Hybrid zone structure and the potential role of selection in hybridizing populations of native westslope cutthroat trout (Oncorhynchus clarki lewisi) and introduced rainbow trout (O. mykiss)

Introgressive hybridization is a common feature of many zones of contact between divergent lineages of fishes. This is particularly common when taxa that are normally allopatric come into artificial (human-induced) secondary contact. We examined 18 native populations of westslope cutthroat trout (Oncorhynchus clarki lewisi, WCT) to determine the extent of introgressive hybridization with introduced rainbow trout (O. mykiss, RBT) and the genetic structure of hybridizing populations in the upper Kootenay River, southeastern British Columbia, Canada. Using four diagnostic nuclear loci we calculated a hybrid index, inbreeding coefficient, FIS, and the linkage disquilibrium correlation coefficient, Rij, for each locality to determine the distribution of genotypes in each population. We also categorized the 142 hybrid individuals found across localities into four hybrid classes based on their genotypes. The majority of localities (11/18) showed a unimodal distribution of genotypes skewed towards genotypes of WCT. Two localities, however (lower Gold Creek and Lodgepole Creek) showed a flat to bimodal distribution and one site (lower Bull River) showed a unimodal distribution skewed towards RBT genotypes. The majority of hybrid individuals were classified genotypically as WCT backcrosses (59%) and post-F1 individuals (24%). We found a skewed ratio of pure WCT to pure RBT (17:1) and only four F1 hybrids (3%), suggesting that the spread of RBT alleles may be facilitated by hybrids straying to neighbouring populations. We also tested for the action of selection in one population using cohort analyses, but found little evidence of differential selection between pure WCT and hybrid individuals. Pooled across age classes there were significant differences in genotypic frequencies among loci suggesting differential introgression. There was no asymmetry to the hybridization between rainbow trout and westslope cutthroat trout because both species' mitochondrial DNA haplotypes were observed at similar frequencies in the hybrids. Our analyses suggest that hybrid swarms are likely to form in the upper Kootenay River drainage and that certain native WCT populations in British Columbia are at risk of local genomic extinction.     

Deep sympatric mitochondrial divergence without reproductive isolation in the common redstart Phoenicurus phoenicurus

Mitochondrial DNA usually shows low sequence variation within and high sequence divergence among species, which makes it a useful marker for phylogenetic inference and DNA barcoding. A previous study on the common redstart (Phoenicurus phoenicurus) revealed two very different mtDNA haplogroups (5% K2P distance). This divergence is comparable to that among many sister species; however, both haplogroups coexist and interbreed in Europe today. Herein, we describe the phylogeographic pattern of these lineages and test hypotheses for how such high diversity in mtDNA has evolved. We found no evidence for mitochondrial pseudogenes confirming that both haplotypes are of mitochondrial origin. When testing for possible reproductive barriers, we found no evidence for lineage‐specific assortative mating and no difference in sperm morphology, indicating that they are not examples of cryptic species, nor likely to reflect the early stages of speciation. A gene tree based on a short fragment of cytochrome c oxidase subunit 1 from the common redstart and 10 other Phoenicurus species, showed no introgression from any of the extant congenerics. However, introgression from an extinct congeneric cannot be excluded. Sequences from two nuclear introns did not show a similar differentiation into two distinct groups. Mismatch distributions indicated that the lineages have undergone similar demographic changes. Taken together, these results confirm that deeply divergent mitochondrial lineages can coexist in biological species. Sympatric mtDNA divergences are relatively rare in birds, but the fact that they occur argues against the use of threshold mtDNA divergences in species delineation.     

Absence of developmental incompatibility in hybrids between rainbow trout and two subspecies of cutthroat trout

We examined the developmental rate of hybrids between rainbow trout (Salmo gairdneri) and two subspecies of cutthroat trout: westslope cutthroat trout (Salmo clarki lewisi) and Yellowstone cutthroat trout (Salmo clarki bouvieri). These taxa show considerable genetic divergence at 42 structural loci encoding enzymes; the mean Nei's d between the rainbow trout and the two species of cutthroat trout is 0.22. We used four measures of developmental rate: time of hatching and yolk resorption, rate of increase in activity of four enzymes, and time of initial detection of seven isozyme loci. The two cutthroat trout subspecies reached hatching and yolk resorption earlier than rainbow trout. Cutthroat trout had higher relative enzyme activities than rainbow trout from deposition of eye pigment to hatching. There was no difference in the rate of increase in enzyme activity or time of initial expression of these loci between these species. Hybrids showed developmental rates intermediate or similar to that of the parental species using all measures. Our results indicate an absence of regulatory and developmental incompatibility between these taxa.This research was supported by NSF Grants ISP-8011449 and BSR-8300039. M.M.F. was supported by a postgraduate scholarship from the Natural Sciences and Engineering Research Council of Canada.     

