Evolution of novel mimicry rings facilitated by adaptive introgression in tropical butterflies

Understanding the genetic basis of phenotypic variation and the mechanisms involved in the evolution of adaptive novelty, especially in adaptive radiations, is a major goal in evolutionary biology. Here, we used whole‐genome sequence data to investigate the origin of the yellow hindwing bar in the Heliconius cydno radiation. We found modular variation associated with hindwing phenotype in two narrow noncoding regions upstream and downstream of the cortex gene, which was recently identified as a pigmentation pattern controller in multiple species of Heliconius. Genetic variation at each of these modules suggests an independent control of the dorsal and ventral hindwing patterning, with the upstream module associated with the ventral phenotype and the downstream module with the dorsal one. Furthermore, we detected introgression between H. cydno and its closely related species Heliconius melpomene in these modules, likely allowing both species to participate in novel mimicry rings. In sum, our findings support the role of regulatory modularity coupled with adaptive introgression as an elegant mechanism by which novel phenotypic combinations can evolve and fuel an adaptive radiation.     

Detecting adaptive trait introgression between Iris fulva and I. brevicaulis in highly selective field conditions

The idea that natural hybridization has served as an important force in evolutionary and adaptive diversification has gained considerable momentum in recent years. By combining genome analyses with a highly selective field experiment, we provide evidence for adaptive trait introgression between two naturally hybridizing Louisiana Iris species, flood-tolerant Iris fulva and dry-adapted I. brevicaulis. We planted reciprocal backcross (BC1) hybrids along with pure-species plants into natural settings that, due to a flooding event, favored I. fulva. As expected, I. fulva plants survived at much higher rates than I. brevicaulis plants. Backcross hybrids toward I. fulva (BCIF) also survived at significantly higher rates than the reciprocal backcross toward I. brevicaulis (BCIB). Survivorship of BCIB hybrids was strongly influenced by the presence of a number of introgressed I. fulva alleles located throughout the genome, while survivorship in the reciprocal BCIF hybrids was heavily influenced by two epistatically acting QTL of opposite effects. These results demonstrate the potential for adaptive trait introgression between these two species and may help to explain patterns of genetic variation observed in naturally occurring hybrid zones.     

Hybridization and mitochondrial genome introgression between Rana chensinensis and R. kukunoris

Mitochondrial genome (mito‐genome) introgression among metazoans is commonplace, and several biological processes may promote such introgression. We examined two proposed processes for the mito‐genome introgression between Rana chensinensis and R. kukunoris: natural hybridization and sex‐biased dispersal. We sampled 477 individuals from 28 sites in the potential hybrid zone in the western Tsinling Mountains. Mitochondrial gene (cyt‐b) trees were used to examine the introgression events. Microsatellite DNA loci, cyt‐b and morphological data were used to identify hybrids and to examine the extent of natural hybridization. We detected rampant bidirectional introgressions, both ancient and recent, between the two species. Furthermore, we found a wide hybrid zone, and frequent and asymmetric hybridization. The hybrid zone cline analysis revealed a clear mitochondrial–nuclear discordance; while most nuclear markers displayed similar and steep clines, cyt‐b had a displaced cline centre and a more gradual and wider cline. We also detected strong and asymmetric historical maternal gene flow across the hybrid zone. This widespread hybridization and detected low mito‐nuclear conflicts may, at least partially, explain the high frequency of introgression. Lastly, microsatellite data and population genetic methods were used to assess sex‐biased dispersal. A weak pattern of female‐biased dispersal was detected in both species, suggesting it may not play an important role in the observed introgression. Our data are consistent with the hybridization hypothesis, but support for the sex‐biased dispersal hypothesis is weak. We further suggest that selective advantages of the R. kukunoris‐type mito‐genome in thermal adaptation may also contribute to the introgression between the two species.     

Nuclear introgression without mitochondrial introgression in two turtle species exhibiting sex‐specific trophic differentiation

Despite the presence of reproductive barriers between species, interspecific gene introgression has been documented in a range of natural systems. Comparing patterns of genetic introgression in biparental versus matrilineal markers can potentially reveal sex‐specific barriers to interspecific gene flow. Hybridization has been documented in the freshwater turtles Graptemys geographica and G. pseudogeographica, whose ranges are largely sympatric. Morphological differentiation between the species is restricted to females, with female G. geographica possessing large heads and jaws compared to the narrow heads of G. pseudogeographica females. If hybrid females are morphologically intermediate, they may be less successful at exploiting parental feeding niches, thereby limiting the introgression of maternally inherited, but not biparental, molecular markers. We paired sequence data with stable isotope analysis and examined sex‐specific genetic introgression and trophic differentiation in sympatric populations of G. geographica and G. pseudogeographica. We observed introgression from G. pseudogeographica into G. geographica at three nuclear loci, but not at the mitochondrial locus. Analysis of ?¹⁵N and ?¹³C was consistent with species differences in trophic positioning in females, but not males. These results suggest that ecological divergence in females may reduce the opportunity for gene flow in this system.     

