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Evolve and re-sequencing (E&R) studies investigate the genomic responses of adaptation during experimental evolution. Because replicate populations evolve in the same controlled environment, consistent responses to selection across replicates are frequently used to identify reliable candidate regions that underlie adaptation to a new environment. However, recent work demonstrated that selection signatures can be restricted to one or a few replicate(s) only. These selection signatures frequently have a weak statistical support, and given the difficulties of functional validation, additional evidence is needed before considering them as candidates for functional analysis. Here, we introduce an experimental procedure to validate candidate loci with weak or replicate-specific selection signature(s). Crossing an evolved population from a primary E&R experiment to the ancestral founder population reduces the frequency of candidate alleles that have reached a high frequency. We hypothesize that genuine selection targets will experience a repeatable frequency increase after the mixing with the ancestral founders if they are exposed to the same environment (secondary E&R experiment). Using this approach, we successfully validate two overlapping selection targets, which showed a mutually exclusive selection signature in a primary E&R experiment of Drosophila simulans adapting to a novel temperature regime. We conclude that secondary E&R experiments provide a reliable confirmation of selection signatures that are either not replicated or show only a low statistical significance in a primary E&R experiment. Such experiments are particularly helpful to prioritize candidate loci for time-consuming functional follow-up investigations.

Significance Adaptation to temperature and drought stress in Drosophila can be experimentally explored as a proxy model for adaptive trait complexes and genomic responses to climate variation. As a snapshot of synchronized adaptive events in a climate gradient, contemporary convergent evolution empowers the detection and understanding of adaptation from population genomic data and advance climate change assessment and forecasting. However, the effects of climate change on living organisms have been shown primarily on regional and global scales, confounding climate-related and climate-unrelated multivariate factors. This study leverages a unique microclimate contrast, known as Evolution Canyon, and a Drosophila model within it to provide a whole-genome perspective of adaptive evolution, convergence under thermal stress, and incipient speciation., Biodiversity refugia formed by unique features of the Mediterranean arid landscape, such as the dramatic ecological contrast of “Evolution Canyon,” provide a natural laboratory in which local adaptations to divergent microclimate conditions can be investigated. Significant insights have been provided by studies of Drosophila melanogaster diversifying along the thermal gradient in Evolution Canyon, but a comparative framework to survey adaptive convergence across sister species at the site has been lacking. To fill this void, we present an analysis of genomic polymorphism and evolutionary divergence of Drosophila simulans, a close relative of Drosophila melanogaster with which it co-occurs on both slopes of the canyon. Our results show even deeper interslope divergence in D. simulans than in D. melanogaster, with extensive signatures of selective sweeps present in flies from both slopes but enhanced in the population from the hotter and drier south-facing slope. Interslope divergence was enriched for genes related to electrochemical balance and transmembrane transport, likely in response to increased selection for dehydration resistance on the hotter slope. Both species shared genomic regions that underwent major selective sweeps, but the overall level of adaptive convergence was low, demonstrating no shortage of alternative genomic solutions to cope with the challenges of the microclimate contrast. Mobile elements were a major source of genetic polymorphism and divergence, affecting all parts of the genome, including coding sequences of mating behavior-related genes.

Sensory perception modulates health and aging across taxa. Understanding the nature of relevant cues and the mechanisms underlying their action may lead to novel interventions that improve the length and quality of life. We found that in the vinegar fly, Drosophila melanogaster, exposure to dead conspecifics in the environment induced cues that were aversive to other flies, modulated physiology, and impaired longevity. The effects of exposure to dead conspecifics on aversiveness and lifespan required visual and olfactory function in the exposed flies. Furthermore, the sight of dead flies was sufficient to produce aversive cues and to induce changes in the head metabolome. Genetic and pharmacologic attenuation of serotonergic signaling eliminated the effects of exposure on aversiveness and lifespan. Our results indicate that Drosophila have an ability to perceive dead conspecifics in their environment and suggest conserved mechanistic links between neural state, health, and aging; the roots of which might be unearthed using invertebrate model systems., Different sensory experiences can affect longevity in Drosophila. Here the authors find that exposure of Drosophila directly to dead conspecifics affects longevity via a serotonergic mechanism, and that Drosophila exposed to dead conspecifics also become an aversive stimulus to naïve choosers.

Meiotic recombination is crucial for chromosomal segregation and facilitates the spread of beneficial and removal of deleterious mutations. Recombination rates frequently vary along chromosomes and Drosophila melanogaster exhibits a remarkable pattern. Recombination rates gradually decrease toward centromeres and telomeres, with a dramatic impact on levels of variation in natural populations. Two close sister species, Drosophila simulans and Drosophila mauritiana do not only have higher recombination rates but also exhibit a much more homogeneous recombination rate that only drops sharply very close to centromeres and telomeres. Because certain sequence motifs are associated with recombination rate variation in D. melanogaster, we tested whether the difference in recombination landscape between D. melanogaster and D. simulans can be explained by the genomic distribution of recombination rate-associated sequence motifs. We constructed the first high-resolution recombination map for D. simulans based on 189 haplotypes from a natural D. simulans population and searched for short sequence motifs linked with higher than average recombination in both sister species. We identified five consensus motifs significantly associated with higher than average chromosome-wide recombination rates in at least one species and present in both. Testing fine resolution associations between motif density and recombination, we found strong and positive associations genome-wide over a range of scales in D. melanogaster, while the results were equivocal in D. simulans. Despite the strong association in D. melanogaster, we did not find a decreasing density of these short-repeat motifs toward centromeres and telomeres. We conclude that the density of recombination-associated repeat motifs cannot explain the large-scale recombination landscape in D. melanogaster, nor the differences to D. simulans. The strong association seen for the sequence motifs in D. melanogaster likely reflects their impact influencing local differences in recombination rates along the genome.