Variation in reproductive isolation across a species range

Reproductive isolation is often variable within species, a phenomenon that while largely ignored by speciation studies, can be leveraged to gain insight into the potential mechanisms driving the evolution of genetic incompatibilities. We used experimental greenhouse crosses to characterize patterns of reproductive isolation among three divergent genetic lineages of Campanulastrum americanum that occur in close geographic proximity in the Appalachian Mountains. Substantial, asymmetrical reproductive isolation for survival due to cytonuclear incompatibility was found among the lineages (up to 94% reduction). Moderate reductions in pollen viability, as well as cytoplasmic male sterility, were also found between some Mountain populations. We then compared these results to previously established patterns of reproductive isolation between these Mountain lineages and a fourth, widespread Western lineage to fully characterize reproductive isolation across the complete geographic and genetic range of C. americanum. Reproductive isolation for survival and pollen viability was consistent across studies, indicating the evolution of the underlying genetic incompatibilities is primarily determined by intrinsic factors. In contrast, reproductive isolation for germination was only found when crossing Mountain populations with the Western lineage, suggesting the underlying genetic incompatibility is likely influenced by environmental or demographic differences between the two lineages. Cytoplasmic male sterility was also limited in occurrence, being restricted to a handful of Mountain populations in a narrow geographic range. These findings illustrate the complexity of speciation by demonstrating multiple, independent genetic incompatibilities that lead to a mosaic of genetic divergence and reproductive isolation across a species range.     

Systematics,reproductive isolation and species boundaries in monogonont rotifers

The typological concept of rotifer species and the morphological basis of rotifer systematics is reviewed and alternatives proposed. Occasional sexuality in the cyclical parthenogenetic life cycle of monogononts permits application of the biological species concept to this group. Data from cross-mating experiments with Asplanchna, Brachionus and Epiphanes illustrate the usefulness of reproductive isolation as a criterion for species boundaries. Populations from different geographic regions are often interfertile indicating that rotifer species are genetically integrated over wide areas. The main types of isolating mechanisms operating in monogononts are reviewed. The role of behavioral reproductive isolation in maintaining species boundaries is examined. The use of a mate recognition bioassay which estimates the probability of copulation and quantifies the degree of isolation is described. Recent work of the mechanism of mate recognition is reviewed. It is concluded that the biological species concept is applicable to rotifers and that a more experimental approach to determining species boundaries is both feasible and desirable.     

Hybridization rapidly reduces fitness of a native trout in the wild

Human-mediated hybridization is a leading cause of biodiversity loss worldwide. How hybridization affects fitness and what level of hybridization is permissible pose difficult conservation questions with little empirical information to guide policy and management decisions. This is particularly true for salmonids, where widespread introgression among non-native and native taxa has often created hybrid swarms over extensive geographical areas resulting in genomic extinction. Here, we used parentage analysis with multilocus microsatellite markers to measure how varying levels of genetic introgression with non-native rainbow trout (Oncorhynchus mykiss) affect reproductive success (number of offspring per adult) of native westslope cutthroat trout (Oncorhynchus clarkii lewisi) in the wild. Small amounts of hybridization markedly reduced fitness of male and female trout, with reproductive success sharply declining by approximately 50 per cent, with only 20 per cent admixture. Despite apparent fitness costs, our data suggest that hybridization may spread due to relatively high reproductive success of first-generation hybrids and high reproductive success of a few males with high levels of admixture. This outbreeding depression suggests that even low levels of admixture may have negative effects on fitness in the wild and that policies protecting hybridized populations may need reconsideration.     

