pathway vary predictably with latitude; this includesfoxo, a conserved gene that regulates insulin signaling and has pleiotropic effects on a variety of fitness-associated traits. We hypothesized that allelic variation atfoxounderlies genetic variance for traits that vary with latitude and reflect local adaptation. To evaluate this, we generated recombinant outbred populations in which the focalfoxoallele was homozygous and fixed for either the allele common at high latitude or low latitude and the genomic background was randomized across 20 inbred lines. After eight generations of recombination, experimental populations were phenotyped for a series of traits related to gene function. Our results demonstrate that natural allelic variation atfoxohas major and predictable effects on body size and starvation tolerance, but not on development time. These patterns mirror those observed in natural populations collected across the latitudinal gradient in the eastern U.S.: clines were observed for starvation tolerance and body size, but development time exhibited no association with latitude. Furthermore, differences in size betweenfoxogenotypes were equivalent to those observed between populations sampled from the latitudinal extremes, although contribution to the genetic variance for starvation tolerance was less pronounced. These results suggest that allelic variation atfoxois a major contributor to adaptive patterns of life history variation in natural populations of this genetic model.

Zones of secondary contact provide a good opportunity to investigate the origin and dynamics of reproductive isolation between related populations. We analyzed genetic and phenotypic patterns and gene fow between two subspecies of the Eurasian magpie Pica pica s.l. which recently came into contact after presumably long periods of isolation. We describe the distribution of subspecies in a young contact zone at Argun’ river basin in southern Siberia where populations occur in parapatry and an older hybrid population in eastern Mongolia. Based on genome-wide SNP data, we analyzed patterns and strength of gene fow between the subspecies. Our results indicate occasional hybridization with backcrossing and asymmetric introgression along a wide range in Transbaikalia and locally in eastern Mongolia. Males of P. p. jankowskii apparently exhibit higher dispersal ability towards the west compared to P. p. leucoptera (towards the east). The former occasionally migrates to eastern Mongolia and Transbaikalia where introgression of nuclear, but not mitochondrial DNA was evident. Bioacoustic investigations showed diferences between the subspecies in speed and structure of vocalization. We discovered intermediate calls of hybrid magpies and bilingual birds alternating calls that are typical for the two taxa. Furthermore, we found dramatically decreased reproductive success in hybridogeneous populations. By complementing our results with established phylogeographic patterns of P. pica s.l. based on a mitochondrial marker sequence, and considering indications of sterility of hybrids in the contact zone, we propose to elevate the two corresponding subspecies to species level: P. pica for the western form and P. serica for the eastern form., Open access funding provided by University of Vienna.

Interspecific gene flow by hybridization may weaken species barriers and adaptive divergence, but can also initiate reinforcement of reproductive isolation trough natural and sexual selection. The extent of interspecific gene flow and its consequences for the initiation and maintenance of species barriers in natural systems remain poorly understood, however. To assess genome-wide patterns of gene flow between the two closely related European dung fly species Sepsis cynipsea and Sepsis neocynipsea (Diptera: Sepsidae), we tested for historical gene flow with the aid of ABBA-BABA test using whole-genome resequencing data from pooled DNA of male specimens originating from natural and laboratory populations. We contrasted genome-wide variation in DNA sequence differences between samples from sympatric populations of the two species in France and Switzerland with that of interspecific differences between pairs of samples involving allopatric populations from Estonia and Italy. In the French Cevennes, we detected a relative excess of DNA sequence identity, suggesting interspecific gene flow in sympatry. In contrast, at two sites in Switzerland, we observed a relative depletion of DNA sequence identity compatible with reinforcement of species boundaries in sympatry. Our results suggest that the species boundaries between S. cynipsea and S. neocynipsea in Europe depend on the eco-geographic context.

Periods of nutrient shortage impose strong selection on animal populations. Experimental studies of genetic adaptation to nutrient shortage largely focus on resistance to acute starvation at adult stage; it is not clear how conclusions drawn from these studies extrapolate to other forms of nutritional stress. We studied the genomic signature of adaptation to chronic juvenile malnutrition in six populations of Drosophila melanogaster evolved for 150 generations on an extremely nutrient-poor larval diet. Comparison with control populations evolved on standard food revealed repeatable genomic differentiation between the two set of population, involving >3,000 candidate SNPs forming >100 independently evolving clusters. The candidate genomic regions were enriched in genes implicated in hormone, carbohydrate, and lipid metabolism, including some with known effects on fitness-related life-history traits. Rather than being close to fixation, a substantial fraction of candidate SNPs segregated at intermediate allele frequencies in all malnutrition-adapted populations. This, together with patterns of among-population variation in allele frequencies and estimates of Tajima’s D, suggests that the poor diet results in balancing selection on some genomic regions. Our candidate genes for tolerance to larval malnutrition showed a high overlap with genes previously implicated in acute starvation resistance. However, adaptation to larval malnutrition in our study was associated with reduced tolerance to acute adult starvation. Thus, rather than reflecting synergy, the shared genomic architecture appears to mediate an evolutionary trade-off between tolerances to these two forms of nutritional stress.