The evolution of sexual dimorphism is constrained by a shared genome, leading to ‘sexual antagonism’, in which different alleles at given loci are favoured by selection in males and females. Despite its wide taxonomic incidence, we know little about the identity, genomic location, and evolutionary dynamics of antagonistic genetic variants. To address these deficits, we use sex-specific fitness data from 202 fully sequenced hemiclonal Drosophila melanogaster fly lines to perform a genome-wide association study (GWAS) of sexual antagonism. We identify approximately 230 chromosomal clusters of candidate antagonistic single nucleotide polymorphisms (SNPs). In contradiction to classic theory, we find no clear evidence that the X chromosome is a hot spot for sexually antagonistic variation. Characterising antagonistic SNPs functionally, we find a large excess of missense variants but little enrichment in terms of gene function. We also assess the evolutionary persistence of antagonistic variants by examining extant polymorphism in wild D. melanogaster populations and closely related species. Remarkably, antagonistic variants are associated with multiple signatures of balancing selection across the D. melanogaster distribution range and in their sister species D. simulans, indicating widespread and evolutionarily persistent (about 1 million years) genomic constraints on the evolution of sexual dimorphism. Based on our results, we propose that antagonistic variation accumulates because of constraints on the resolution of sexual conflict over protein coding sequences, thus contributing to the long-term maintenance of heritable fitness variation., This study characterises antagonistic loci across the genome of the fruit fly Drosophila melanogaster, finding them to be preferentially associated with variation in coding sequences and to be selectively maintained across worldwide populations of D. melanogaster, and even its sister species D. simulans.

Species of the Drosophila melanogaster species subgroup, including the species D. simulans, D. mauritiana, D. yakuba, and D. santomea, have long served as model systems for studying evolution. However, studies in these species have been limited by a paucity of genetic and transgenic reagents. Here, we describe a collection of transgenic and genetic strains generated to facilitate genetic studies within and between these species. We have generated many strains of each species containing mapped piggyBac transposons including an enhanced yellow fluorescent protein (EYFP) gene expressed in the eyes and a φC31 attP site-specific integration site. We have tested a subset of these lines for integration efficiency and reporter gene expression levels. We have also generated a smaller collection of other lines expressing other genetically encoded fluorescent molecules in the eyes and a number of other transgenic reagents that will be useful for functional studies in these species. In addition, we have mapped the insertion locations of 58 transposable elements in D. virilis that will be useful for genetic mapping studies. Fil: Stern, David L.. Janelia Research Campus; Estados Unidos Fil: Crocker, Justin. Janelia Research Campus; Estados Unidos Fil: Ding, Yun. Janelia Research Campus; Estados Unidos Fil: Frankel, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Ecología, Genética y Evolución de Buenos Aires. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Ecología, Genética y Evolución de Buenos Aires; Argentina Fil: Kappes, Gretchen. No especifica; Fil: Kim, Elizabeth. Janelia Research Campus; Estados Unidos Fil: Kuzmickas, Ryan. Brigham and Women's Hospital; Estados Unidos Fil: Lemire, Andrew. Janelia Research Campus; Estados Unidos Fil: Mast, Joshua D.. California State University; Estados Unidos Fil: Picard, Serge. University of Princeton; Estados Unidos

Male fitness is dependent on sexual traits that influence mate acquisition (pre-copulatory sexual selection) and paternity (post-copulatory sexual selection), and while many studies have documented the form of selection in one or the other of these arenas, fewer have done it for both. Nonetheless, it appears that the dominant form of sexual selection is directional, although theoretically, populations should converge on peaks in the fitness surface, where selection is stabilizing. Many factors, however, can prevent populations from reaching adaptive peaks. Genetic constraints can be important if they prevent the development of highest fitness phenotypes, as can the direction of selection if it reverses across episodes of selection. In this study, we examine the evidence that these processes influence the evolution of the multivariate sex comb morphology of male Drosophila simulans. To do this, we conduct a quantitative genetic study together with a multivariate selection analysis to infer how the genetic architecture and selection interact. We find abundant genetic variance and covariance in elements of the sex comb. However, there was little evidence for directional selection in either arena. Significant nonlinear selection was detected prior to copulation when males were mated to non-virgin females, and post-copulation during sperm offence (again with males mated to non-virgins). Thus contrary to our predictions, the evolution of the D. simulans sex comb is limited neither by genetic constraints nor by antagonistic selection between pre- and post-copulatory arenas, but nonlinear selection on the multivariate phenotype may prevent sex combs from evolving to reach some fitness maximising optima. This article is protected by copyright. All rights reserved.

Robustness in development allows for the accumulation of neutral genetically based variation in expression, and here will be termed ‘genetic stochasticity‘. This largely neutral variation is potentially important for both evolution and complex disease phenotypes. However, it has generally only been investigated as variation exhibited in the response to large genetic perturbations. In addition, work on variation in gene expression has similarly generally been limited to being spatial, or quantitative, but because of technical restrictions not both. Here we bridge these gaps by investigating replicated quantitative spatial gene expression using rigorous statistical models, in different genotypes, sexes, and species (Drosophila melanogasterandD. simulans). Using this type of quantitative approach with developmental data allows for effective comparison among conditions, including health versus disease. We apply this approach to the morphogenetic furrow, a wave of differentiation that sweeps across the developing eye disc. Within the morphogenetic furrow, we focus on four conserved morphogens,hairy, atonal, hedgehog, andDelta. Hybridization chain reaction quantitatively measures spatial gene expression, co-staining for all four genes simultaneously and with minimal effort. We find considerable variation in the spatial expression pattern of these genes in the eye between species, genotypes, and sexes. We also find that there has been evolution of the regulatory relationship between these genes. Lastly, we show that the spatial interrelationships of these genes evolved between species in the morphogenetic furrow. This is essentially the first ‘population genetics of development’ as we are able to evaluate wild type differences in spatial and quantitative gene expression at the level of genotype, species and sex.

