Intrinsic reproductive isolation between Trinidadian populations of the guppy, Poecilia reticulata

Although Trinidadian populations of the guppy, Poecilia reticulata, show considerable adaptive genetic differentiation, they have been assumed to show little or no reproductive isolation. We tested this assumption by crossing Caroni (Tacarigua River) and Oropuche (Oropuche R.) drainage populations from Trinidad's Northern Range, and by examining multiple aspects of reproductive compatibility in the F1, F2 and BC1 generations. In open-aquarium experiments, F1 males performed fewer numbers of mating behaviours relative to parental population controls. This is the first documentation of hybrid behavioural sterility within a species, and it suggests that such sterility may feasibly be involved in causing speciation. The crosses also uncovered hybrid breakdown for embryo viability, brood size and sperm counts. In contrast, no reductions in female fertility were detected, indicating that guppies obey Haldane's rule for sterility. Intrinsic isolation currently presents a much stronger obstacle to gene flow than behavioural isolation, and our results indicate that Trinidadian populations constitute a useful model for investigating incipient speciation.     

Genetic dissection of hybrid incompatibilities between Drosophila simulans and D. mauritiana. III. Heterogeneous accumulation of hybrid incompatibilities, degree of dominance, and implications for Haldane's rule

The genetic basis of Haldane's rule was investigated through estimating the accumulation of hybrid incompatibilities between Drosophila simulans and D. mauritiana by means of introgression. The accumulation of hybrid male sterility (HMS) is at least 10 times greater than that of hybrid female sterility (HFS) or hybrid lethality (HL). The degree of dominance for HMS and HL in a pure D. simulans background is estimated as 0.23-0.29 and 0.33-0.39, respectively; that for HL in an F1 background is unlikely to be very small. Evidence obtained here was used to test the Turelli-Orr model of Haldane's rule. Composite causes, especially, faster-male evolution and recessive hybrid incompatibilities, underlie Haldane's rule in heterogametic male taxa such as Drosophila (XY male and XX female). However, if faster-male evolution is driven by sexual selection, it contradicts Haldane's rule for sterility in heterogametic-female taxa such as Lepidoptera (ZW female and ZZ male). The hypothesis of a faster-heterogametic-sex evolution seems to fit the current data best. This hypothesis states that gametogenesis in the heterogametic sex, instead of in males per se, evolves much faster than in the homogametic sex, in part because of sex-ratio selection. This hypothesis not only explains Haldane's rule in a simple way, but also suggests that genomic conflicts play a major role in evolution and speciation.     

EPISTASIS MODIFIES THE DOMINANCE OF LOCI CAUSING HYBRID MALE STERILITY IN THE DROSOPHILA PSEUDOOBSCURA SPECIES GROUP

Speciation, the evolution of reproductive isolation between populations, serves as the driving force for generating biodiversity. Postzygotic barriers to gene flow, such as F ₁ hybrid sterility and inviability, play important roles in the establishment and maintenance of biological species. F ₁ hybrid incompatibilities in taxa that obey Haldane's rule, the observation that the heterogametic sex suffers greater hybrid fitness problems than the homogametic sex, are thought to often result from interactions between recessive-acting X-linked loci and dominant-acting autosomal loci. Because they play such prominent roles in producing hybrid incompatibilities, we examine the dominance and nature of epistasis between alleles derived from Drosophila persimilis that confer hybrid male sterility in the genetic background of its sister species, D. pseudoobscura bogotana . We show that epistasis elevates the apparent dominance of individually recessive-acting QTL such that they can contribute to F ₁ hybrid sterility. These results have important implications for assumptions underlying theoretical models of hybrid incompatibilities and may offer a possible explanation for why, to date, identification of dominant-acting autosomal "speciation genes" has been challenging.     