Much has been learned about the genetics of aging from studies in model organisms, but still little is known about naturally occurring alleles that contribute to variation in longevity. For example, analysis of mutants and transgenes has identified insulin signaling as a major regulator of longevity, yet whether standing variation in this pathway underlies microevolutionary changes in lifespan and correlated fitness traits remains largely unclear. Here, we have analyzed the genomes of a set of Drosophila melanogaster lines that have been maintained under direct selection for postponed reproduction and indirect selection for longevity, relative to unselected control lines, for over 35 years. We identified many candidate loci shaped by selection for longevity and late-life fertility, but – contrary to expectation – we did not find overrepresentation of canonical longevity genes. Instead, we found an enrichment of immunity genes, particularly in the Toll pathway, suggesting that evolutionary changes in immune function might underpin – in part – the evolution of late-life fertility and longevity. To test whether this genomic signature is causative, we performed functional experiments. In contrast to control flies, long-lived flies tended to downregulate the expression of antimicrobial peptides upon infection with age yet survived fungal, bacterial, and viral infections significantly better, consistent with alleviated immunosenescence. To examine whether genes of the Toll pathway directly affect longevity, we employed conditional knockdown using in vivo RNAi. In adults, RNAi against the Toll receptor extended lifespan, whereas silencing the pathway antagonist cactus-–causing immune hyperactivation – dramatically shortened lifespan. Together, our results suggest that genetic changes in the age-dependent regulation of immune homeostasis might contribute to the evolution of longer life.

Background: The black soldier fly (Hermetia illucens) is the most promising insect candidate for nutrient-recycling through bioconversion of organic waste into biomass, thereby improving sustainability of protein supplies for animal feed and facilitating transition to a circular economy. Contrary to conventional livestock, genetic resources of farmed insects remain poorly characterised. We present the first comprehensive population genetic characterisation of H. illucens. Based on 15 novel microsatellite markers, we genotyped and analysed 2862 individuals from 150 wild and captive populations originating from 57 countries on seven subcontinents. Results: We identified 16 well-distinguished genetic clusters indicating substantial global population structure. The data revealed genetic hotspots in central South America and successive northwards range expansions within the indigenous ranges of the Americas. Colonisations and naturalisations of largely unique genetic profiles occurred on all non-native continents, either preceded by demographically independent founder events from various single sources or involving admixture scenarios. A decisive primarily admixed Polynesian bridgehead population serially colonised the entire Australasian region and its secondarily admixed descendants successively mediated invasions into Africa and Europe. Conversely, captive populations from several continents traced back to a single North American origin and exhibit considerably reduced genetic diversity, although some farmed strains carry distinct genetic signatures. We highlight genetic footprints characteristic of progressing domestication due to increasing socio-economic importance of H. illucens, and ongoing introgression between domesticated strains globally traded for large-scale farming and wild populations in some regions. Conclusions: We document the dynamic population genetic history of a cosmopolitan dipteran of South American origin shaped by striking geographic patterns. These reflect both ancient dispersal routes, and stochastic and heterogeneous anthropogenic introductions during the last century leading to pronounced diversification of worldwide structure of H. illucens. Upon the recent advent of its agronomic commercialisation, however, current human-mediated translocations of the black soldier fly largely involve genetically highly uniform domesticated strains, which meanwhile threaten the genetic integrity of differentiated unique local resources through introgression. Our in-depth reconstruction of the contemporary and historical demographic trajectories of H. illucens emphasises benchmarking potential for applied future research on this emerging model of the prospering insect-livestock sector. This work was primarily supported by the Swiss Federal Office for Agriculture (grant no. 627000824 to CS), as well as by the R4D programme of the Swiss Agency for Development and Cooperation and the Swiss National Science Foundation (grant no. IZ01Z0_147278 to CS, and grant no. 400540_152154 to MKen), the Austrian Science Foundation (grant no. FWF P32275 to MKap), the Canadian International Development Research Centre (IDRC) and the Australian Centre for International Agricultural Research (ACIAR) (grant no. 108866-001 to CMT), and the National Research Foundation, South Africa (reference no. CSRP170506229933 to CR).

Wolbachiabacteria are maternally transmitted intracellular microbes that are not only restricted to the reproductive organs but also found in various somatic tissues of their native hosts. The abundance of the endosymbiont in somatic tissues, usually a dead end for vertically transmitted bacteria, causes a multitude of effects on life history traits of their hosts, which are still not well understood. Thus, deciphering the host-symbiont interactions on a cellular level throughout a host’s lifecycle is of great importance to understand their homeostatic nature, persistence and spreading success. Using fluorescent and transmission electron microscopy, we conducted a comprehensive analysis ofWolbachiatropism in somatic and reproductive tissues of sixDrosophilaspecies at the intracellular level during host development. Our data uncovered diagnostic patterns of infections to embryonic primordial germ cells and to particular cells of somatic tissues in three different neotropicalDrosophilaspecies of the willistoni and saltans groups that have apparently evolved in both independently. We further found that restricted patterns ofWolbachiatropism are already determined in early fly embryogenesis. This is achieved via selective autophagy, and the restriction of infection is preserved through larval hatching and metamorphosis. We further uncovered tight interactions ofWolbachiawith membranes of the endoplasmic reticulum, which might play a scaffolding role for autophagosome formation and subsequent elimination of the endosymbiont. Finally, by analyzingD. simulanslines transinfected with non-nativeWolbachia, we uncovered that the host genetic background regulates tissue tropism of infection. Our data demonstrate a peculiar and novel mechanism to limit and spatially restrict bacterial infection in somatic tissues during a very early stage of host development.