All genomes contain repeated sequences that are known as transposable elements (TEs). Among these are endogenous retroviruses (ERVs), which are sequences similar to retroviruses and are transmitted across generations from parent to progeny. These sequences are controlled in genomes through epigenetic mechanisms. At the center of the epigenetic control of TEs are small interfering RNAs of the piRNA class, which trigger heterochromatinization of TE sequences. The tirant ERV of Drosophila simulans displays intra-specific variability in copy numbers, insertion sites, and transcription levels, providing us with a well-suited model to study the dynamic relationship between a TE family and the host genome through epigenetic mechanisms. We show that tirant transcript amounts and piRNA amounts are positively correlated in ovaries in normal conditions, unlike what was previously described following divergent crosses. In addition, we describe tirant insertion polymorphism in the genomes of three D. simulans wild-type strains, which reveals a limited number of insertions that may be associated with gene transcript level changes through heterochromatin spreading and have phenotypic impacts. Taken together, our results participate in the understanding of the equilibrium between the host genome and its TEs.

Reproductive isolation is a key component of speciation. In many insects, a major driver of this isolation is cuticular hydrocarbon pheromones, which help to identify potential intraspecific mates [1-3]. When the distributions of related species overlap, there may be strong selection on mate choice for intraspecific partners [4-9] because interspecific hybridization carries significant fitness costs [10]. Drosophila has been a key model for the investigation of reproductive isolation; although both male and female mate choices have been extensively investigated [6, 11-16], the genes underlying species recognition remain largely unknown. To explore the molecular mechanisms underlying Drosophila speciation, we measured tissue-specific cis-regulatory divergence using RNA sequencing (RNA-seq) in D. simulans × D. sechellia hybrids. By focusing on cis-regulatory changes specific to female oenocytes, the tissue that produces cuticular hydrocarbons, we rapidly identified a small number of candidate genes. We found that one of these, the fatty acid elongase eloF, broadly affects the hydrocarbons present on D. sechellia and D. melanogaster females, as well as the propensity of D. simulans males to mate with them. Therefore, cis-regulatory changes in eloF may be a major driver in the sexual isolation of D. simulans from multiple other species. Our RNA-seq approach proved to be far more efficient than quantitative trait locus (QTL) mapping in identifying candidate genes; the same framework can be used to pinpoint candidate drivers of cis-regulatory divergence in traits differing between any interfertile species.

During speciation, sex chromosomes often accumulate interspecific genetic incompatibilities faster than the rest of the genome. The drive theory posits that sex chromosomes are susceptible to recurrent bouts of meiotic drive and suppression, causing the evolutionary build-up of divergent cryptic sex-linked drive systems and, incidentally, genetic incompatibilities. To assess the role of drive during speciation, we combine high-resolution genetic mapping of X-linked hybrid male sterility with population genomics analyses of divergence and recent gene flow between the fruitfly species,Drosophila mauritianaandD. simulans. Our findings reveal a high density of genetic incompatibilities and a corresponding dearth of gene flow on the X chromosome. Surprisingly, we find that, rather than contributing to interspecific divergence, a known drive element has recently migrated between species, caused a strong reduction in local divergence, and undermined the evolution of hybrid sterility. Gene flow can therefore mediate the effects of selfish genetic elements during speciation.

How dietary selection affects genome evolution to define the optimal range of nutrient intake is a poorly understood question with medical relevance. We have addressed this question by analyzing Drosophila simulans and sechellia, recently diverged species with differential diet choice. D. sechellia larvae, specialized to a nutrient scarce diet, did not survive on sugar-rich conditions, while the generalist species D. simulans was sugar tolerant. Sugar tolerance in D. simulans was a tradeoff for performance on low-energy diet and was associated with global reprogramming of metabolic gene expression. Hybridization and phenotype-based introgression revealed the genomic regions of D. simulans that were sufficient for sugar tolerance. These regions included genes that are involved in mitochondrial ribosome biogenesis and intracellular signaling, such as PPP1R15/Gadd34 and SERCA, which contributed to sugar tolerance. In conclusion, genomic variation affecting genes involved in global metabolic control defines the optimal range for dietary macronutrient composition., eLife digest Animals meet their nutritional needs in a variety of ways. Some animals are specialists feeding only on one type of food; others are generalists that can choose many different kinds of food depending on the situation. Despite these differences in diet, animals have similar needs for basic cellular metabolism. This suggests that generalist and specialist species likely process the foods they eat in different ways in order to meet their basic needs. For example, the metabolism of generalist species may be more flexible to adapt to changing food sources. To learn more about how metabolism evolves to respond to diet, scientists can study closely related species that eat different foods. For example, a species of fruit fly called Drosophila simulans is a generalist and its larvae can grow and develop by feeding on different kinds of decaying fruits and vegetables. Larvae of a closely related fruit fly called Drosophila sechellia are specialized to eat only the nutrient-poor Morinda fruit. Looking at how genetic differences between these species affect metabolism may provide scientists with clues about how these feeding strategies evolved. Melvin et al. grew larvae of D. sechellia and D. simulans in different conditions. D. sechellia larvae thrived in low nutrient conditions, but died when exposed to high sugar foods. By contrast, D. simulans larvae tolerated high sugar levels, but did poorly in low-nutrient conditions. Melvin et al. then bred the two species with each other, selecting flies that are genetically similar to D. sechellia but have the genes necessary for larvae to tolerate sugar. Analyzing the selected hybrid flies revealed genetic changes that explain the different survival abilities of each species. These changes suggest that D. sechellia rapidly evolved to thrive in low nutrient conditions, but the trade-off was losing their ability to tolerate high sugar levels. Overall, the results presented by Melvin et al. suggest that genetic adaptions to food sources can occur quickly and drastically change metabolism. Further research will be needed to confirm if similar metabolic trade-offs developed as part of human evolution. If so, human populations that survived with limited nutrition for many generations may have a harder time adapting to high-sugar modern diets.

