Speciation with gene flow and the genetics of habitat transitions

Whether speciation can advance to completion in the face of initially high levels of gene flow is a very controversial topic in evolutionary biology. Extensive gene exchange is generally considered to homogenize populations and counteract divergence. Moreover, the role of introgressive hybridization in evolution remains largely unexplored in animals, particularly in freshwater zooplankton in which allopatric speciation is considered to be the norm. Our work investigates the genetic structure of two young ecological species: the pond species, Daphnia pulex and the lake species, Daphnia pulicaria. Phylogenetic and population genetics analyses were conducted on mitochondrial NADH dehydrogenase 5 (ND5) gene, the nuclear Lactate dehydrogenase (Ldh) gene and 21 nuclear microsatellite markers in 416 individuals from habitats with various degrees of permanence. The strong and consistent phylogenetic discordance between nuclear and mitochondrial markers suggests a complex evolutionary history of multiple independent habitat transition events that involved hybridization and introgression between lake and pond Daphnia. On the other hand, the low level of contemporary gene flow between adjacent populations indicates the presence of effective habitat isolating barriers. The Daphnia system provides strong evidence for a divergence-with-gene flow speciation model that involves multiple habitat transition events.     

Nuclear gene sequences resolve species phylogeny and mitochondrial introgression in Leptocarabus beetles showing trans-species polymorphisms

We studied the phylogenetic relationships among Japanese Leptocarabus ground beetles, which show extensive trans-species polymorphisms in mitochondrial gene genealogies. Simultaneous analysis of combined nuclear data with partial sequences from the long-wavelength rhodopsin, wingless, phosphoenolpyruvate carboxykinase, and 28S rRNA genes resolved the relationships among the five species, although separate analyses of these genes provided topologies with low resolution. For both the nuclear gene tree resulting from the combined data from four genes and a mitochondrial cytochrome oxidase subunit I (COI) gene tree, we applied a Bayesian divergence time estimation using a common calibration method to identify mitochondrial introgression events that occurred after speciation. Three mitochondrial lineages shared by two or three species were likely subject to introgression due to interspecific hybridization because the coalescent times for these lineages were much shorter than the corresponding speciation times estimated from nuclear gene sequences. We demonstrated that when species phylogeny is fully resolved with nuclear gene sequence data, comparative analysis of nuclear and mitochondrial gene trees can be used to infer introgressive hybridization events that might cause trans-species polymorphisms in mitochondrial gene trees.     

Hybrid populations selectively filter gene introgression between species

Hybrids have long been recognized as a potential pathway for gene flow between species that can have important consequences for evolution and conservation biology. However, few studies have demonstrated that genes from one species can introgress or invade another species over a broad geographic area. Using 35 genetically mapped restriction fragment length polymorphism (RFLP) markers of two species of cottonwoods (Populus fremontii x P. angustifolia) and their hybrids (n = 550 trees), we showed that the majority of the genome is prohibited from introgressing from one species into the other. However, this barrier was not absolute; Fremont cpDNA and mtDNA were found throughout the geographic range of narrowleaf cottonwood, and 20% of the nuclear markers of Fremont cottonwood introgressed varying distances (some over 100 km) into the recipient species' range. Rates of nuclear introgression were variable, but two nuclear markers introgressed as fast as the haploid, cytoplasmically inherited chloroplast and mitochondrial markers. Our genome-wide analysis provides evidence for positive, negative, and neutral effects of introgression. For example, we predict that DNA fragments that introgress through several generations of backcrossing will be small, because small fragments are less likely to contain deleterious genes. These results argue that recombination will be important, that introgression can be very selective, and that evolutionary forces within the hybrid population to effectively "filter" gene flow between species. A strong filter may make introgression adaptive, prevent genetic assimilation, lead to relaxed isolating mechanisms, and contribute to the stability of hybrid zones. Thus, rather than hybridization being a negative factor as is commonly argued, natural hybridization between native species may provide important genetic variation that impacts both ecological and evolutionary processes. Finally, we propose two hypotheses that contrast the likelihood of contemporary versus ancient introgression in this system.     