Sex-linked hybrid sterility in a butterfly

Recent studies, primarily in Drosophila, have greatly advanced our understanding of Haldane's rule, the tendency for hybrid sterility or inviability to affect primarily the heterogametic sex (Haldane 1922). Although dominance theory (Turelli and Orr 1995) has been proposed as a general explanation of Haldane's rule, this remains to be tested in female-heterogametic taxa, such as the Lepidoptera. Here we describe a novel example of Haldane's rule in Heliconius melpomene (Lepidoptera; Nymphalidae). Female F1 offspring are sterile when a male from French Guiana is crossed to a female from Panama, but fertile in the reciprocal cross. Male F1s are fertile in both directions. Similar female F1 sterility occurs in crosses between French Guiana and eastern Colombian populations. Backcrosses and linkage analysis show that sterility results from an interaction between gene(s) on the Z chromosome of the Guiana race with autosomal factors in the Panama genome. Large X (or Z) effects are commonly observed in Drosophila, but to our knowledge have not been previously demonstrated for hybrid sterility in Lepidoptera. Differences in the abundance of male versus female or Z-linked versus autosomal sterility factors cannot be ruled out in our crosses as causes of Haldane's rule. Nonetheless, the demonstration that recessive Z-linked loci cause hybrid sterility in a female heterogametic species supports the contention that dominance theory provides a general explanation of Haldane's rule (Turelli and Orr 2000).     

The Genetics of Postzygotic Isolation in the Drosophila Virilis Group

In a genetic study of postzygotic reproductive isolation among species of the Drosophila virilis group, we find that the X chromosome has the largest effect on male and female hybrid sterility and inviability. The X alone has a discernible effect on postzygotic isolation between closely related species. Hybridizations involving more distantly related species also show large X-effects, although the autosomes may also play a role. In the only hybridization yet subjected to such analysis, we show that hybrid male and female sterility result from the action of different X-linked loci. Our results accord with genetic studies of other taxa, and support the view that both Haldane's rule (heterogametic F1 sterility or inviability) and the large effect of the X chromosome on reproductive isolation result from the accumulation by natural selection of partially recessive or underdominant mutations. We also describe a method that allows genetic analysis of reproductive isolation between species that produce completely sterile or inviable hybrids. Such species pairs, which represent the final stage of speciation, cannot be analyzed by traditional methods. The X chromosome also plays an important role in postzygotic isolation between these species.     

Hybrid incompatibilities in the parasitic wasp genus Nasonia: negative effects of hemizygosity and the identification of transmission ratio distortion loci

The occurrence of hybrid incompatibilities forms an important stage during the evolution of reproductive isolation. In early stages of speciation, males and females often respond differently to hybridization. Haldane's rule states that the heterogametic sex suffers more from hybridization than the homogametic sex. Although haplodiploid reproduction (haploid males, diploid females) does not involve sex chromosomes, sex-specific incompatibilities are predicted to be prevalent in haplodiploid species. Here, we evaluate the effect of sex/ploidy level on hybrid incompatibilities and locate genomic regions that cause increased mortality rates in hybrid males of the haplodiploid wasps Nasonia vitripennis and Nasonia longicornis. Our data show that diploid F(1) hybrid females suffer less from hybridization than haploid F(2) hybrid males. The latter not only suffer from an increased mortality rate, but also from behavioural and spermatogenic sterility. Genetic mapping in recombinant F(2) male hybrids revealed that the observed hybrid mortality is most likely due to a disruption of cytonuclear interactions. As these sex-specific hybrid incompatibilities follow predictions based on Haldane's rule, our data accentuate the need to broaden the view of Haldane's rule to include species with haplodiploid sex determination, consistent with Haldane's original definition.     

Haldane's rule and its legacy: Why are there so many sterile males?

A general pattern of animal hybridization, known as Haldane's rule, is that the XY (ZW) sex is more severely affected in its viability or fertility than the XX (ZZ) sex. Recent evidence suggests that three different forces have shaped this pattern: (1) the X chromosome and autosomes are in greater disharmony in the XY sex; (2) evolution of hybrid male sterility is greatly accelerated, at least in species with XY males; and (3) maternal-zygotic incompatibility preferentially affects the viability of the XX sex. In species with XY males, the rapid evolution toward hybrid male sterility may be responsible for the bulk of observations pertaining to Haldane's rule. One interesting and testable hypothesis is that sexual selection drives this rapid evolution.     

