Drosophila Americana Research Articles

High-Quality Genome Assemblies Reveal Evolutionary Dynamics of Repetitive DNA and Structural Rearrangements in the Drosophila virilis Subgroup.

High-quality genome assemblies across a range of nontraditional model organisms can accelerate the discovery of novel aspects of genome evolution. The Drosophila virilis group has several attributes that distinguish it from more highly studied species in the Drosophila genus, such as an unusual abundance of repetitive elements and extensive karyotype evolution, in addition to being an attractive model for speciation genetics. Here, we used long-read sequencing to assemble five genomes of three virilis group species and characterized sequence and structural divergence and repetitive DNA evolution. We find that our contiguous genome assemblies allow characterization of chromosomal arrangements with ease and can facilitate analysis of inversion breakpoints. We also leverage a small panel of resequenced strains to explore the genomic pattern of divergence and polymorphism in this species and show that known demographic histories largely predicts the extent of genome-wide segregating polymorphism. We further find that a neo-X chromosome in Drosophila americana displays X-like levels of nucleotide diversity. We also found that unusual repetitive elements were responsible for much of the divergence in genome composition among species. Helitron-derived tandem repeats tripled in abundance on the Y chromosome in D. americana compared to Drosophila novamexicana, accounting for most of the difference in repeat content between these sister species. Repeats with characteristics of both transposable elements and satellite DNAs expanded by 3-fold, mostly in euchromatin, in both D. americana and D. novamexicana compared to D. virilis. Our results represent a major advance in our understanding of genome biology in this emerging model clade.

Drosophilids with darker cuticle have higher body temperature under light

Cuticle pigmentation was shown to be associated with body temperature for several relatively large species of insects, but it was questioned for small insects. Here we used a thermal camera to assess the association between drosophilid cuticle pigmentation and body temperature increase when individuals are exposed to light. We compared mutants of large effects within species (Drosophila melanogaster ebony and yellow mutants). Then we analyzed the impact of naturally occurring pigmentation variation within species complexes (Drosophila americana/Drosophila novamexicana and Drosophila yakuba/Drosophila santomea). Finally we analyzed lines of D. melanogaster with moderate differences in pigmentation. We found significant differences in temperatures for each of the four pairs we analyzed. The temperature differences appeared to be proportional to the differently pigmented area: between Drosophila melanogaster ebony and yellow mutants or between Drosophila americana and Drosophila novamexicana, for which the whole body is differently pigmented, the temperature difference was around 0.6 °C ± 0.2 °C. By contrast, between D. yakuba and D. santomea or between Drosophila melanogaster Dark and Pale lines, for which only the posterior abdomen is differentially pigmented, we detected a temperature difference of about 0.14 °C ± 0.10 °C. This strongly suggests that cuticle pigmentation has ecological implications in drosophilids regarding adaptation to environmental temperature.

A shift to shorter cuticular hydrocarbons accompanies sexual isolation among Drosophila americana group populations.

Because sensory signals often evolve rapidly, they could be instrumental in the emergence of reproductive isolation between species. However, pinpointing their specific contribution to isolating barriers, and the mechanisms underlying their divergence, remains challenging. Here, we demonstrate sexual isolation due to divergence in chemical signals between two populations of Drosophila americana (SC and NE) and one population of D. novamexicana, and dissect its underlying phenotypic and genetic mechanisms. Mating trials revealed strong sexual isolation between Drosophila novamexicana males and SC Drosophila americana females, as well as more moderate bi‐directional isolation between D. americana populations. Mating behavior data indicate SC D. americana males have the highest courtship efficiency and, unlike males of the other populations, are accepted by females of all species. Quantification of cuticular hydrocarbon (CHC) profiles—chemosensory signals that are used for species recognition and mate finding in Drosophila—shows that the SC D. americana population differs from the other populations primarily on the basis of compound carbon chain‐length. Moreover, manipulation of male CHC composition via heterospecific perfuming—specifically perfuming D. novamexicana males with SC D. americana males—abolishes their sexual isolation from these D. americana females. Of a set of candidates, a single gene—elongase CG17821—had patterns of gene expression consistent with a role in CHC differences between species. Sequence comparisons indicate D. novamexicana and our Nebraska (NE) D. americana population share a derived CG17821 truncation mutation that could also contribute to their shared “short” CHC phenotype. Together, these data suggest an evolutionary model for the origin and spread of this allele and its consequences for CHC divergence and sexual isolation in this group.

