Sex‐ratio in natural and experimental populations of Drosophila subobscura from North America

The sex‐ratio (SR), defined as the proportion of males, has been studied in three North American colonizing populations of Drosophila subobscura (Eureka, Davis and Gilroy). The proportion of sexes under laboratory conditions was studied using the one‐generation serial transfer technique in one‐ and two‐species populations, to infer whether biased SR affects the outcome when competing with Drosophila pseudoobscura, another member of the same group now in sympatry with D. subobscura in North America. The wild samples of D. subobscura yielded a significantly higher number of males than females during those months where the species is more abundant. However, there was no significant deviation in the 1 : 1 proportion of sexes in the descendants of D. subobscura at any of the experimental conditions. On the contrary, D. pseudoobscura produced a higher proportion of females which could be responsible for the exclusion of D. subobscura in laboratory competition experiments with overlapping generations. Thus, if sexes are equal at birth and survival is similar, the preponderance of males of D. subobscura in our wild collections could indicate greater activity and probably greater chance of dispersal of males versus females especially under favourable conditions.     

Preadult competition between Drosophila subobscura and Drosophila pseudoobscura

The Palaearctic species Drosophila subobscura has recently colonized a large area of North America where it coexists with Drosophila pseudoobscura. The viability and developmental rate of these species were studied at 13 d?C, 18d?C and 23 d?C and at densities of 10, 50, 100 and 200 eggs per vial. The two species were differently affected by density and temperature in the ranges studied. Both intra- and interspecific cultures showed that D. pseudoobscura was best adapted to 23 d?C, where it was clearly the dominant species. On the other hand, at 18 d?C and especially at 13 d?C, although D. subobscura was less viable than D. pseudoobscura, its developmental time was shorter, which may give advantage to this species. Results reported here agree with the observed distribution of these species in North America.     

FOUNDER-FLUSH SPECIATION IN DROSOPHILA PSEUDOOBSCURA: A LARGE-SCALE EXPERIMENT

A founder-flush-crash model of speciation has been proposed that may particularly apply to island and other colonizations. Previous laboratory experiments testing the model have given inconsistent results. We have conducted a large experiment with Drosophila pseudoobscura designed to meet the essential postulates of the model and to separately test some of the postulates. Forty-five experimental and 12 control populations have been studied during seven successive founder-flush-crash cycles, or about 50 generations. Sexual isolation tests yield significantly positive assortative mating in a few tests between pairs of experimental populations. Populations with fewer founders (N = 1 or 3) yield more significant instances of assortative mating than those with more founders (n = 5, 7, or 9), and this difference becomes statistically significant for pooled data. Only one of 15 population pairs tested three times (generations 25, 32, and 46) shows positive assortative mating in all three tests. No significant assortative mating occurs between control populations, including highly inbred ones. We conclude that although founder events may occasionally lead to the evolution of assortative mating and hence to speciation, our results do not support the claim that the founder-flush-crash model identifies conditions very likely to result in speciation events.     

MITOCHONDRIAL DNA PHYLOGENIES FOR THE DROSOPHILA OBSCURA GROUP

Species belonging to the obscura group of the genus Drosophila have long held a central position in evolutionary studies, especially in experimental population genetics. Despite the considerable amount of accumulated knowledge, many of the phylogenetic relationships of the species in the group remain unclear. Here we present DNA sequence data for the mitochondrial gene cytochrome oxidase I (COI) for 13 species native to both the Old and New Worlds. We combine these data with seven other mitochondrial gene sequences from previous studies, for a total of over 3 kb per species. Strongly supported conclusions include: (1) the two North American subgroups, pseudoobscura and affinis, are each monophyletic; and (2) among Eurasian species two unambiguous clades are identified, one containing D. tristis, D. ambigua, and D. obscura and the other containing D. guanche, D. subobscura, and D. madeirensis. Constructing firm hypotheses connecting these four major clades is problematic with all datasets. Major ambiguities are the number of invasions giving rise to the North American obscura species and the relationships among the Eurasian species. The inadequacy of the mtDNA data to resolve these ambiguities does not reside in lack of changes; the transversions-only parsimony tree has 283 informative characters. Rather, the problems are likely intrinsic to the history of the group: while radiating in temperate Eurasia, North America was colonized once or twice, followed by one or two radiations in the New World.     

