Evidence for bimodal hybrid zones between two species of char (Pisces: Salvelinus) in northwestern North America

Dolly Varden (Salvelinus malma, Pisces: Salmonidae) and bull trout (Salvelinus confluentus) have widely overlapping, but largely parapatric ranges in watersheds in northwestern North America from Washington State to northern British Columbia. Genetic analysis of natural populations using diagnostic molecular markers revealed widespread local sympatry and hybridization with hybrids comprising 0-25% of the local samples. In a detailed analysis of hybridization using four nuclear DNA markers and mitochondrial DNA within the Thutade Lake watershed, northcentral British Columbia, hybrid genotypes constituted up to 9% of the population of juvenile char. There were significant deviations from Hardy-Weinberg, gametic, and cytonuclear equilibria, and local samples showed bimodal frequency distributions of genotypes. Pure parental and inferred backcross genotypes were most common, and F1 and F(n) hybrids were comparatively rare. Interspecific hybridization was asymmetrical, with most F1 hybrids (five of six) bearing S. confluentus mtDNA. The introgression of nuclear and mitochondrial alleles was asymmetrical, with S. confluentus mtDNA and Growth Hormone 2 introgressing into S. malma significantly more than either introgression of the three other nuclear loci, or introgression of S. malma alleles into S. confluentus. Substantial prezygotic isolation between the species likely depends on the large body size difference between them in sympatry: S. malma have small bodies and a stream resident life history (12-21 cm adult fork length at maturity), while S. confluentus are larger and adfluvial, i.e., they migrate to Thutade Lake where they grow to maturity before returning to tributary streams to spawn (40-90 cm at maturity). These traits may limit interspecific pairings because of size assortative pairing and size-dependent reproductive habitat use.     

Geographical extent of Arctic char (Salvelinus alpinus) mtDNA introgression in brook char populations (S. fontinalis) from eastern Québec, Canada

The geographical extent of Arctic char ( Salvelinus alpinus ) mitochondrial DNA introgression into brook char ( Salvelinus fontinalis ) populations found in eastern Québec was determined by analysing a total of 598 fish from 29 lakes. The nuclear genome was analysed by protein electrophoresis, whereas the ND-5,6 portion of the mitochondrial genome was analysed by restriction fragment length polymorphism. This survey revealed that introgressed S. fontinalis populations are restricted to only one river subdrainage of the Portneuf basin, where Arctic char is completely absent. Elsewhere, nonintrogressed pure S. fontinalis populations populate the lakes. These findings suggest that the initial hybridization event between the species is ancient and probably occurred shortly after recolonization of the area. At that time, the species would have been in contact and the chances of reproductive isolation mechanisms breaking down would have been high. We discuss the possibility that a combination of biogeographical conditions coupled with positive selection for mtDNA introgression led to the present-day distribution of introgressed S. fontinalis in northeastern North America.     

Population structure in two sympatric species of the Lake Tanganyika cichlid tribe Eretmodini: evidence for introgression

Patterns of genetic differentiation were analysed and compared in two sympatric species of the endemic Lake Tanganyika cichlid tribe Eretmodini by means of mitochondrial DNA (mtDNA) sequences of the control region and six microsatellite DNA loci. The sample area covers a total of 138 km of mostly uninterrupted rocky shoreline in the Democratic Republic of Congo and includes the entire distribution range of Tanganicodus cf. irsacae that stretches over a distance of 35 km. Both markers detected significant genetic differentiation within and between the two species. T. cf. irsacae contained lower overall genetic variation than Eretmoduscyanostictus, possibly due to its more restricted range of distribution and its smaller effective population sizes. Complete fixation of Tanganicodus mtDNA haplotypes was observed in Eretmodus at two localities, while at two other localities some Tanganicodus individuals possessed Eretmodus mtDNA haplotypes. Taking into account the relatively large average sequence divergence of 6.2% between the two species, as well as the geographical distribution of mtDNA haplotypes in the lake, the observed pattern is more likely to be a consequence of asymmetric introgression than of shared ancestral polymorphism. As there is significant population differentiation between sympatric Tanganicodus and Eretmodus populations, the events of introgressions may have happened after secondary contact, but our data provide no evidence for ongoing gene flow and suggest that both species are reproductively isolated at present time.     