Differences in Postmating Transcriptional Responses between Conspecific and Heterospecific Matings in Drosophila.

In many animal species, females undergo physiological and behavioral changes after mating. Some of these changes are driven by male-derived seminal fluid proteins and are critical for fertilization success. Unfortunately, our understanding of the molecular interplay between female and male reproductive proteins remains inadequate. Here, we analyze the postmating response in a Drosophila species that has evolved strong gametic incompatibility with its sister species; Drosophila novamexicana females produce only ∼1% fertilized eggs in crosses with Drosophila americana males, compared to ∼98% produced in within-species crosses. This incompatibility is likely caused by mismatched male and female reproductive molecules. In this study, we use short-read RNA sequencing to examine the evolutionary dynamics of female reproductive genes and the postmating transcriptome response in crosses within and between species. First, we found that most female reproductive tract genes are slow-evolving compared to the genome average. Second, postmating responses in con- and heterospecific matings are largely congruent, but heterospecific matings induce expression of additional stress-response genes. Some of those are immunity genes that are activated by the Imd pathway. We also identify several genes in the JAK/STAT signaling pathway that are induced in heterospecific, but not conspecific mating. While this immune response was most pronounced in the female reproductive tract, we also detect it in the female head and ovaries. These results show that the female’s postmating transcriptome-level response is determined in part by the genotype of the male, and that divergence in male reproductive genes and/or traits can have immunogenic effects on females.

Genetic architecture of a body colour cline in Drosophila americana.

Phenotypic variation within a species is often structured geographically in clines. In Drosophila americana, a longitudinal cline for body colour exists within North America that appears to be due to local adaptation. The tan and ebony genes have been hypothesized to contribute to this cline, with alleles of both genes that lighten body colour found in D.americana. These alleles are similar in sequence and function to the allele fixed in D.americana's more lightly pigmented sister species, Drosophila novamexicana. Here, we examine the frequency and geographic distribution of these D.novamexicana-like alleles in D.americana. Among alleles from over 100 strains of D.americana isolated from 21 geographic locations, we failed to identify additional alleles of tan or ebony with as much sequence similarity to D.novamexicana as the D.novamexicana-like alleles previously described. However, using genetic analysis of 51 D.americana strains derived from 20 geographic locations, we identified one new allele of ebony and one new allele of tan segregating in D.americana that are functionally equivalent to the D.novamexicana allele. An additional 5 alleles of tan also showed marginal evidence of functional similarity. Given the rarity of these alleles, however, we conclude that they are unlikely to be driving the pigmentation cline. Indeed, phenotypic distributions of the 51 backcross populations analysed indicate a more complex genetic architecture, with diversity in the number and effects of loci altering pigmentation observed both within and among populations of D.americana. This genetic heterogeneity poses a challenge to association studies and genomic scans for clinal variation, but might be common in natural populations.

Constitutive and Plastic Gene Expression Variation Associated with Desiccation Resistance Differences in the Drosophila americana Species Group.