The genetics of Drosophila subobscura populations. 3. Inversion polymorphism and climatic factors

Summary The natural population of Drosophila subobscura of Mt. Parnes has been followed for two successive years and the frequency of the gene arrangements determined. Some changes occured in the first year but do not explain the existence of the geographical clines for these frequencies. Within the second year no changes were observed. For chromosomes J and U there is evidence for a greater fitness of the heterozygotes.Experiments on temperature effect on viability of adult male flies, on their temporal sterility, on the viability of flies during all their biological cycle and experiments on the effect of dryness on the viability of adult male flies did not show an important change of gene arrangement frequencies in the resistant individuals. Nor did diapausing females have very different gene arrangements, except for chromosome O. Cage experiments with temperature as a selective agent did not elucidate the action of this factor. It is concluded that temperature or dryness do not affect very much the polymorphic system, nor explain the existence of these clines.This polymorphic system is much more stable in D. subobscura than in D. pseudoobscura, its nearctic relative. It is also richer. It is concluded that it is historically older and helps the populations to cope with many environmental changes.     

GENETICS OF SEXUAL ISOLATION AND COURTSHIP DYSFUNCTION IN MALE HYBRIDS OF DROSOPHILA PSEUDOOBSCURA AND DROSOPHILA PERSIMILIS

Despite the importance of sexual isolation to speciation, few studies have analyzed the genetic basis of interspecific mating discrimination, particularly using hybrid males. In this study, I investigated the genetic basis of sexual isolation using male hybrids of Drosophila pseudoobscura and D. persimilis. Hybrid male mating success was caused by interactions between the X-chromosome and autosomes (or Y-chromosome), and different arms of the X-chromosome contributed to mating success with females of each species. Further, although there was an X-chromosome component to mating success, its magnitude was not disproportionately large when compared with the proportion of the genome contained on this chromosome. Some hybrid males courted with an anomalously low intensity, so I simultaneously mapped the genetic basis of this “courtship dysfunction.” The courtship dysfunction was caused by an interaction between the left arm of the X-chromosome in D. persimilis with the autosomes or Y-chromosome from D. pseudoobscura. Anomalous courtship behavior in interspecific hybrids can obscure the conclusions of studies of the genetics of sexual isolation, so courtship intensity should be evaluated in all such investigations.     

Clan Lethals and Inter-Year Allelism in Death Valley DROSOPHILA PSEUDOOBSCURA

A chromosome 2 lethal allelism rate of about 3% was found in the 1974 population of D. pseudoobscura in Death Valley, California. This rate was significantly higher than allelism rates in other Southern California populations. The Death Valley population was sampled again in 1975 and 1977, with allelism rates of 1% and 0.5%, respectively. In 1974, several lethals were in high frequencies (about 1%), a pattern that reappeared in 1975 and 1977. However, none of the lethals in high frequency one year were in high frequency another year; the particular lethal alleles present in this ephemeral population appear to be due to their random presence in the flies which refound the population every winter. The results for the Death Valley population are compared with a Japanese population of D. melanogaster in which lethals in high frequency one year are also in high frequency in succeeding years and with earlier work on chromosome 3 of D. pseudoobscura, which showed a lower lethal frequency and higher allelism rate.     

Analysis of inbreeding in a colonizing population of Drosophila subobscura

An analysis of the effects of inbreeding on the genetic structure of a colonizing population of Drosophila subobscura has been carried out. Species of Drosophila, particularly D. subobscura, may have lethal alleles associated with chromosomal inversions and our aim was to assess the extent to which the genome is balanced in this way. The frequencies of chromosomal inversions were compared between a large population and a set of 72 lines that were maintained by brother-sister mating for 10 generations. Fisher's matrix method was used to calculate the expected homozygosity in these inbred lines for 5 allozyme loci (Aph, Hk-1, Lap, Odh, and Pept-1) used as markers of large chromosomal segments. Furthermore, the expected rates of fixation corresponding to these allozyme loci were also calculated. The results show that the amount of homozygosis observed did not differ significantly from expectations (with the corresponding loss of lines as a consequence of the reduction in viability). However, two deviations from strict neutrality were observed: there was a heterozygote excess at the Lap locus, and the frequency of the O ₅ inversion (always associated with a lethal gene in colonizing populations) was higher than expected.     

