Characterizing the processes underlying reproductive isolation between diverging lineages is central to understanding speciation. Here, we present RIDGE-Reproductive Isolation Detection using Genomic polymorphisms-a tool tailored for quantifying gene flow barrier proportion and identifying the relevant genomic regions. RIDGE relies on an Approximate Bayesian Computation with a model-averaging approach to accommodate diverse scenarios of lineage divergence. It captures heterogeneity in effective migration rate along the genome while accounting for variation in linked selection and recombination. The barrier detection test relies on numerous summary statistics to compute a Bayes factor, offering a robust statistical framework that facilitates cross-species comparisons. Simulations revealed RIDGE\'s efficiency in capturing signals of ongoing migration. Model averaging proved particularly valuable in scenarios of high model uncertainty where no migration or migration homogeneity can be wrongly assumed, typically for recent divergence times <0.1 2Ne generations. Applying RIDGE to four published crow data sets, we first validated our tool by identifying a well-known large genomic region associated with mate choice patterns. Second, while we identified a significant overlap of outlier loci using RIDGE and traditional genomic scans, our results suggest that a substantial portion of previously identified outliers are likely false positives. Outlier detection relies on allele differentiation, relative measures of divergence and the count of shared polymorphisms and fixed differences. Our analyses also highlight the value of incorporating multiple summary statistics including our newly developed outlier ones that can be useful in challenging detection conditions.

Delimiting and naming biodiversity is a vital step toward wildlife conservation and research. However, species delimitation must be consistent across biota so that the limited resources available for nature protection can be spent effectively and objectively. To date, newly discovered lineages typically are either left undescribed and thus remain unprotected or are being erroneously proposed as new species despite mixed evidence for completed speciation, in turn contributing to the emerging problem of taxonomic inflation. Inspired by recent conceptual and methodological progress, we propose a standardized workflow for species delimitation that combines phylogenetic and hybrid zone analyses of genomic datasets (\"genomic taxonomy\"), in which phylogeographic lineages that do not freely admix are ranked as species, while those that have remained fully genetically compatible are ranked as subspecies. In both cases, we encourage their formal taxonomic naming, diagnosis, and description to promote social awareness toward biodiversity. The use of loci throughout the genome overcomes the unreliability of widely used barcoding genes when phylogeographic patterns are complex, while the evaluation of divergence and reproductive isolation unifies the long-opposed concepts of lineage species and biological species. We suggest that a shift in conservation assessments from a single level (species) toward a two-level hierarchy (species and subspecies) will lead to a more balanced perception of biodiversity in which both intraspecific and interspecific diversity are valued and more adequately protected.

Sex chromosomes are popularized as a special role in driving speciation. However, the empirical evidence from natural population processes has been limited to organisms with degenerated sex chromosomes, where hemizygosity is mainly considered to act as the driver of reproductive isolation. Here, we examined several hybrid zones of torrent frog Amolops mantzorum species complex, using an approach by mapping species-diagnostic loci onto the reference genome to compare sex-linked versus autosomal patterns of introgression. We find little support in sex-linked incompatibilities for large X-effects for these populations in hybrid zones with homomorphic sex chromosomes, due to the absence of the hemizygous effects. As expected, the large X-effects were not found in those with heteromorphic but newly evolved sex chromosomes, owing to the absence of strong genetic differences between X and Y chromosomes. The available data so far on amphibians suggest little role for sex-linked genes in speciation. The large X-effects in those with nascent sex chromosomes may not be as ubiquitous as presumed across the animal kingdom.

Evaluating whether hybrid zones are stable or mobile can provide novel insights for evolution and conservation biology. Butterflies exhibit high sensitivity to environmental changes and represent an important model system for the study of hybrid zone origins and maintenance. Here, we review the literature exploring butterfly hybrid zones, with a special focus on their spatiotemporal dynamics and the potential mechanisms that could lead to their movement or stability. We then compare different lines of evidence used to investigate hybrid zone dynamics and discuss the strengths and weaknesses of each approach. Our goal with this review is to reveal general conditions associated with the stability or mobility of butterfly hybrid zones by synthesizing evidence obtained using different types of data sampled across multiple regions and spatial scales. Finally, we discuss spatiotemporal dynamics in the context of a speciation/divergence continuum, the relevance of hybrid zones for conservation biology, and recommend key topics for future investigation.

BACKGROUND: Microsatellite markers were developed for distylous Linum suffruticosum and tested in the monomorphic sister species Linum tenuifolium. These species are perennial herbs endemic to the western and northwestern Mediterranean, respectively, with a partially overlapping distribution area.
RESULTS: We developed 12 microsatellite markers for L. suffruticosum using next generation sequencing, and assessed their polymorphism and genetic diversity in 152 individuals from seven natural populations. The markers displayed high polymorphism, with two to 16 alleles per locus and population, and average observed and expected heterozygosities of 0.833 and 0.692, respectively. All loci amplified successfully in the sister species L. tenuifolium, and 150 individuals from seven populations were also screened. The polymorphism exhibited was high, with two to ten alleles per locus and population, and average observed and expected heterozygosities of 0.77 and 0.62, respectively.
CONCLUSIONS: The microsatellite markers identified in L. suffruticosum and tested in L. tenuifolium are a powerful tool to facilitate future investigations of the population genetics, mating patterns and hybridization between both Linum species in their contact zone.