Hybrid sterility,Haldane's rule and speciation in Heliconius cydno and H. melpomene

Most genetic studies of Haldane's rule, in which hybrid sterility or inviability affects the heterogametic sex preferentially, have focused on Drosophila. It therefore remains unclear to what extent the conclusions of that work apply more generally, particularly in female-heterogametic taxa such as birds and Lepidoptera. Here we present a genetic analysis of Haldane's rule in Heliconius butterflies. Female F(1) hybrids between Heliconius melpomene and H. cydno are completely sterile, while males have normal to mildly reduced fertility. In backcrosses of male F(1) hybrids, female offspring range from completely sterile to fully fertile. Linkage analysis using the Z-linked triose-phosphate isomerase locus demonstrates a "large X" (Z) effect on sterility. Expression of female sterility varies among crosses in this and a previous study of Heliconius. Sterility may result from the production of normal but infertile eggs, production of small infertile eggs, or from a complete failure to develop ovarioles, which suggests multiple routes to the evolution of hybrid sterility in these Heliconius species. These results conform to the expectations of the "dominance" rather than "faster male" theories of Haldane's rule and suggest that relatively few loci are responsible. The two species are broadly sympatric and hybridize in the wild, so that female hybrid sterility forms one of several strong but incomplete barriers to gene flow in nature. The effect of female sterility is comparable to that of selection against non-mimetic hybrids, while mate choice forms a much stronger barrier to gene transfer.     

Patterns of postzygotic isolation in Lepidoptera

I present patterns characterizing the evolution of intrinsic postzygotic isolation in Lepidoptera by analyzing data from the literature on genetic distance, strength of hybrid sterility and inviability, biogeography, and natural hybridization. Using genetic distance as a proxy for time, I investigate the time-course of the evolution of postzygotic isolation and the waiting times to particular hybrid fitness problems. The results show that postzygotic isolation increases gradually as species diverge, but that hybrid sterility evolves faster than hybrid inviability. The overwhelming preponderance of female-specific hybrid problems in Lepidoptera shows that Haldane's rule (the preferential sterility or inviability of the heterogametic sex) is well obeyed. Together the rates and patterns characterizing the accumulation of postzygotic isolation allow several tests of the composite theory of Haldane's rule. Interestingly, comparing these data with those from Drosophila reveals that Haldane's rule for sterility evolves as fast (if not faster) in Lepidoptera. Finally, I show that a substantial fraction of sympatric species hybridizes in nature and that the majority of these suffer some level of hybrid sterility or inviability.     

RAPID EVOLUTION OF ASYMMETRIC REPRODUCTIVE INCOMPATIBILITIES IN STALK‐EYED FLIES

The steps by which isolated populations acquire reproductive incompatibilities remain poorly understood. One potentially important process is postcopulatory sexual selection because it can generate divergence between populations in traits that influence fertilization success after copulation. Here we present a comprehensive analysis of this form of reproductive isolation by conducting reciprocal crosses between variably diverged populations of stalk‐eyed flies (Teleopsis dalmanni). First, we measure seven types of reproductive incompatibility between copulation and fertilization. We then compare fertilization success to hatching success to quantify hybrid inviability. Finally, we determine if sperm competition acts to reinforce or counteract any incompatibilities. We find evidence for multiple incompatibilities in most crosses, including failure to store sperm after mating, failure of sperm to reach the site of fertilization, failure of sperm to fertilize eggs, and failure of embryos to develop. Local sperm have precedence over foreign sperm, but this effect is due mainly to differences in sperm transfer and reduced hatching success. Crosses between recently diverged populations are asymmetrical with regard to the degree and type of incompatibility. Because sexual conflict in these flies is low, postcopulatory sexual selection, rather than antagonistic coevolution, likely causes incompatibilities due to mismatches between male and female reproductive traits.     