Genetic panmixia within a narrow contact zone between chromosomally and ecologically distinct black fly sibling species (Diptera: Simuliidae)

Hybrid zones are windows into the speciation process, and their study can give clues into the maintenance and breakdown of species boundaries. Using both genetic and ecological tools, we investigate lineage diversification across a contact zone characterized by chromosome rearrangements. We show that black fly sibling species, Simulium arcticum sensu stricto (s.s.) and Simulium saxosum, lack genetic differentiation at both microsatellite and mtDNA loci in allopatry and sympatry, as well as exhibit high levels of gene flow and continuous chromosome variation in sympatry. Furthermore, hybrid frequencies at the contact zone are similar to those seen between races, rather than species. In contrast, S. arcticum s.s. and S. saxosum maintain ecological differences and distinct habitat associations ‐ the contact zone situated at the margin of suitable habitat for each sibling species. Moreover, gene flow occurs only in a narrow band along an ecological transition. Except for the contact zone, S. arcticum s.s. and S. saxosum hybrids do not occur elsewhere within the sibling species' ranges. Although S. arcticum s.s. and S. saxosum maintain the potential to interbreed freely, we conclude that habitat associations and, perhaps, chromosome systems prevent expansion of ranges and assimilation of lineages.     

A hybrid zone between two subspecies of Chorthippus parallelus. X-chromosome variation through a contact zone

Two grasshopper subspecies, Chorthippus parallelus parallelus (Cpp) and Chorthippus parallelus erythropus (Cpe), meet along the Pyrenees where they hybridize and produce a hybrid zone. A contact zone located in the Col de Portalet has been analyzed for the distribution of chromosome markers on the sex (X) chromosome in pure and hybrid populations. C-banding allowed us to distinguish both pure subspecific forms and recombinant forms, and to demonstrate their different frequencies through the contact zone. Interestingly, the distal C-band (P) on the X chromosome that characterizes Cpp individuals occurs at very high frequency through the zone and then drops sharply, close to where pure Cpe populations are found. A novel interstitial C-band (H) has been found, probably derived from that characterizing Cpe individuals (E). This marker band is only present in hybrid populations. These data are discussed in the light of the dynamics of the hybrid zone.     

Molecular and morphological patterns of introgression between two large white-headed gull species in a zone of recent secondary contact

Incomplete reproductive isolation promotes gene flow between diverging taxa. However, any gene encoding for traits involved in the reproductive barriers will be less prone to introgression than neutral markers. Comparing introgression rates among loci is thus informative of the number and functions of loci involved in the reproductive barriers. This study aimed at identifying possible mechanisms of restriction to gene flow across a zone of recent secondary contact between Larus argentatus and Larus cachinnans by comparing introgression patterns for nine microsatellite loci, a fragment of mitochondrial DNA and a set of phenotypic traits. The low linkage disequilibrium between neutral nuclear markers indicated introgression without any barrier to gene flow. However, asymmetric introgression of mitochondrial DNA suggested that interspecific crosses may be more successful in one direction. The introgression rate for phenotypic traits was variable and low compared to neutral molecular markers. This was particularly evident in colouration of bare parts: individuals with intermediate colouration were scarcer in sympatry than expected if the genomes recombined freely. We hypothesized that one of these variables, the orbital ring colour, may play a role in mate choice, acting as an incomplete premating barrier through assortative mating. This study emphasizes that multilocus approaches are useful to discriminate among possible mechanisms responsible for the maintenance of hybrid zones.     