Stress response mechanisms are ubiquitous and important for adaptation to heterogenous environments and could be based on constitutive or plastic responses to environmental stressors. Here we quantify constitutive and plastic gene expression differences under ambient and desiccation stress treatments, in males and females of three species of Drosophila known to differ in desiccation resistance. Drosophila novamexicana survives desiccation trials significantly longer than the two subspecies of Drosophila americana, consistent with its natural species range in the desert southwest USA. We found that desiccation stress reduces global expression differences between species—likely because many general stress response mechanisms are shared among species—but that all species showed plastic expression changes at hundreds of loci during desiccation. Nonetheless, D. novamexicana had the fewest genes with significant plastic expression changes, despite having the highest desiccation resistance. Of the genes that were significantly differentially expressed between species—either within each treatment (>200 loci), constitutively regardless of treatment (36 loci), or with different species-specific plasticity (26 loci)—GO analysis did not find significant enrichment of any major gene pathways or broader functions associated with desiccation stress. Taken together, these data indicate that if gene expression changes contribute to differential desiccation resistance between species, these differences are likely shaped by a relatively small set of influential genes rather than broad genome-wide differentiation in stress response mechanisms. Finally, among the set of genes with the greatest between-species plasticity, we identified an interesting set of immune-response genes with consistent but opposing reaction norms between sexes, whose potential functional role in sex-specific mechanisms of desiccation resistance remains to be determined.

Change in the magnitude of meiotic drive due to increased temperature

While major changes to chromosomal form is generally deleterious, karyotypes are not static. Major chromosomal rearrangements contribute to genetic isolation and divergence within and between species. As mechanisms that drive these divergences have not been fully elucidated. We hypothesize that meiotic drive, ie. bias transmission from parent to offspring, has contributed to the reshaping of karyotypes. Drosophila americana and Drosophila novamexicana are sister species that are an excellent model to study such evolutionary forces. Since diverging from D. novamexicana, D. americana has had two major chromosomal rearrangements: centromere fusions between the 2nd and 3rd chromosomes and the X and 4th chromosome. The X-4 chromosome has remained polymorphic. It is found in a latitudinal gradient along the Mississippi River. Northern populations contain more of the fused arrangement while southern populations have more of the unfused arrangement. With meiotic drive favoring the X-4 chromosome, it should fix in the population. There are two leading hypotheses explaining the polymorphic nature of the X-4 fused chromosome. First, the meiotic drive is only activated in times of stress, which is more consistent in Northern populations. Second, Meiotic drive occurs in all flies, but selection against the fused X-4 chromosome is high in climates that are more associated with the Southern United States. This study was intended to identify if increased temperature affects meiotic drive. We measured inheritance patterns from heterozygous females to offspring at an increased temperature. Then the offspring’s DNA was extracted. PCR and Gel electrophoresis was done to identify which chromosome was inherited.

Desiccation resistance and pigmentation variation reflects bioclimatic differences in the Drosophila americana species complex

BackgroundDisentangling the selective factors shaping adaptive trait variation is an important but challenging task. Many studies—especially in Drosophila—have documented trait variation along latitudinal or altitudinal clines, but frequently lack resolution about specific environmental gradients that could be causal selective agents, and often do not investigate covariation between traits simultaneously. Here we examined variation in multiple macroecological factors across geographic space and their associations with variation in three physiological traits (desiccation resistance, UV resistance, and pigmentation) at both population and species scales, to address the role of abiotic environment in shaping trait variation.ResultsUsing environmental data from collection locations of three North American Drosophila species—D. americana americana, D. americana texana and D. novamexicana—we identified two primary axes of macroecological variation; these differentiated species habitats and were strongly loaded for precipitation and moisture variables. In nine focal populations (three per species) assayed for each trait, we detected significant species-level variation for both desiccation resistance and pigmentation, but not for UV resistance. Species-level trait variation was consistent with differential natural selection imposed by variation in habitat water availability, although patterns of variation differed between desiccation resistance and pigmentation, and we found little evidence for pleiotropy between traits.ConclusionsOur multi-faceted approach enabled us to identify potential agents of natural selection and examine how they might influence the evolution of multiple traits at different evolutionary scales. Our findings highlight that environmental factors influence functional trait variation in ways that can be complex, and point to the importance of studies that examine these relationships at both population- and species-levels.

