Addicted? Reduced host resistance in populations with defensive symbionts

Heritable symbionts that protect their hosts from pathogens have been described in a wide range of insect species. By reducing the incidence or severity of infection, these symbionts have the potential to reduce the strength of selection on genes in the insect genome that increase resistance. Therefore, the presence of such symbionts may slow down the evolution of resistance. Here we investigated this idea by exposing Drosophila melanogaster populations to infection with the pathogenic Drosophila C virus (DCV) in the presence or absence of Wolbachia, a heritable symbiont of arthropods that confers protection against viruses. After nine generations of selection, we found that resistance to DCV had increased in all populations. However, in the presence of Wolbachia the resistant allele of pastrel—a gene that has a major effect on resistance to DCV—was at a lower frequency than in the symbiont-free populations. This finding suggests that defensive symbionts have the potential to hamper the evolution of insect resistance genes, potentially leading to a state of evolutionary addiction where the genetically susceptible insect host mostly relies on its symbiont to fight pathogens.     

Virulence variability of the Drosophila C virus and effects of the microparasite on demographic parameters of the host (Drosophila melanogaster)

We carried out experiments with the Drosophila C virus (DCV), a nonhereditary virus acting on demographic parameters of infected Drosophila host populations. It is well known that DCV increases mortality rate, decreases developmental time, and increases daily fecundity. As usual for Drosophila viruses, the DCV was multiplied in vivo. In this study we tested the hypothesis of virulence variability in DCV strains by isolating different stocks of the virus. The flies were tested for susceptibility to injection of such isolates and for virulence variability. Possible interactions between demographic parameters in three Drosophila host populations and injected isolates were studied under two egg densities (low and high). The hypothesis of virulence variability of DCV was supported by significant differences in mortality rates, depending on whether virus isolates were ingested or injected. When DCV was ingested, differences between host mortality rates were independent of the Drosophila host populations. Nevertheless, the developmental time was equally decreased by each virus isolate, independent of the host population. Moreover, the two viral stocks strongly increased the egg production of the flies. This experimental approach clearly showed that DCV could be considered a polymorphic virus. The phenotypic interactions between DCV and host flies varied according to parasite genotype.     

Genome-Wide Association Studies Reveal a Simple Genetic Basis of Resistance to Naturally Coevolving Viruses in Drosophila melanogaster

Variation in susceptibility to infectious disease often has a substantial genetic component in animal and plant populations. We have used genome-wide association studies (GWAS) in Drosophila melanogaster to identify the genetic basis of variation in susceptibility to viral infection. We found that there is substantially more genetic variation in susceptibility to two viruses that naturally infect D. melanogaster (DCV and DMelSV) than to two viruses isolated from other insects (FHV and DAffSV). Furthermore, this increased variation is caused by a small number of common polymorphisms that have a major effect on resistance and can individually explain up to 47% of the heritability in disease susceptibility. For two of these polymorphisms, it has previously been shown that they have been driven to a high frequency by natural selection. An advantage of GWAS in Drosophila is that the results can be confirmed experimentally. We verified that a gene called pastrel—which was previously not known to have an antiviral function—is associated with DCV-resistance by knocking down its expression by RNAi. Our data suggest that selection for resistance to infectious disease can increase genetic variation by increasing the frequency of major-effect alleles, and this has resulted in a simple genetic basis to variation in virus resistance.     

Genetics of dispersal

Dispersal is a process of central importance for the ecological and evolutionary dynamics of populations and communities, because of its diverse consequences for gene flow and demography. It is subject to evolutionary change, which begs the question, what is the genetic basis of this potentially complex trait? To address this question, we (i) review the empirical literature on the genetic basis of dispersal, (ii) explore how theoretical investigations of the evolution of dispersal have represented the genetics of dispersal, and (iii) discuss how the genetic basis of dispersal influences theoretical predictions of the evolution of dispersal and potential consequences. Dispersal has a detectable genetic basis in many organisms, from bacteria to plants and animals. Generally, there is evidence for significant genetic variation for dispersal or dispersal‐related phenotypes or evidence for the micro‐evolution of dispersal in natural populations. Dispersal is typically the outcome of several interacting traits, and this complexity is reflected in its genetic architecture: while some genes of moderate to large effect can influence certain aspects of dispersal, dispersal traits are typically polygenic. Correlations among dispersal traits as well as between dispersal traits and other traits under selection are common, and the genetic basis of dispersal can be highly environment‐dependent. By contrast, models have historically considered a highly simplified genetic architecture of dispersal. It is only recently that models have started to consider multiple loci influencing dispersal, as well as non‐additive effects such as dominance and epistasis, showing that the genetic basis of dispersal can influence evolutionary rates and outcomes, especially under non‐equilibrium conditions. For example, the number of loci controlling dispersal can influence projected rates of dispersal evolution during range shifts and corresponding demographic impacts. Incorporating more realism in the genetic architecture of dispersal is thus necessary to enable models to move beyond the purely theoretical towards making more useful predictions of evolutionary and ecological dynamics under current and future environmental conditions. To inform these advances, empirical studies need to answer outstanding questions concerning whether specific genes underlie dispersal variation, the genetic architecture of context‐dependent dispersal phenotypes and behaviours, and correlations among dispersal and other traits.     