Chromosomal inversion polymorphism of Drosophila subobscura populations from Jastrebac Mountain shows temporal and habitat-related changes

Inversion polymorphism in populations of D. subobscura from a beech forest on Jastrebac mountain was studied in June 1990, 1993, and 1994, respectively. The same analysis was performed in 1990 for D. subobscura populations in a beech forest and an oak forest in the same region. Statistically significant differences in the frequencies of the gene arrangements of A₁, J and U chromosome were observed during the period of investigation. A tendency towards a decrease in the frequency of the standard gene arrangements was found for all chromosomes, but was particularly evident with chromosomes A and J. The frequency of the gene arrangements A₁ A₂, J₁ and U_1–2+6 increased at the same time. Differences in the frequency of the gene arrangements of A, J and U chromosomes were also observed when the populations from two ecologically different habitats (beech and oak forest) were compared in 1990.     

STRUCTURE AND POPULATION GENETICS OF THE BREAKPOINTS OF A POLYMORPHIC INVERSION IN DROSOPHILA SUBOBSCURA

Drosophila subobscura is a paleartic species of the obscura group with a rich chromosomal polymorphism. To further our understanding on the origin of inversions and on how they regain variation, we have identified and sequenced the two breakpoints of a polymorphic inversion of D. subobscura—inversion 3 of the O chromosome—in a population sample. The breakpoints could be identified as two rather short fragments (～300 bp and 60 bp long) with no similarity to any known transposable element family or repetitive sequence. The presence of the ～300‐bp fragment at the two breakpoints of inverted chromosomes implies its duplication, an indication of the inversion origin via staggered double‐strand breaks. Present results and previous findings support that the mode of origin of inversions is neither related to the inversion age nor species‐group specific. The breakpoint regions do not consistently exhibit the lower level of variation within and stronger genetic differentiation between arrangements than more internal regions that would be expected, even in moderately small inversions, if gene conversion were greatly restricted at inversion breakpoints. Comparison of the proximal breakpoint region in species of the obscura group shows that this breakpoint lies in a small high‐turnover fragment within a long collinear region (～300 kb).     

Contest competition inDrosophila subobscura

The role of larval intraspecific competition in laboratory populations ofDrosophila subobscura was investigated. Mortality is density-independent during the first 3 days after hatching but becomes density dependent as development proceeds to pupation. Although total biomass per patch was independent of initial egg density, competition betweenDrosophila larvae leads to the formation of smaller pupae. This resulted in a population that was dominated by suppressed individuals. Development rate ofD. subobscura larvae was not affected by high larval densities. Smaller pupae give rise to females with fewer eggs in their ovarioles. A simple simulation model, predicting the effects of intraspecific competition on the fecundity of the nextDrosophila generation is described.     

P1 clones from Drosophila melanogaster as markers to study the chromosomal evolution of Muller's A element in two species of the obscura group of Drosophila

Thirty P1 clones from the X chromosome (Muller's A element) of Drosophila melanogaster were cross-hybridized in situ to Drosophila subobscura and Drosophila pseudoobscura polytene chromosomes. An additional recombinant phage Dsuby was also used as a marker. Twenty-three (77%) of the P1 clones gave positive hybridization on D. pseudoobscura chromosomes bat only 16 (53%) did so with those of D. subobscura. Eight P1 clones gave more than one hybridization signal on D. pseudoobscura and/or D. subobscura chromosomes. All P1 clones and Dsuby hybridized on Muller's A element (X chromosome) of D. subobscura. In contrast, only 18 P1 clones and Dsuby hybridized on Muller's D element (XR chromosomal arm) of D. pseudoobscura; 4 additional P1 clones hybridized on Muller's D element (XR chromosomal arm) of this species and the remaining P1 clone gave on hybridization signal on each arm of the X chromosome. This latter clone may contain one breakpoint of a pericentric inversion that may account for the interchange of genetic material between Muller's A and D elements in D. pseudoobscura. In contrast to the rare interchange of genetic material between chromosomal elements, profound differences in the order and spacing of markers were detected between D. melanogaster, D. pseudoobscura and D. subobscura. In fact, the number of chromosomal segments delimited by identical markers and conserved between pairwise comparisons is small. Therefore, extensive reorganization within Muller's A element has been produced during the divergence of the three species. Rough estimates of the number of cytologically detectable inversions contributing to differentiation of Muller's A element were obtained. The most reliable of these estimates is that obtained from the D. pseudoobscura and D. melanogaster comparison since a greater number of markers have been mapped in both species. Tentatively, one inversion breakpoint about every 200 kb has been produced and fixed during the divergence of D. pseudoobscura and D. melanogaster.     