Drosophila (Sophophora) simulans Sturtevant, 1919 Drosophila simulans Sturtevant, 1919: 153. Holotype, male. USA (Florida: Lakeland) (AMNH). Distribution: Cosmopolitan. Egyptian localities: Coastal Strip: Abu Sir (Mourad & Mallah 1960), Alexandria, Burg El-Arab, Janaleese. Fayoum:?. Eastern Desert: Esmailiya. Lower Nile Valley & Delta: Abu- Hommos, Beni Suef, Cairo, Fakoos, Kafr El-Sheikh, Mahalla El-Kubra, Tanta (Mourad & Mallah 1960; Megeed et al. 2015), Mansoura. Upper Nile Valley: Assiout, Kom Ombo, Luxur (Mourad & Mallah 1960; B��chli et al. 1982). Western Desert: El-Nubariya, Siwa, Wadi El-Natroun (Mourad & Mallah 1960; Yassin & Abou-Youssef 2004). Dates of collection: March to December (Mourad & Mallah 1960; B��chli et al. 1982 and Yassin & Abou- Youssef 2004)., Published as part of El-Hawagry, Magdi S., Zatwarnicki, Tadeusz & Ebrahim, Ayman M., 2018, Catalogue of the Egyptian Ephydroidea (Diptera: Schizophora: Acalyptratae), pp. 201-246 in Zootaxa 4444 (3) on page 236, DOI: 10.11646/zootaxa.4444.3.1, http://zenodo.org/record/1309626, {"references":["Sturtevant, A. H. (1919) A new species closely resembling Drosophila melanogaster. Psyche, 26, 153 - 155. https: // doi. org / 10.1155 / 1919 / 97402","Mourad, A. M. & Mallah, G. S. (1960) Chromosomal polymorphism in Egyptian populations of Drosophila melanogaster. Evolution, 14, 166 - 170. https: // doi. org / 10.1111 / j. 1558 - 5646.1960. tb 03075. x","Megeed, M. S, Galal, O. A. & Abdel-Razek, M. M. (2015) Phylogenetic relationship of an invasive drosophilid, Zaprionus indianus and closely related species of Drosophilidae (Diptera) based on esterase patterns. Egyptian Journal of Genetics and Cytology, 44, 281 - 290.","Yassin, A. & Abou-Youssef, A. Y. (2004) A new front for a global invasive drosophilid: the colonization of the Northern- Western desert of Egypt by Zaprionus indianus Gupta, 1970. Drosophila Information Service, 87, 67 - 68."]}

The genetic architecture of adaptive traits is of key importance to predict evolutionary responses. Most adaptive traits are polygenic—i.e., result from selection on a large number of genetic loci—but most molecularly characterized traits have a simple genetic basis. This discrepancy is best explained by the difficulty in detecting small allele frequency changes (AFCs) across many contributing loci. To resolve this, we use laboratory natural selection to detect signatures for selective sweeps and polygenic adaptation. We exposed 10 replicates of a Drosophila simulans population to a new temperature regime and uncovered a polygenic architecture of an adaptive trait with high genetic redundancy among beneficial alleles. We observed convergent responses for several phenotypes—e.g., fitness, metabolic rate, and fat content—and a strong polygenic response (99 selected alleles; mean s = 0.059). However, each of these selected alleles increased in frequency only in a subset of the evolving replicates. We discerned different evolutionary paradigms based on the heterogeneous genomic patterns among replicates. Redundancy and quantitative trait (QT) paradigms fitted the experimental data better than simulations assuming independent selective sweeps. Our results show that natural D. simulans populations harbor a vast reservoir of adaptive variation facilitating rapid evolutionary responses using multiple alternative genetic pathways converging at a new phenotypic optimum. This key property of beneficial alleles requires the modification of testing strategies in natural populations beyond the search for convergence on the molecular level., Replicate populations of the fruit fly Drosophila simulans exposed to a novel temperature environment showed convergent evolution on the phenotypic level. On the genetic level, however, no convergence was observed. These results support the “trait optimum” paradigm as the predominant mode of adaptation., Author summary It is widely assumed that adaptation is mainly polygenic, with the underlying frequency changes being so small that they are difficult to detect and characterize. Taking advantage of laboratory-based natural selection of replicated populations of the fruit fly Drosophila simulans exposed to a high temperature regime, we identified a polygenic response with many (99) selected alleles having pronounced allele frequency changes (AFCs). Despite phenotypic convergence across the 10 replicates, the genomic response was highly heterogeneous, with different subsets of the selected alleles contributing to the convergent phenotype in each of the replicate populations. The observed genetic redundancy not only indicates lack of genetic constraint but also demonstrates that multiple genetic pathways lead to convergent phenotypic outcomes.