Drosophila melanogaster is a leading model in population genetics and genomics, and a growing number of whole-genome datasets from natural populations of this species have been published over the last 20 years. A major challenge is the integration of these disparate datasets, often generated using different sequencing technologies and bioinformatic pipelines, which hampers our ability to address questions about the evolution and population structure of this species. Here we address these issues by developing a bioinformatics pipeline that maps pooled sequencing (Pool-Seq) reads from D. melanogaster to a hologenome consisting of fly and symbiont genomes and estimates allele frequencies using either a heuristic (PoolSNP) or a probabilistic variant caller (SNAPE-pooled). We use this pipeline to generate the largest data repository of genomic data available for D. melanogaster to date, encompassing 271 population samples from over 100 locations in >20 countries on four continents based on a combination of 121 unpublished and 150 previously published genomic datasets. Several of these locations have been sampled at different seasons across multiple years. This dataset, which we call Drosophila Evolution over Space and Time (DEST), is coupled with sampling and environmental meta-data. A web-based genome browser and web portal provide easy access to the SNP dataset. Our aim is to provide this scalable platform as a community resource which can be easily extended via future efforts for an even more extensive cosmopolitan dataset. Our resource will enable population geneticists to analyze spatio-temporal genetic patterns and evolutionary dynamics of D. melanogaster populations in unprecedented detail.

This note details the consortium’s rationale behind its decision to modify the metadata for putatively misidentified European samples in the DrosRTEC dataset. In brief, we use PCA on published datasets from North America and Europe to generate phylogeographic clusters reflective of worldwide D. melanogaster demography. We used this PCA to train a DAPC model in order the predict the group membership. Our results indicate that 4 out of 73 samples were misclassified and the metadata was updated accordingly. These samples are a spring-fall pair from Spain and Austria.

Drosophila melanogaster is an important model for antiviral immunity in arthropods, but very few DNA viruses have been described from the family Drosophilidae. This deficiency limits our opportunity to use natural host-pathogen combinations in experimental studies, and may bias our understanding of the Drosophila virome. Here, we report fourteen DNA viruses detected in a metagenomic analysis of 6668 pool-sequenced Drosophila, sampled from forty-seven European locations between 2014 and 2016. These include three new nudiviruses, a new and divergent entomopoxvirus, a virus related to Leptopilina boulardi filamentous virus, and a virus related to Musca domestica salivary gland hypertrophy virus. We also find an endogenous genomic copy of galbut virus, a double-stranded RNA partitivirus, segregating at very low frequency. Remarkably, we find that Drosophila Vesanto virus, a small DNA virus previously described as a bidnavirus, may be composed of up to twelve segments and thus represent a new lineage of segmented DNA viruses. Two of the DNA viruses, Drosophila Kallithea nudivirus and Drosophila Vesanto virus are relatively common, found in 2 per cent or more of wild flies. The others are rare, with many likely to be represented by a single infected fly. We find that virus prevalence in Europe reflects the prevalence seen in publicly available datasets, with Drosophila Kallithea nudivirus and Drosophila Vesanto virus the only ones commonly detectable in public data from wild-caught flies and large population cages, and the other viruses being rare or absent. These analyses suggest that DNA viruses are at lower prevalence than RNA viruses in D.melanogaster, and may be less likely to persist in laboratory cultures. Our findings go some way to redressing an earlier bias toward RNA virus studies in Drosophila, and lay the foundation needed to harness the power of Drosophila as a model system for the study of DNA viruses., M.W. was supported by the UK Natural Environmental Research Council through the E3 doctoral training programme (NE/L002558/1), and S.R. was supported by Wellcome Trust PhD programme (108905/Z/15/Z). A.B. received funding from BBSRC (grant number BB/P00685X/1). T.F. received funding from Swiss National Science Foundation (grant numbers 31003A-182262, PP00P3_165836, and PP00P3_133641/1). C.G. received funding from Agence Nationale de la Recherche (grant number ANR-15-CE32-0011-01). J.G. received funding from the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation programme (H2020-ERC-2014-CoG-647900) and from the Fundación Española para la Ciencia y la Tecnologia-Ministerio de Economía y Competitividad (FCT-15-10187). S.G. received funding from Deutsche Forschungsgemeinschaft (grant number GR 4495/2). M.K. received funding from Academy of Finland projects (268214 and 322980). M.K. received funding from Austrian Science Fund (FWF; grant number P32275). V.L. received funding from Danish Research council for natural Sciences (FNU; grant number 4002-00113B). B.S.O. received funding from the Scientific and Technological Research Council of Turkey (TUBITAK; grant number 214Z238). J.P. received funding from Deutsche Forschungsgemeinschaft (grant number PA 903/8). M.S.-R., M.S.V., and M.J. received funding from the Ministry of Education, Science and Technological Development of the Republic of Serbia (grant number 451-03-68/2020-14/200178). F.S. received funding from Deutsche Forschungsgemeinschaft (grant number STA1154/4-1; Projektnummer 408908608). M.T., A.P., and K.E. received funding from the Ministry of Education, Science and Technological Development of the Republic of Serbia (grant number 451-03-68/2020-14/200007). The DrosEU consortium has been funded by a Special Topics Network (STN) grant by the European Society of Evolutionary Biology (ESEB).