Limited gene exchange between two sister species of leaf beetles within a hybrid zone in the Alps

Analysing genomic variation within and between sister species is a first step towards understanding species boundaries. We focused on two sister species of cold‐resistant leaf beetles, Gonioctena quinquepunctata and G. intermedia, whose ranges overlap in the Alps. A previous study of DNA sequence variation had revealed multiple instances of mitochondrial genome introgression in this region, suggesting recent hybridization between the two species. To evaluate the extent of gene exchange resulting from these hybridization events, we sampled individuals of both species inside and outside the hybrid zone and analysed genomic variation among them using RAD‐seq markers. Individual levels of introgression in the nuclear genome were estimated first by defining species‐specific SNPs (displaying a fixed difference between species) a priori and second by using model‐based methods. Both types of analyses indicated little gene exchange, if any, between species at the level of the nuclear genome. Whereas the first method suggested slightly more gene flow, we argue that it has likely overestimated introgression in the phylogeographic context of this study. We conclude that strong intrinsic barriers prevent genetic exchange at the level of the nuclear genome between the two species. The apparent discrepancy observed between introgression occurring in the nuclear and mitochondrial genomes could be explained by selection acting in favour of the latter. Also, these results have consequences for the phylogeographic study of each species, since we can assume that genetic diversity in the overlapping portion of their ranges is not the product of introgression.     

Chlorophyll-binding proteins revisited - a multigenic family of light-harvesting and stress proteins from a brown algal perspective

Background

Mitochondrial introgression may result in the mitochondrial genome of one species being replaced by that of another species without leaving any trace of past hybridization in its nuclear genome. Such introgression can confuse the species genealogy estimates and lead to absurd inferences of species history. We used a phylogenetic approach to explore the potential mitochondrial genome introgression event(s) between two closely related green pond frog species, Pelophylax nigromaculatus and P. plancyi.

Results

DNA sequence data of one mitochondrial and two nuclear genes from an extensive sampling of the two species were collected, and the genealogies of the three genes were constructed and compared. While the two nuclear genes congruently showed mutual reciprocal monophyly of both species, the mitochondrial phylogeny separated a Korean P. nigromaculatus clade, a paraphyletic central China P. plancyi assemblage, and a large well-supported introgression clade. Within the introgression clade, the mitochondrial haplotypes of the two species were mixed together. This reticulated pattern can be most parsimoniously explained by an ancient mitochondrial introgression event from P. plancyi to P. nigromaculatus that occurred at least 1.36 MYA, followed by multiple recent introgression events from P. nigromaculatus back to P. plancyi within the last 0.63 MY. The re-constitution of previously co-adapted genomes in P. plancyi may be responsible for the recent rampant introgression events. The Korean P. nigromaculatus clade likely represents the only surviving "true" mitochondrial lineage of P. nigromaculatus, and the central China P. plancyi assemblage likely represents the "original" P. plancyi mitochondrial lineage. Refugia in the Korean Peninsula and central China may have played a significant role in preserving these ancient lineages.

Conclusions

The majority of individuals in the two species have either introgressed (P. nigromaculatus) or reclaimed (P. plancyi) mitochondrial genomes while no trace of past hybridization in their nuclear genomes was detected. Asymmetrical reproductive ability of hybrids and continuous backcrossing are likely responsible for the observed mitochondrial introgression. This case is unique in that it includes an ancient "forward" introgression and many recent "backward" introgressions, which re-constitutes the original nuclear and mitochondrial genomes of P. plancyi. This hybrid system provides an excellent opportunity to study cyto-nuclear interaction and co-adaptation.     

