Molecular evolution of duplicated amylase gene regions in Drosophila melanogaster: evidence of positive selection in the coding regions and selective constraints in the cis-regulatory regions

In this study, we randomly sampled Drosophila melanogaster from Japanese and Kenyan natural populations. We sequenced duplicated (proximal and distal) Amy gene regions to test whether the patterns of polymorphism were consistent with neutral molecular evolution. F(st) between the two geographically distant populations, estimated from Amy gene regions, was 0.084, smaller than reported values for other loci, comparing African and Asian populations. Furthermore, little genetic differentiation was found at a microsatellite locus (DROYANETSB) in these samples (G'st = -0.018). The results of several tests (Tajima's, Fu and Li's, and Wall's tests) were not significantly different from neutrality. However, a significantly higher level of fixed replacement substitutions was detected by a modified McDonald and Kreitman test for both populations. This indicates that positive selection occurred during or immediately after the speciation of D. melanogaster. Sliding-window analysis showed that the proximal region 1, a part of the proximal 5' flanking region, was conserved between D. melanogaster and its sibling species, D. simulans. An HKA test was significant when the proximal region 1 was compared with the 5' flanking region of Alcohol dehydrogenase (Adh), indicating a severe selective constraint on the Amy proximal region 1. These results suggest that natural selection has played an important role in the molecular evolution of Amy gene regions in D. melanogaster.     

Comparative population genomics of latitudinal variation in Drosophila simulans and Drosophila melanogaster

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.     

Phylogeny of the Oriental Drosophila melanogaster species group: a multilocus reconstruction

The melanogaster species group of Drosophila (subgenus Sophophora) has long been a favored model for evolutionary studies because of its morphological and ecological diversity and wide geographic distribution. However, phylogenetic relationships among species and subgroups within this lineage are not well understood. We reconstructed the phylogeny of 17 species representing 7 "oriental" species subgroups, which are especially closely related to D. melanogaster. We used DNA sequences of four nuclear and two mitochondrial loci in an attempt to obtain the best possible estimate of species phylogeny and to assess the extent and sources of remaining uncertainties. Comparison of trees derived from single-gene data sets allowed us to identify several strongly supported clades, which were also consistently seen in combined analyses. The relationships among these clades are less certain. The combined data set contains data partitions that are incongruent with each other. Trees reconstructed from the combined data set and from internally homogenous data sets consisting of three or four genes each differ at several deep nodes. The total data set tree is fully resolved and strongly supported at most nodes. Statistical tests indicated that this tree is compatible with all individual and combined data sets. Therefore, we accepted this tree as the most likely model of historical relationships. We compared the new molecular phylogeny to earlier estimates based on morphology and chromosome structure and discuss its taxonomic and evolutionary implications.     

Quantitative genetic variation and selection of enzyme activities in Drosophila melanogaster

The nature of the genetic variation for the activity of three enzymes (α-GPD, ME, and SOD) was studied by means of analyses of variance among full-sib and half-sib families. The results presented here indicate that the genetic variation of activity of these enzymes consist primarily of non-additive genetic variance. A moderate level of additive genetic variation was found only for α-GPD activity. We also examined the question whether an association exists between enzyme activities and selection for preadult developmental time. Using the method developed by Lande and Arnold (1983), significant directional selection was observed for α-GPD activity.     

Demography and natural selection have shaped genetic variation in Drosophila melanogaster: a multi-locus approach

Demography and selection have been recognized for their important roles in shaping patterns of nucleotide variability. To investigate the relative effects of these forces in the genome of Drosophila melanogaster, we used a multi-locus scan (105 fragments) of X-linked DNA sequence variation in a putatively ancestral African and a derived European population. Surprisingly, we found evidence for a recent size expansion in the African population, i.e., a significant excess of singletons at a chromosome-wide level. In the European population, such an excess was not detected. In contrast to the African population, we found evidence for positive natural selection in the European sample: (i) a large number of loci with low levels of variation and (ii) a significant excess of derived variants at the low-variation loci that are fixed in the European sample but rare in the African population. These results are consistent with the hypothesis that the European population has experienced frequent selective sweeps in the recent past during its adaptation to new habitats. Our study shows the advantages of a genomic approach (over a locus-specific analysis) in disentangling demographic and selective forces.     