Drosophila melanogaster is a leading model in population genetics and genomics, and a growing number of whole-genome datasets from natural populations of this species have been published over the last years. A major challenge is the integration of disparate datasets, often generated using different sequencing technologies and bioinformatic pipelines, which hampers our ability to address questions about the evolution of this species. Here we address these issues by developing a bioinformatics pipeline that maps pooled sequencing (Pool-Seq) reads from D. melanogaster to a hologenome consisting of fly and symbiont genomes and estimates allele frequencies using either a heuristic (PoolSNP) or a probabilistic variant caller (SNAPE-pooled). We use this pipeline to generate the largest data repository of genomic data available for D. melanogaster to date, encompassing 271 previously published and unpublished population samples from over 100 locations in > 20 countries on four continents. Several of these locations have been sampled at different seasons across multiple years. This dataset, which we call Drosophila Evolution over Space and Time (DEST), is coupled with sampling and environmental meta-data. A web-based genome browser and web portal provide easy access to the SNP dataset. We further provide guidelines on how to use Pool-Seq data for model-based demographic inference. Our aim is to provide this scalable platform as a community resource which can be easily extended via future efforts for an even more extensive cosmopolitan dataset. Our resource will enable population geneticists to analyze spatio-temporal genetic patterns and evolutionary dynamics of D. melanogaster populations in unprecedented detail., DrosEU is funded by a Special Topic Networks (STN) grant from the European Society for Evolutionary Biology (ESEB). MK (M. Kapun) was supported by the Austrian Science Foundation (grant no. FWF P32275); JG by the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation programme (H2020-ERC-2014-CoG-647900) and by the Spanish Ministry of Science and Innovation (BFU-2011-24397); TF by the Swiss National Science Foundation (SNSF grants PP00P3_133641, PP00P3_165836, and 31003A_182262) and a Mercator Fellowship from the German Research Foundation (DFG), held as a EvoPAD Visiting Professor at the Institute for Evolution and Biodiversity, University of Münster; AOB by the National Institutes of Health (R35 GM119686); MK (M. Kankare) by Academy of Finland grant 322980; VL by Danish Natural Science Research Council (FNU) grant 4002-00113B; FS Deutsche Forschungsgemeinschaft (DFG) grant STA1154/4-1, Project 408908608; JP by the Deutsche Forschungsgemeinschaft Projects 274388701 and 347368302; AU by FPI fellowship (BES-2012-052999); ET Israel Science Foundation (ISF) grant 1737/17; MSV, MSR and MJ by a grant from the Ministry of Education, Science and Technological Development of the Republic of Serbia (451-03-68/2020-14/200178); AP, KE and MT by a grant from the Ministry of Education, Science and Technological Development of the Republic of Serbia (451-03-68/2020-14/200007); and TM NSERC grant RGPIN-2018-05551.

Periods of nutrient shortage impose strong selection on animal populations. Experimental studies of genetic adaptation to nutrient shortage largely focus on resistance to acute starvation at adult stage; it is not clear how conclusions drawn from these studies extrapolate to other forms of nutritional stress. We studied the genomic signature of adaptation to chronic juvenile malnutrition in six populationsof Drosophila melanogasterevolved for 150 generations on an extremely nutrient-poor larval diet. Comparison with control populations evolved on standard food revealed repeatable genomic differentiation between the two set of population, involving >3,000 candidate SNPs forming >100 independently evolving clusters. The candidate genomic regions were enriched in genes implicated in hormone, carbohydrate, and lipid metabolism, including some with known effects on fitness-related life-history traits. Rather than being close to fixation, a substantial fraction of candidate SNPs segregated at intermediate allele frequencies in all malnutrition-adapted populations. This, together with patterns of among-population variation in allele frequencies and estimates of Tajima’sD, suggests that the poor diet results in balancing selection on some genomic regions. Our candidate genes for tolerance to larval malnutrition showed a high overlap with genes previously implicated in acute starvation resistance. However, adaptation to larval malnutrition in our study was associated with reduced tolerance to acute adult starvation. Thus, rather than reflecting synergy, the shared genomic architecture appears to mediate an evolutionary trade-off between tolerances to these two forms of nutritional stress.

European and African natural populations of Drosophila melanogaster have been the focus of several studies aiming at inferring demographic and adaptive processes based on genetic variation data. However, in these analyses little attention has been given to gene flow between African and European samples. Here we present a dataset consisting of 14 fully sequenced haploid genomes sampled from a natural population from the northern species range (Umeå, Sweden). We co-analyzed this new data with an African population to compare the likelihood of several competing demographic scenarios for European and African populations and show that gene flow improves the fit of demographic models to data.