Ancient hybridization and mitochondrial capture between two species of chipmunks

Models that posit speciation in the face of gene flow are replacing classical views that hybridization is rare between animal species. We use a multilocus approach to examine the history of hybridization and gene flow between two species of chipmunks ( Tamias ruficaudus and T. amoenus ). Previous studies have shown that these species occupy different ecological niches and have distinct genital bone morphologies, yet appear to be incompletely isolated reproductively in multiple areas of sympatry. We compared data from four sequenced nuclear loci and from seven microsatellite loci to published cytochrome b sequences. Interspecific gene flow was primarily restricted to introgression of the T. ruficaudus mitochondrial genome into a sympatric subspecies of T. amoenus , T. a. canicaudus , with the four sequenced nuclear loci showing little to no interspecific allele sharing. Microsatellite data were consistent with high levels of differentiation between the species and also showed no current gene flow between broadly sympatric populations of T. a. canicaudus and T. ruficaudus . Coalescent analyses date the mtDNA introgression event from the mid-Pleistocene to late Pliocene. Overall, these data indicate that introgression has had a minimal impact on the nuclear genomes of T. amoenus and T. ruficaudus despite multiple independent hybridization events. Our findings challenge long-standing assumptions on patterns of reproductive isolation in chipmunks and suggest that there may be other examples of hybridization among the 23 species of Tamias that occur in western North America.     

Radiation and reticulation: extensive introgressive hybridization in the carabid beetles Ohomopterus inferred from mitochondrial gene genealogy

 In the evolutionary process of an animal lineage, interactions in secondary contacts of differentiated populations and introgressive hybridization may play an important role. In the Japanese islands, the carabid subgenus Ohomopterus (genus Carabus) exhibits a marked differentiation in body size and genital morphology. Although geographical differentiation is apparent, two or three species usually coexist at many localities. Their reproductive isolation relies on body size differences, chemical cues for mate recognition, and a species-specific genital lock-and-key system. However, these isolation mechanisms are not always effective enough to prevent interspecific hybridization. An initial assessment of the species-level phylogeny with mitochondrial gene sequences revealed that the gene genealogy is highly inconsistent with the morphology-based taxonomy. A comparison of mitochondrial and nuclear gene genealogies showed that these are strongly incongruent with each other, while the nuclear gene genealogy is more consistent with traditional taxonomy, indicating the repeated occurrence of introgression of mitochondria across species. Here, two different cases of mitochondrial introgression among Ohomopterus species are described in detail, one for parapatric species and the other for sympatric species. First, mitochondrial haplotypes and sequences were studied in Carabus insulicola and three taxa parapatric with C. insulicola, at least two of which hybridize with C. insulicola naturally. Among the four species studied, directional introgressions of mitochondria across boundary zones were detected. Second, in the Mt. Kongo area in central Honshu, which harbors five species, introgression of mitochondria among four out of the five species was detected, despite the apparent absence of on-going natural hybridization. These inferred cases of mitochondrial introgression indicate that species interactions through hybridization could have played an important role at various stages in the evolution of Ohomopterus. Received: April 12, 2002 / Accepted: October 17, 2002 Acknowledgments I am grateful to Alfried P. Vogler for a long-lasting collaboration in the molecular phylogenetic study of Ohomopterus. R. Ishikawa, K. Kubota, M. Ujiie, Y. Takami, and F. Kusumoto have also collaborated at various stages of this study. Thanks are also due to K. Miyashita, T. Funakoshi, H. Fujimoto, T. Dejima, Y. Nagahata, T. Miyagawa, K. Yodoe, H. Kadowaki, S. Nakamine, Y. Oka, H. Tanaka, T. Tanabe, K. Kusakari, and T. Okumura for their care of specimens. Supported by grants-in-aid from the Japan Society for the Promotion of Science (Nos. 09640748, 11304056).     

The role of mitochondrial introgression in illuminating the evolutionary history of Nearctic treefrogs

Inferring the evolutionary history of lineages often becomes difficult when gene histories are in conflict with each other. Introgression, for example, can cause DNA sequences from one species to be more similar to sequences of a different species and lead to incongruence amongst gene trees. However, incorporating congruent and incongruent locus‐specific phylogenetic estimates with the geographical distribution of lineages may provide valuable insight into evolutionary processes important to speciation. In this study, we investigated mitochondrial introgression within the Hyla eximia group to better understand its role in illuminating the evolutionary history and phylogeography of these treefrogs. We reconstructed and compared the matrilineal history of the Hyla eximia group with estimates of evolutionary history inferred from nuclear genes. We tested for introgression within the mitochondrial and nuclear genes using a posterior predictive checking approach. Reconstructions of the species tree based on the mitochondrial DNA (mtDNA) and nuclear DNA data were strongly discordant. Introgression between lineages was widespread in the mtDNA data set (145 occurrences amongst 11 of the 16 lineages), but uncommon in the nuclear genes (12 occurrences amongst four of the 16 lineages). Nonetheless, the geographical structuring of mtDNA within species provides valuable information on biogeographical areas, ancient areas of hybridization, and unique histories of lineages within the H. eximia group. These results suggest that the combination of nuclear, mitochondrial, and spatial information can provide a more complete picture of ‘how evolutionary history played out’, particularly in cases where mitochondrial introgression is known to occur. © 2014 The Linnean Society of London     