Achiasmy and sex chromosome evolution

Abstract Achiasmy is a phenomenon whereby chiasma (“chromosomal cross–over”) formation is absent during meiosis in one or both sexes. It is one of the strategies that have evolved to cease recombination between sex chromosomes in various taxa. Absence of recombination makes Y (W) chromosome diverge from X (Z) chromosome via natural selection on linked genes. In the present review, we first review the research history of achiasmy in Drosophila species and summarize the prevalence of achiasmy in a wide range of animal taxa and the rapid turnover of achiasmy. We then briefly describe the mechanism of achiasmatic meiosis in Drosophila melanogaster and the evolution of meiotic genes in Drosophila species. Second, we discuss sex chromosome evolution in achiasmatic species. If meiotic recombination had been suppressed between sex chromosomes, heterogametic sex chromosomes have no chance of meiotic recombination. Therefore, the neo–Y chromosome which originated from Y–autosome fusion in achiasmatic species has no chance of recombination and no pre-existing genetic diversity since its origination. On the contrary, if the neo-Y chromosome had chance of recombination, it can maintain ancestral polymorphisms. We compare three different evolutionary trajectories of neo–Y chromosome in Drosophila americana, Drosophila miranda, and Drosophila albomicans. Lastly, we discuss the origin of Y chromosome in achiasmatic species. In particular, we introduce hypotheses proposed in three insect species that achiasmatic meiosis may facilitate the recruitment of an accessory B chromosome to a Y (W) chromosome.

Origin and Consequences of Chromosomal Inversions in the virilis Group of Drosophila.

In Drosophila, large variations in rearrangement rate have been reported among different lineages and among Muller’s elements. Nevertheless, the mechanisms that are involved in the generation of inversions, their increase in frequency, as well as their impact on the genome are not completely understood. This is in part due to the lack of comparative studies on species distantly related to Drosophila melanogaster. Therefore, we sequenced and assembled the genomes of two species of the virilis phylad (Drosophila novamexicana [15010-1031.00] and Drosophila americana [SF12]), which are diverging from D. melanogaster for more than 40 Myr. Based on these data, we identified the precise location of six novel inversion breakpoints. A molecular characterization provided clear evidence that DAIBAM (a miniature inverted–repeat transposable element) was involved in the generation of eight out of the nine inversions identified. In contrast to what has been previously reported for D. melanogaster and close relatives, ectopic recombination is thus the prevalent mechanism of generating inversions in species of the virilis phylad. Using pool-sequencing data for three populations of D. americana, we also show that common polymorphic inversions create a high degree of genetic differentiation between populations for chromosomes X, 4, and 5 over large physical distances. We did not find statistically significant differences in expression levels between D. americana (SF12) and D. novamexicana (15010-1031.00) strains for the three genes surveyed (CG9588, Fig 4, and fab1) flanking three inversion breakpoints.

Genes Belonging to the Insulin and Ecdysone Signaling Pathways Can Contribute to Developmental Time, Lifespan and Abdominal Size Variation in Drosophila americana

Even within a single genus, such as Drosophila, cases of lineage-specific adaptive evolution have been found. Therefore, the molecular basis of phenotypic variation must be addressed in more than one species group, in order to infer general patterns. In this work, we used D. americana, a species distantly-related to D. melanogaster, to perform an F2 association study for developmental time (DT), chill-coma recovery time (CRT), abdominal size (AS) and lifespan (LS) involving the two strains (H5 and W11) whose genomes have been previously sequenced. Significant associations were found between the 43 large indel markers developed here and DT, AS and LS but not with CRT. Significant correlations are also found between DT and LS, and between AS and LS, that might be explained by variation at genes belonging to the insulin and ecdysone signaling pathways. Since, in this F2 association study a single marker, located close to the Ecdysone receptor (EcR) gene, explained as much as 32.6% of the total variation in DT, we performed a second F2 association study, to determine whether large differences in DT are always due to variation in this genome region. No overlapping signal was observed between the two F2 association studies. Overall, these results illustrate that, in D. americana, pleiotropic genes involved in the highly-conserved insulin and ecdysone signaling pathways are likely responsible for variation observed in ecologically relevant phenotypic traits, although other genes are also involved.