History repeated: recent and historical mitochondrial introgression between the current darter Etheostoma uniporum and rainbow darter Etheostoma caeruleum (Teleostei: Percidae)

Incongruence between recognized taxonomy and phylogenetic relationships between two species from a diverse clade (Percidae: Etheostomatinae) of stream fishes was found in a mitochondrial (mt) DNA gene tree. Two darters in subgenus Oligocephalus , Etheostoma uniporum current darter and Etheostoma caeruleum rainbow darter were sampled throughout their sympatric distribution in the Ozark Highlands of the central United States. Sequences from cytochrome (cyt) b and the first intron of the nuclear marker S7 were analysed separately using maximum parsimony and Bayesian methods. Cyt b recovered both species as polyphyletic; E. uniporum haplotypes were interspersed within E. caeruleum . However, both species were monophyletic and non-sister taxa based on S7. The cyt b gene tree pattern is caused by introgressive hybridization resulting in the mtDNA replacement of E. uniporum haplotypes by those of E. caeruleum . Some E. uniporum haplotypes are shared with geographically proximate E. caeruleum , and this is consistent with recent hybridization, while other E. uniporum haplotypes indicate historical sorting of introgressed lineages. The mechanism of introgression is likely asymmetric sneaking behaviour by male E. uniporum , a mating tactic observed in related species. MtDNA replacement may have occurred in E. uniporum due to drift fixation in a historically small female effective population. Additional evidence for darter hybridization will likely be discovered in future molecular genetic surveys of the nearly 200 species in eastern North America.     

Allozyme and mitochondrial DNA analysis of a hybrid zone between white-tailed deer and mule deer (Odocoileus) in west texas

Thirty allozyme loci and 35 mitochondrial DNA (mtDNA) restriction sites were examined in 24 white-tailed deer and 46 mule deer from a hybrid zone in West Texas. A common mtDNA genotype is shared by all of the mule deer with 67% of the white-tailed deer. At the albumin locus, 13% of the white-tailed deer and 24% of the mule deer are heterozygous, sharing alleles that are otherwise species-specific in allopatric populations; 7% of the mule deer are homozygous for the allele that is characteristic of allopatric white-tailed deer. Gene flow appears to have been bidirectional, with greater genetic introgression into mule deer. The mtDNA data suggest that matings between white-tailed and mule deer have occurred in the past. Despite evidence of genetic introgression, analysis of multilocus genotypes indicates that none of the deer examined is an F₁ hybrid. Production of such hybrids appears to be generally uncommon in North American deer; management plans that assume otherwise should be reconsidered.This work was supported by an NIH Biomedical Research Support Grant, Texas Agricultural Experiment Station Program Development and Expanded Research Awards, the Caesar Kleberg Research Program in Wildlife Ecology, and a Natural Sciences and Engineering Research Council Operating Grant.     

A hybrid zone and bidirectional introgression between two catadromous species: Australian bass Macquaria novemaculeata and estuary perch Macquaria colonorum

The presence and distribution of hybrid individuals and the existence of a hybrid zone between the catadromous Australian bass Macquaria novemaculeata and estuary perch Macquaria colonorum were investigated throughout the range of both species in Australia. Bayesian analyses and genotypic simulations identified 140 putative hybrids (11·5% of the total sample) with varying levels of introgression. Most hybrids were observed in an area extending from the Snowy River to the Albert River suggesting a hybrid zone in the eastern Bass Strait region. Sixteen hybrids, however, were found outside this zone, possibly reflecting the movement of hybrid offspring between estuaries or their inadvertent release during fish stocking programmes. Biparental backcrossing was found to occur suggesting that hybrids were fertile. These results have implications for the management of the extensive stocking programme in M. novemaculeata and for understanding the potential role of habitat degradation and reduced water flow in facilitating hybridization in species with migratory life histories.     

Gene flow and differential mortality in a contact zone between two Albinaria species (Gastropoda; Clausiliidae)

This paper reports on the disruption of normally effective ecological and behavioural barriers, at a contact zone on Monemvasia peninsula (Peloponnese, Greece), between parapatric populations of the land snail species Albinaria discolor and Albinaria campylauchen. Detected outcomes were the increase of gene flow between these two species at the contact zone, the occurrence of rare alleles, as well as the occurrence of morphologically and ethologically intermediate A. campylauchen specimens. Furthermore, there was high genetic variability along with deficiency of heterozygotes, probably due to inbreeding. Additionally, the study of mortality of these populations, during the adverse summer aestivation period, indicated that A. campylauchen individuals with an inappropriate combination of morphological and behavioural characteristics suffered heavily. The maladaptation and disadvantage of these specimens revealed the possible action of a postzygotic isolation mechanism between these two morphologically, molecularly and electrophoretically distinct species, that can, however, mate under laboratory conditions. Finally, the above situation is discussed in terms of exogenous versus endogenous selection.     