Cryptic postzygotic isolation in an eruptive species of bark beetle (Dendroctonus ponderosae)

Studies of postzygotic isolation often involve well-differentiated taxa that show a consistent level of incompatibility, thereby limiting our understanding of the initial stages and development of reproductive barriers. Dendroctonus ponderosae provides an informative system because recent evidence suggests that distant populations produce hybrids with reproductive incompatibilities. Dendroctonus ponderosae shows an isolation-by-distance gene flow pattern allowing us to characterize the evolution of postzygotic isolation (e.g., hybrid inviability, hybrid sterility) by crossing populations along a continuum of geographic/genetic divergence. We found little evidence of hybrid inviability among these crosses. However, crosses between geographically distant populations produced sterile males (consistent with Haldane's rule). This effect was not consistent with the fixation of mutations in an isolation-by-distance pattern, but instead is spatially localized. These reproductive barriers are uncorrelated with a reduction in gene flow suggesting their recent development. Crosses between geographically proximal populations bounding the transition from compatibility to hybrid male sterility showed evidence of unidirectional reduction in hybrid male fecundity. Our study describes significant postzygotic isolation occurring across a narrow and molecularly cryptic geographic zone between the states of Oregon and Idaho. This study provides a view of the early stages of postzygotic isolation in a geographically widespread species.     

High-resolution genome-wide dissection of the two rules of speciation in Drosophila

Postzygotic reproductive isolation is characterized by two striking empirical patterns. The first is Haldane's rule—the preferential inviability or sterility of species hybrids of the heterogametic (XY) sex. The second is the so-called large X effect—substitution of one species's X chromosome for another's has a disproportionately large effect on hybrid fitness compared to similar substitution of an autosome. Although the first rule has been well-established, the second rule remains controversial. Here, we dissect the genetic causes of these two rules using a genome-wide introgression analysis of Drosophila mauritiana chromosome segments in an otherwise D. sechellia genetic background. We find that recessive hybrid incompatibilities outnumber dominant ones and that hybrid male steriles outnumber all other types of incompatibility, consistent with the dominance and faster-male theories of Haldane's rule, respectively. We also find that, although X-linked and autosomal introgressions are of similar size, most X-linked introgressions cause hybrid male sterility (60%) whereas few autosomal introgressions do (18%). Our results thus confirm the large X effect and identify its proximate cause: incompatibilities causing hybrid male sterility have a higher density on the X chromosome than on the autosomes. We evaluate several hypotheses for the evolutionary cause of this excess of X-linked hybrid male sterility.     

Haldane's rule: hybrid sterility affects the heterogametic sex first because sexual differentiation is on the path to species differentiation

Prevention of recombination is needed to preserve both phenotypic differentiation between species and sexual phenotypic differentiation within species. For species differentiation (speciation), isolating barriers preventing recombination may be pre-zygotic (gamete transfer barriers), or post-zygotic (either a developmental barrier resulting in hybrid inviability, or a chromosomal-pairing barrier resulting in hybrid sterility). The sterility barrier is usually the first to appear and, although often initially only manifest in the heterogametic sex (Haldane's rule), is finally manifest in both sexes. For sexual differentiation, the first and only barrier is chromosomal-pairing, and always applies to the heterogametic sex. For regions of sex chromosomes affecting sexual differentiation there must be something analogous to the process generating the hybrid sterility seen when allied species cross. Explanations for Haldane's rule have generally assumed that the chromosomal-pairing barrier initiating evolutionary divergence into species is due to incompatibilities between gene products ("genic), or sets of gene products ("polygenic), rather than between chromosomes per se ("chromosomal"). However, if chromosomal incompatibilities promoting incipient sexual differentiation could also contribute to the process of incipient speciation, then a step towards speciation would have been taken in the heterogametic sex. Thus, incipient speciation, manifest as hybrid sterility when "varieties" are crossed, would appear at the earliest stage in the heterogametic sex, even in genera with homomorphic sex chromosomes (Haldane's rule for hybrid sterility). In contrast, it has been proposed that Haldane's rule for hybrid inviability needs differences in dosage compensation, so could not apply to genera with homomorphic sex chromosomes.     