Phylogeny of the Drosophila obscura group as inferred from one- and two-dimensional protein electrophoresis

The phylogenetic relationships of 15 species of the obscura group of Drosophila were analysed by use of one- and two-dimensional electrophoresis. Genetic distances based on two-dimensional data are five times smaller than those based on native proteins. From the data, it is proposed that the species radiation of the obscura group happened in two evolutionary bursts, the first one giving rise to at least four palearctic proto-lineages (bifasciata, obscura (including D. subsilvestris), subobscura, and microlabis) and one or two proto-nearctic lineages (affinis, pseudoobscura), and the second, more recent burst giving rise to the current speciation within lineages.     

DNA variation at the rp49 gene region in Drosophila madeirensis and D. subobscura from Madeira: inferences about the origin of an insular endemic species

An ~1.6 ‐ kb fragment spanning the rp49 gene was sequenced in 16 lines of Drosophila subobscura from Madeira and in 22 lines of the endemic species D. madeirensis. Nucleotide diversity in D. subobscura from Madeira (π=0.0081) was similar to that in lines from Spain carrying the O₃₊₄ chromosomal arrangement (π=0.0080). No significant genetic differentiation was detected between insular and continental O₃₊₄ lines of D. subobscura. These results are compatible both with a rather recent and massive colonization, and with multiple colonization events from the continent. Nucleotide diversity in D. madeirensis (π=0.0076) was similar to that in D. subobscura, which deviates from the expectation, under strict neutrality, of a lower level of variation in an insular species with a small population size. The observed numbers of shared polymorphisms and of fixed differences between D. madeirensis and D. subobscura are compatible with the isolation model of speciation, where shared polymorphisms are due to common ancestry.     

AN INTERACTION BETWEEN ENVIRONMENTAL TEMPERATURE AND GENETIC VARIATION FOR BODY SIZE FOR THE FITNESS OF ADULT FEMALE DROSOPHILA MELANOGASTER

Abstract. — Drosophila and other ectotherms show geographic genetic variation in body size, with larger individuals at higher latitudes and altitudes. Temperature is implicated as an important selective agent because long-term laboratory culture of Drosophila leads to the evolution of larger body size at lower temperatures. In this paper, we tested the hypothesis that, in Drosophila melanogaster, larger size is favored at lower temperatures in part because of selection on adult females. We used replicated lines of D. melanogaster artificially selected for increased and decreased wing area with constant cell area. The resulting size differences between the selected lines were due solely to differences in cell number, and thereby were similar to the cellular basis of clinal variation in body size in nature. We examined life-history traits of adult females at 18 and 25°C. Rearing for two generations at the two temperatures did not affect the extent of the size differences between lines from the different selection regimes. There was a strong interaction between temperature and size selection for both survival and lifetime reproductive success, with larger females living significantly longer and producing more offspring over their lifetime only when reared and tested in the colder environment. There was also an increase in average daily progeny production in large-line females relative to the control and small lines again, only in the colder environment. Thus, the females from the large selection lines were relatively fitter at the colder temperature. At both experimental temperatures, especially the lower one, the small- line females rescheduled their progeny production to later ages. Larger body size may have evolved at higher latitudes and altitudes because of the advantages to the adult female of being larger at lower temperatures.     

Selection by Fertility in DROSOPHILA PSEUDOOBSCURA

Fertility, the component of selection due to female fecundity and male mating success, differed significantly among the ST/ST, ST/AR, and AR/AR karyotypes in experimental populations and varied with karyotypic frequency. In relation to ST/AR, ST/ST females and males had higher fertilities at low frequency; AR/AR males and females were at a significant fertility disadvantage at intermediate frequency, while at low and at high frequencies their fertilities matched or exceeded that of the heterokaryotype. These fertility differences were comparable in size to viability differences previously reported for D. pseudoobscura karyotypes. Differential fertility seems likely to be an important element, perhaps just as important as differential viability, in the balancing selection that maintains the chromosomal polymorphism in this species.     