Drosophila melanogasteris an important model for antiviral immunity in arthropods, but very few DNA viruses have been described from the family Drosophilidae. This deficiency limits our opportunity to use natural host-pathogen combinations in experimental studies, and may bias our understanding of theDrosophilavirome. Here we report fourteen DNA viruses detected in a metagenomic analysis of approximately 6500 pool-sequencedDrosophila, sampled from 47 European locations between 2014 and 2016. These include three new Nudiviruses, a new and divergent Entomopox virus, a virus related toLeptopilina boulardifilamentous virus, and a virus related toMusca domesticasalivary gland hypertrophy virus. We also find an endogenous genomic copy of Galbut virus, a dsRNA Partitivirus, segregating at very low frequency. Remarkably, we find thatDrosophilaVesanto virus, a small DNA virus previously described as a Bidnavirus, may be composed of up to 12 segments and represent a new lineage of segmented DNA viruses. Two of the DNA viruses,DrosophilaKallithea nudivirus andDrosophilaVesanto virus are relatively common, found in 2% or more of wild flies. The others are rare, with many likely to be represented by a single infected fly. We find that virus prevalence in Europe reflects the prevalence seen in publicly-available datasets, withDrosophilaKallithea nudivirus andDrosophilaVesanto virus the only ones commonly detectable in public data from wild-caught flies and large population cages, and the other viruses being rare or absent. These analyses suggest that DNA viruses are at lower prevalence than RNA viruses inD. melanogaster, and may be less likely to persist in laboratory cultures. Our findings go some way to redressing an earlier bias toward RNA virus studies inDrosophila, and lay the foundation needed to harness the power ofDrosophilaas a model system for the study of DNA viruses.

Summary The relative importance of host control, environmental effects and stochasticity in the assemblage of host-associated microbiomes is being debated. We analysed the microbiome among fly populations that were sampled across Europe by the European Drosophila Population Genomics Consortium (DrosEU). In order to better understand the structuring principles of the natural D. melanogaster microbiome, we combined environmental data on climate and food-substrate with dense genomic data on host populations and microbiome profiling. Food-substrate, temperature, and host population structure correlated with microbiome structure. Microbes, whose abundance was co-structured with host populations, also differed in abundance between flies and their substrate in an independent survey. This finding suggests common, host-related structuring principles of the microbiome on different spatial scales.

Drosophila melanogaster, a small dipteran of African origin, represents one of the best-studied model organisms. Early work in this system has uniquely shed light on the basic principles of genetics and resulted in a versatile collection of genetic tools that allow to uncover mechanistic links between genotype and phenotype. Moreover, given its worldwide distribution in diverse habitats and its moderate genome-size, Drosophila has proven very powerful for population genetics inference and was one of the first eukaryotes whose genome was fully sequenced. In this book chapter, we provide a brief historical overview of research in Drosophila and then focus on recent advances during the genomic era. After describing different types and sources of genomic data, we discuss mechanisms of neutral evolution including the demographic history of Drosophila and the effects of recombination and biased gene conversion. Then, we review recent advances in detecting genome-wide signals of selection, such as soft and hard selective sweeps. We further provide a brief introduction to background selection, selection of noncoding DNA and codon usage and focus on the role of structural variants, such as transposable elements and chromosomal inversions, during the adaptive process. Finally, we discuss how genomic data helps to dissect neutral and adaptive evolutionary mechanisms that shape genetic and phenotypic variation in natural populations along environmental gradients. In summary, this book chapter serves as a starting point to Drosophila population genomics and provides an introduction to the system and an overview to data sources, important population genetic concepts and recent advances in the field.

Drosophila melanogaster, a small dipteran of African origin, represents one of the best-studied model organisms. Early work in this system has uniquely shed light on the basic principles of genetics and resulted in a versatile collection of genetic tools that allow to uncover mechanistic links between genotype and phenotype. Moreover, given its world-wide distribution in diverse habitats and its moderate genome-size, Drosophila has proven very powerful for population genetics inference and was one of the first eukaryotes whose genome was fully sequenced. In this book-chapter, we provide a brief historical overview of research in Drosophila and then focus on recent advances during the genomic era. After describing different types and sources of genomic data, we discuss mechanisms of neutral evolution including the demographic history of Drosophila and the effects of recombination and biased gene conversion. Then, we review recent advances in detecting genome-wide signals of selection, such as soft and hard selective sweeps. We further provide a brief introduction to background selection, selection of non-coding DNA and codon usage and focus on the role of structural variants, such as transposable elements and chromosomal inversions, during the adaptive process. Finally, we discuss how genomic data helps to dissect neutral and adaptive evolutionary mechanisms that shape genetic and phenotypic variation in natural populations along environmental gradients. In summary, this book chapter serves as a starting point to Drosophila population genomics and provides an introduction to the system and an overview to data sources, important population genetic concepts and recent advances in the field., Comment: book chapter, 69 pages (ref included), 2 figures

Chromosomal inversions, structural mutations that reverse a segment of a chromosome, cause suppression of recombination in the heterozygous state. Several studies have shown that inversion polymorphisms can form clines or fluctuate predictably in frequency over seasonal time spans. These observations prompted the hypothesis that chromosomal rearrangements might be subject to spatially and/or temporally varying selection. Here, we review what has been learned about the adaptive significance of inversion polymorphisms in the vinegar fly Drosophila melanogaster, the species in which they were first discovered by Sturtevant in 1917. A large body of work provides compelling evidence that several inversions in this system are adaptive; however, the precise selective mechanisms that maintain them polymorphic in natural populations remain poorly understood. Recent advances in population genomics, modelling and functional genetics promise to greatly improve our understanding of this long-standing and fundamental problem in the near future.