Mitochondrial DNA variation in bull trout (Salvelinus confluentus) from northwestern North America: implications for zoogeography and conservation

Bull trout, Salvelinus confluentus (Salmonidae), are distributed in northwestern North America from Nevada to Yukon Territory, largely in interior drainages. The species is of conservation concern owing to declines in abundance, particularly in southern portions of its range. To investigate phylogenetic structure within bull trout that might form the basis for the delineation of major conservation units, we conducted a mitochondrial DNA (mtDNA) survey in bull trout from throughout its range. Restriction fragment length polymorphism (RFLP) analysis of four segments of the mtDNA genome with 11 restriction enzymes resolved 21 composite haplotypes that differed by an average of 0.5% in sequence. One group of haplotypes predominated in 'coastal' areas (west of the coastal mountain ranges) while another predominated in 'interior' regions (east of the coastal mountains). The two putative lineages differed by 0.8% in sequence and were also resolved by sequencing a portion of the ND1 gene in a representative of each RFLP haplotype. Significant variation existed within individual sample sites (12% of total variation) and among sites within major geographical regions (33%), but most variation (55%) was associated with differences between coastal and interior regions. We concluded that: (i) bull trout are subdivided into coastal and interior lineages; (ii) this subdivision reflects recent historical isolation in two refugia south of the Cordilleran ice sheet during the Pleistocene: the Chehalis and Columbia refugia; and (iii) most of the molecular variation resides at the interpopulation and inter-region levels. Conservation efforts, therefore, should focus on maintaining as many populations as possible across as many geographical regions as possible within both coastal and interior lineages.     

Evidence for gene flow in parasitic nematodes between two host species of shrews

We describe the genetic structure of populations of the intestinal nematode Longistriata caudabullata (Trichostrongyloidea: Heligmosomidae), a common parasite of short-tailed shrews (genus Blarina, Insectivora: Soricidae). Parasites and hosts were collected from a transect across a contact zone between two species of hosts, Blarina brevicauda and B. hylophaga, in central North America. An 800-base pairs (bp) fragment of the ND4 mitochondrial DNA (mtDNA) gene was sequenced for 28 worms and a 783-bp fragment of the mtDNA control region was analysed for 16 shrews. Phylogenetic analyses of mtDNA sequences revealed reciprocal monophyly for the shrew species, concordant with morphological diagnosis, and supported the idea that the transect cuts through a secondary contact zone between well-differentiated B. brevicauda and B. hylophaga. In contrast to this pattern, the parasitic nematode mtDNA phylogeny was not subdivided according to host affiliation. Genealogical discordance between parasite and host phylogenies suggests extensive gene flow among parasites across the host species boundary.     

Asymmetric and differential gene introgression at a contact zone between two highly divergent lineages of field voles (Microtus agrestis)

Secondary contact zones have the potential to shed light on the mode and rate at which reproductive isolation accumulates during allopatric speciation. We investigated the population genetics of a contact zone between two highly divergent lineages of field voles (Microtus agrestis) in the Swiss Jura mountains. To shed light on the processes underlying introgression, we used maternally, paternally, and bi-parentally inherited markers. Though the two lineages maintained a strong genetic structure, we found some hybrids and evidence of gene flow. The extent of introgression varied with the mode of inheritance, being highest for mtDNA and absent for the Y chromosome. In addition, introgression was asymmetric, occurring only from the Northern to the Southern lineage. Both patterns seem parsimoniously explained by neutral processes linked to differences in effective sizes and sex-biased dispersal rates. The lineage with lower effective population size was also the more introgressed, and the mode-of-inheritance effect correlated with the male-biased dispersal rate of microtine rodents. We cannot exclude, however, that Haldane's effect contributed to the latter, as we found a marginally significant deficit in males (the heterogametic sex) among hybrids. We propose a possible demographic scenario to account for the patterns documented, and empirical extensions to further investigate this contact zone.     