Drosophila americana as a Model Species for Comparative Studies on the Molecular Basis of Phenotypic Variation

Understanding the molecular basis of within and between species phenotypic variation is one of the main goals of Biology. In Drosophila, most of the work regarding this issue has been performed in D. melanogaster, but other distantly related species must also be studied to verify the generality of the findings obtained for this species. Here, we make the case for D. americana, a species of the virilis group of Drosophila that has been diverging from the model species, D. melanogaster, for approximately 40 Myr. To determine the suitability of this species for such studies, polymorphism and recombination estimates are presented for D. americana based on the largest nucleotide sequence polymorphism data set so far analyzed (more than 100 data sets) for this species. The polymorphism estimates are also compared with those obtained from the comparison of the genome assembly of two D. americana strains (H5 and W11) here reported. As an example of the general utility of these resources, we perform a preliminary study on the molecular basis of lifespan differences in D. americana. First, we show that there are lifespan differences between D. americana populations from different regions of the distribution range. Then, we perform five F2 association experiments using markers for 21 candidate genes previously identified in D. melanogaster. Significant associations are found between polymorphism at two genes (hep and Lim3) and lifespan. For the F2 association study involving the two sequenced strains (H5 and W11), we identify amino acid differences at Lim3 and Hep that could be responsible for the observed changes in lifespan. For both genes, no large gene expression differences were observed between the two strains.

Local adaptation for body color in Drosophila americana: commentary on Wittkopp et al.

Wittkopp et al. (2010) report clinal variation of cuticle coloration among populations of Drosophila americana despite the high levels of gene flow across collection sites, providing evidence for local (darwinian) adaptation of body coloration in this species. Several mechanisms have been proposed to explain the evolutionary patterns of pigment variation in ectothermic organisms such as D. americana. For example, the occurrence of dark-pigmented populations of several Drosophila spp. has been associated with dry environments (for example, Parkash et al., 2010). This relationship represents the main hypothesis tested by Wittkopp et al. (2010), and the authors conclude that pigmentation and desiccation resistance are not unequivocally linked among the populations investigated. However, other climatic explanations that are not addressed in the study may explain these color patterns. For example, the thermal melanism hypothesis (TMH) suggests that if all else is equal, darker individuals may heat up faster and reach higher equilibrium temperatures than lighter individuals, providing a benefit in cooler conditions (Clusella-Trullas et al., 2007). Recently, the TMH has received support at several spatial scales in reptiles (Clusella-Trullas et al., 2008, 2009) and has also been demonstrated in many insect species (for example, Brakefield and Willmer, 1985; reviewed in Clusella-Trullas et al., 2007). Wittkopp et al. (2010) showed that longitude was a better explanatory variable than environmental relative humidity. However, latitude and longitude are typically used as proxies of climate (for example, temperature and humidity) and site (topography) variation. The use of longitude as a predictor of pigment variation does not indicate which climate variables (or interactions thereof) might best explain the patterns observed, and therefore provides little insight into the potential evolutionary mechanisms at work.

Local adaptation for body color in Drosophila americana.

Pigmentation is one of the most variable traits within and between Drosophila species. Much of this diversity appears to be adaptive, with environmental factors often invoked as selective forces. Here, we describe the geographic structure of pigmentation in Drosophila americana and evaluate the hypothesis that it is a locally adapted trait. Body pigmentation was quantified using digital images and spectrometry in up to 10 flies from each of 93 isofemale lines collected from 17 locations across the United States and found to correlate most strongly with longitude. Sequence variation at putatively neutral loci showed no evidence of population structure and was inconsistent with an isolation-by-distance model, suggesting that the pigmentation cline exists despite extensive gene flow throughout the species range, and is most likely the product of natural selection. In all other Drosophila species examined to date, dark pigmentation is associated with arid habitats; however, in D. americana, the darkest flies were collected from the most humid regions. To investigate this relationship further, we examined desiccation resistance attributable to an allele that darkens pigmentation in D. americana. We found no significant effect of pigmentation on desiccation resistance in this experiment, suggesting that pigmentation and desiccation resistance are not unequivocally linked in all Drosophila species.