A molecular analysis of hybridization between native westslope cutthroat trout and introduced rainbow trout in southeastern British Columbia, Canada

Restriction site variation in the Ikaros gene intron was used to assess the incidence of westslope cutthroat trout ( Oncorhynchus clarki lewisi ), rainbow trout ( O. mykiss ) and interspecific hybrids at 11 localities among eight streams tributary to the upper Kootenay River system in south-eastern British Columbia, Canada. Out of 356 fish assayed by this technique, hybrids ( n =16) were found at seven of the 11 sites across five different streams. Rainbow trout ( n =6) were found at two of the 11 sites. Analysis of hybrids with a second genetic marker (heat shock 71 intron) indicated that most represented either backcrosses to both westslope cutthroat and rainbow trout, or post F₁ hybrids. Mitochondrial DNA analysis indicated that hybrid matings occur between male rainbow trout and female westslope cutthroat trout and vice versa. Comparison of present hybridization in five tributaries relative to an allozyme-based analysis in the mid-1980s, that documented hybrids in only a single tributary of seven that were common to the two studies, suggests that hybridization and introgression has increased in upper Kootenay River tributaries. The present analysis is a conservative estimate of genetic interaction between the species because introgression was not tested in the majority of samples. Identification of genetically pure westslope cutthroat trout populations, and why they might be resistant to introgression from rainbow trout, are crucial conservation priorities for this unique subspecies of cutthroat trout.     

The role of salivary androgen-binding protein in reproductive isolation between two subspecies of house mouse: Mus musculus musculus and Mus musculus domesticus

Previous behavioural studies using inbred lines have suggested that the gene ( Abpa ) for the alpha subunit of salivary androgen-binding protein (ABP) plays a role in prezygotic isolation between house mouse Mus musculus subspecies. We tested this hypothesis in animals from wild allopatric (121 individuals from four samples) and parapatric (320 animals from 15 samples) populations sampled on the Czech–Bavarian transect across the hybrid zone between M. m. domesticus and M. m. musculus . The study did not reveal a consistent statistically significant trend of homosubspecific preferences in individual allopatric and parapatric populations. Nonetheless, the whole pattern of preference was skewed toward homosubspecific preference mostly on the M. m. musculus side of the hybrid zone. The pattern of homosubspecific preferences was stronger for the time spent sniffing than it was for the first choice of the signal (the ratio of homosubspecific vs. heterosubspecific preferences for both sexes was 6 : 2 in allopatric and 21 : 9 in parapatric populations, while the same rates were 4 : 4 and 16 : 14 for the first choice). To the extent that Y-maze tests reflect preference under wild conditions, we suggest that this slight preference may not in itself be sufficient to impede gene flow between the two subspecies and thus act as a reproductive barrier. ABP most probably participates in a complex system of subspecies-specific recognition in the hybrid zone, but the picture is far too complex at this time to allow a conclusive evaluation of the importance of this role.  © 2005 The Linnean Society of London, Biological Journal of the Linnean Society , 2005, 84 , 349–361.     

Hybridization, species collapse, and species reemergence after disturbance to premating mechanisms of reproductive isolation

There are now a number of well-studied cases in which hybridization between closely related sympatric species has increased, sometimes resulting in the replacement of species pairs by hybrid swarms. Many of these cases have been linked to anthropogenic environmental change, but the mechanisms leading from environmental change to species collapse, and the long-term effects of hybridization on species pairs, remain poorly understood. We used an individual-based stochastic simulation model to explore the conditions under which disturbances that weaken premating barriers to reproduction patterns between sympatric species might lead to increased hybridization and to species collapse. Disturbances often resulted in bouts of hybridization, but in many cases strong reproductive isolation spontaneously reemerged. This was sometimes true even after hybrid swarms had replaced parental species. The reemergence of species pairs was most likely when disturbances were of short duration. Counterintuitively, incipient species pairs were more likely to reemerge after strong but temporary disturbances than after weaker disturbances of the same duration. Even temporary bouts of hybridization often led to substantial homogenization of species pairs. This suggests that ecosystem managers may be able to refill ecological niches, but in general will not be able to resurrect lost species after species collapse.     