Genetics of host–parasite interactions: towards a comprehensive dissection of Drosophila resistance to viral infection

One of the major challenges in evolutionary biology is to unravel the genetic basis of adaptation. This issue has been gaining momentum in recent years with the accelerated development of novel genetic and genomic techniques and resources. In this issue of Molecular Ecology, Cogni et al. (2016) address the genetic basis of resistance to two viruses in Drosophila melanogaster using a panel of recombinant inbred lines with unprecedented resolution allowing detection of rare alleles and/or alleles of small effect. The study confirms the role of previously identified genes of major effect and adds novel regions with minor effect to the genetic basis of Drosophila resistance to the Drosophila C virus or the sigma virus. Additional analyses reveal the absence of cross‐resistance and of epistasis between the various genomic regions. This detailed information on the genetic architecture of host resistance constitutes an important step towards the understanding of both the physiology of antiviral immunity and the evolution of host–parasite interactions.     

Ethanol resistance in Drosophila melanogaster has increased in parallel cold‐adapted populations and shows a variable genetic architecture within and between populations

Understanding the genetic properties of adaptive trait evolution is a fundamental crux of biological inquiry that links molecular processes to biological diversity. Important uncertainties persist regarding the genetic predictability of adaptive trait change, the role of standing variation, and whether adaptation tends to result in the fixation of favored variants. Here, we use the recurrent evolution of enhanced ethanol resistance in Drosophila melanogaster during this species’ worldwide expansion as a promising system to add to our understanding of the genetics of adaptation. We find that elevated ethanol resistance has evolved at least three times in different cooler regions of the species’ modern range—not only at high latitude but also in two African high‐altitude regions. Applying a bulk segregant mapping framework, we find that the genetic architecture of ethanol resistance evolution differs substantially not only between our three resistant populations, but also between two crosses involving the same European population. We then apply population genetic scans for local adaptation within our quantitative trait locus regions, and we find potential contributions of genes with annotated roles in spindle localization, membrane composition, sterol and alcohol metabolism, and other processes. We also apply simulation‐based analyses that confirm the variable genetic basis of ethanol resistance and hint at a moderately polygenic architecture. However, these simulations indicate that larger‐scale studies will be needed to more clearly quantify the genetic architecture of adaptive evolution and to firmly connect trait evolution to specific causative loci.     

Genetic variation affecting host-parasite interactions: different genes affect different aspects of sigma virus replication and transmission in Drosophila melanogaster

In natural populations, genetic variation affects resistance to disease. Knowing how much variation exists, and understanding the genetic architecture of this variation, is important for medicine, for agriculture, and for understanding evolutionary processes. To investigate the extent and nature of genetic variation affecting resistance to pathogens, we are studying a tractable model system: Drosophila melanogaster and its natural pathogen the vertically transmitted sigma virus. We show that considerable genetic variation affects transmission of the virus from parent to offspring. However, maternal and paternal transmission of the virus is affected by different genes. Maternal transmission is a simple Mendelian trait: most of the genetic variation is explained by a polymorphism in ref(2)P, a gene already well known to affect resistance to sigma. In contrast, there is considerable genetic variation in paternal transmission that cannot be explained by ref(2)P and is caused by other loci on chromosome 2. Furthermore, we found no genetic correlation between paternal transmission of the virus and resistance to infection by the sigma virus following injection. This suggests that different loci affect viral replication and paternal transmission.     