Mitonuclear discordance is caused by rampant mitochondrial introgression in Neodiprion (Hymenoptera: Diprionidae) sawflies

We investigate the pervasiveness of hybridization and mitochondrial introgression in Neodiprion Rohwer (Hymenoptera; Diprionidae), a Holarctic genus of conifer-feeding sawflies. A phylogenetic analysis of the lecontei species group revealed extensive discordance between a contiguous mitochondrial region spanning three genes (COI, tRNA-leucine, and COII) and three nuclear loci (EF1alpha, CAD, and an anonymous nuclear locus). Bayesian tests of monophyly and Shimodaira-Hasegawa (SH) tests of topological congruence were consistent with mitochondrial introgression; however, these patterns could also be explained by lineage sorting (i.e., deep coalescence). Therefore, to explicitly test the mitochondrial introgression hypothesis, we used a novel application of coalescent-based isolation with migration (IM) models to measure interspecific gene flow at each locus. In support of our hypothesis, mitochondrial gene flow was consistently higher than nuclear gene flow across 120 pairwise species comparisons (P < 1 x 10(-12)). We combine phylogenetic and coalescent evidence to identify likely cases of recent and ancient introgression in Neodiprion, and based on these observations, we hypothesize that shared hosts and/or pheromones facilitate hybridization, whereas disparate abundances between hybridizing species promote mitochondrial introgression. Our results carry implications for phylogenetic analysis, and we advocate the separation of high and low gene flow regions to inform analyses of hybridization and speciational history, respectively.     

Utility of nuclear allele networks for the analysis of closely related species in the genus Carabus, subgenus Ohomopterus

Nuclear DNA sequence data for diploid organisms are potentially a rich source of phylogenetic information for disentangling the evolutionary relationships of closely related organisms, but present special phylogenetic problems owing to difficulties arising from heterozygosity and recombination. We analyzed allelic relationships for two nuclear gene regions (phosphoenolpyruvate carboxykinase and elongation factor-1a), along with a mitochondrial gene region (NADH dehydrogenase subunit 5), for an assemblage of closely related species of carabid beetles (Carabus subgenus Ohomopterus). We used a network approach to examine whether the nuclear gene sequences provide substantial phylogenetic information on species relationships and evolutionary history. The mitochondrial gene genealogy strongly contradicted the morphological species boundary as a result of introgression of heterospecific mitochondria. Two nuclear gene regions showed high allelic diversity within species, and this diversity was partially attributable to recombination between various alleles and high variability in the intron region. Shared nuclear alleles among species were rare and were considered to represent shared ancestral polymorphism. Despite the presence of recombination, nuclear allelic networks recovered species monophyly more often and presented genetic differentiation patterns (low to high) among species more clearly. Overall, nuclear gene networks provide clear evidence for separate biological species and information on the phylogenetic relationships among closely related carabid beetles.     

Interspecific hybridization causes long‐term phylogenetic discordance between nuclear and mitochondrial genomes in freshwater fishes

Classification, phylogeography and the testing of evolutionary hypotheses rely on correct estimation of species phylogeny. Early molecular phylogenies often relied on mtDNA alone, which acts as a single linkage group with one history. Over the last decade, the use of multiple nuclear sequences has often revealed conflict among gene trees. This observation can be attributed to hybridization, lineage sorting, paralogy or selection. Here, we use 54 groups of fishes from 48 studies to estimate the degree of concordance between mitochondrial and nuclear gene trees in two ecological grades of fishes: marine and freshwater. We test the hypothesis that freshwater fish phylogenies should, on average, show more discordance because of their higher propensity for hybridization in the past. In keeping with this idea, concordance between mitochondrial and nuclear gene trees (as measured by proportion of components shared) is on average 50% higher in marine fishes. We discuss why this difference almost certainly results from introgression caused by greater historical hybridization among lineages in freshwater groups, and further emphasize the need to use multiple nuclear genes, and identify conflict among them, in estimation of species phylogeny.     