Natural populations often exist in spatially diverse environments and may experience variation in the strength and targets of natural selection across their ranges. Drosophila provides an excellent opportunity to study the effects of spatially varying selection in natural populations, as both Drosophila melanogaster and Drosophila simulans live across a wide range of environments in North America. Here, we characterize patterns of variation in transposable elements (TEs) from six populations of D. melanogaster and nine populations of D. simulans sampled from multiple latitudes across North America. We find a nearly twofold excess of TEs in D. melanogaster relative to D. simulans, with this difference largely driven by TEs segregating at the lowest and highest allele frequencies. We find no effect of latitude on either total TE abundance or average TE allele frequencies in either species. Moreover, we show that, as a class of mutations, the most common patterns of TE variation do not coincide with the sampled latitudinal gradient, nor are they consistent with local adaptation acting on environmental differences found in the most extreme latitudes. We also do not find a cline in ancestry for North American D. melanogaster-for either TEs or single nucleotide polymorphisms-suggesting a limited role for demography in shaping patterns of TE variation. Though we find little evidence for widespread clinality among TEs in Drosophila, this does not necessarily imply a limited role for TEs in adaptation. We discuss the need for improved models of adaptation to large-scale environmental heterogeneity, and how these might be applied to TEs.

Hybridization and gene flow between species appears to be common. Even though it is clear that hybridization is widespread across all surveyed taxonomic groups, the magnitude and consequences of introgression are still largely unknown. Thus it is crucial to develop the statistical machinery required to uncover which genomic regions have recently acquired haplotypes via introgression from a sister population. We developed a novel machine learning framework, called FILET (Finding Introgressed Loci via Extra-Trees) capable of revealing genomic introgression with far greater power than competing methods. FILET works by combining information from a number of population genetic summary statistics, including several new statistics that we introduce, that capture patterns of variation across two populations. We show that FILET is able to identify loci that have experienced gene flow between related species with high accuracy, and in most situations can correctly infer which population was the donor and which was the recipient. Here we describe a data set of outbred diploid Drosophila sechellia genomes, and combine them with data from D. simulans to examine recent introgression between these species using FILET. Although we find that these populations may have split more recently than previously appreciated, FILET confirms that there has indeed been appreciable recent introgression (some of which might have been adaptive) between these species, and reveals that this gene flow is primarily in the direction of D. simulans to D. sechellia., Author summary Understanding the extent to which species or diverged populations hybridize in nature is crucially important if we are to understand the speciation process. Accordingly numerous research groups have developed methodology for finding the genetic evidence of such introgression. In this report we develop a supervised machine learning approach for uncovering loci which have introgressed across species boundaries. We show that our method, FILET, has greater accuracy and power than competing methods in discovering introgression, and in addition can detect the directionality associated with the gene flow between species. Using whole genome sequences from Drosophila simulans and Drosophila sechellia we show that FILET discovers quite extensive introgression between these species that has occurred mostly from D. simulans to D. sechellia. Our work highlights the complex process of speciation even within a well-studied system and points to the growing importance of supervised machine learning in population genetics.

Geographic patterns of genetic differentiation have long been used to understand population history and to learn about the biological mechanisms of adaptation. Here we present an examination of genomic patterns of differentiation between northern and southern populations of Australian and North American Drosophila simulans, with an emphasis on characterizing signals of parallel differentiation. We report on the genomic scale of differentiation and functional enrichment of outlier SNPs. While, overall, signals of shared differentiation are modest, we find the strongest support for parallel differentiation in genomic regions that are associated with regulation. Comparisons to Drosophila melanogaster yield potential candidate genes involved in local adaptation in both species, providing insight into common selective pressures and responses. In contrast to D. melanogaster, in D. simulans we observe patterns of variation that are inconsistent with a model of temperate adaptation out of a tropical ancestral range, highlighting potential differences in demographic and colonization histories of this cosmopolitan species pair.

Examples of clinal variation in phenotypes and genotypes across latitudinal transects have served as important models for understanding how spatially varying selection and demographic forces shape variation within species. Here, we examine the selective and demographic contributions to latitudinal variation through the largest comparative genomic study to date of Drosophila simulans and Drosophila melanogaster, with genomic sequence data from 382 individual fruit flies, collected across a spatial transect of 19 degrees latitude and at multiple time points over 2 years. Consistent with phenotypic studies, we find less clinal variation in D. simulans than D. melanogaster, particularly for the autosomes. Moreover, we find that clinally varying loci in D. simulans are less stable over multiple years than comparable clines in D. melanogaster. D. simulans shows a significantly weaker pattern of isolation by distance than D. melanogaster and we find evidence for a stronger contribution of migration to D. simulans population genetic structure. While population bottlenecks and migration can plausibly explain the differences in stability of clinal variation between the two species, we also observe a significant enrichment of shared clinal genes, suggesting that the selective forces associated with climate are acting on the same genes and phenotypes in D. simulans and D. melanogaster.

Essential genes and species incompatibilities Crosses between two fruit fly species, Drosophila melanogaster and D. simulans , result in hybrid progeny that are all female. Although some of the genes responsible for this species barrier are known, the full complement of molecular determinants that lead to inviable males has remained mysterious. Phadnis et al. used mutagenesis and a sequencing-based genomic screen to link hybrid inviability to the cell cycle. The inviable males result from an interaction between three genes, one of which is essential, which precluded its identification with standard genetic screens. This strategy to identify speciation genes can be applied to other model and nonmodel systems. Science , this issue p. 1552

Hypermutable strains of Drosophila simulans have been studied for 20 years. Several mutants were isolated and characterized, some of which had phenotypes associated with alteration in development; for example, showing ectopic legs with eyes being expressed in place of antennae. The causal agent of this hypermutability is a non-autonomous hobo-related sequence (hoboVA). Around 100 mobilizable copies of this element are present in the D. simulans genome, and these are likely mobilized by the autonomous and canonical hobo element. We have shown that hoboVA has transcription factor binding sites for the developmental genes, hunchback and evenskipped, and that this transposon is expressed in embryos, following the patterns of these genes. We suggest that hobo and hobo-related elements can be material for the emergence of new regulatory networks.