Evolution of reproductive isolation within a parasitic fungal species complex

Despite important advances in the last few years, the evolution of reproductive isolation (RI) remains an unresolved and critical gap in our understanding of speciation processes. In this study, we investigated the evolution of RI among species of the parasitic fungal species complex Microbotryum violaceum, which is responsible for anther smut disease of the Caryophyllaceae. We found no evidence for significant positive assortative mating by M. violaceum even over substantial degrees of genetic divergence, suggesting a lack of prezygotic isolation. In contrast, postzygotic isolation increased with the genetic distance between mating partners when measured as hyphal growth. Total RI, measured as the ability of the pathogen to infect and produce a diploid progeny in the host plant, was significantly and positively correlated with genetic distance, remaining below complete isolation for most of the species pairs. The results of this study, the first one on the time course of speciation in a fungus, are therefore consistent with previous works showing that RI generally evolves gradually with genetic distance, and thus presumably with time. Interestingly, prezygotic RI due to gamete recognition did not increase with genetic distance, in contrast to the pattern found in plants and animals.     

Lineages of Silene nutans developed rapid,strong, asymmetric postzygotic reproductive isolation in allopatry

Reproductive isolation can rise either as a consequence of genomic divergence in allopatry or as a byproduct of divergent selection in parapatry. To determine whether reproductive isolation in gynodioecious Silene nutans results from allopatric divergence or from ecological adaptation following secondary contact, we investigated the pattern of postzygotic reproductive isolation and hybridization in natural populations using two phylogeographic lineages, western (W1) and eastern (E1). Experimental crosses between the lineages identified strong, asymmetric postzygotic isolation between the W1 and the E1 lineages, independent of geographic overlap. The proportion of ovules fertilized, seeds aborted, and seeds germinated revealed relatively little effect on the fitness of hybrids. In contrast, hybrid mortality was high and asymmetric: while half of the hybrid seedlings with western lineage mothers died, nearly all hybrid seedlings with E1 mothers died. This asymmetric mortality mirrored the proportion of chlorotic seedlings, and is congruent with cytonuclear incompatibility. We found no evidence of hybridization between the lineages in regions of co‐occurrence using nuclear and plastid markers. Together, our results are consistent with the hypothesis that strong postzygotic reproductive isolation involving cytonuclear incompatibilities arose in allopatry. We argue that the dynamics of cytonuclear gynodioecy could facilitate the evolution of reproductive isolation.     

A test of ecologically dependent postmating isolation between sympatric sticklebacks

Ecological speciation occurs when reproductive isolation evolves ultimately as a result of divergent natural selection between populations inhabiting different environments or exploiting alternative resources. I tested a prediction of the ecological model concerning the fitness of hybrids between two young, sympatric species of threespine sticklebacks (Benthics and Limnetics). The two species are ecologically and morphologically divergent: the Benthic is adapted to feeding on invertebrates in the littoral zone of the lake whereas the Limnetic is adapted to feeding on zooplankton in the open water. The growth rate of two types of hybrids, the Benthic backcross and the Limnetic backcross, as well as both parent species, was evaluated in enclosures in both parental habitats in the lake. The use of backcrosses is ideal because a comparison of their growth rates in the two habitats estimates an ecologically dependent component of their fitness while controlling for any intrinsic genetic incompatibilities that may exist between the Benthic and Limnetic genomes. The backcross results revealed a striking pattern of ecological dependence: in the littoral zone, Benthic backcrosses grew at approximately twice the rate of Limnetic backcrosses, while in the open water, Limnetic backcrosses grew at approximately twice the rate of Benthic backcrosses. Such a reversal of relative fitness of the two cross-types in the two environments provides strong evidence that divergent natural selection has played a central role in the evolution of postmating isolation between Benthics and Limnetics. Although the rank order of growth rates of all cross-types in the littoral zone was Benthic > Benthic backcross > Limnetic backcross > Limnetic, neither backcross differed significantly from the parent from which it was mainly derived. Implications of this result are discussed in terms of ecological speciation and possible introgressive hybridization between the species. Results in the open water were less clear and were not fully consistent with the ecological model of speciation, mainly as a result of the low growth rate of Limnetics. However, analysis of the diet of the fish in the open water suggests that these enclosures may not have been fully successful at replicating the food regimes characteristic of this habitat.     