Reproduction and diet are two major factors controlling the physiology of aging and life history, but how they interact to affect the evolution of longevity is unknown. Moreover, although studies of large-effect mutants suggest an important role of nutrient sensing pathways in regulating aging, the genetic basis of evolutionary changes in lifespan remains poorly understood. To address these questions, we analyzed the genomes of experimentally evolved Drosophila melanogaster populations subjected to a factorial combination of two selection regimes: reproductive age (early versus postponed), and diet during the larval stage (“low,” “control,” “high”), resulting in six treatment combinations with four replicate populations each. Selection on reproductive age consistently affected lifespan, with flies from the postponed reproduction regime having evolved a longer lifespan. In contrast, larval diet affected lifespan only in early-reproducing populations: flies adapted to the “low” diet lived longer than those adapted to control diet. Here, we find genomic evidence for strong independent evolutionary responses to either selection regime, as well as loci that diverged in response to both regimes, thus representing genomic interactions between the two. Overall, we find that the genomic basis of longevity is largely independent of dietary adaptation. Differentiated loci were not enriched for “canonical” longevity genes, suggesting that naturally occurring genic targets of selection for longevity differ qualitatively from variants found in mutant screens. Comparing our candidate loci to those from other “evolve and resequence” studies of longevity demonstrated significant overlap among independent experiments. This suggests that the evolution of longevity, despite its presumed complex and polygenic nature, might be to some extent convergent and predictable.

Background Population genetic theory predicts that rapid adaptation is largely driven by complex traits encoded by many loci of small effect. Because large-effect loci are quickly fixed in natural populations, they should not contribute much to rapid adaptation. Results To investigate the genetic architecture of thermal adaptation — a highly complex trait — we performed experimental evolution on a natural Drosophila simulans population. Transcriptome and respiration measurements reveal extensive metabolic rewiring after only approximately 60 generations in a hot environment. Analysis of genome-wide polymorphisms identifies two interacting selection targets, Sestrin and SNF4Aγ, pointing to AMPK, a central metabolic switch, as a key factor for thermal adaptation. Conclusions Our results demonstrate that large-effect loci segregating at intermediate allele frequencies can allow natural populations to rapidly respond to selection. Because SNF4Aγ also exhibits clinal variation in various Drosophila species, we suggest that this large-effect polymorphism is maintained by temporal and spatial temperature variation in natural environments. Electronic supplementary material The online version of this article (10.1186/s13059-018-1503-4) contains supplementary material, which is available to authorized users.

A fundamental aim of adaptation genomics is to identify polymorphisms that underpin variation in fitness traits. InD. melanogasterlatitudinal life-history clines exist on multiple continents and make an excellent system for dissecting the genetics of adaptation. We have previously identified numerous clinal SNPs in insulin/insulin-like growth factor signaling (IIS), a pathway known from mutant studies to affect life history. However, the effects of natural variants in this pathway remain poorly understood. Here we investigate how two clinal alternative alleles atfoxo, a transcriptional effector of IIS, affect fitness components (viability, size, starvation resistance, fat content). We assessed this polymorphism from the North American cline by reconstituting outbred populations, fixed for either the low- or high-latitude allele, from inbred DGRP lines. Since diet and temperature modulate IIS, we phenotyped alleles across two temperatures (18°C, 25°C) and two diets differing in sugar source and content. Consistent with clinal expectations, the high-latitude allele conferred larger body size and reduced wing loading. Alleles also differed in starvation resistance and expression ofInR, a transcriptional target of FOXO. Allelic reaction norms were mostly parallel, with few GxE interactions. Together, our results suggest that variation in IIS makes a major contribution to clinal life-history adaptation.

Background: The Drosophila C virus (DCV) is a common and well-studied Drosophila pathogen. Although natural infections are known from Drosophila melanogaster and D. simulans, and artificial infections have been reported from several. Drosophila species and other insects, it remains unclear to date whether DCV infections also occur naturally in other Drosophila species. Methods/Principal Findings: Using reverse transcription PCR, we detected natural infections in six Drosophila species, which have not been previously known as natural hosts. By subsequent Sanger sequencing we compared DCV haplotypes among eight Drosophila host species. Our data suggest that cross-infections might be frequent both within and among species within the laboratory environment. Moreover, we find that some lines exhibit multiple infections with distinct DCV haplotypes. Conclusions: Our results suggest that the natural host range of DCV is much broader than previously assumed and that cross-infections might be a common phenomenon in the laboratory, even among different Drosophila hosts.

