Melanogaster Populations Research Articles

Strong epistatic and additive effects of linked candidate SNPs for Drosophila pigmentation have implications for analysis of genome-wide association studies results

BackgroundThe mapping resolution of genome-wide association studies (GWAS) is limited by historic recombination events and effects are often assigned to haplotype blocks rather than individual SNPs. It is not clear how many of the SNPs in the block, and which ones, are causative. Drosophila pigmentation is a powerful model to dissect the genetic basis of intra-specific and inter-specific phenotypic variation. Three tightly linked SNPs in the t-MSE enhancer have been identified in three D. melanogaster populations as major contributors to female abdominal pigmentation. This enhancer controls the expression of the pigmentation gene tan (t) in the abdominal epidermis. Two of the three SNPs were confirmed in an independent study using the D. melanogaster Genetic Reference Panel established from a North American population.ResultsWe determined the functional impact of SNP1, SNP2, and SNP3 using transgenic lines to test all possible haplotypes in vivo. We show that all three candidate SNPs contribute to female Drosophila abdominal pigmentation. Interestingly, only two SNPs agree with the effect predicted by GWAS; the third one goes in the opposite direction because of linkage disequilibrium between multiple functional SNPs. Our experimental design uncovered strong additive effects for the three SNPs, but we also found significant epistatic effects explaining up to 11% of the total variation.ConclusionsOur results suggest that linked causal variants are important for the interpretation of GWAS and functional validation is needed to understand the genetic architecture of traits.

Reproductive Isolation through Experimental Manipulation of Sexually Antagonistic Coevolution in Drosophila melanogaster

Promiscuity can drive the evolution of sexual conflict before and after mating occurs. Post mating, the male ejaculate can selfishly manipulate female physiology, leading to a chemical arms race between the sexes. Theory suggests that drift and sexually antagonistic coevolution can cause allopatric populations to evolve different chemical interactions between the sexes, thereby leading to postmating reproductive barriers and speciation. There is, however, little empirical evidence supporting this form of speciation. We tested this theory by creating an experimental evolutionary model of Drosophila melanogaster populations undergoing different levels of interlocus sexual conflict. We found that allopatric populations under elevated sexual conflict show assortative mating, indicating premating reproductive isolation. Further, these allopatric populations also show reduced copulation duration and sperm defense ability when mating happens between individuals across populations compared to that within the same population, indicating postmating prezygotic isolation. Sexual conflict can cause reproductive isolation in allopatric populations through the coevolution of chemical (postmating prezygotic) as well as behavioural (premating) interactions between the sexes. Thus, to our knowledge, we provide the first comprehensive evidence of postmating (as well as premating) reproductive isolation due to sexual conflict.

Population Dynamics and Genetic Homogeneity in Natural Populations of Drosophila melanogaster from Faisalabad, Pakistan

Drosophila melanogaster has been considered an important model species to explore the patterns of diversity in natural populations. The current study was conducted to study population dynamics and genetic variability, gene flow and rate of migration in Faisalabad. The samples were collected from 17 different collection sites and cultured in the laboratory. The data for temperature and humidity were also recorded. It was observed that a decrease in temperature and increase in relative humidity caused an increase in population number and vice versa. Ten RAPD primers were used for PCR. Four primers were further selected on the basis of their efficiency and reproducibility. The number of amplification products varied from 4 to 10 with an average of 7.0 bands per primer. Mostly, all the samples showed monomorphic bands except for few samples from the south of the city. The allele and genotype frequencies were measured at each RAPD loci. Average observed heterozygosity (Ht 0.1886) and the genetic variation (G ST 0.129) showed that D. melanogaster natural populations had low level of genetic diversity. The dendrogram based on genetic similarities revealed three main groups of D. melanogaster populations. This pattern of genetic homogeneity due to low genetic variability in local populations was attributed to the gene flow with no genetic drift in D. melanogaster populations in Faisalabad.