Influence of hydrogeographic history and hybridization on the distribution of genetic variation in the pupfishes Cyprinodon atrorus and C. bifasciatus

The evolutionary importance of hybridization in animals has been subject of much debate. In this study, we examined the influence of hydrogeographic history and hybridization on the present distribution of nuclear and mitochondrial DNA variation in two pupfish species, Cyprinodon atrorus and Cyprinodon bifasciatus. Results presented here indicate that there has been limited introgression of nuclear genes; however, mtDNA introgression has been substantial, with complete replacement of the C. bifasciatus mitochondrial genome by that of C. atrorus. Subsequent to this replacement, there has been diversification of mitochondrial haplotypes along major geographic regions in the basin. Evidence was also found that mitochondrial replacement follows a predictable, cyclical pattern in this system, with isolation and diversification followed by re-contact and replacement of C. bifasciatus mitochondrial haplotypes by those of C. atrorus. This pattern is best explained by a combination of a numeric bias towards C. atrorus and mating site selection rather than selection for C. atrorus mitochondrial genome. These results demonstrate the important role hybridization can play in evolution.     

Negligible nuclear introgression despite complete mitochondrial capture between two species of chipmunks

The idea that species boundaries can be semipermeable to gene flow is now widely accepted but the evolutionary importance of introgressive hybridization remains unclear. Here we examine the genomic contribution of gene flow between two hybridizing chipmunk species, Tamias ruficaudus and T. amoenus. Previous studies have shown that ancient hybridization has resulted in complete fixation of introgressed T. ruficaudus mitochondrial DNA (mtDNA) in some populations of T. amoenus, but the extent of nuclear introgression is not known. We used targeted capture to sequence over 10,500 gene regions from multiple individuals of both species. We found that most of the nuclear genome is sorted between these species and that overall genealogical patterns do not show evidence for introgression. Our analysis rules out all but very minor levels of interspecific gene flow, indicating that introgressive hybridization has had little impact on the overall genetic composition of these species outside of the mitochondrial genome. Given that much of the evidence for introgression in animals has come from mtDNA, our results underscore that unraveling the importance introgressive hybridization during animal speciation will require a genome‐wide perspective that is still absent for many species.     

Leaky prezygotic isolation and porous genomes: rapid introgression of maternally inherited DNA

Accurate phylogenies are crucial for understanding evolutionary processes, especially species diversification. It is commonly assumed that "good" species are sufficiently isolated genetically that gene genealogies represent accurate phylogenies. However, it is increasingly clear that good species may continue to exchange genetic material through hybridization (introgression). Many studies of closely related species reveal introgression of some genes without others, often with more rapid introgression of maternally inherited chloroplast or mitochondrial DNA (cpDNA, mtDNA). We seek a general explanation for this biased introgression using simple models of common reproductive isolating barriers (RIBs). We compare empirically informed models of prezygotic isolation (for pre- and postinsemination mechanisms of both female choice and male competition) with postzygotic isolation and demonstrate that rate of introgression depends critically upon type of RIB and mode of genetic inheritance (maternal versus biparental versus paternal). Our frequency-dependent prezygotic RIBs allow much more rapid introgression of biparentally and maternally inherited genes than do commonly modeled postzygotic RIBs (especially maternally inherited DNA). After considering the specific predictions in the context of empirical observations, we conclude that our model of prezygotie RIBs is a general explanation for biased introgression of maternally inherited genomic components. These findings suggest that we should use extreme caution when interpreting single gene genealogies as species phylogenies, especially for cpDNA and mtDNA.     

