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.

Large-scale genomic resources can place genetic variation into an ecologically informed context. To advance our understanding of the population genetics of the fruit fly Drosophila melanogaster, we present an expanded release of the community-generated population genomics resource Drosophila Evolution over Space and Time (DEST 2.0; https://dest.bio/). This release includes 530 high-quality pooled libraries from flies collected across six continents over more than a decade (2009 to 2021), most at multiple time points per year; 211 of these libraries are sequenced and shared here for the first time. We used this enhanced resource to elucidate several aspects of the species' demographic history and identify novel signs of adaptation across spatial and temporal dimensions. For example, we showed that the spatial genetic structure of populations is stable over time, but that drift due to seasonal contractions of population size causes populations to diverge over time. We identified signals of adaptation that vary between continents in genomic regions associated with xenobiotic resistance, consistent with independent adaptation to common pesticides. Moreover, by analyzing samples collected during spring and fall across Europe, we provide new evidence for seasonal adaptation related to loci associated with pathogen response. Furthermore, we have also released an updated version of the DEST genome browser. This is a useful tool for studying spatiotemporal patterns of genetic variation in this classic model system.

We analyzed the developmental time, egg-to-adult viability, and developmental stability (fluctuating wing size asymmetry) in Drosophila subobscura, maintained for six generations on different concentrations of lead. Development time is significantly affected by generation and lead concentration, but interaction of these factors is not a significant source of variability for this fitness component. Generation and the interaction generation x concentration of lead significantly affect egg-to-adult viability. Levene's test of heterogeneity of variance showed that variability of FA is not significant in any of the samples. Within both lead concentrations females showed significantly higher FA indices for the wing width than males. Within sexes, a significantly higher FA was found only in females for wing width FA between the control and the lower concentration of lead. The results show that if strong relationship between FA and the studied fitness components exists, it results in a stronger selection of unstable genotypes under lead as a stress factor and, consequently, FA needs to be used with caution as a biomarker in natural populations under environmental stress.

The genetic structure of Drosophila subobscura from the Balkan Peninsula was studied with respect to restriction site polymorphism of mitochondrial DNA in populations from the Derventa River Gorge and Sicevo Gorge (Serbia). To investigate the role of cytonuclear interactions in shaping mitochondrial DNA variability in natural populations of this species, the study was complemented with the analysis of linkage disequilibria between mitochondrial haplotypes and chromosomal inversion arrangements. Similar to other populations of D. subobscura, two main haplotypes (I and II) were found, as well as a series of less common ones. The frequencies of haplotypes I and II accounted for 25.8% and 71.0%, respectively, in the population from the Derventa River Gorge, and for 32.4% and 58.1%, respectively, in the population from Sicevo Gorge. One of the haplotypes harbored a large insertion (2.7 kb) in the A+T rich region. The frequency distribution of both haplotypes did not depart from neutrality. Contrary to prior studies, we did not detect any significant linkage disequilibrium between the two most frequent mtDNA haplotypes and any of the chromosomal arrangements in either of the populations. We conclude that linkage disequilibrium is not a general occurrence in natural populations of D. subobscura, and we discuss how transient coadaptations, ecologically specific selective pressures, and demographics could contribute to population-specific patterns of linkage disequilibrium.

The adaptive significance of sympatric mitochondrial (mtDNA) variation and the role of selective mechanisms that maintain it are debated to this day. Isofemale lines of Drosophila obscura collected from four populations were backcrossed within populations to construct experimental lines, with all combinations of mtDNA Cyt b haplotypes and nuclear genetic backgrounds (nuDNA). Individuals of both sexes from these lines were then subjected to four fitness assays (desiccation resistance, developmental time, egg-to-adult viability and sex ratio) on two experimental temperatures to examine the role of temperature fluctuations and sex-specific selection, as well as the part that interactions between the two genomes play in shaping mtDNA variation. The results varied across populations and fitness components. In the majority of comparisons, they show that sympatric mitochondrial variants affect fitness. However, their effect should be examined in light of interactions with nuDNA, as mito-nuclear genotype was even more influential on fitness across all components. We found both sex-specific and temperature-specific differences in mitochondrial and mito-nuclear genotype ranks in all fitness components. The effect of temperature-specific selection was found to be more prominent, especially in desiccation resistance. From the results of different components tested, we can also infer that temperature-specific mito-nuclear interactions rather than sex-specific selection on mito-nuclear genotypes have a more substantial role in preserving mtDNA variation in this model species.

A particularly well-studied evolutionary model is the vinegar fly Drosophila melanogaster, a cosmopolitan insect of ancestral southern-central African origin. Recent work suggests that it expanded out of Africa ∼9,000 years ago, and spread from the Middle East into Europe ∼1,800 years ago. During its global expansion, this human commensal adapted to novel climate zones and habitats. Despite much work on phenotypic differentiation and adaptation on several continents (especially North America and Australia), typically in the context of latitudinal clines, little is known about phenotypic divergence among European populations. Here, we sought to provide a continent-wide study of phenotypic differentiation among European populations of D. melanogaster. In a consortium-wide phenomics effort, we assayed 16 fitness-related traits on a panel of 173 isofemale lines from 9 European populations, with the majority of traits measured by several groups using semi-standardized protocols. For most fitness-related traits, we found significant differentiation among populations on a continental scale. Despite inevitable differences in assay conditions among labs, the reproducibility and hence robustness of our measurements were overall remarkably good. Several fitness components (e.g., viability, development time) exhibited significant latitudinal or longitudinal clines, and populations differed markedly in multivariate trait structure. Notably, populations experiencing higher humidity/rainfall and lower maximum temperature showed higher viability, fertility, starvation resistance, and lifespan at the expense of lower heat-shock survival, suggesting a pattern of local adaptation. Our results indicate that derived populations of this tropical fly have been shaped by pervasive spatially varying multivariate selection and adaptation to different climates on the European continent.