Does increased heat resistance result in higher susceptibility to predation? A test using Drosophila melanogaster selection and hardening

Heat resistance of ectotherms can be increased both by plasticity and evolution, but these effects may have trade-offs resulting from biotic interactions. Here, we test for predation costs in Drosophila melanogaster populations with altered heat resistance produced by adult hardening and directional selection for increased heat resistance. In addition, we also tested for genetic trade-offs by testing heat resistance in lines that have evolved under increased predation risk. We show that while 35/37°C hardening increases heat resistance as expected, it does not increase predation risk from jumping spiders or mantids; in fact, there was an indication that survival may have increased under predation following a triple 37°C compared to a single 35°C hardening treatment. Flies that survived a 39°C selection cycle showed lower survival under predation, suggesting a predation cost of exposure to a more severe heat stress. There was, however, no correlated response to selection because survival did not differ between control and selected lines after selection was relaxed for one or two generations. In addition, lines selected for increased predation risk did not differ in heat resistance. Our findings suggest independent evolutionary responses to predation and heat as measured in laboratory assays, and no costs of heat hardening on susceptibility to predation.

Genetic Architecture of Natural Variation Underlying Adult Foraging Behavior That Is Essential for Survival of Drosophila melanogaster

Foraging behavior is critical for the fitness of individuals. However, the genetic basis of variation in foraging behavior and the evolutionary forces underlying such natural variation have rarely been investigated. We developed a systematic approach to assay the variation in survival rate in a foraging environment for adult flies derived from a wild Drosophila melanogaster population. Despite being such an essential trait, there is substantial variation of foraging behavior among D. melanogaster strains. Importantly, we provided the first evaluation of the potential caveats of using inbred Drosophila strains to perform genome-wide association studies on life-history traits, and concluded that inbreeding depression is unlikely a major contributor for the observed large variation in adult foraging behavior. We found that adult foraging behavior has a strong genetic component and, unlike larval foraging behavior, depends on multiple loci. Identified candidate genes are enriched in those with high expression in adult heads and, demonstrated by expression knock down assay, are involved in maintaining normal functions of the nervous system. Our study not only identified candidate genes for foraging behavior that is relevant to individual fitness, but also shed light on the initial stage underlying the evolution of the behavior.

Evolution of circadian rhythms in Drosophila melanogaster populations reared in constant light and dark regimes for over 330 generations

ABSTRACTOrganisms are believed to have evolved circadian clocks as adaptations to deal with cyclic environmental changes, and therefore it has been hypothesized that evolution in constant environments would lead to regression of such clocks. However, previous studies have yielded mixed results, and evolution of circadian clocks under constant conditions has remained an unsettled topic of debate in circadian biology. In continuation of our previous studies, which reported persistence of circadian rhythms in Drosophila melanogaster populations evolving under constant light, here we intended to examine whether circadian clocks and the associated properties evolve differently under constant light and constant darkness. In this regard, we assayed activity-rest, adult emergence and oviposition rhythms of D. melanogaster populations which have been maintained for over 19 years (~330 generations) under three different light regimes – constant light (LL), light–dark cycles of 12:12 h (LD) and constant darkness (DD). We observed that while circadian rhythms in all the three behaviors persist in both LL and DD stocks with no differences in circadian period, they differed in certain aspects of the entrained rhythms when compared to controls reared in rhythmic environment (LD). Interestingly, we also observed that DD stocks have evolved significantly higher robustness or power of free-running activity-rest and adult emergence rhythms compared to LL stocks. Thus, our study, in addition to corroborating previous results of circadian clock evolution in constant light, also highlights that, contrary to the expected regression of circadian clocks, rearing in constant darkness leads to the evolution of more robust circadian clocks which may be attributed to an intrinsic adaptive advantage of circadian clocks and/or pleiotropic functions of clock genes in other traits.

Is diapause an ancient adaptation in Drosophila?