Phylogeny estimation of the radiation of western North American chipmunks (Tamias) in the face of introgression using reproductive protein genes

The causes and consequences of rapid radiations are major unresolved issues in evolutionary biology. This is in part because phylogeny estimation is confounded by processes such as stochastic lineage sorting and hybridization. Because these processes are expected to be heterogeneous across the genome, comparison among marker classes may provide a means of disentangling these elements. Here we use introns from nuclear-encoded reproductive protein genes expected to be resistant to introgression to estimate the phylogeny of the western chipmunks (Tamias: subgenus: Neotamias), a rapid radiation that has experienced introgressive hybridization of mitochondrial DNA (mtDNA). We analyze the nuclear loci using coalescent-based species-tree estimation methods and concatenation to estimate a species tree and we use parametric bootstraps and coalescent simulations to differentiate between phylogenetic error, coalescent stochasticity and introgressive hybridization. Results indicate that the mtDNA gene tree reflects several introgression events that have occurred between taxa of varying levels of divergence and at different time points in the tree. T. panamintinus and T. speciosus appear to be fixed for ancient mitochondrial introgressions from T. minimus. A southern Rocky Mountains clade appears well sorted (i.e., species are largely monophyletic) at multiple nuclear loci, while five of six taxa are nonmonophyletic based on cytochrome b. Our simulations reject phylogenetic error and coalescent stochasticity as causes. The results represent an advance in our understanding of the processes at work during the radiation of Tamias and suggest that sampling reproductive-protein genes may be a viable strategy for phylogeny estimation of rapid radiations in which reproductive isolation is incomplete. However, a genome-scale survey that can statistically compare heterogeneity of genealogical process at many more loci will be necessary to test this conclusion.     

HALDANE'S RULE AND SEX BIASSED GENE FLOW BETWEEN TWO HYBRIDIZING FLYCATCHER SPECIES (FICEDULA ALBICOLLIS AND F. HYPOLEUCA,AVES: MUSCICAPIDAE)

The collared flycatcher (Ficedula albicollis) and the pied flycatcher (F. hypoleuca) hybridize where their geographic ranges overlap. Restriction fragment comparison of 5% of the mitochondrial genome showed a sequence divergence of 10% between these flycatcher species. This degree of sequence divergence between a closely related pair of bird species is unusually high and contrasts with the low level of divergence between F. albicollis and F. hypoleuca in nuclear genes (Nei's D = 0.0006) revealed by enzyme electrophoresis. The low nuclear differentiation is explained by sex biassed gene flow and introgression in nuclear genes (via fertile male hybrids), while the high mitochondrial DNA sequence divergence is preserved by sterility of female hybrids, which prevents mitochondrial introgression. This pattern is in accordance with Haldane's rule and is supported by field data on hybrid fertility. The high mtDNA differentiation could be explained by transfer of mitochondrial DNA from a third species during a past period of hybridization.     

Morphology and nuclear markers reveal extensive mitochondrial introgressions in the Iberian Wall Lizard species complex

Mitochondrial markers are still often used alone to identify evolutionary units, despite widespread evidence for processes such as incomplete lineage sorting or introgressive hybridization that may blur past population history. The combination of mitochondrial DNA data with other sources of information (morphology, nuclear genes) is a powerful tool to reveal when and why mitochondrial markers are potentially misleading. In this study, we evaluate the performance of mtDNA markers to unravel the evolutionary history of Spanish lizards from the Podarcis hispanicus species complex. We first uncover several cases of discordance between morphological and mitochondrial data in delimitation of taxa. To assess the origin of these discordances, we analysed the same populations using several independent nuclear loci. Both morphological and nuclear markers identified the same three evolutionary units in the region, while mitochondrial data revealed four deeply divergent lineages. We suggest here that the most likely scenario to explain this discordance is ancient mitochondrial introgression originating from a fourth evolutionary unit presently absent from the study area. Notably, this resulted in a complete replacement of the original lineage in a large part of the distribution of one of the taxa investigated. We discuss the potential evolutionary scenarios leading to this complete mitochondrial replacement and suggest why the previous studies have failed to recover the correct history of this species complex.     