Massive introgression of major histocompatibility complex (MHC) genes in newt hybrid zones

Variation in the vertebrate major histocompatibility complex (MHC) genes is crucial for fighting pathogen assault. Because new alleles confer a selective advantage, MHC should readily introgress between species, even under limited hybridization. Using replicated transects through two hybrid zones between strongly reproductively isolated European newts, Lissotriton montandoni and L. vulgaris, we demonstrated recent and ongoing MHC class I and II introgression in the Carpathian region. The extent of introgression correlated with the age of contact. In the older zone, MHC similarity between species within transects exceeded similarity between transects within species, implying pervasive introgression ‐ a massive exchange of MHC genes, not limited to specific variants. In simulations, the observed pattern emerged under the combined action of balancing selection and hybridization, but not when these processes acted separately. Thus, massive introgression at advanced stages of divergence can introduce novel and restore previously lost MHC variation, boosting the adaptive potential of hybridizing taxa. In consequence, MHC genes may be the last to stop introgressing between incipient species.     

Hybridization between two parapatric ranid frog species in the northern Sierra Nevada,California, USA

Contact zones between species provide a unique opportunity to test whether taxa can hybridize or not. Cross‐breeding or hybridization between closely related taxa can promote gene flow (introgression) between species, adaptation, or even speciation. Though hybridization events may be short‐lived and difficult to detect in the field, genetic data can provide information about the level of introgression between closely related taxa. Hybridization can promote introgression between species, which may be an important evolutionary mechanism for either homogenization (reversing initial divergence between species) or reproductive isolation (potentially leading to speciation). Here, we used thousands of genetic markers from nuclear DNA to detect hybridization between two parapatric frog species (Rana boylii and Rana sierrae) in the Sierra Nevada of California. Based on principal components analysis, admixture, and analysis of heterozygosity at species diagnostic SNPs, we detected two F1 hybrid individuals in the Feather River basin, as well as a weak signal of introgression and gene flow between the frog species compared with frog populations from two other adjacent watersheds. This study provides the first documentation of hybridization and introgression between these two species, which are of conservation concern.     

Outlier loci highlight the direction of introgression in oaks

Loci considered to be under selection are generally avoided in attempts to infer past demographic processes as they do not fit neutral model assumptions. However, opportunities to better reconstruct some aspects of past demography might thus be missed. Here we examined genetic differentiation between two sympatric European oak species with contrasting ecological dynamics (Quercus robur and Quercus petraea) with both outlier (i.e. loci possibly affected by divergent selection between species or by hitchhiking effects with genomic regions under selection) and nonoutlier loci. We sampled 855 individuals in six mixed forests in France and genotyped them with a set of 262 SNPs enriched with markers showing high interspecific differentiation, resulting in accurate species delimitation. We identified between 13 and 74 interspecific outlier loci, depending on the coalescent simulation models and parameters used. Greater genetic diversity was predicted in Q. petraea (a late‐successional species) than in Q. robur (an early successional species) as introgression should theoretically occur predominantly from the resident species to the invading species. Remarkably, this prediction was verified with outlier loci but not with nonoutlier loci. We suggest that the lower effective interspecific gene flow at loci showing high interspecific divergence has better preserved the signal of past asymmetric introgression towards Q. petraea caused by the species' contrasting dynamics. Using markers under selection to reconstruct past demographic processes could therefore have broader potential than generally recognized.     

Differential introgression from a sister species explains high FST outlier loci within a mussel species