Humid forest refugia, speciation and secondary introgression between evolutionary lineages: differentiation in a Near Eastern brown frog, Rana macrocnemis

Brown frogs of the complex Rana macrocnemis demonstrate various degrees of differentiation between the two widespread forms, macrocnemis and camerani , in different parts of Anatolia, the Caucasus Isthmus and northern Iran. In order to reveal whether or not the forms represent monophyletic evolutionary lineages, we analysed the geographic distribution of characters of external morphology, alleles at three polymorphic allozyme loci and mitochondrial DNA haplotypes. All three character sets showed a highly congruent pattern in a limited area of humid forests in the south-west Caucasus. Frogs from this area (form macrocnemis) represent a monophyletic lineage which forms a narrow hybrid zone with populations of camerani inhabiting the southern Caucasus and north-east Turkey. However, in other parts of the group's range, hybridization between the two lineages resulted in the merger of forms and the formation of a clinal pattern of variations. The role of landscape-dependent diversifying selection in the evolution of the group is discussed. Conceptual difficulties in delimiting the borders of subspecies are pointed out.     

Asymmetrical introgression between two Morus species (M. alba, M. rubra) that differ in abundance

Asymmetrical introgression is an expected genetic consequence of hybridization when parental taxa differ in abundance; however, evidence for such effects in small populations is scarce. To test this prediction, we estimated the magnitude and direction of hybridization between red mulberry (Morus rubra L.), an endangered species in Canada, and the introduced and more abundant white mulberry (Morus alba L.) using nuclear (randomly amplified polymorphic DNA) and cytoplasmic (chloroplast DNA sequence) markers. Parentage of 184 trees (n = 42 using cpDNA) from four sympatric populations was estimated using a hybrid index and related to six morphological characters and population frequencies of the parental classes. Overall, the frequency of nuclear hybrids was 53.7% (n = 99) and ranged from 43% to 67% among populations. The parental and hybrid taxa differed with respect to all of the morphological traits. Sixty-seven percent of all hybrids contained more nuclear markers from M. alba than M. rubra (hybrid index x = 0.46); among populations, the degree of M. alba bias was correlated with the frequency of M. alba. In addition, the majority of hybrids (68%) contained the chloroplast genome of white mulberry. These results suggest that introgression is bidirectional but asymmetrical and is related, in part, to the relative frequency of parental taxa.     

Limited hybridization along a large contact zone between two genetic lineages of the butterfly Erebia medusa (Satyrinae, Lepidoptera) in Central Europe

Genetic lineages evolving during glacial isolation frequently come into contact as the result of postglacial range expansions. Hybridization often occurs along these contact zones. In Europe, the high mountain systems of the Alps and Pyrenees are well known for their hybrid belts. This article studies the contact zone of the Woodland Ringlet Erebia medusa in a Hercynian mountain area in the Czech-German border region not exceeding 1500 m a.s.l. Hybrid populations between an eastern and a western genetic lineage were detected by amova -based tests, principal component analysis and neighbour joining analysis. Over most of the range of the Czech-German Border Mts, the ridges separate the western and the eastern genetic lineage from each other. However, two important hybrid areas were detected: (1) the watershed of the Ohře river in the north-west of that area, a major valley system passing through these mountains and (2) the high plateaux of the Šumava Mts in the south-east, an extended area of high elevation. The location of hybrid populations in geographical vicinity to non-hybrid populations and the generally low F _IS (2.1%) make reduced fitness of hybrid individuals little likely. The hybrid populations have intermediate genetic diversity between the genetically poor western and the genetically rich eastern lineage populations.     