Clinal variation is widely regarded as strong evidence for adaptation in natural populations. One of the best-studied clinal phenotypes is the latitudinal melanism cline in ectotherms. Owing to the repeatability of clinal pigmentation across species and continents, this trait is frequently considered as a classic example for an adaptive trait. Nevertheless, how selection operates on this trait is still debated. The thermal budget hypothesis, one popular explanation, suggests that pigmentation intensity modulates the use of sunlight for thermoregulation. Here, we challenge this hypothesis and show how misleading the evolutionary interpretation of phenotypic variation can be. Using highly replicated experimental evolution setup, we exposed natural Drosophila simulans populations to novel thermal environments. After more than 100 generations of adaptation to hot temperature, the key components of melanin metabolism evolved in the same direction as in natural clines. Because temperature adaptation was not mediated by exposure to sun, we searched for alternative explanations. Melanin, the key ingredient for pigmentation, is synthesized from dopamine, a highly pleiotropic neurotransmitter. Using RNA- seq, metabolomics, and transgenic and pharmacologic modulation of dopamine, we uncovered the adaptive role of dopamine levels. Temperature strongly modulates signaling activity and in our experiment and natural populations, evolution modulates dopaminergic signaling to maintain synaptic homeostasis. We demonstrate that the evolutionary modulation of dopaminergic signaling helps to maintain normal locomotion even at high temperatures. We conclude that the pigmentation cline in Drosophila is a pleiotropic read-out of adaptive changes in dopamine metabolism, rather than an adaptive phenotype. Our results demonstrate that pleiotropy is a major challenge for the interpretation of phenotypic variation in a causative evolutionary framework.

Autophagy and phagocytosis are cellular immune mechanisms for internalization and elimination of intracellular and extracellular pathogens. Some pathogens have evolved the ability to inhibit or manipulate these processes, raising the prospect of adaptive reciprocal co-evolution by the host. We performed population genetic analyses on phagocytosis and autophagy genes in Drosophila melanogaster and D. simulans to test for molecular evolutionary signatures of immune adaptation. We found that phagocytosis and autophagy genes as a whole exhibited an elevated level of haplotype homozygosity in both species. In addition, we detected signatures of recent selection, notably in the Atg14 and Ykt6 genes in D. melanogaster and a pattern of elevated sequence divergence in the genderblind (gb) gene on the D. simulans lineage. These results suggest that the evolution of the host cellular immune system as a whole may be shaped by a dynamic conflict between Drosophila and its pathogens even without pervasive evidence of strong adaptive evolution at the individual gene level.

The rapidly expanding availability of large NGS data sets provides an opportunity to investigate population genetics at an unprecedented scale. Drosophila simulans is the sister species of the model organism Drosophila melanogaster, and is often presumed to share similar demographic history. However, previous population genetic and ecological work suggests very different signatures of selection and demography. Here, we sequence a new panel of 170 inbred genotypes of a North American population of D. simulans, a valuable complement to the DGRP and other D. melanogaster panels. We find some unexpected signatures of demography, in the form of excess intermediate frequency polymorphisms. Simulations suggest that this is possibly due to a recent population contraction and selection. We examine the outliers in the D. simulans genome determined by a haplotype test to attempt to parse the contribution of demography and selection to the patterns observed in this population. Untangling the relative contribution of demography and selection to genomic patterns of variation is challenging, however, it is clear that although D. melanogaster was thought to share demographic history with D. simulans different forces are at work in shaping genomic variation in this population of D. simulans.

Two interesting unanswered questions are the extent to which both the broad patterns and genetic details of adaptive divergence are repeatable across species, and the timescales over which parallel adaptation may be observed. Drosophila melanogaster is a key model system for population and evolutionary genomics. Findings from genetics and genomics suggest that recent adaptation to latitudinal environmental variation (on the timescale of hundreds or thousands of years) associated with Out-of-Africa colonization plays an important role in maintaining biological variation in the species. Additionally, studies of interspecific differences between D. melanogaster and its sister species D. simulans have revealed that a substantial proportion of proteins and amino acid residues exhibit adaptive divergence on a roughly few million years long timescale. Here we use population genomic approaches to attack the problem of parallelism between D. melanogaster and a highly diverged conger, D. hydei, on two timescales. D. hydei, a member of the repleta group of Drosophila, is similar to D. melanogaster, in that it too appears to be a recently cosmopolitan species and recent colonizer of high latitude environments. We observed parallelism both for genes exhibiting latitudinal allele frequency differentiation within species and for genes exhibiting recurrent adaptive protein divergence between species. Greater parallelism was observed for long-term adaptive protein evolution and this parallelism includes not only the specific genes/proteins that exhibit adaptive evolution, but extends even to the magnitudes of the selective effects on interspecific protein differences. Thus, despite the roughly 50 million years of time separating D. melanogaster and D. hydei, and despite their considerably divergent biology, they exhibit substantial parallelism, suggesting the existence of a fundamental predictability of adaptive evolution in the genus.

Wolbachia pipientis is an intracellular endosymbiont infecting many arthropods and filarial nematodes. Little is known about the short-term evolution of Wolbachia or its interaction with its host. Wolbachia is maternally inherited, resulting in co-inheritance of mitochondrial organelles such as mtDNA. Here I explore the evolution of Wolbachia, and the relationship between Wolbachia and mtDNA, using a large inbred panel of Drosophila simulans. I compare this to the only other large population genomic Wolbachia dataset from D. melanogaster. I find reduced diversity relative to expectation in both Wolbachia and mtDNA, but only mtDNA shows evidence of a recent selective sweep or population bottleneck. I estimate Wolbachia and mtDNA titre in each genotype, and I find considerable variation in both phenotypes, despite low genetic diversity in Wolbachia and mtDNA. A phylogeny of Wolbachia and of mtDNA suggest a recent origin of the infection derived from a single origin. Using Wolbachia and mtDNA titre as a phenotype, I perform the first association analysis using this phenotype with the nuclear genome and find several implicated regions, including one which contains four CAAX-box protein processing genes. CAAX-box protein processing can be an important part of host-pathogen interactions in other systems, suggesting interesting directions for future research.