The relative importance of host-control, environmental effects, and stochasticity in the assemblage of host-associated microbiomes has been much debated. With recent sampling efforts, the underpinnings of D. melanogaster’s microbiome structure have become tractable on larger spatial scales. We analyzed the microbiome among fly populations that were sampled across Europe by the European Drosophila Population Genomics Consortium (DrosEU). We combined environmental data on climate and food-substrate, dense genomic data on host population structure, and microbiome profiling. Food-substrate, temperature, and host population-structure correlated with microbiome-structure. The microbes, whose abundance was co-structured with host populations, also differed in abundance between flies and their substrate in an independent survey, suggesting host-control. Patterns of enrichment and depletion of microbes between host and substrate were consistent with a model of host-control, where the host manipulates its microbiome for its benefit. Putative host-control was bacterial strain specific, supporting recent evidence for high specificity of D. melanogaster-microbe interaction.

International audience; Size trade-offs of visual versus olfactory organs is a pervasive feature of animal evolution. This could result from genetic or functional constraints. We demonstrate that head sensory organ size trade-offs in Drosophila are genetically encoded and arise through differential subdivision of the head primordium into visual versus non-visual fields. We discover that changes in the temporal regulation of the highly conserved eyeless/Pax6 gene expression during development is a conserved mechanism for sensory trade-offs within and between Drosophila species. We identify a natural single nucleotide polymorphism in the cis-regulatory region of eyeless in a binding site of its repressor Cut that is sufficient to alter its temporal regulation and eye size. Because eyeless/Pax6 is a conserved regulator of head sensory placode subdivision, we propose that its temporal regulation is key to define the relative size of head sensory organs.

Chromosomal inversions often contribute to local adaptation across latitudinal clines, but the underlying selective mechanisms remain poorly understood. We and others have previously shown that a clinal inversion polymorphism in Drosophila melanogaster, In(3R)Payne, underpins body size clines along the North American and Australian east coasts. Here, we ask whether this polymorphism also contributes to clinal variation in other fitness-related traits, namely survival traits (lifespan, survival upon starvation and survival upon cold shock). We generated homokaryon lines, either carrying the inverted or standard chromosomal arrangement, isolated from populations approximating the endpoints of the North American cline (Florida, Maine) and phenotyped the flies at two growth temperatures (18 °C, 25 °C). Across both temperatures, high-latitude flies from Maine lived longer and were more stress resistant than low-latitude flies from Florida, as previously observed. Interestingly, we find that this latitudinal pattern is partly explained by the clinal distribution of the In(3R)P polymorphism, which is at 50% frequency in Florida but absent in Maine: inverted karyotypes tended to be shorter-lived and less stress resistant than uninverted karyotypes. We also detected an interaction between karyotype and temperature on survival traits. As In(3R)P influences body size and multiple survival traits, it can be viewed as a ‘supergene’, a cluster of tightly linked loci affecting multiple complex phenotypes. We conjecture that the inversion cline is maintained by fitness trade-offs and balancing selection across geography; elucidating the mechanisms whereby this inversion affects alternative, locally adapted phenotypes across the cline is an important task for future work.

SUMMARYSize trade-offs of visual versus olfactory organs is a pervasive feature of animal evolution. ComparingDrosophilaspecies, we find that larger eyes correlate with smaller antennae, where olfactory organs reside, and narrower faces. We demonstrate that this tradeoff arises through differential subdivision of the head primordium into visual versus non-visual fields. Specification of the visual field requires a highly-conserved eye development gene calledeyelessin flies and Pax6 in humans. We discover that changes in the temporal regulation ofeyelessexpression during development is a conserved mechanism for sensory trade-offs within and betweenDrosophilaspecies. We identify a natural single nucleotide polymorphism in the cis-regulatory region ofeyelessthat is sufficient to alter its temporal regulation and eye size. Because Pax6 is a conserved regulator of sensory placode subdivision, we propose that alterations in the mutual repression between sensory territories is a conserved mechanism for sensory trade-offs in animals.

Much has been learned about the genetics of aging from studies in model organisms, but still little is known about naturally occurring alleles that contribute to variation in longevity. For example, analysis of mutants and transgenes has identified insulin signaling as a major regulator of longevity, yet whether standing variation in this pathway underlies microevolutionary changes in lifespan and correlated fitness traits remains largely unclear. Here, we have analyzed the genomes of a set of Drosophila melanogaster lines that have been maintained under direct selection for postponed reproduction and indirect selection for longevity, relative to unselected control lines, for over 35 years. We identified many candidate loci shaped by selection for longevity and late‐life fertility, but – contrary to expectation – we did not find overrepresentation of canonical longevity genes. Instead, we found an enrichment of immunity genes, particularly in the Toll pathway, suggesting that evolutionary changes in immune function might underpin – in part – the evolution of late‐life fertility and longevity. To test whether this genomic signature is causative, we performed functional experiments. In contrast to control flies, long‐lived flies tended to downregulate the expression of antimicrobial peptides upon infection with age yet survived fungal, bacterial, and viral infections significantly better, consistent with alleviated immunosenescence. To examine whether genes of the Toll pathway directly affect longevity, we employed conditional knockdown using in vivo RNAi. In adults, RNAi against the Toll receptor extended lifespan, whereas silencing the pathway antagonist cactus‐–causing immune hyperactivation – dramatically shortened lifespan. Together, our results suggest that genetic changes in the age‐dependent regulation of immune homeostasis might contribute to the evolution of longer life.