Replicated analysis of the genetic architecture of quantitative traits in two wild great tit populations

Currently, there is much debate on the genetic architecture of quantitative traits in wild populations. Is trait variation influenced by many genes of small effect or by a few genes of major effect? Where is additive genetic variation located in the genome? Do the same loci cause similar phenotypic variation in different populations? Great tits (Parus major) have been studied extensively in long‐term studies across Europe and consequently are considered an ecological ‘model organism’. Recently, genomic resources have been developed for the great tit, including a custom SNP chip and genetic linkage map. In this study, we used a suite of approaches to investigate the genetic architecture of eight quantitative traits in two long‐term study populations of great tits—one in the Netherlands and the other in the United Kingdom. Overall, we found little evidence for the presence of genes of large effects in either population. Instead, traits appeared to be influenced by many genes of small effect, with conservative estimates of the number of contributing loci ranging from 31 to 310. Despite concordance between population‐specific heritabilities, we found no evidence for the presence of loci having similar effects in both populations. While population‐specific genetic architectures are possible, an undetected shared architecture cannot be rejected because of limited power to map loci of small and moderate effects. This study is one of few examples of genetic architecture analysis in replicated wild populations and highlights some of the challenges and limitations researchers will face when attempting similar molecular quantitative genetic studies in free‐living populations.     

Evidence for genetic hitchhiking effect associated with insecticide resistance in Aedes aegypti.

Information on genetic variation within and between populations is critical for understanding the evolutionary history of mosquito populations and disease epidemiology. Previous studies with Drosophila suggest that genetic variation of selectively neutral loci in a large fraction of genome may be constrained by fixation of advantageous mutations associated with hitchhiking effect. This study examined restriction fragment length polymorphisms of four natural Aedes aegypti mosquito populations from Trinidad and Tobago, at 16 loci. These populations have been subjected to organophosphate (OP) insecticide treatments for more than two decades, while dichlor-diphenyltrichlor (DDT) was the insecticide of choice prior to this period. We predicted that genes closely linked to the OP target loci would exhibit reduced genetic variation as a result of the hitchhiking effect associated with intensive OP insecticide selection. We also predicted that genetic variability of the genes conferring resistance to DDT and loci near the target site would be similar to other unlinked loci. As predicted, reduced genetic variation was found for loci in the general chromosomal region of a putative OP target site, and these loci generally exhibited larger F(ST) values than other random loci. In contrast, the gene conferring resistance to DDT and its linked loci show polymorphisms and genetic differentiation similar to other random loci. The reduced genetic variability and apparent gene deletion in some regions of chromosome 1 likely reflect the hitchhiking effect associated with OP insecticide selection.     

Drosophila-Host Genetic Control of Susceptibility to Drosophila C Virus

Interactions between Drosophila C virus (DCV) and its natural host, Drosophila melanogaster, were investigated using 15 geographical population samples infected by intraabdominal inoculation. These strains derived from natural populations of D. melanogaster differed in susceptibility to the DCV(C). One strain was ``partially tolerant'. Isofemale lines obtained from one susceptible and one partially tolerant strain were studied. The partially tolerant phenotype was dominant, and there was no difference between F(1) progeny of direct and reciprocal crosses. Analysis of F(2) progeny showed that neither sex-linked genes nor maternal effects are involved in susceptibility to DCV(C). The partially tolerant strain phenotype was dominant and segregated with chromosome III. Two nonexclusive hypotheses are proposed to explain chromosome III gene action.     

Costs and benefits of sublethal Drosophila C virus infection

Viruses are major evolutionary drivers of insect immune systems. Much of our knowledge of insect immune responses derives from experimental infections using the fruit fly Drosophila melanogaster. Most experiments, however, employ lethal pathogen doses through septic injury, frequently overwhelming host physiology. While this approach has revealed several immune mechanisms, it is less informative about the fitness costs hosts may experience during infection in the wild. Using both systemic and oral infection routes, we find that even apparently benign, sublethal infections with the horizontally transmitted Drosophila C virus (DCV) can cause significant physiological and behavioural morbidity that is relevant for host fitness. We describe DCV‐induced effects on fly reproductive output, digestive health and locomotor activity, and we find that viral morbidity varies according to the concentration of pathogen inoculum, host genetic background and sex. Notably, sublethal DCV infection resulted in a significant increase in fly reproduction, but this effect depended on host genotype. We discuss the relevance of sublethal morbidity for Drosophila ecology and evolution, and more broadly, we remark on the implications of deleterious and beneficial infections for the evolution of insect immunity.     