Genetic variation in odorant receptors contributes to variation in olfactory behavior in a natural population of Drosophila melanogaster

Chemoreception is a principle modality by which organisms gain information from their environment, and extensive variation in odor-mediated behavior has been documented within and among species. To examine the mechanisms by which sensory systems mediate these responses, we ask to what extent variation in Drosophila melanogaster odorant receptor genes contributes to variation in odor-mediated behavior. Significant differences in behavioral responses to structurally similar odorants, methyl hexanoate and ethyl hexanoate, were found in a natural population. Polymorphisms in 3 genomic regions (Or22a/Or22b, Or35a, and Or47a) were identified and associated with variation in behavior to these esters. Overall similarity in association profiles for both odorants was observed, except for Or47a in which polymorphisms were associated solely with variation in responses to ethyl hexanoate. Our analyses were then extended to examine polymorphisms in 3 odorant receptors previously reported to contribute to variation in olfactory behavior for the chemically distinct odorants benzaldehyde and acetophenone. Two Or10a polymorphisms were associated with variation in response to ethyl hexanoate. Finally, differences in Or35a and Or47a expression were associated with variation in responses to ethyl hexanoate. These results demonstrate that the genetic variation at the peripheral sensory stage plays a role in mediating differences in odor-mediated behavior.     

Correlated responses to selection for stress resistance and longevity in a laboratory population of Drosophila melanogaster

Laboratory studies on Drosophila have revealed that resistance to one environmental stress often correlates with resistance to other stresses. There is also evidence on genetic correlations between stress resistance, longevity and other fitness-related traits. The present work investigates these associations using artificial selection in Drosophila melanogaster. Adult flies were selected for increased survival after severe cold, heat, desiccation and starvation stresses as well as increased heat-knockdown time and lifespan (CS, HS, DS, SS, KS and LS line sets, respectively). The number of selection generations was 11 for LS, 27 for SS and 21 for other lines, with selection intensity being around 0.80. For each set of lines, the five stress-resistance parameters mentioned above as well as longevity (in a nonstressful environment) were estimated. In addition, preadult developmental time, early age productivity and thorax length were examined in all lines reared under nonstressful conditions. Comparing the selection lines with unselected control revealed clear-cut direct selection responses for the stress-resistance traits. Starvation resistance increased as correlated response in all sets of selection lines, with the exception of HS. Positive correlated responses were also found for survival after cold shock (HS and DS) and heat shock (KS and DS). With regard to values of resistance across different stress assays, the HS and KS lines were most similar. The resistance values of the SS lines were close to those of the LS lines and tended to be the lowest among all selection lines. Developmental time was extended in the SS and KS lines, whereas the LS lines showed a reduction in thorax length. The results indicate a possibility of different multiple-stress-resistance mechanisms for the examined traits and fitness costs associated with stress resistance and longevity.     

Nonclinality of molecular variation implicates selection in maintaining a morphological cline of Drosophila melanogaster

One general approach for assessing whether phenotypic variation is due to selection is to test its correlation with presumably neutral molecular variation. Neutral variation is determined by population history, the most likely alternative explanation of spatial genetic structure, whereas phenotypic variation may be influenced by the spatial pattern of selection pressure. Several methods for comparing the spatial apportionment of molecular and morphological variation have been used. Here, we present an analysis of variance framework that compares the magnitudes of latitudinal effects for molecular and morphological variation along a body size cline in Australian Drosophila populations. Explicit incorporation of the relevant environmental gradient can result in a simple and powerful test of selection. For the Australian cline, our analysis provides strong internal evidence that the cline is due to selection.     

Temporal dynamics and variation of multilocus ISSR-PCR DNA markers in the Uman' population of Drosophila melanogaster over two decades (1984-2004)

The temporal dynamics of genomic variation in the Uman' (Ukraine) population of Drosophila melanogaster over the period 1984-2004 was studied using multilocus ISSR markers. Considerable polymorphism of the genomic DNA fragments corresponding to ISSR markers was found in the D. melanogaster population studied: the values of average heterozygosity varied from 0.085 to 0.127 depending on the year. Significant differences in the frequencies of dominant alleles between the samples of different years were recorded for 12 of the 30 DNA fractions detected. These changes are nondirectional and random. The pattern of detected variation suggests the determining influence of gene drift and migration process on the variation of noncoding DNA sequences in the Uman' population of D. melanogaster.     

Analysis of allozymic and quantitative variation produced by artificial selection in Drosophila melanogaster

In order to study a possible connection between allozymic and quantitative variation in D. melanogaster, three selection experiments were carried out, using founder strains of known genetic and chromosomal composition.The Adh and Gpdh-1 enzyme loci have been used as genetic markers and the maximum wing length has been the quantitative trait chosen. Two selected lines (high and low) were maintained and also one without selection (drift), to estimate the effect of random fluctuation on gene frequency variation. The allozymic variation was analysed by means of a polynomial regression, and a normal linear model allowed to make pairwise comparisons.The allelic combination ((F), A(-)) was favoured in the low lines of the selection experiments; the selection acted in favour of homozygotes, with a correlated loss of genetic homoeostasis. The similar behaviour of the drift and the control lines shows that random fluctuations in the gene frequencies in selected lines are negligible.     