Summary Environmental variation can have profound and direct effects on fitness, fecundity, and host–symbiont interactions. Replication rates of microbes within arthropod hosts, for example, are correlated with incubation temperature but less is known about the influence of host–symbiont dynamics on environmental preference. Hence, we conducted thermal preference (T p) assays and tested if infection status and genetic variation in endosymbiont bacterium Wolbachia affected temperature choice of Drosophila melanogaster. We demonstrate that isogenic flies infected with Wolbachia preferred lower temperatures compared with uninfected Drosophila. Moreover, T p varied with respect to three investigated Wolbachia variants (wMel, wMelCS, and wMelPop). While uninfected individuals preferred 24.4°C, we found significant shifts of −1.2°C in wMel‐ and −4°C in flies infected either with wMelCS or wMelPop. We, therefore, postulate that Wolbachia‐associated T p variation within a host species might represent a behavioural accommodation to host–symbiont interactions and trigger behavioural self‐medication and bacterial titre regulation by the host.

Size trade-offs of visual versus olfactory organs is a pervasive feature of animal evolution. Such tradeoffs could result from genetic or functional constraints. We demonstrate that head sensory organ size trade-offs in Drosophila are genetically encoded and arise through differential subdivision of the head primordium into visual versus non-visual fields. We discover that changes in the temporal regulation of the highly-conserved eyeless/Pax6 gene expression during development is a conserved mechanism for sensory trade-offs within and between Drosophila species. We identify a natural single nucleotide polymorphism in the cis-regulatory region of eyeless that is sufficient to alter its temporal regulation and eye size. Because eyeless/Pax6 is a conserved regulator of head sensory placode subdivision, we propose that its temporal regulation is key to define the relative size of head sensory organs.

Chromosomal inversion polymorphisms are common in animals and plants, and recent models suggest that alternative arrangements spread by capturing different combinations of alleles acting additively or epistatically to favour local adaptation. It is also thought that inversions typically maintain favoured combinations for a long time by suppressing recombination between alternative chromosomal arrangements. Here, we consider patterns of linkage disequilibrium and genetic divergence in an old inversion polymorphism in Drosophila melanogaster (In(3R)Payne) known to be associated with climate change adaptation and a recent invasion event into Australia. We extracted, karyotyped and sequenced whole chromosomes from two Australian populations, so that changes in the arrangement of the alleles between geographically separated tropical and temperate areas could be compared. Chromosome-wide linkage disequilibrium (LD) analysis revealed strong LD within the region spanned by In(3R)Payne. This genomic region also showed strong differentiation between the tropical and the temperate populations, but no differentiation between different karyotypes from the same population, after controlling for chromosomal arrangement. Patterns of differentiation across the chromosome arm and in gene ontologies were enhanced by the presence of the inversion. These data support the notion that inversions are strongly selected by bringing together combinations of genes, but it is still not clear if such combinations act additively or epistatically. Our data suggest that climatic adaptation through inversions can be dynamic, reflecting changes in the relative abundance of different forms of an inversion and ongoing evolution of allelic content within an inversion.

Understanding the rate of evolutionary change and the genetic architecture that facilitates rapid adaptation is a current challenge in evolutionary biology. Comparative studies show that genes with immune function are among the most rapidly evolving genes in a range of taxa. Here, we use immune defense in natural populations of D. melanogaster to understand the rate of evolution in natural populations and the genetics underlying the rapid change. We probed the immune system using the natural pathogens Enterococcus faecalis and Providencia rettgeri to measure post-infection survival and bacterial load of wild D. melanogaster populations collected across seasonal time along a latitudinal transect on the eastern North America (Massachusetts, Pennsylvania, and Virginia). There are pronounced and repeatable changes in the immune response over approximately 10 generations between the spring and fall populations with a significant but less distinct difference among geographic locations. Genes with known immune function are not enriched among alleles that cycle with seasonal time, but the immune function of a subset of seasonally cycling alleles in immune genes was tested using reconstructed outbred populations. We find that flies containing seasonal alleles in Thioester-containing protein 3 (Tep3) have different functional responses to infection and that epistatic interactions among seasonal Tep3 and Drosomycin-like 6 (Dro6) alleles produce the immune phenotypes observed in natural populations. This rapid, cyclic response to seasonal environmental pressure broadens our understanding of the complex ecological and genetic interactions determining the evolution of immune defense in natural populations.

Genome-wide association studies (GWAS) are designed to identify the portion of single-nucleotide polymorphisms (SNPs) in genome sequences associated with a complex trait. Strategies based on the gene list enrichment concept are currently applied for the functional analysis of GWAS, according to which a significant overrepresentation of candidate genes associated with a biological pathway is used as a proxy to infer overrepresentation of candidate SNPs in the pathway. Here we show that such inference is not always valid and introduce the program SNP2GO, which implements a new method to properly test for the overrepresentation of candidate SNPs in biological pathways.