Diversity of genetic backgrounds modulating the durability of a major resistance gene. Analysis of a core collection of pepper landraces resistant to Potato virus Y

The evolution of resistance‐breaking capacity in pathogen populations has been shown to depend on the plant genetic background surrounding the resistance genes. We evaluated a core collection of pepper (Capsicum annuum) landraces, representing the worldwide genetic diversity, for its ability to modulate the breakdown frequency by Potato virus Y of major resistance alleles at the pvr2 locus encoding the eukaryotic initiation factor 4E (eIF4E). Depending on the pepper landrace, the breakdown frequency of a given resistance allele varied from 0% to 52.5%, attesting to their diversity and the availability of genetic backgrounds favourable to resistance durability in the plant germplasm. The mutations in the virus genome involved in resistance breakdown also differed between plant genotypes, indicating differential selection effects exerted on the virus population by the different genetic backgrounds. The breakdown frequency was positively correlated with the level of virus accumulation, confirming the impact of quantitative resistance loci on resistance durability. Among these loci, pvr6, encoding an isoform of eIF4E, was associated with a major effect on virus accumulation and on the breakdown frequency of the pvr2‐mediated resistance. This exploration of plant genetic diversity delivered new resources for the control of pathogen evolution and the increase in resistance durability.     

Host–parasite coevolution: genetic variation in a virus population and the interaction with a host gene

Host–parasite coevolution is considered to be an important factor in maintaining genetic variation in resistance to pathogens. Drosophila melanogaster is naturally infected by the sigma virus, a vertically transmitted and host‐specific pathogen. In fly populations, there is a large amount of genetic variation in the transmission rate from parent to offspring, much of which is caused by major‐effect resistance polymorphisms. We have found that there are similarly high levels of genetic variation in the rate of paternal transmission among 95 different isolates of the virus as in the host. However, when we examined a transmission‐blocking gene in the host, we found that it was effective across virus isolates. Therefore, the high levels of genetic variation observed in this system do not appear to be maintained because of coevolution resulting from interactions between this host gene and parasite genes.     

Evaluating the within-host fitness effects of mutations fixed during virus adaptation to different ecotypes of a new host

The existence of genetic variation for resistance in host populations is assumed to be essential to the spread of an emerging virus. Models predict that the rate of spread slows down with the increasing frequency and higher diversity of resistance alleles in the host population. We have been using the experimental pathosystem Arabidopsis thaliana—tobacco etch potyvirus (TEV) to explore the interplay between genetic variation in host''s susceptibility and virus diversity. We have recently shown that TEV populations evolving in A. thaliana ecotypes that differ in susceptibility to infection gained within-host fitness, virulence and infectivity in a manner compatible with a gene-for-gene model of host–parasite interactions: hard-to-infect ecotypes were infected by generalist viruses, whereas easy-to-infect ecotypes were infected by every virus. We characterized the genomes of the evolved viruses and found cases of host-driven convergent mutations. To gain further insights in the mechanistic basis of this gene-for-gene model, we have generated all viral mutations individually as well as in specific combinations and tested their within-host fitness effects across ecotypes. Most of these mutations were deleterious or neutral in their local ecotype and only a very reduced number had a host-specific beneficial effect. We conclude that most of the mutations fixed during the evolution experiment were so by drift or by selective sweeps along with the selected driver mutation. In addition, we evaluated the ruggedness of the underlying adaptive fitness landscape and found that mutational effects were mostly multiplicative, with few cases of significant epistasis.     

Genomic dissection of variation in clutch size and egg mass in a wild great tit (Parus major) population

Clutch size and egg mass are life history traits that have been extensively studied in wild bird populations, as life history theory predicts a negative trade‐off between them, either at the phenotypic or at the genetic level. Here, we analyse the genomic architecture of these heritable traits in a wild great tit (Parus major) population, using three marker‐based approaches – chromosome partitioning, quantitative trait locus (QTL) mapping and a genome‐wide association study (GWAS). The variance explained by each great tit chromosome scales with predicted chromosome size, no location in the genome contains genome‐wide significant QTL, and no individual SNPs are associated with a large proportion of phenotypic variation, all of which may suggest that variation in both traits is due to many loci of small effect, located across the genome. There is no evidence that any regions of the genome contribute significantly to both traits, which combined with a small, nonsignificant negative genetic covariance between the traits, suggests the absence of genetic constraints on the independent evolution of these traits. Our findings support the hypothesis that variation in life history traits in natural populations is likely to be determined by many loci of small effect spread throughout the genome, which are subject to continued input of variation by mutation and migration, although we cannot exclude the possibility of an additional input of major effect genes influencing either trait.     