D. melanogaster enters a state of reproductive arrest when exposed to low temperatures (12°C) and shorter photoperiods. A number of studies have suggested that diapause has recently evolved in European D. melanogaster populations, that it is not present in the sibling species D. simulans, that it is non-photoperiodic in American D. melanogaster populations, and that it spontaneously terminates after 6–8weeks. We have studied the overwintering phenotype under different conditions and observe that American, European and, surprisingly, African D. melanogaster populations can show photoperiodic diapause, as can European, but not African D. simulans. Surprisingly other Drosophila species from pan-tropical regions can also show significant levels of photoperiodic diapause. We observe that spontaneous termination of diapause after a few weeks can be largely avoided with a more realistic winter simulation for D. melanogaster, but not D. simulans. Examining metabolite accumulation during diapause reveals that the shallow diapause of D. melanogaster has similar features to that of other more robustly-diapausing species. Our results suggest that diapause may be an ancient character that emerged in the tropics to resist unfavourable seasonal conditions and which has been enhanced during D. melanogaster’s colonisation of temperate regions. Our results also highlight how different methodologies to quantify diapause can lead to apparently conflicting results that we believe can now largely be resolved.

A Hidden Markov Model Approach for Simultaneously Estimating Local Ancestry and Admixture Time Using Next Generation Sequence Data in Samples of Arbitrary Ploidy.

Admixture—the mixing of genomes from divergent populations—is increasingly appreciated as a central process in evolution. To characterize and quantify patterns of admixture across the genome, a number of methods have been developed for local ancestry inference. However, existing approaches have a number of shortcomings. First, all local ancestry inference methods require some prior assumption about the expected ancestry tract lengths. Second, existing methods generally require genotypes, which is not feasible to obtain for many next-generation sequencing projects. Third, many methods assume samples are diploid, however a wide variety of sequencing applications will fail to meet this assumption. To address these issues, we introduce a novel hidden Markov model for estimating local ancestry that models the read pileup data, rather than genotypes, is generalized to arbitrary ploidy, and can estimate the time since admixture during local ancestry inference. We demonstrate that our method can simultaneously estimate the time since admixture and local ancestry with good accuracy, and that it performs well on samples of high ploidy—i.e. 100 or more chromosomes. As this method is very general, we expect it will be useful for local ancestry inference in a wider variety of populations than what previously has been possible. We then applied our method to pooled sequencing data derived from populations of Drosophila melanogaster on an ancestry cline on the east coast of North America. We find that regions of local recombination rates are negatively correlated with the proportion of African ancestry, suggesting that selection against foreign ancestry is the least efficient in low recombination regions. Finally we show that clinal outlier loci are enriched for genes associated with gene regulatory functions, consistent with a role of regulatory evolution in ecological adaptation of admixed D. melanogaster populations. Our results illustrate the potential of local ancestry inference for elucidating fundamental evolutionary processes.

Hooked on Wolbachia

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.

Robust and scalable inference of population history from hundreds of unphased whole genomes.

It has recently been demonstrated that inference methods based on genealogical processes with recombination can uncover past population history in unprecedented detail. However, these methods scale poorly with sample size, limiting resolution in the recent past, and they require phased genomes, which contain switch errors that can catastrophically distort the inferred history. Here we present SMC++, a new statistical tool capable of analyzing orders of magnitude more samples than existing methods while requiring only unphased genomes (its results are independent of phasing). SMC++ can jointly infer population size histories and split times in diverged populations, and it employs a novel spline regularization scheme that greatly reduces estimation error. We apply SMC++ to analyze sequence data from over a thousand human genomes in Africa and Eurasia, hundreds of genomes from a Drosophila melanogaster population in Africa, and tens of genomes from zebra finch and long-tailed finch populations in Australia.