No evidence for faster male hybrid sterility in population crosses of an intertidal copepod (Tigriopus californicus)

Two different forces are thought to contribute to the rapid accumulation of hybrid male sterility that has been observed in many inter-specific crosses, namely the faster male and the dominance theories. For male heterogametic taxa, both faster male and dominance would work in the same direction to cause the rapid evolution of male sterility; however, for taxa lacking differentiated sex chromosomes only the faster male theory would explain the rapid evolution of male hybrid sterility. It is currently unknown what causes the faster evolution of male sterility, but increased sexual selection on males and the sensitivity of genes involved in male reproduction are two hypotheses that could explain the observation. Here, patterns of hybrid sterility in crosses of genetically divergent copepod populations are examined to test potential mechanisms of faster male evolution. The study species, Tigriopus californicus, lacks differentiated, hemizygous sex chromosomes and appears to have low levels of divergence caused by sexual selection acting upon males. Hybrid sterility does not accumulate more rapidly in males than females in these crosses suggesting that in this taxon male reproductive genes are not inherently more prone to disruption in hybrids.     

Haldane's rule by sexual transformation in Caenorhabditis

Haldane's rule in C. briggsae x C. remanei broods was caused by sexual transformation; XX and XO hybrids were female. C. briggsae and C. remanei variants that partially suppress hybrid sexual transformation were identified. Effects of variant strains were cumulative. Hence, aberrant sex determination is a reproductive isolation mechanism in Caenorhabditis.     

The Genetics of Reproductive Isolation in the Drosophila Simulans Clade: X Vs. Autosomal Effects and Male Vs. Female Effects

A strong effect of homozygous autosomal regions on reproductive isolation was found for crosses between the species in the Drosophila simulans clade. Second chromosome regions were introgressed from D. mauritiana and D. sechellia into D. simulans and tested for their homozygous effects on hybrid male and hybrid female sterility and inviability. Most introgressions are fertile as heterozygotes, yet produce sterile male offspring when made homozygous. The density of homozygous autosomal factors contributing to hybrid male sterility is comparable to the density of X chromosome factors for this level of resolution. Female sterility was also revealed, yet the disparity between male and female levels of sterility was great, with male sterility being up to 23 times greater than female sterility. Complete hybrid inviability was also associated with some regions of the second chromosome, yet there were no strong sex differences. In conclusion, we find no evidence to support a strong X chromosome bias in the evolution of hybrid sterility or inviability but do find a very strong sex bias in the evolution of hybrid sterility. In light of these findings, we reevaluate the current models proposed to explain the genetic pattern of reproductive isolation.     

Haldane's rule in marsupials: what happens when both sexes are functionally hemizygous?

During the process of speciation, diverging taxa often hybridize and produce offspring wherein the heterogametic sex (i.e., XY or ZW) is unfit (Haldane's rule). Dominance theory seeks to explain Haldane's rule in terms of the difference in X-linked dominance regimes experienced by the sexes. However, X inactivation in female mammals extends the effects of hemizygosity to both sexes. Here, we highlight where the assumptions of dominance theory are particularly problematic in marsupials, where X inactivation uniformly results in silencing the paternal X. We then present evidence of Haldane's rule for sterility but not for viability in marsupials, as well as the first violations of Haldane's rule for these traits among all mammals. Marsupials represent a large taxonomic group possessing heteromorphic sex chromosomes, where the dominance theory cannot explain Haldane's rule. In this light, we evaluate alternative explanations for the preponderance of male sterility in interspecific hybrids, including faster male evolution, X-Y interactions, and genomic conflict hypotheses.     