Reduced Effectiveness of Selection Caused by a Lack of Recombination

Genetic recombination associated with sexual reproduction is expected to have important consequences for the effectiveness of natural selection. These effects may be evident within genomes, in the form of contrasting patterns of molecular variation and evolution in regions with different levels of recombination. Previous work reveals patterns that are consistent with a benefit of recombination for adaptation at the level of protein sequence: both positive selection for adaptive variants and purifying selection against deleterious ones appear to be compromised in regions of low recombination [1-11]. Here, we re-examine these patterns by using polymorphism and divergence data from the Drosophila dot chromosome, which has a long history of reduced recombination. To avoid confounding selection and demographic effects, we collected these data from a species with an apparently stable demographic history, Drosophila americana. We find that D. americana dot loci show several signatures of ineffective purifying and positive selection, including an increase in the rate of protein evolution, an increase in protein polymorphism, and a reduction in the proportion of amino acid substitutions attributable to positive selection.

An X chromosome effect responsible for asymmetric reproductive isolation between maleDrosophila virilisand heterospecific females

Reproductive isolation between closely related species is expressed through uncoordinated courtship, failed fertilization, and (or) postzygotic barriers. Behavioural components of mating often form an initial barrier to hybridization between species. In many animals, females are responsible for mating discrimination in both intra- and interspecific crosses; males of Drosophila virilis group represent an exception to this trend. Using overall productivity tests, we show that a lower proportion of D. virilis males sire progeny when paired with a heterospecific female (Drosophila novamexicana or Drosophila americana texana) for 2 weeks. This suggests male mate discrimination or some other kind of asymmetrical incompatibility in courtship and mating or early zygote mortality. We used males from D. virilis-D. novamexicana and from D. virilis-D. a. texana backcross populations to map chromosome effects responsible for male reproductive isolation. Results from the analysis of both backcross male populations indicate a major X chromosome effect. Further, we conduct a male behavioural analysis to show that D. virilis males significantly fail to continue courtship after the first step of courtship, when they tap heterospecific females. The combined results of a major X chromosome effect and the observation that D. virilis males walk away from females after tapping suggest that future studies should concentrate on the identification of X-linked genes affecting the ability of males to recognize conspecific females.

An old bilbo-like non-LTR retroelement insertion provides insight into the relationship of species of the virilis group

In Drosophila, at the population and species level, fixation of a TE insertion is an unlikely fate. Of the few reported fixations at the species level most involve non-LTR retroelements. In this work we report the fixation of a non-LTR retroelement in five species ( Drosophila littoralis, Drosophila virilis, Drosophila lummei, Drosophila americana and Drosophila novamexicana) of the virilis group of Drosophila. In most species, this TE insertion is being lost through the accumulation of small deletions, but there is also evidence for the accumulation of large deletions. In the americana lineage an insertion of about 900 bp of the non-LTR retroelement is a marker for the Xc inversion. This insertion is, at most, 80 kb away from the basal Xc inversion breakpoint. The presence of a bilbo-like element in D. littoralis but not in D. kanekoi, suggests that D. littoralis is more closely related to species of the virilis phylad than to species of the montana phylad, which is in contrast with the traditional view. Nevertheless, the phylogenetic analyses here performed using a 7 gene dataset suggest that D. littoralis is indeed more closely related to species of the virilis phylad than to species of the montana phylad. The re-evaluation of the phylogenetic relationship of the species of the virilis group, under the assumption of a relaxed molecular clock, results in an estimated age of the bilbo-like element insertion of at least 7.5 Mya.

CLINAL DISTRIBUTION OF A CHROMOSOMAL REARRANGEMENT: A PRECURSOR TO CHROMOSOMAL SPECIATION?