Post-zygotic reproductive isolation between sympatric taxa in the Chamaecrista desvauxii complex (Leguminosae-Caesalpinioideae)

BACKGROUND AND AIMS: Differences in the mating systems and the mechanisms of reproductive isolation between Chamaecrista desvauxii var. graminea and C. desvauxii var. latistipula were examined in the Chapada Diamantina, Brazil. These taxa occur sympatrically, and their populations demonstrate marked morphological differences. The objective of the present work was to determine if reproductive isolation mechanisms exist between these two populations of C. desvauxii, and to determine the influence of these putative mechanisms on their genetic differentiation. METHODS: Field observations were made of floral biology, phenology and floral visitation, and experiments on intra- and interpopulation pollination and germination rates of the resultant seeds were performed. A genetic examination of the populations was undertaken using four allozyme loci. KEY RESULTS: The varieties examined demonstrated overlapping of flowering periods during the months of June to September. The main pollinator for both varieties was the bee Bombus brevivillus. Both varieties are self-compatible, and a large number of fruits are formed in cross-pollinations with high seed germination rates. Inter-taxa pollinations result in high levels of fruit production, but no seeds are formed. Two of the four loci examined were diagnostic for the varieties, and exclusive high-frequency alleles were encountered at the other loci, leading to a high genetic distance between the two populations (0.495). CONCLUSIONS: Pre-zygotic barriers were not found between the two varieties, and these remain isolated due to post-zygotic events. The two varieties demonstrate marked differences in their morphology, floral biology, phenology and genetic make-up, all of which indicate that they should be treated as two distinct species. A complete revision involving the other varieties of the C. desvauxii complex will be necessary in order to define these two taxa formally.     

Lock‐and‐key structural isolation between sibling Drosophila species

Drosophila santomea Lachaise & Harry, which is endemic to the African island of São Tomé, and its sibling D. yakuba Burla comprise a new model system of speciation. They are morphologically distinguishable only by slight differences in the male genitalia and body coloration. As a previously undescribed difference, the aedeagus of D. yakuba bears a pair of stout spines (the ventral branches of the basal processes (VB)), instead of the paired humps found in D. santomea. Here, we show that this difference works as a lock‐and‐key isolating mechanism between the siblings. During conspecific copulation, D. yakuba females receive the spines in a pair of pocket‐shaped structures, which are protected by hardened plates, in the genitalia. The females of D. santomea, which lack such pockets, are wounded by the spines of the VB when mated with D. yakuba males. This genital mismatching resulted in leakage of the ejaculate, making 80% of the matings infertile and causing a prolonged struggle to separate pairs glued together by the ejaculate.     

Mechanical and tactile incompatibilities cause reproductive isolation between two young damselfly species

External male reproductive structures have received considerable attention as a cause of reproductive isolation (RI), because the morphology of these structures often evolves rapidly between populations. This rapid evolution presents the potential for mechanical incompatibilities with heterospecific female structures during mating and could thus prevent interbreeding between nascent species. Although such mechanical incompatibilities have received little empirical support as a common cause of RI, the potential for mismatch of reproductive structures to cause RI due to incompatible species‐specific tactile cues has not been tested. We tested the importance of mechanical and tactile incompatibilities in RI between Enallagma anna and E. carunculatum, two damselfly species that diverged within the past ～250,000 years and currently hybridize in a sympatric region. We quantified 19 prezygotic and postzygotic RI barriers using both naturally occurring and laboratory‐reared damselflies. We found incomplete mechanical isolation between the two pure species and between hybrid males and pure species females. Interestingly, in mating pairs for which mechanical isolation was incomplete, females showed greater resistance and refusal to mate with hybrid or heterospecific males compared to conspecific males. This observation suggests that tactile incompatibilities involving male reproductive structures can influence female mating decisions and form a strong barrier to gene flow in early stages of speciation.     

赤拟谷盗和杂拟谷盗的生殖隔离研究

赤拟谷盗和杂拟谷盗是两个同域发生的近缘储粮害虫。为明确它们的生殖隔离程度和机制,本研究比较分析了赤拟谷盗和杂拟谷盗雄虫对同种和异种雌虫的交配选择;赤拟谷盗与杂拟谷盗交配后,测定了精子在异种雌体内的存活情况;将赤拟谷盗和杂拟谷盗进行正反杂交,研究其F1代、F2代和回交代杂种是否产生及其雌雄比。结果显示,赤拟谷盗和杂拟谷盗雄虫对同种雌虫的爬跨率和交配率高于其对异种雌虫的爬跨率和交配率,表明赤拟谷盗和杂拟谷盗种间性隔离不完全;赤拟谷盗和杂拟谷盗交配后精子在异种雌体内是存活的,杂交所产的F1代卵为受精卵,说明交配后完成了精子传送和受精过程,表明种间机械隔离和配子隔离不完全;赤拟谷盗和杂拟谷盗的杂种F1代自交和回交产生了F2代和回交代,表明种间不存在杂种不育;一些杂交组合产生的F2代和回交代数量少且存在雌雄性比偏离,表明种间存在部分杂种不活和杂种衰败。本研究明确了赤拟谷盗和杂拟谷盗的生殖隔离机制,这对于揭示它们的物种进化关系有着重要意义。     