Whole genome resequencing of a laboratory-adapted Drosophila melanogaster population sample

As part of a study into the molecular genetics of sexually dimorphic complex traits, we used high-throughput sequencing to obtain data on genomic variation in an outbred laboratory-adapted fruit fly (Drosophila melanogaster) population. We successfully resequenced the whole genome of 220 hemiclonal females that were heterozygous for the same Berkeley reference line genome (BDGP6/dm6), and a unique haplotype from the outbred base population (LHM). The use of a static and known genetic background enabled us to obtain sequences from whole-genome phased haplotypes. We used a BWA-Picard-GATK pipeline for mapping sequence reads to the dm6 reference genome assembly, at a median depth-of coverage of 31X, and have made the resulting data publicly-available in the NCBI Short Read Archive (Accession number SRP058502). We used Haplotype Caller to discover and genotype 1,726,931 small genomic variants (SNPs and indels, <200bp). Additionally we detected and genotyped 167 large structural variants (1-100Kb in size) using GenomeStrip/2.0. Sequence and genotype data are publicly-available at the corresponding NCBI databases: Short Read Archive, dbSNP and dbVar (BioProject PRJNA282591). We have also released the unfiltered genotype data, and the code and logs for data processing and summary statistics (https://zenodo.org/communities/sussex_drosophila_sequencing/).

Whole genome resequencing of a laboratory-adapted Drosophila melanogaster.

As part of a study into the molecular genetics of sexually dimorphic complex traits, we used high-throughput sequencing to obtain data on genomic variation in an outbred laboratory-adapted fruit fly ( Drosophila melanogaster) population. We successfully resequenced the whole genome of 220 hemiclonal females that were heterozygous for the same Berkeley reference line genome (BDGP6/dm6), and a unique haplotype from the outbred base population (LH M). The use of a static and known genetic background enabled us to obtain sequences from whole-genome phased haplotypes. We used a BWA-Picard-GATK pipeline for mapping sequence reads to the dm6 reference genome assembly, at a median depth-of coverage of 31X, and have made the resulting data publicly-available in the NCBI Short Read Archive (Accession number SRP058502). We used Haplotype Caller to discover and genotype 1,726,931 small genomic variants (SNPs and indels, <200bp). Additionally we detected and genotyped 167 large structural variants (1-100Kb in size) using GenomeStrip/2.0. Sequence and genotype data are publicly-available at the corresponding NCBI databases: Short Read Archive, dbSNP and dbVar (BioProject PRJNA282591). We have also released the unfiltered genotype data, and the code and logs for data processing and summary statistics ( https://zenodo.org/communities/sussex_drosophila_sequencing/).

Bacterial Communities Differ among Drosophila melanogaster Populations and Affect Host Resistance against Parasitoids.

In Drosophila, diet is considered a prominent factor shaping the associated bacterial community. However, the host population background (e.g. genotype, geographical origin and founder effects) is a factor that may also exert a significant influence and is often overlooked. To test for population background effects, we characterized the bacterial communities in larvae of six genetically differentiated and geographically distant D. melanogaster lines collected from natural populations across Europe. The diet for these six lines had been identical for ca. 50 generations, thus any differences in the composition of the microbiome originates from the host populations. We also investigated whether induced shifts in the microbiome—in this case by controlled antibiotic administration—alters the hosts’ resistance to parasitism. Our data revealed a clear signature of population background on the diversity and composition of D. melanogaster microbiome that differed across lines, even after hosts had been maintained at the same diet and laboratory conditions for over 4 years. In particular, the number of bacterial OTUs per line ranged from 8 to 39 OTUs. Each line harboured 2 to 28 unique OTUs, and OTUs that were highly abundant in some lines were entirely missing in others. Moreover, we found that the response to antibiotic treatment differed among the lines and significantly altered the host resistance to the parasitoid Asobara tabida in one of the six lines. Wolbachia, a widespread intracellular endosymbiont associated with parasitoid resistance, was lacking in this line, suggesting that other components of the Drosophila microbiome caused a change in host resistance. Collectively, our results revealed that lines that originate from different population backgrounds show significant differences in the established Drosophila microbiome, outpacing the long-term effect of diet. Perturbations on these naturally assembled microbiomes to some degree influenced the hosts’ resistance against natural parasites.