Gene flow between Drosophila yakuba and Drosophila santomea in subunit V of cytochrome c oxidase: A potential case of cytonuclear cointrogression

Introgression is the effective exchange of genetic information between species through natural hybridization. Previous genetic analyses of the Drosophila yakuba—D. santomea hybrid zone showed that the mitochondrial genome of D. yakuba had introgressed into D. santomea and completely replaced its native form. Since mitochondrial proteins work intimately with nuclear‐encoded proteins in the oxidative phosphorylation (OXPHOS) pathway, we hypothesized that some nuclear genes in OXPHOS cointrogressed along with the mitochondrial genome. We analyzed nucleotide variation in the 12 nuclear genes that form cytochrome c oxidase (COX) in 33 Drosophila lines. COX is an OXPHOS enzyme composed of both nuclear‐ and mitochondrial‐encoded proteins and shows evidence of cytonuclear coadaptation in some species. Using maximum‐likelihood methods, we detected significant gene flow from D. yakuba to D. santomea for the entire COX complex. Interestingly, the signal of introgression is concentrated in the three nuclear genes composing subunit V, which shows population migration rates significantly greater than the background level of introgression in these species. The detection of introgression in three proteins that work together, interact directly with the mitochondrial‐encoded core, and are critical for early COX assembly suggests this could be a case of cytonuclear cointrogression.     

Widespread decoupling of mtDNA variation and species integrity in Grammia tiger moths (Lepidoptera: Noctuidae)

Abstract. We investigate the diversity of the North American tiger moth genus Grammia Rambur (Lepidoptera: Noctuidae) by comparing mitochondrial DNA (mtDNA) ‘barcode’ fragments of cytochrome oxidase I with non‐molecular characters such as morphology, ecology, behaviour and distribution. Mitochondrial DNA genealogy is strikingly at odds with morpho‐species taxonomy for most of the 28 sampled species, as haplotypic polyphyly not only is taxonomically widespread, but involves multiple shared haplotypes among two to four species. Morpho‐ecological traits show that those species sharing haplotypes are often not closely related. Furthermore, high mtDNA divergences occur within species. Haplotypic variation is highly discordant with species taxonomy, but variation at a continental scale reveals significant geographic structuring of haplogroups, transcending morpho‐species boundaries. A nested clade analysis and comparison of non‐molecular with mtDNA data indicate that most discordance between mtDNA and taxonomy in Grammia is explained best by taxonomically and geographically widespread ongoing hybridization events resulting in mtDNA introgression. We hypothesize that broad areas of sympatry, interspecifically compatible genitalic structure, and species overlap in pheromone components facilitate hybridization, with disparate interspecies abundances promoting mitochondrial introgression. The molecular evolution of Grammia challenges the view that interspecific gene exchange occurs rarely and is restricted to recently diverged species. These results show the value of mtDNA in detecting cryptic hybridization, while highlighting the inherent dangers of drawing taxonomic conclusions based solely on mtDNA.     

Subterranean termite phylogeography reveals multiple postglacial colonization events in southwestern Europe

A long‐standing goal of evolutionary biology is to understand how paleoclimatic and geological events shape the geographical distribution and genetic structure within and among species. Using a diverse set of markers (cuticular hydrocarbons, mitochondrial and nuclear gene sequences, microsatellite loci), we studied Reticulitermes grassei and R. banyulensis, two closely related termite species in southwestern Europe. We sought to clarify the current genetic structure of populations that formed following postglacial dispersal from refugia in southern Spain and characterize the gene flow between the two lineages over the last several million years. Each marker type separately provided a fragmented picture of the evolutionary history at different timescales. Chemical analyses of cuticular hydrocarbons and phylogenetic analyses of mitochondrial and nuclear genes showed clear separation between the species, suggesting they diverged following vicariance events in the Late Miocene. However, the presence of intermediate chemical profiles and mtDNA introgression in some Spanish colonies suggests ongoing gene flow. The current genetic structure of Iberian populations is consistent with alternating isolation and dispersal events during Quaternary glacial periods. Analyses of population genetic structure revealed postglacial colonization routes from southern Spain to France, where populations underwent strong genetic bottlenecks after traversing the Pyrenees resulting in parapatric speciation.