Scanning genomes for loci with high levels of population differentiation has become a standard of population genetics. F_ST outlier loci are most often interpreted as signatures of local selection, but outliers might arise for many other reasons too often left unexplored. Here, we tried to identify further the history and genetic basis underlying strong differentiation at F_ST outlier loci in a marine mussel. A genome scan of genetic differentiation has been conducted between Atlantic and Mediterranean populations of Mytilus galloprovincialis. The differentiation was low overall (F_ST = 0.03), but seven loci (2%) were strong F_ST outliers. We then analysed DNA sequence polymorphism at two outlier loci. The genetic structure proved to be the consequence of differential introgression of alleles from the sister‐hybridizing species Mytilus edulis. Surprisingly, the Mediterranean population was the most introgressed at these two loci, although the contact zone between the two species is nowadays localized along the Atlantic coasts of France and the British Isles. A historical contact between M. edulis and Mediterranean M. galloprovincialis should have happened during glacial periods. It proved difficult to disentangle two hypotheses: (i) introgression was adaptive, implying edulis alleles have been favoured in Mediterranean populations, or (ii) the genetic architecture of the barrier to edulis gene flow is different between the two M. galloprovincialis backgrounds. Five of the seven outliers between M. galloprovincialis populations were also outliers between M. edulis and Atlantic M. galloprovincialis, which would support the latter hypothesis. Differential introgression across semi‐permeable barriers to gene flow is a neglected scenario to interpret outlying loci that may prove more widespread than anticipated.     

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.     

Repeated adaptive introgression at a gene under multiallelic balancing selection

Recently diverged species typically have incomplete reproductive barriers, allowing introgression of genetic material from one species into the genomic background of the other. The role of natural selection in preventing or promoting introgression remains contentious. Because of genomic co-adaptation, some chromosomal fragments are expected to be selected against in the new background and resist introgression. In contrast, natural selection should favor introgression for alleles at genes evolving under multi-allelic balancing selection, such as the MHC in vertebrates, disease resistance, or self-incompatibility genes in plants. Here, we test the prediction that negative, frequency-dependent selection on alleles at the multi-allelic gene controlling pistil self-incompatibility specificity in two closely related species, Arabidopsis halleri and A. lyrata, caused introgression at this locus at a higher rate than the genomic background. Polymorphism at this gene is largely shared, and we have identified 18 pairs of S-alleles that are only slightly divergent between the two species. For these pairs of S-alleles, divergence at four-fold degenerate sites (K = 0.0193) is about four times lower than the genomic background (K = 0.0743). We demonstrate that this difference cannot be explained by differences in effective population size between the two types of loci. Rather, our data are most consistent with a five-fold increase of introgression rates for S-alleles as compared to the genomic background, making this study the first documented example of adaptive introgression facilitated by balancing selection. We suggest that this process plays an important role in the maintenance of high allelic diversity and divergence at the S-locus in flowering plant families. Because genes under balancing selection are expected to be among the last to stop introgressing, their comparison in closely related species provides a lower-bound estimate of the time since the species stopped forming fertile hybrids, thereby complementing the average portrait of divergence between species provided by genomic data.     

Bidirectional mitochondrial introgression between Korean cobitid fish mediated by hybridogenetic hybrids

Genomic introgression through interspecific hybridization has been observed in some species of the freshwater fish family Cobitidae. Within this family, a Cobitis hankugensis–Iksookimia longicorpa diploid–triploid hybrid species complex on the Korean peninsula is unique in displaying hybridogenesis, a unisexual reproduction mode that allows hybrids to mediate the transfer of mitochondrial DNA (but not nuclear DNA) between the two parent species. However, populations of the parental species in the wild have never been examined for the potential effect of introgression on their genomes. To address the genetic consequences of unisexual hybridization on the parental species, we examined genetic structure of the two parental species, C. hankugensis and I. longicorpa, in three independent natural habitats where they coexist with their hybrid complex using DNA sequence data of one mitochondrial gene and three nuclear genes. We found that mitochondrial introgression between the two species was extensive in all the examined localities, while there was no evidence of nuclear introgression across the species boundary. This result indicates that the hybridogenetic individuals mediate mitochondrial introgression from one species to the other, producing mito‐nuclear mosaic genomes such as C. hankugensis nuclear genomes associated with I. longicorpa mitochondrial DNA and the reverse. The direction and degree of introgression varied among the three localities, but the underlying mechanisms for this observation proved elusive. Introgression might depend on which species serves as the predominant sperm or ovum donor or the environmental conditions of the localities. The present study suggests that introgressive hybridization between pure C. hankugensis and I. longicorpa species is highly likely where the two species co‐occur with hybridogenetic individuals, but the consequence of introgression could be variable due to the history and environmental characteristics of particular populations across the parental species’ ranges.     