Geographically structured genetic variants provide an effective means to assess sources of natural selection and mechanisms of adaptation to local environments. Correlated selection pressures along environmental gradients favor subdivision of genomes through chromosomal rearrangement. This study examines populations of Drosophila americana to evaluate selection pressures affecting chromosomal forms distinguished by a centromeric fusion. Analyses of chromosomal polymorphism throughout the Mississippi River Valley in the central United States reveal the presence of a distinct latitudinal cline for the chromosomal rearrangement. The cline has a width of 623 km centered at 35.97 degrees N and displays a characteristic sigmoid shape consistent with a balance between selection and dispersal. Extreme low temperature during January, an indicator of winter severity, was identified as the environmental variable that most accurately predicts arrangement frequency. An intriguing relationship identified between the chromosomal cline and operational sex ratio indicates that these alternative arrangements of the X chromosome may influence sex-specific survival. A hypothesis for the cline is presented wherein variation associated with the alternative chromosome forms influences distinct overwintering strategies. The resulting subdivision within the genome embodies a transitory stage of a speciation process in which locally adapted gene complexes provide a foundation for species formation.

Inferring the evolutionary history ofDrosophila americanaandDrosophila novamexicanausing a multilocus approach and the influence of chromosomal rearrangements in single gene analyses

The evolutionary history of closely related organisms can prove sometimes difficult to infer. Hybridization and incomplete lineage sorting are the main concerns; however, genome rearrangements can also influence the outcome of analyses based on nuclear sequences. In the present study, DNA sequences from 12 nuclear genes, for which the approximate chromosomal locations are known, have been used to estimate the evolutionary history of two forms of Drosophila americana (Drosophila americana americana and Drosophila americana texana) and Drosophila novamexicana (virilis group of species). The phylogenetic analysis of the combined data set resulted in a phylogeny showing reciprocal monophyly for D. novamexicana and D. americana. Single gene analyses, however, resulted in incongruent phylogenies influenced by chromosomal rearrangements. Genetic differentiation estimates indicated a significant differentiation between the two species for all genes. Within D. americana, however, there is no evidence for differentiation between the chromosomal forms except at genes located near the X/4 fusion and Xc inversion breakpoint. Thus, the specific status of D. americana and D. novamexicana is confirmed, but there is no overall evidence for genetic differentiation between D. a. americana and D. a. texana, not supporting a subspecific status. Based on levels of allele and nucleotide diversity found in the strains used, it is proposed that D. americana has had a stable, large population during the recent past while D. novamexicana has speciated from a peripheral southwestern population having had an ancestral small effective population size. The influence of chromosomal rearrangements in single gene analyses is also examined.

Genetic basis of reduced eyes in the hybrids of Drosophila virilis phylad species.

Structural anomalies were detected in F1 and backcross hybrids of Drosophila virilis and three of its sibling species Drosophila novamexicana, Drosophila americana texana and Drosophila americana americana. Similar but not identical anomalies have been described in D. virilis x D. lummei hybrids. F1 (D. virilis x D. novamexicana) hybrids had only slight disorders in the abdominal cuticula, but 5.3% of the first backcross generation hybrids had reduced eyes. Eye defects in the second generation were due to interaction of the heterozygous fourth chromosome of D. novamexicana with the homozygous D. virilis chromosomes 2 and 3. Reciprocal F1 hybrids between D. virilis and D. a. americana and between D. virilis and D. a. texana had normal eyes, but in the second hybrid generation 4.9% of D. virilis x D. a. americana and 1.6% of D. virilis x D. a. texana hybrids had anomalous eyes. In D. virilis x D. a. texana hybrids, the heterozygous fourth chromosome of D. a. texana was incompatible with homozygous D. virilis chromosomes 2 and 3. Of the D. virilis x D. a. americana hybrids homozygous for D. virilis chromosomes 2 and 3, 10.2% had the eye defect. The incompatibility system of non-conspecific chromosomes causing the eye anomalies in the hybrids of these three species pairs was different from those in D. virilis x D. lummei hybrids and has evolved independently after the diverging of the American lineage from D. lummei.