The wild side of life

Drosophila species vary in the rates at which females remate and the number of sperm they receive in the laboratory. In species such as D. melanogaster and D. pseudoobscura, in which females receive thousands of sperm and remate infrequently compared with species such as D. hydei and D. nigrospiracula, where females receive only a few hundred sperm and remate many times in a day, wild caught females should produce far more progeny. We tested this prediction by collecting, directly from nature, females of six species whose remating rates and number of sperm received vary from high to low and assessing the proportion of females with sperm and the number of progeny females produce. Over 95% of D. pseudoobscura and D. melanogaster females were inseminated while far fewer of the other species contained any sperm. In addition, D, pseudoobscura females produced progeny for over two weeks, D. melanogaster for over a week, while D. hydei and D. nigrospiracula females ran out of sperm after 1–2 d. These observations suggest extreme sperm limitation in these latter species.     

Evolution of total net fitness in thermal lines: Drosophila subobscura likes it 'warm'

Fisher's fundamental theorem states that heritable variation for net fitness sets a limit to the rate of response to natural selection. How will temperate (i.e. cold‐tolerant) species cope with contemporary rapid global warming? Using three‐fold replicated lines of Drosophila subobscura that had been allowed to evolve for 4 years (between 32 and 59 generations) at 13 °C (cold), 18 °C (the supposed optimum temperature), and 22 °C (warm) I assess here how net fitness changes according to thermal environments. Net fitness was estimated following the classical approach in population genetics of competing over a number of generation in outbred experimental populations multiple wild‐type O chromosomes (homologous to arm 3R in D. melanogaster) independently derived from each base thermal stock in an otherwise homogeneous genetic background against a balancer chromosome. Warm‐adapted populations (‘warm‐adapted O chromosomes’) performed comparatively well at all tested temperatures. However, net fitness was severely reduced in cold‐adapted populations when transferred to warmer conditions. It seems, therefore, that thermal fitness breath for D. subobscura flies is positively associated to temperature. These findings are discussed in relation to the fast world‐wide clinal shifts in the frequency of genetic markers correlated with current climate change.     

Nucleotide divergence of the rp49 gene region between Drosophila melanogaster and two species of the Obscura group of Drosophila

Summary A 2.1-kb SStI fragment including the rp49 gene and the 3 end of the -serendipity gene has been cloned and sequenced in Drosophila pseudoobscura. rp49 maps at region 62 on the tip of chromosome II of this species. Both the coding and flanking regions have been aligned and compared with those of D. subobscura. There is no evidence for heterogeneity in the rate of silent substitution between the rp49 coding region and the rate of substitutions in flanking regions, the overall silent divergence per site being 0.19. Noncoding regions also differ between both species by different insertions/deletions, some of which are related to repeated sequences. The rp49 region of D. pseudoobscura shows a strong codon bias similar to those of D. subobscura and D. melanogaster. Comparison of the rates of silent (K _S ) and nonsilent (K _a ) substitutions of the rp49 gene and other genes completely sequenced in D. pseudoobscura and D. melanogaster confirms previous results indicating that rp49 is evolving slowly both at silent and nonsilent sites. According to the data for the rp49 region, D. pseudoobscura and D. subobscura lineages would have diverged some 9 Myr ago, if one assumes a divergence time of 30 Myr for the melanogaster and obscura groups.Offprint requests to: C. Segarra     

SPERM PRODUCTION AND STERILITY IN HYBRIDS BETWEEN TWO SUBSPECIES OF DROSOPHILA PSEUDOOBSCURA

Subspecies of Drosophila pseudoobscura, one occurring in the United States and the other in Bogota, Columbia, exhibit Haldane's Rule in one direction of the cross. Additionally, D. pseudoobscura produces two sperm types: short, sterile sperm and long, fertile, sperm. Here I examine the relationship between the production of short and long sperm and hybrid sterility. Fertile and sterile hybrid males produce a greater proportion of short sperm compared to parental males with sterile hybrids producing mainly short, immotile sperm. Sperm transfer and storage patterns were similar between fertile hybrid and parental strains; and unexpectedly, short, immotile sperm from sterile hybrids were stored. These findings raise the question of whether different genetic mechanisms disrupt both sperm heteromorphic production and sperm motility and whether this indicates that females exert some control over sperm storage.