Genetic variation in Drosophila melanogaster resistance to infection: a comparison across bacteria

Insects use a generalized immune response to combat bacterial infection. We have previously noted that natural populations of D. melanogaster harbor substantial genetic variation for antibacterial immunocompetence and that much of this variation can be mapped to genes that are known to play direct roles in immunity. It was not known, however, whether the phenotypic effects of variation in these genes are general across the range of potentially infectious bacteria. To address this question, we have reinfected the same set of D. melanogaster lines with Serratia marcescens, the bacterium used in the previous study, and with three additional bacteria that were isolated from the hemolymph of wild-caught D. melanogaster. Two of the new bacteria, Enterococcus faecalis and Lactococcus lactis, are gram positive. The third, Providencia burhodogranaria, is gram negative like S. marcescens. Drosophila genotypes vary highly significantly in bacterial load sustained after infection with each of the four bacteria, but mean loads are largely uncorrelated across bacteria. We have tested statistical associations between immunity phenotypes and nucleotide polymorphism in 21 candidate immunity genes. We find that molecular variation in some genes, such as Tehao, contributes to phenotypic variation in the suppression of only a subset of the pathogens. Variation in SR-CII and 18-wheeler, however, has effects that are more general. Although markers in SR-CII and 18-wheeler explain >20% of the phenotypic variation in resistance to L. lactis and E. faecalis, respectively, most of the molecular polymorphisms tested explain <10% of the total variance in bacterial load sustained after infection.     

Chromosomal polymorphism and extra bristles of Drosophila melanogaster: joint variation under selection in isofemale lines

Selection for increased numbers of dorsocentral and scutellar bristles produced an increase of In(3R)C heterozygotes in isofemale lines of D. melanogaster. The influence of polygenic selection in the maintenance of the chromosomal polymorphism is discussed.     

Multiple signatures of positive selection downstream of notch on the X chromosome in Drosophila melanogaster

To identify genomic regions affected by the rapid fixation of beneficial mutations (selective sweeps), we performed a scan of microsatellite variability across the Notch locus region of Drosophila melanogaster. Nine microsatellites spanning 60 kb of the X chromosome were surveyed for variation in one African and three non-African populations of this species. The microsatellites identified an approximately 14-kb window for which we observed relatively low levels of variability and/or a skew in the frequency spectrum toward rare alleles, patterns predicted at regions linked to a selective sweep. DNA sequence polymorphism data were subsequently collected within this 14-kb region for three of the D. melanogaster populations. The sequence data strongly support the initial microsatellite findings; in the non-African populations there is evidence of a recent selective sweep downstream of the Notch locus near or within the open reading frames CG18508 and Fcp3C. In addition, we observe a significant McDonald-Kreitman test result suggesting too many amino acid fixations species wide, presumably due to positive selection, at the unannotated open reading frame CG18508. Thus, we observe within this small genomic region evidence for both recent (skew toward rare alleles in non-African populations) and recurring (amino acid evolution at CG18508) episodes of positive selection.     

Adaptation to fluctuating environments in a selection experiment with Drosophila melanogaster

A fundamental question in life‐history evolution is how organisms cope with fluctuating environments, including variation between stressful and benign conditions. For short‐lived organisms, environments commonly vary between generations. Using a novel experimental design, we exposed wild‐derived Drosophila melanogaster to three different selection regimes: one where generations alternated between starvation and benign conditions, and starvation was always preceded by early exposure to cold; another where starvation and benign conditions alternated in the same way, but cold shock sometimes preceded starvation and sometimes benign conditions; and a third where conditions were always benign. Using six replicate populations per selection regime, we found that selected flies increased their starvation resistance, most strongly for the regime where cold and starvation were reliably combined, and this occurred without decreased fecundity or extended developmental time. The selected flies became stress resistant, displayed a pronounced increase in early life food intake and resource storage. In contrast to previous experiments selecting for increased starvation resistance in D. melanogaster, we did not find increased storage of lipids as the main response, but instead that, in particular for females, storage of carbohydrates was more pronounced. We argue that faster mobilization of carbohydrates is advantageous in fluctuating environments and conclude that the phenotype that evolved in our experiment corresponds to a compromise between the requirements of stressful and benign environments.     

The effect of directed and stabilizing selection on the population size of Drosophila melanogaster

Population size as a measure of population fitness was assayed in laboratory lines of Drosophila melanogaster subjected to directional selection for positive photoactivity and stabilizing selection for a complex of metric wing characters. Three experimental designs were used: directional and stabilizing selection (DS); directional selection (D); unselected controls (C). Population size increased under all three experimental designs. The population size of DS and C lines as well as the rate of increase of the DS lines were higher as compared to the D lines. The results suggest that stabilizing selection maintains population fitness at the higher level.