Experimental Evolution and Heart Function in Drosophila.

Drosophila melanogaster is a good model species for the study of heart function. However, most previous work on D. melanogaster heart function has focused on the effects of large-effect genetic variants. We compare heart function among 18 D. melanogaster populations that have been selected for altered development time, aging, or stress resistance. We find that populations with faster development and faster aging have increased heart dysfunction, measured as percentage heart failure after electrical pacing. Experimental evolution of different triglyceride levels, by contrast, has little effect on heart function. Evolved differences in heart function correlate with allele frequency changes at many loci of small effect. Genomic analysis of these populations produces a list of candidate loci that might affect cardiac function at the intersection of development, aging, and metabolic control mechanisms.

Whole genome resequencing of a laboratory-adapted Drosophila melanogaster population sample.

As part of a study into the molecular genetics of sexually dimorphic complex traits, we used next-generation sequencing to obtain data on genomic variation in an outbred laboratory-adapted fruit fly (Drosophila melanogaster) population. We successfully resequenced the whole genome of 220 hemiclonal females that were heterozygous for the same Berkeley reference line genome (BDGP6/dm6), and a unique haplotype from the outbred base population (LHM). The use of a static and known genetic background enabled us to obtain sequences from whole genome phased haplotypes. We used a BWA-Picard-GATK pipeline for mapping sequence reads to the dm6 reference genome assembly, at a median depth of coverage of 31X, and have made the resulting data publicly-available in the NCBI Short Read Archive (Accession number SRP058502). We used Haplotype Caller to discover and genotype 1,726,931 small genomic variants (SNPs and indels, <200bp). Additionally we detected and genotyped 167 large structural variants (1-100Kb in size) using GenomeStrip/2.0. Sequence and genotype data are publicly-available at the corresponding NCBI databases: Short Read Archive, dbSNP and dbVar (BioProject PRJNA282591). We have also released the unfiltered genotype data, and the code and logs for data processing and summary statistics ( https://zenodo.org/communities/sussex_drosophila_sequencing/).

Patterns of physiological decline due to age and selection in Drosophila melanogaster.

In outbred sexually reproducing populations, age-specific mortality rates reach a plateau in late life following the exponential increase in mortality rates that marks aging. Little is known about what happens to physiology when cohorts transition from aging to late life. We measured age-specific values for starvation resistance, desiccation resistance, time-in-motion, and geotaxis in ten Drosophila melanogaster populations: five populations selected for rapid development and five control populations. Adulthood was divided into two stages, the aging phase and the late-life phase according to demographic data. Consistent with previous studies, we found that populations selected for rapid development entered the late-life phase at an earlier age than the controls. Age-specific rates of change for all physiological phenotypes showed differences between the aging phase and the late-life phase. This result suggests that late life is physiologically distinct from aging. The ages of transitions in physiological characteristics from aging to late life statistically match the age at which the demographic transition from aging to late life occurs, in all cases but one. These experimental results support evolutionary theories of late life that depend on patterns of decline and stabilization in the forces of natural selection.

Reconstruction of Haplotype-Blocks Selected during Experimental Evolution.

The genetic analysis of experimentally evolving populations typically relies on short reads from pooled individuals (Pool-Seq). While this method provides reliable allele frequency estimates, the underlying haplotype structure remains poorly characterized. With small population sizes and adaptive variants that start from low frequencies, the interpretation of selection signatures in most Evolve and Resequencing studies remains challenging. To facilitate the characterization of selection targets, we propose a new approach that reconstructs selected haplotypes from replicated time series, using Pool-Seq data. We identify selected haplotypes through the correlated frequencies of alleles carried by them. Computer simulations indicate that selected haplotype-blocks of several Mb can be reconstructed with high confidence and low error rates, even when allele frequencies change only by 20% across three replicates. Applying this method to real data from D. melanogaster populations adapting to a hot environment, we identify a selected haplotype-block of 6.93 Mb. We confirm the presence of this haplotype-block in evolved populations by experimental haplotyping, demonstrating the power and accuracy of our haplotype reconstruction from Pool-Seq data. We propose that the combination of allele frequency estimates with haplotype information will provide the key to understanding the dynamics of adaptive alleles.