Wolbachia pipientis from Drosophila melanogaster (wMel) is an endosymbiotic bacterium that restricts transmission of human pathogenic flaviviruses and alphaviruses, including dengue, Zika, and chikungunya viruses, when introduced into the mosquito vector Aedes aegypti. To date, wMel-infected Ae. aegypti have been released in field trials in 5 countries to evaluate the effectiveness of this strategy for disease control. Despite the success in establishing wMel-infected mosquitoes in wild populations, and the well-characterized antiviral capabilities of wMel, transinfecting different or additional Wolbachia strains into Ae. aegypti may improve disease impact, and perhaps more importantly, could provide a strategy to account for the possible evolution of resistant arboviruses. Here, we report the successful transinfection of Ae. aegypti with the Wolbachia strains wMelCS (D. melanogaster), wRi (D. simulans) and wPip (Culex quinquefasciatus) and assess the effects on Ae. aegypti fitness, cytoplasmic incompatibility, tissue tropism and pathogen blocking in a laboratory setting. The results demonstrate that wMelCS provides a similar degree of protection against dengue virus as wMel following an infectious blood meal, and significantly reduces viral RNA levels beyond that of wMel following a direct challenge with infectious virus in mosquitoes, with no additional fitness cost to the host. The protection provided by wRi is markedly weaker than that of wMelCS, consistent with previous characterisations of these lines in Drosophila, while wPip was found to substantially reduce the fitness of Ae. aegypti. Thus, we determine wMelCS as a key candidate for further testing in field-relevant fitness tests and viremic blood feeding challenges in a clinical setting to determine if it may represent an alternative Wolbachia strain with more desirable attributes than wMel for future field testing., Author summary Dengue viruses are transmitted by the Aedes aegypti mosquito, with an estimated 390 million human infections occurring per year worldwide. There is no approved antiviral therapeutic, and vaccines described so far have had limited efficacy. Recently, the endosymbiotic bacterium Wolbachia from Drosophila melanogaster (wMel) has been used to infect Ae. aegypti populations as a novel technology for reducing dengue virus transmission. Here we report the generation of three new mosquito lines infected with the Wolbachia strains wMelCS, wRi and wPip. Each line induced cytoplasmic incompatibility and was effectively maternally transmitted, as required for rapid spread through uninfected mosquito populations. Each Wolbachia strain was also found to reside in the salivary glands; a key tissue involved in viral transmission. Perhaps most importantly, wMelCS inhibited dengue virus replication and dissemination in mosquitoes following an infectious blood meal or intrathoracic injection, providing a similar level of protection as that described for wMel. wMelCS therefore warrants further investigation as a potential release strain in future field trials.

Transposable elements (TEs) are widespread genomic parasites, and their evolution has remained a critical question in evolutionary genomics. Here, we study the relatively unexplored epigenetic impacts of TEs and provide the first genome-wide quantification of such effects in D. melanogaster and D. simulans. Surprisingly, the spread of repressive epigenetic marks (histone H3K9me2) to nearby DNA occurs at >50% of euchromatic TEs, and can extend up to 20 kb. This results in differential epigenetic states of genic alleles and, in turn, selection against TEs. Interestingly, the lower TE content in D. simulans compared to D. melanogaster correlates with stronger epigenetic effects of TEs and higher levels of host genetic factors known to promote epigenetic silencing. Our study demonstrates that the epigenetic effects of euchromatic TEs, and host genetic factors modulating such effects, play a critical role in the evolution of TEs both within and between species. DOI: http://dx.doi.org/10.7554/eLife.25762.001, eLife digest The DNA inside an organism encodes all the instructions needed for the organism to develop and work properly. Organisms carefully organize and maintain their DNA (collectively known as the genome) so that the genetic information remains intact and the cell can understand the instructions. However, there are some pieces of DNA that are capable of moving around the genome. For example, pieces known as transposable elements can make new copies of themselves and jump into new locations in the genome. Most transposons do not appear to have any important roles, and in fact they are usually harmful to organisms. Despite this, transposons are present in the genomes of almost all species. The number of transposons in a genome varies greatly between individuals and species, but it is not clear why this is the case. Organisms have evolved ways to limit the damage caused by transposons. For example, many cells package regions of DNA containing transposons into a tightly packed structure known as heterochromatin. However, this type of DNA packaging sometimes spreads to neighboring sections of DNA. This is a problem because cells are not usually able to read the information contained within heterochromatin. This means that transposons can prevent some instructions from being produced when they should be. Lee and Karpen used fruit flies to investigate to what extent transposons harm organisms by changing the way DNA is packaged, and whether this influences how transposons evolve. The experiments show that that more than half of the transposons in fruit flies cause neighboring sections of DNA to be packaged into heterochromatin. This can negatively impact up to 20% of genes in the genome. As a result, transposons that have harmful effects on DNA packaging are more likely to be lost from the fly population during evolution than transposons that do not have harmful effects. Fruit fly species containing transposons that tend to package more neighboring sections of DNA into heterochromatin generally have fewer transposons than genomes containing less harmful transposons. The findings of Lee and Karpen provide new insight as to why the numbers of transposons vary among organisms. The next challenge is to find out whether transposons that alter how DNA is packaged are also common in primates and other animals. DOI: http://dx.doi.org/10.7554/eLife.25762.002