Hybridization patterns and the evolution of reproductive isolation in ducks

Much of our knowledge of the evolution of reproductive isolation comes from studies of Drosophila . This body of work has revealed the following patterns: (1) reproductive isolation increases with phylogenetic distance between hybridizing species; (2) reproductive isolation is greater between sympatric than allopatric species with the same level of divergence; and (3) hybrid crosses conform to Haldane's rule. We tested for the existence of these patterns in ducks (subfamily Anatinae, sensu Livezey, 1997b ) based on 1037 hybrids of known parentage. Our analyses of the number of interspecific crosses in relation to phylogenetic distance found a significant deviation between the observed and expected distribution of crosses controlling for the topology of the Anatinae phylogeny. In particular, we found both an excess of hybrid crosses among closely related species and a scarcity among distantly related species. The number of hybrid males also decreased with increasing phylogenetic distance between parental species, although the number of hybrid females remained low and constant. Sympatric species produced higher numbers of hybrid males than allopatric ones, despite no difference in phylogenetic distance among parental species in compared groups. The number of hybrid males exceeded the number of hybrid females, consistent with Haldane's rule. This was evident even though the analysis was restricted to a reduced set of phylogenetically independent crosses. However, the pattern was no longer significant after correction for the number of hybrid males by the male-biased sex ratio of adult ducks.  © 2002 The Linnean Society of London, Biological Journal of the Linnean Society , 2002, 77 , 193−200.     

Evolution of premating reproductive isolation among conspecific populations of the sea rock-pool beetle Ochthebius urbanelliae driven by reinforcing natural selection

How natural selection might be involved in speciation remains a fundamental question in evolutionary biology. When two or more species co-occur in the same areas, natural selection may favor divergence in mating traits. By acting in sympatric but not allopatric populations, natural selection can also affect mate choice within species and ultimately initiate speciation among conspecific populations. Here, we address this potential effect in the sea rock-pool beetles Ochthebius quadricollis and O. urbanelliae. The two species, which inhabit the Mediterranean coasts, co-occurr syntopically in an area along the Italian Tyrrhenian coast and completed reproductive isolation by reinforcement. In this article, through mating trials under laboratory conditions between conspecific populations, we found in O. quadricollis no deviations from random mating. Conversely, in O. urbanelliae, we found a clear pattern of premating isolation between the reinforced populations sympatric with O. quadricollis and those nonreinforced allopatric. This pattern is consistent with the view that natural selection, which completed the reproductive isolation between the two species in sympatry, led incidentally also to partial premating reproductive isolation (I(PSI) estimator from 0.683 to 0.792) between conspecific populations of O. urbanelliae. This case study supports an until recently underappreciated role of natural selection resulting from species interactions in initiating speciation.     

Spawning behavior and reproductive isolation of two species ofPseudolabrus

The reproductive behavior of two wrasses,Pseudolabrus eoethinus (Richardson, 1846) andP. sieboldi Mabuchi & Nakabo, 1997, was studied on a rocky coast in Shikoku, Japan, where the two species occur sympatrically. In the study area, individual males of each species established mating territories within which pair spawnings occurred. Pair spawnings were observed only between conspecifics, indicating that the two coexisting species were reproductively isolated. Both species usually spawned during the afternoon in early winter and had similar reproductive behavior. Although habitat, temporal and behavioral isolation mechanisms were not apparent, reproductive isolation seemed to occur via premating isolation through visual recognition of conspecific mates because the two species differed in body size and coloration. On one occasion a maleP. sieboldi was observed to perform streaking behavior to join a spawning pair ofP. eoethinus. Thus, postmating isolation between the two species is also discussed.