Drosophila melanogaster males evolve increased courtship as a correlated response to larval crowding

A competitive resource-based environment such as larval crowding is an important stress factor in the life history of holometabolous insects such as Drosophila melanogaster and can have major consequences for adult fitness through its direct effects on body size. We investigated precopulatory sexual behaviour, which is an important subset of adult fitness, using D. melanogaster populations adapted to larval crowding for 100+ generations. We found that selected males evolved significantly higher courtship frequency than their ancestral controls. While we found a negative correlation between larval density and adult courtship frequency using low, moderate and high larval rearing densities, we also found that supplementation of live yeast, which is an important dietary component in their life cycle, caused a significant rise in courtship frequency. We further dissected out the male and female behavioural components that contribute to overall courtship activity by factorially combining selected and ancestral male and female types, reared in both low and high larval densities, and found that courtship frequency of selected males was higher with selected females than with control females. We also quantified the mating frequency, a crude measure for mating success, and found that increased courtship frequency in selected populations did not lead to increased mating success. From the results of this and a previous study, we conclude that precopulatory sexual activity in these populations is not traded for life span.

Drosophila Adaptation to Viral Infection through Defensive Symbiont Evolution.

Microbial symbionts can modulate host interactions with biotic and abiotic factors. Such interactions may affect the evolutionary trajectories of both host and symbiont. Wolbachia protects Drosophila melanogaster against several viral infections and the strength of the protection varies between variants of this endosymbiont. Since Wolbachia is maternally transmitted, its fitness depends on the fitness of its host. Therefore, Wolbachia populations may be under selection when Drosophila is subjected to viral infection. Here we show that in D. melanogaster populations selected for increased survival upon infection with Drosophila C virus there is a strong selection coefficient for specific Wolbachia variants, leading to their fixation. Flies carrying these selected Wolbachia variants have higher survival and fertility upon viral infection when compared to flies with the other variants. These findings demonstrate how the interaction of a host with pathogens shapes the genetic composition of symbiont populations. Furthermore, host adaptation can result from the evolution of its symbionts, with host and symbiont functioning as a single evolutionary unit.

An Intronic Polymorphism in couch potato Is Not Distributed Clinally in European Drosophila melanogaster Populations nor Does It Affect Diapause Inducibility.

couch potato (cpo) encodes an RNA binding protein that has been reported to be expressed in the peripheral and central nervous system of embryos, larvae and adults, including the major endocrine organ, the ring gland. A polymorphism in the D. melanogaster cpo gene coding region displays a latitudinal cline in frequency in North American populations, but as cpo lies within the inversion In(3R)Payne, which is at high frequencies and itself shows a strong cline on this continent, interpretation of the cpo cline is not straightforward. A second downstream SNP in strong linkage disequilibrium with the first has been claimed to be primarily responsible for the latitudinal cline in diapause incidence in USA populations.Here, we investigate the frequencies of these two cpo SNPs in populations of Drosophila throughout continental Europe. The advantage of studying cpo variation in Europe is the very low frequency of In(3R)Payne, which we reveal here, does not appear to be clinally distributed. We observe a very different geographical scenario for cpo variation from the one in North America, suggesting that the downstream SNP does not play a role in diapause. In an attempt to verify whether the SNPs influence diapause we subsequently generated lines with different combinations of the two cpo SNPs on known timeless (tim) genetic backgrounds, because polymorphism in the clock gene tim plays a significant role in diapause inducibility. Our results reveal that the downstream cpo SNP does not seem to play any role in diapause induction in European populations in contrast to the upstream coding cpo SNP. Consequently, all future diapause studies on strains of D. melanogaster should initially determine their tim and cpo status.