The Y chromosome should degenerate because it cannot recombine. However, male-limited transmission increases selection efficiency for male-benefit alleles on the Y, and therefore, Y chromosomes should contribute significantly to variation in male fitness. This means that although the Drosophila Y chromosome is small and gene-poor, Y-linked genes are vital for male fertility in Drosophila melanogaster and the Y chromosome has large male fitness effects. It is unclear whether the same pattern is seen in the closely related Drosophila simulans. We backcrossed Y chromosomes from three geographic locations into five genetic backgrounds and found strong Y and genetic background effects on male fertility. There was a significant Y-background interaction, indicating substantial epistasis between the Y and autosomal genes affecting male fertility. This supports accumulating evidence that interactions between the Y chromosome and the autosomes are key determinants of male fitness.

Drosophila simulans is a close relative of the genetic model D. melanogaster. Its worldwide distribution in combination with the absence of segregating chromosomal inversions makes this species an increasingly attractive model to study the molecular signatures of adaptation in natural and experimental populations. In an effort to improve the genomic resources for D. simulans, we assembled and annotated the genome of a strain originating from Madagascar (M252), the ancestral range of D. simulans. The comparison of the M252 genome to other available D. simulans assemblies confirmed its high quality, but also highlighted genomic regions that are difficult to assemble with NGS data. The annotation of M252 provides a clear improvement with alternative splicing for 52% of the multiple-exon genes, UTRs for 70% of the genes, 225 novel genes and 781 pseudogenes being reported. We anticipate that the M252 genome will be a valuable resource for many research questions.

The seemingly transparent wings of many insects have recently been found to display unexpected structural coloration. These structural colours (wing interference patterns: WIPs) may be involved in species recognition and mate choice, yet little is known about the evolutionary processes that shape them. Furthermore, to date investigations of WIPs have not fully considered how they are actually perceived by the viewers' colour vision. Here, we use multispectral digital imaging and a model ofDrosophilavision to compare WIPs of male and femaleDrosophila simulansfrom replicate populations forced to evolve with or without sexual selection for 68 generations. We show that WIPs modelled inDrosophilavision evolve in response to sexual selection and provide evidence that WIPs correlate with male sexual attractiveness. These findings add a new element to the otherwise well-describedDrosophilacourtship display and confirm that wing colours evolve through sexual selection.

simulans Sturtevant, 1919: 153. Type locality: USA, Florida, Lakeland. HT M (AMNH). NT NE PA AF OR AU Distr.: cosmopolitan. Colombia (Bogota, D:C, Amazonas (Leticia), Cundinamarca (Fusagasug��)). Refs.: Hunter, 1960: 86; Hunter, 1964: 39; Hunter, 1966: 423; Hunter & Navarro, 1969: 1102; Yoon, 1996: 14., Published as part of Tidon, Rosana & Almeida, Jonathan Mendes De, 2016, FAMILY DROSOPHILIDAE, pp. 719-751 in Zootaxa 4122 (1) on page 734, DOI: 10.11646/zootaxa.4122.1.63, http://zenodo.org/record/263272, {"references":["Sturtevant, A. H. (1919) A new species closely resembling Drosophila melanogaster. Psyche, 26, 153 - 155. http: // dx. doi. org / 10.1155 / 1919 / 97402","Hunter, A. S. (1960) Drosophilidae of Bogota, Colombia. Drosophila Information Service, 34, 86.","Hunter, A. S. & Hunter, R. A. (1964) The mesophragmatica species group of Drosophila in Colombia. Annals of the Entomological Society of America, 57, 732 - 736. http: // dx. doi. org / 10.1093 / aesa / 57.6.732","Hunter, A. S. (1966) High-altitude Drosophila of Colombia. (Diptera, Drosophilidae). Annals of the Entomological Society of America, 59, 413 - 423. http: // dx. doi. org / 10.1093 / aesa / 59.3.413","Hunter, A. S. & Navarro, A. (1969) A natural population of Drosophila (Diptera: Drosophilidae) in Colombia. Annals of the Entomological Society of America, 62, 1101 - 1105. http: // dx. doi. org / 10.1093 / aesa / 62.5.1101","Yoon, Z. (1996) Stock List. National Drosophila Species Resource Center. Drosophila Information Service, 77, 1 - 44."]}

Mechanisms of reproductive isolation inhibit gene flow between species and can be broadly sorted into two categories: prezygotic and postzygotic. While comparative studies suggest that prezygotic barriers tend to evolve first, postzygotic barriers are crucial for maintaining species boundaries and impeding gene flow that might otherwise cause incipient species to merge. Most, but not all, postzygotic barriers result from genetic incompatibilities between two or more loci from different species, and occur due to divergent evolution in allopatry. Hybrid defects result from improper allelic interactions between these loci. While some postzygotic barriers are environmentally-independent, the magnitude of others has been shown to vary in penetrance depending on environmental factors. We crossed Drosophila melanogaster mutants to two other species, D. simulans and D. santomea, and collected fitness data of the hybrids at two different temperatures. Our goal was to examine the effect of temperature on recessive incompatibility alleles in their genomes. We found that temperature has a stronger effect on the penetrance of recessive incompatibility alleles in the D. simulans genome than on those in the D. santomea genome. These results suggest that the penetrance of hybrid incompatibilities can be strongly affected by environmental context, and that the magnitude of such gene-by-environment interactions can be contingent on the genotype of the hybrid.

The evolution of a sex-linked meiotic drive system in natural and experimental populations of [i]Drosophila simulans[/i] . Réunion du GDR Approche Interdisciplinaire de l’Evolution Moléculaire