Understanding the genetic targets of natural selection is one of the most challenging goals of population genetics. Some of the earliest candidate genes were identified from associations between allozyme allele frequencies and environmental variation. One such example is the clinal polymorphism in the arginine kinase (Ak) gene in the marine snail Littorina fabalis. While other enzyme loci do not show differences in allozyme frequencies among populations, the Ak alleles are near differential fixation across repeated wave exposure gradients in Europe. Here, we use this case to illustrate how a new sequencing toolbox can be employed to characterize the genomic architecture associated with historical candidate genes. We found that the Ak alleles differ by nine nonsynonymous substitutions, which perfectly explain the different migration patterns of the allozymes during electrophoresis. Moreover, by exploring the genomic context of the Ak gene, we found that the three main Ak alleles are located on different arrangements of a putative chromosomal inversion that reaches near fixation at the opposing ends of two transects covering a wave exposure gradient. This shows Ak is part of a large (3/4 of the chromosome) genomic block of differentiation, in which Ak is unlikely to be the only target of divergent selection. Nevertheless, the nonsynonymous substitutions among Ak alleles and the complete association of one allele with one inversion arrangement suggest that the Ak gene is a strong candidate to contribute to the adaptive significance of the inversion.

How new species evolve is one of the most fundamental questions in biology. Population divergence, which may lead to speciation, may be occurring in the Eastern Yellow Robin, a common passerine that lives along the eastern coast of Australia. This species is composed of 2 parapatric lineages that have highly divergent mitochondrial DNA; however, similar levels of divergence have not been observed in the nuclear genome. Here we re-examine the nuclear genomes of these mitolineages to test potential mechanisms underlying the discordance between nuclear and mitochondrial divergence. We find that nuclear admixture occurs in a narrow hybrid zone, although the majority of markers across the genome show evidence of reproductive isolation between populations of opposing mitolineages. There is an 8 MB section of a previously identified putative neo-sex chromosome that is highly diverged between allopatric but not parapatric populations, which may be the result of a chromosomal inversion. The neo-sex chromosomal nature of this region, as well as the geographic patterns in which it exhibits divergence, suggest it is unlikely to be contributing to reproductive isolation through mitonuclear incompatibilities as reported in earlier studies. In addition, there are sex differences in the number of markers that are differentiated between populations of opposite mitolineages, with greater differentiation occurring in females, which are heterozygous, than males. These results suggest that, despite the absence of previously observed assortative mating, mitolineages of Eastern Yellow Robin experience at least some postzygotic isolation from each other, in a pattern consistent with Haldane\'s Rule.

Natural hybrid zones have provided important insights into the evolutionary process, and their geographic dynamics over time can help to disentangle the underlying biological processes that maintain them. Here, we leverage replicated sampling of an identical transect across the hybrid zone between yellow-shafted and red-shafted flickers in the Great Plains to assess its stability over ∼60 years (1955-1957 to 2016-2018). We identify a ∼73-km westward shift in the hybrid zone center toward the range of the red-shafted flicker, but find no associated changes in width over our sampling period. In fact, the hybrid zone remains remarkably narrow, suggesting some kind of selective pressure maintains the zone. By comparing to previous work in the same geographic region, it appears likely that the movement in the hybrid zone has occurred in the years since the early 1980s. This recent movement may be related to changes in climate or land management practices that have allowed westward movement of yellow-shafted flickers into the Great Plains.

Dusky Salamanders (genus Desmognathus) currently comprise only 22 described, extant species. However, recent mitochondrial and nuclear estimates indicate the presence of up to 49 candidate species based on ecogeographic sampling. Previous studies also suggest a complex history of hybridization between these lineages. Studies in other groups suggest that disregarding admixture may affect both phylogenetic inference and clustering-based species delimitation. With a dataset comprising 233 Anchored Hybrid Enrichment (AHE) loci sequenced for 896 Desmognathus specimens from all 49 candidate species, we test three hypotheses regarding (i) species-level diversity, (ii) hybridization and admixture, and (iii) misleading phylogenetic inference. Using phylogenetic and population-clustering analyses considering gene flow, we find support for at least 47 candidate species in the phylogenomic dataset, some of which are newly characterized here while others represent combinations of previously named lineages that are collapsed in the current dataset. Within these, we observe significant phylogeographic structure, with up to 64 total geographic genetic lineages, many of which hybridize either narrowly at contact zones or extensively across ecological gradients. We find strong support for both recent admixture between terminal lineages and ancient hybridization across internal branches. This signal appears to distort concatenated phylogenetic inference, wherein more heavily admixed terminal specimens occupy apparently artifactual early-diverging topological positions, occasionally to the extent of forming false clades of intermediate hybrids. Additional geographic and genetic sampling and more robust computational approaches will be needed to clarify taxonomy, and to reconstruct a network topology to display evolutionary relationships in a manner that is consistent with their complex history of reticulation.

Under incomplete reproductive isolation, secondary contact of diverged allopatric lineages may lead to the formation of hybrid zones that allow to study recombinants over several generations as excellent systems of genomic interactions resulting from the evolutionary forces acting on certain genes and phenotypes. Hybrid phenotypes may either exhibit intermediacy or, alternatively, transgressive traits, which exceed the extremes of their parents due to epistasis and segregation of complementary alleles. While transgressive morphotypes have been examined in fish, reptiles, birds, and mammals, studies in amphibians are rare. Here, we associate microsatellite-based genotypes with morphometrics-based morphotypes of two tree frog species of the Hyla arborea group, sampled across a hybrid zone in Poland, to understand whether the genetically differentiated parental species also differ in morphology between each other and their hybrids and whether secondary contact leads to the evolution of intermediate or transgressive morphotypes. Using univariate approaches, explorative multivariate methods (principal component analyses) as well as techniques with prior grouping (discriminant function analyses), we find that morphotypes of both parental species and hybrids differ from each other. Importantly, hybrid morphotypes are neither intermediate nor transgressive but found to be more similar to H. orientalis than to H. arborea.