Genetic studies of two sister species in the Drosophila melanogaster subgroup, D. yakuba and D. santomea

We performed genetic analysis of hybrid sterility and of one morphological difference (sex-comb tooth number) on D. yakuba and D. santomea, the former species widespread in Africa and the latter endemic to the oceanic island of S?o Tomé, on which there is a hybrid zone. The sterility of hybrid males is due to at least three genes on the X chromosome and at least one on the Y, with the cytoplasm and large sections of the autosomes having no effect. F1 hybrid females carrying two X chromosomes from either species are perfectly fertile despite their genetic similarity to completely sterile F1 hybrid males. This implies that the appearance of Haldane's rule in this cross is at least partially due to the faster accumulation of genes causing male than female sterility. The larger effects of the X and Y chromosomes than of the autosomes, however, also suggest that the genes causing male sterility are recessive in hybrids. Some female sterility is also seen in interspecific crosses, but this does not occur between all strains. This is seen in pure-species females inseminated by heterospecific males (probably reflecting incompatibility between the sperm of one species and the female reproductive tract of the other) as well as in inseminated F1 and backcross females, probably reflecting genetically based incompatibilities in hybrids that affect the reproductive system. The latter 'innate' sterility appears to involve deleterious interactions between D. santomea chromosomes and D. yakuba cytoplasm. The difference in male sex-comb tooth number appears to involve fairly large effects of the X chromosome. We discuss the striking evolutionary parallels in the genetic basis of sterility, in the nature of sexual isolation, and in morphological differences between the D. santomea/D. yakuba divergence and two other speciation events in the D. melanogaster subgroup involving island colonization.     

Sexual conflict does not drive reproductive isolation in experimental populations of Drosophila pseudoobscura

Sexual conflict has been predicted to drive reproductive isolation by generating arbitrary but rapid coevolutionary changes in reproductive traits among allopatric populations. A testable prediction of this proposal is that allopatric populations experiencing different levels of sexual conflict should exhibit different levels of reproductive isolation. We tested this prediction using experimentally evolved populations of the promiscuous Drosophila pseudoobscura. We manipulated sexual conflict by enforcing either monogamy, maintaining natural levels of promiscuity, or elevating promiscuity. Within each treatment, we carried out sympatric and allopatric crosses using replicated populations and examined pre-zygotic (number of mating pairs, mating speed and copulation duration) and post-zygotic (hybrid inviability and sterility) indicators of reproductive isolation. After 50 generations of selection, none of the measures conformed to predictions of sexual conflict driving reproductive isolation. Our results cannot be explained by lack of genetic variation or weak selection and suggest that sexual conflict may not be a widespread engine of speciation.     

Postzygotic isolation involves strong mitochondrial and sex-specific effects in Tigriopus californicus,a species lacking heteromorphic sex chromosomes

Detailed studies of the genetics of speciation have focused on a few model systems, particularly Drosophila. The copepod Tigriopus californicus offers an alternative that differs from standard animal models in that it lacks heteromorphic chromosomes (instead, sex determination is polygenic) and has reduced opportunities for sexual conflict, because females mate only once. Quantitative trait loci (QTL) mapping was conducted on reciprocal F₂ hybrids between two strongly differentiated populations, using a saturated linkage map spanning all 12 autosomes and the mitochondrion. By comparing sexes, a possible sex ratio distorter was found but no sex chromosomes. Although studies of standard models often find an excess of hybrid male sterility factors, we found no QTL for sterility and multiple QTL for hybrid viability (indicated by non-Mendelian adult ratios) and other characters. Viability problems were found to be stronger in males, but the usual explanations for weaker hybrid males (sex chromosomes, sensitivity of spermatogenesis, sexual selection) cannot fully account for these male viability problems. Instead, higher metabolic rates may amplify deleterious effects in males. Although many studies of standard speciation models find the strongest genetic incompatibilities to be nuclear–nuclear (specifically X chromosome–autosome), we found the strongest deleterious interaction in this system was mito–nuclear. Consistent with the snowball theory of incompatibility accumulation, we found that trigenic interactions in this highly divergent cross were substantially more frequent (>6 × ) than digenic interactions. This alternative system thus allows important comparisons to studies of the genetics of reproductive isolation in more standard model systems.