Large-scale asymmetric introgression of cytoplasmic DNA reveals Holocene range displacement in a North American boreal pine complex

Jack pine (Pinus banksiana) and lodgepole pine (Pinus contorta var. latifolia) are two North American boreal hard pines that hybridize in their zone of contact in western Canada. The main objective of this study was to characterize their patterns of introgression resulting from past and recent gene flow, using cytoplasmic markers having maternal or paternal inheritance. Mitochondrial DNA (mtDNA) and chloroplast DNA (cpDNA) diversity was assessed in allopatric populations of each species and in stands from the current zone of contact containing morphological hybrids. Cluster analyses were used to identify genetic discontinuities among groups of populations. A canonical analysis was also conducted to detect putative associations among cytoplasmic DNA variation, tree morphology, and site ecological features. MtDNA introgression was extensive and asymmetric: it was detected in P. banksiana populations from the hybrid zone and from allopatric areas, but not in P. contorta populations. Very weak cpDNA introgression was observed, and only in P. banksiana populations. The mtDNA introgression pattern indicated that central Canada was first colonized by migrants from a P. contorta glacial population located west of the Rocky Mountains, before being replaced by P. banksiana migrating westward during the Holocene. In contrast, extensive pollen gene flow would have erased the cpDNA traces of this ancient presence of P. contorta. Additional evidence for this process was provided by the results of canonical analysis, which indicated that the current cpDNA background of trees reflected recent pollen gene flow from the surrounding dominant species rather than historical events that took place during the postglacial colonization.     

Wolbachia have made it twice: Hybrid introgression between two sister species of Eurema butterflies

Wolbachia, cytoplasmically inherited endosymbionts of arthropods, are known to hijack their host reproduction in various ways to increase their own vertical transmission. This may lead to the selective sweep of associated mitochondria, which can have a large impact on the evolution of mitochondrial lineages. In Japan, two different Wolbacahia strains (wCI and wFem) are found in two sister species of pierid butterflies, Eurema mandarina and Eurema hecabe. In both species, females infected with wCI (C females) produce offspring with a nearly 1:1 sex ratio, while females infected with both wCI and wFem (CF females) produce all‐female offspring. Previous studies have suggested the historical occurrence of hybrid introgression in C individuals between the two species. Furthermore, hybrid introgression in CF individuals is suggested by the distinct mitochondrial lineages between C females and CF females of E. mandarina. In this study, we performed phylogenetic analyses based on nuclear DNA and mitochondrial DNA markers of E. hecabe with previously published data on E. mandarina. We found that the nuclear DNA of this species significantly diverged from that of E. mandarina. By contrast, mitochondrial DNA haplotypes comprised two clades, mostly reflecting Wolbachia infection status rather than the individual species. Collectively, our results support the previously suggested occurrence of two independent historical events wherein the cytoplasms of CF females and C females moved between E. hecabe and E. mandarina through hybrid introgression.     

Distinct patterns of hybridization across a suture zone in a coral reef fish (Dascyllus trimaculatus)

Hybrid zones are natural laboratories for investigating the dynamics of gene flow, reproductive isolation, and speciation. A predominant marine hybrid (or suture) zone encompasses Christmas Island (CHR) and Cocos (Keeling) Islands (CKE), where 15 different instances of interbreeding between closely related species from Indian and Pacific Oceans have been documented. Here, we report a case of hybridization between genetically differentiated Pacific and Indian Ocean lineages of the three‐spot dascyllus, Dascyllus trimaculatus (Rüppell, 1829). Field observations indicate there are subtle color differences between Pacific and Indian Ocean lineages. Most importantly, population densities of color morphs and genetic analyses (mitochondrial DNA and SNPs obtained via RADSeq) suggest that the pattern of hybridization within the suture zone is not homogeneous. At CHR, both color morphs were present, mitochondrial haplotypes of both lineages were observed, and SNP analyses revealed both pure and hybrid genotypes. Meanwhile, in CKE, the Indian Ocean color morphs were prevalent, only Indian Ocean mitochondrial haplotypes were observed, and SNP analysis showed hybrid individuals with a large proportion (~80%) of their genotypes assigning to the Indian Ocean lineage. We conclude that CHR populations are currently receiving an influx of individuals from both ocean basins, with a greater influence from the Pacific Ocean. In contrast, geographically isolated CKE populations appear to be self‐recruiting and with more influx of individuals from the Indian Ocean. Our research highlights how patterns of hybridization can be different at scales of hundreds of kilometers, due to geographic isolation and the history of interbreeding between lineages.