A DNA virus of Drosophila

Little is known about the viruses infecting most species. Even in groups as well-studied as Drosophila, only a handful of viruses have been well-characterized. A viral metagenomic approach was used to explore viral diversity in 83 wild-caught Drosophila innubila, a mushroom feeding member of the quinaria group. A single fly that was injected with, and died from, Drosophila C Virus (DCV) was added to the sample as a control. Two-thirds of reads in the infected sample had DCV as the best BLAST hit, suggesting that the protocol developed is highly sensitive. In addition to the DCV hits, several sequences had Oryctes rhinoceros Nudivirus, a double-stranded DNA virus, as a best BLAST hit. The virus associated with these sequences was termed Drosophila innubila Nudivirus (DiNV). PCR screens of natural populations showed that DiNV was both common and widespread taxonomically and geographically. Electron microscopy confirms the presence of virions in fly fecal material similar in structure to other described Nudiviruses. In 2 species, D. innubila and D. falleni, the virus is associated with a severe (～80-90%) loss of fecundity and significantly decreased lifespan.     

Genetics of alcohol consumption in Drosophila melanogaster

Individual variation in alcohol consumption in human populations is determined by genetic, environmental, social and cultural factors. In contrast to humans, genetic contributions to complex behavioral phenotypes can be readily dissected in Drosophila, where both the genetic background and environment can be controlled and behaviors quantified through simple high‐throughput assays. Here, we measured voluntary consumption of ethanol in ～3000 individuals of each sex from an advanced intercross population derived from 37 lines of the Drosophila melanogaster Genetic Reference Panel. Extreme quantitative trait loci mapping identified 385 differentially segregating allelic variants located in or near 291 genes at P < 10^?8. The effects of single nucleotide polymorphisms associated with voluntary ethanol consumption are sex‐specific, as found for other alcohol‐related phenotypes. To assess causality, we used RNA interference knockdown or P{MiET1} mutants and their corresponding controls and functionally validated 86% of candidate genes in at least one sex. We constructed a genetic network comprised of 23 genes along with a separate trio and a pair of connected genes. Gene ontology analyses showed enrichment of developmental genes, including development of the nervous system. Furthermore, a network of human orthologs showed enrichment for signal transduction processes, protein metabolism and developmental processes, including nervous system development. Our results show that the genetic architecture that underlies variation in voluntary ethanol consumption is sexually dimorphic and partially overlaps with genetic factors that control variation in feeding behavior and alcohol sensitivity. This integrative genetic architecture is rooted in evolutionarily conserved features that can be extrapolated to human genetic interaction networks.     

Identification of candidate loci for adaptive phenotypic plasticity in natural populations of spadefoot toads

Phenotypic plasticity allows organisms to alter their phenotype in direct response to changes in the environment. Despite growing recognition of plasticity's role in ecology and evolution, few studies have probed plasticity's molecular bases—especially using natural populations. We investigated the genetic basis of phenotypic plasticity in natural populations of spadefoot toads (Spea multiplicata). Spea tadpoles normally develop into an “omnivore” morph that is favored in long‐lasting, low‐density ponds. However, if tadpoles consume freshwater shrimp or other tadpoles, they can alternatively develop (via plasticity) into a “carnivore” morph that is favored in ephemeral, high‐density ponds. By combining natural variation in pond ecology and morph production with population genetic approaches, we identified candidate loci associated with each morph (carnivores vs. omnivores) and loci associated with adaptive phenotypic plasticity (adaptive vs. maladaptive morph choice). Our candidate morph loci mapped to two genes, whereas our candidate plasticity loci mapped to 14 genes. In both cases, the identified genes tended to have functions related to their putative role in spadefoot tadpole biology. Our results thereby form the basis for future studies into the molecular mechanisms that mediate plasticity in spadefoots. More generally, these results illustrate how diverse loci might mediate adaptive plasticity.     

GENETIC ARCHITECTURE OF SKELETAL EVOLUTION IN EUROPEAN LAKE AND STREAM STICKLEBACK

Advances in genomic techniques are greatly facilitating the study of molecular signatures of selection in diverging natural populations. Connecting these signatures to phenotypes under selection remains challenging, but benefits from dissections of the genetic architecture of adaptive divergence. We here perform quantitative trait locus (QTL) mapping using 488 F₂ individuals and 2011 single nucleotide polymorphisms (SNPs) to explore the genetic architecture of skeletal divergence in a lake‐stream stickleback system from Central Europe. We find QTLs for gill raker, snout, and head length, vertebral number, and the extent of lateral plating (plate number and height). Although two large‐effect loci emerge, QTL effect sizes are generally small. Examining the neighborhood of the QTL‐linked SNPs identifies several genes involved in bone formation, which emerge as strong candidate genes for skeletal evolution. Finally, we use SNP data from the natural source populations to demonstrate that some SNPs linked to QTLs in our cross also exhibit striking allele frequency differences in the wild, suggesting a causal role of these QTLs in adaptive population divergence. Our study paves the way for comparative analyses across other (lake‐stream) stickleback populations, and for functional investigations of the candidate genes.