Cryptic diversity, polyploidy, and intraspecific divergence in <i>Erysimum</i> (Brassicaceae) species endemic to the western Balkans

BACKGROUND AND AIMS: Several evolutionary drivers have been proposed to explain the exceptional species and genetic diversity of the Balkan flora, including the moderate impact of glaciations enabling long-term in situ persistence, ecological and topographic heterogeneity promoting vicariant differentiation, and polyploidy. Despite these shared drivers, plant lineages often show contrasting evolutionary histories, suggesting lineage-specific rather than broadly generalizable responses. The genus Erysimum, notable for its high endemism, karyological variation, recent and rapid speciation, and signatures of reticulate evolution, provides an excellent model for exploring this diversity of evolutionary trajectories and speciation drivers in the Balkans. METHODS: Focusing on five Balkan endemics of the E. odoratum species complex sampled from 47 populations, we examined chromosomal and genome size variation and applied two complementary genomic approaches, target enrichment (Hyb-Seq) and RADseq, along with morphometric analyses. These datasets were used to circumscribe species, uncover intraspecific variation patterns, and assess the extent and evolutionary consequences of interspecific gene flow and polyploidy. KEY RESULTS: Genetic and morphological evidence was largely congruent, clarifying species boundaries but also revealing cryptic diversity, leading to the recognition of eight entities. Most diversity was concentrated in the southern Dinarides, where we detected multiple cytotypes, an undescribed lineage, a polyphyletic species resulting from two independent autopolyploidization events, and allotetraploids arising in a contact zone. Geographic distance and biogeographic barriers across the mountainous Balkan landscape likely shaped patterns of divergence and promoted speciation. Discrepancies between chromosome numbers and genome size indicate dynamic chromosomal evolution. CONCLUSIONS: Vicariant differentiation, auto- and allopolyploidy, genome size and chromosomal dynamics together account for the pronounced diversity of Erysimum in the Balkans, illustrating how speciation is driven within this biodiversity hotspot. Importantly, substantial evolutionary potential is retained within species, as the coexistence of multiple intraspecific genetic lineages and cytotypes can reinforce reproductive barriers and promote future diversification.