Adaptive introgression of the beta-globin cluster in two Andean waterfowl

Introgression of beneficial alleles has emerged as an important avenue for genetic adaptation in both plant and animal populations. In vertebrates, adaptation to hypoxic high-altitude environments involves the coordination of multiple molecular and cellular mechanisms, including selection on the hypoxia-inducible factor (HIF) pathway and the blood-O₂ transport protein hemoglobin (Hb). In two Andean duck species, a striking DNA sequence similarity reflecting identity by descent is present across the ~20 kb β-globin cluster including both embryonic (HBE) and adult (HBB) paralogs, though it was yet untested whether this is due to independent parallel evolution or adaptive introgression. In this study, we find that identical amino acid substitutions in the β-globin cluster that increase Hb-O₂ affinity have likely resulted from historical interbreeding between high-altitude populations of two different distantly-related species. We examined the direction of introgression and discovered that the species with a deeper mtDNA divergence that colonized high altitude earlier in history (Anas flavirostris) transferred adaptive genetic variation to the species with a shallower divergence (A. georgica) that likely colonized high altitude more recently possibly following a range shift into a novel environment. As a consequence, the species that received these β-globin variants through hybridization might have adapted to hypoxic conditions in the high-altitude environment more quickly through acquiring beneficial alleles from the standing, hybrid-origin variation, leading to faster evolution.Subject terms: Evolutionary genetics, Evolutionary biology     

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.     

Integrating Bayesian genomic cline analyses and association mapping of morphological and ecological traits to dissect reproductive isolation and introgression in a Louisiana Iris hybrid zone

Hybrid zones provide unique opportunities to examine reproductive isolation and introgression in nature. We utilized 45,384 single nucleotide polymorphism (SNP) loci to perform association mapping of 14 floral, vegetative and ecological traits that differ between Iris hexagona and Iris fulva, and to investigate, using a Bayesian genomic cline (BGC) framework, patterns of genomic introgression in a large and phenotypically diverse hybrid zone in southern Louisiana. Many loci of small effect size were consistently found to be associated with phenotypic variation across all traits, and several individual loci were revealed to influence phenotypic variation across multiple traits. Patterns of genomic introgression were quite heterogeneous throughout the Louisiana Iris genome, with I. hexagona alleles tending to be favoured over those of I. fulva. Loci that were found to have exceptional patterns of introgression were also found to be significantly associated with phenotypic variation in a small number of morphological traits. However, this was the exception rather than the rule, as most loci that were associated with morphological trait variation were not significantly associated with excess ancestry. These findings provide insights into the complexity of the genomic architecture of phenotypic differences and are a first step towards identifying loci that are associated with both trait variation and reproductive isolation in nature.     

Tracking the progression of speciation: variable patterns of introgression across the genome provide insights on the species delimitation between progenitor–derivative spruces (Picea mariana × P. rubens)

The genic species concept implies that while most of the genome can be exchanged somewhat freely between species through introgression, some genomic regions remain impermeable to interspecific gene flow. Hence, interspecific differences can be maintained despite ongoing gene exchange within contact zones. This study assessed the heterogeneous patterns of introgression at gene loci across the hybrid zone of an incipient progenitor–derivative species pair, Picea mariana (black spruce) and Picea rubens (red spruce). The spruce taxa likely diverged in geographic isolation during the Pleistocene and came into secondary contact during late Holocene. A total of 300 SNPs distributed across the 12 linkage groups (LG) of black spruce were genotyped for 385 individual trees from 33 populations distributed across the allopatric zone of each species and within the zone of sympatry. An integrative framework combining three population genomic approaches was used to scan the genomes, revealing heterogeneous patterns of introgression. A total of 23 SNPs scattered over 10 LG were considered impermeable to introgression and putatively under diverging selection. These loci revealed the existence of impermeable genomic regions forming the species boundary and are thus indicative of ongoing speciation between these two genetic lineages. Another 238 SNPs reflected selectively neutral diffusion across the porous species barrier. Finally, 39 highly permeable SNPs suggested ancestral polymorphism along with balancing selection. The heterogeneous patterns of introgression across the genome indicated that the speciation process between black spruce and red spruce is young and incomplete, albeit some interspecific differences are maintained, allowing ongoing species divergence even in sympatry. The approach developed in this study can be used to track the progression of ongoing speciation processes.     

The genetic legacy of the 19th‐century decline of the British polecat: evidence for extensive introgression from feral ferrets

In the 19th century, the British polecat suffered a demographic contraction, as a consequence of direct persecution, reaching its lowest population in the years that preceded the First World War. The polecat is now recovering and expanding throughout Britain, but introgressive hybridization with feral ferrets has been reported, which could be masking the true range of the polecat and introducing domestic genes into the species. We used a fragment of the mitochondrial DNA control region and 11 microsatellite loci to characterize the frequency and extent of hybridization and introgression between the two species and assess whether the 19th‐century decline corresponded to a genetic bottleneck in the polecat. The proportion of admixture detected in the wild was high (31%) and hybrids were more frequently found outside Wales, suggesting that hybridization is more likely to occur along the eastern edge of the polecat's range expansion. The patterns observed in the mitochondrial and nuclear DNA data show that introgression was mediated by crosses between male polecats and female ferrets, whose offspring backcrossed with polecats. No first‐generation (F₁) hybrids were identified, and the broad range of observed admixture proportions agrees with a scenario of past extensive hybridization between the two species. Using several different methods to investigate demographic history, we did not find consistent evidence for a genetic bottleneck in the British polecat, a result that could be interpreted as a consequence of hybridization with ferrets. Our results highlight the importance of the Welsh polecat population for the conservation and restoration of the genetic identity of the British polecat.     

