Background and Aims Southwestern Asia is a significant centre of biodiversity and a cradle of diversification for many plant groups, especially xerophytic elements. In contrast, little is known about the evolution and diversification of hygrophytic flora. To fill this gap, we focus on Cardamine (Brassicaceae) species that grow in wetlands over a wide altitudinal range. We aimed to elucidate their evolution, assess the extent of presumed historical gene flow between species, and draw inferences about intraspecific structure. Methods We applied the phylogenomic Hyb-Seq approach, ecological niche analyses, and multivariate morphometrics to a total of 85 Cardamine populations from the target region of Anatolia-Caucasus, usually treated as four to six species, and supplemented them with close relatives from Europe. Key Results Five diploids are recognised in the focus area, three of which occur in regions adjacent to the Black and/or Caspian Seas (C. penzesii, C. tenera, C. lazica), one species widely distributed from the Caucasus to Lebanon and Iran (C. uliginosa), and one western Anatolian entity (provisionally as C. cf. uliginosa). Phylogenomic data suggest recent speciation during the Pleistocene, likely driven by both geographical separation (allopatry) and ecological divergence. With the exception of a single hybrid (allotetraploid) speciation event proven for C. wiedemanniana, an endemic of southern Turkey, no significant traces of past or present interspecific gene flow were observed. Genetic variation within the studied species is spatially structured, suggesting reduced gene flow due to geographical and ecological barriers, but also glacial survival in different refugia. Conclusions This study highlights the importance of the refugial regions of the Black and Caspian Seas for both harbouring and generating hygrophytic species diversity in SW Asia. It also supports the significance of evolutionary links between Anatolia and the Balkan Peninsula. Reticulation and polyploidisation played a minor evolutionary role here in contrast to the European relatives.

Mountains of the Balkan Peninsula are significant biodiversity hotspots with great species richness and a large proportion of narrow endemics. Processes that have driven the evolution of the rich Balkan mountain flora, however, are still insufficiently explored and understood. Here we focus on a group of Cardamine (Brassicaceae) perennials growing in wet, mainly mountainous habitats. It comprises several Mediterranean endemics, including those restricted to the Balkan Peninsula. We used target enrichment with genome skimming (Hyb-Seq) to infer their phylogenetic relationships, and, along with genomic in situ hybridization (GISH), to resolve the origin of tetraploid Cardamine barbaraeoides endemic to the Southern Pindos Mts. (Greece). We also explored the challenges of phylogenomic analyses of polyploid species and developed a new approach of allele sorting into homeologs that allows identifying subgenomes inherited from different progenitors. We obtained a robust phylogenetic reconstruction for diploids based on 1,168 low-copy nuclear genes, which suggested both allopatric and ecological speciation events. In addition, cases of plastid–nuclear discordance, in agreement with divergent nuclear ribosomal DNA (nrDNA) copy variants in some species, indicated traces of interspecific gene flow. Our results also support biogeographic links between the Balkan and Anatolian–Caucasus regions and illustrate the contribution of the latter region to high Balkan biodiversity. An allopolyploid origin was inferred for C. barbaraeoides, which highlights the role of mountains in the Balkan Peninsula both as refugia and melting pots favoring species contacts and polyploid evolution in response to Pleistocene climate-induced range dynamics. Overall, our study demonstrates the importance of a thorough phylogenomic approach when studying the evolution of recently diverged species complexes affected by reticulation events at both diploid and polyploid levels. We emphasize the significance of retrieving allelic and homeologous variation from nuclear genes, as well as multiple nrDNA copy variants from genome skim data.