Bayesian analysis of hybridization and introgression between the endangered european mink (Mustela lutreola) and the polecat (Mustela putorius)

Human-mediated global change will probably increase the rates of natural hybridization and genetic introgression between closely related species, and this will have major implications for conservation of the taxa involved. In this study, we analyse both mitochondrial and nuclear data to characterize ongoing hybridization and genetic introgression between two sympatric sister species of mustelids, the endangered European mink (Mustela lutreola) and the more abundant polecat (M. putorius). A total of 317 European mink, 114 polecats and 15 putative hybrid individuals were collected from different localities in Europe and genotyped with 13 microsatellite nuclear markers. Recently developed Bayesian methods for assigning individuals to populations and identifying admixture proportions were applied to the genetic data. To identify the direction of hybridization, we additionally sequenced mtDNA and Y chromosomes from 78 individuals and 29 males respectively. We found that both hybridization and genetic introgression occurred at low levels (3% and 0.9% respectively) and indicated that hybridization is asymmetric, as only pure polecat males mate with pure European mink females. Furthermore, backcrossing and genetic introgression was detected only from female first-generation (F1) hybrids of European mink to polecats. This latter result implies that Haldane's rule may apply. Our results suggest that hybridization and genetic introgression between the two species should be considered a rather uncommon event. However, the current low densities of European mink might be changing this trend.     

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.     

Next-generation hybridization and introgression

Hybridization has a major role in evolution-from the introgression of important phenotypic traits between species, to the creation of new species through hybrid speciation. Molecular studies of hybridization aim to understand the class of hybrids and the frequency of introgression, detect the signature of ancient hybridization, and understand the behaviour of introgressed loci in their new genomic background. This often involves a large investment in the design and application of molecular markers, leading to a compromise between the depth and breadth of genomic data. New techniques designed to assay a large sub-section of the genome, in association with next-generation sequencing (NGS) technologies, will allow genome-wide hybridization and introgression studies in organisms with no prior sequence data. These detailed genotypic data will unite the breadth of sampling of loci characteristic of population genetics with the depth of sequence information associated with molecular phylogenetics. In this review, we assess the theoretical and methodological constraints that limit our understanding of natural hybridization, and promote the use of NGS for detecting hybridization and introgression between non-model organisms. We also make recommendations for the ways in which emerging techniques, such as pooled barcoded amplicon sequencing and restriction site-associated DNA tags, should be used to overcome current limitations, and enhance our understanding of this evolutionary significant process.     

Maintaining their genetic distance: Little evidence for introgression between widely hybridizing species of Geum with contrasting mating systems

Within the plant kingdom, many genera contain sister lineages with contrasting outcrossing and inbreeding mating systems that are known to hybridize. The evolutionary fate of these sister lineages is likely to be influenced by the extent to which they exchange genes. We measured gene flow between outcrossing Geum rivale and selfing Geum urbanum, sister species that hybridize in contemporary populations. We generated and used a draft genome of G. urbanum to develop dd‐RAD data scorable in both species. Coalescent analysis of RAD data from allopatric populations indicated that the species diverged 2–3 Mya, and that historical gene flow between them was extremely low (1 migrant every 25 generations). Comparison of genetic divergence between species in sympatry and allopatry, together with an analysis of allele frequencies in potential parental and hybrid populations, provided no evidence of contemporary introgression in sympatric populations. Cluster‐ and species‐specific marker analyses revealed that, apart from four early‐generation hybrids, individuals in sympatric populations fell into two genetically distinct groups that corresponded exactly to their morphological species classification with maximum individual admixture estimates of only 1–3%. However, we did observe joint segregation of four putatively introgressed SNPs across two scaffolds in the G. urbanum population that was associated with significant morphological variation, interpreted as tentative evidence for rare, recent interspecific gene flow. Overall, our results indicate that despite the presence of hybrids in contemporary populations, genetic exchange between G. rivale and G. urbanum has been extremely limited throughout their evolutionary history.