The Balkan Peninsula is recognized as one of the hotspots of biodiversity in Europe. This area has shown since the Last Glacial Maximum appropriate conditions for species diversification and hybridization, which has led to the existence of numerous taxonomically unresolved entities. Here, we focus on the Western Balkans and explore the genetic structure and relationships among species belonging to the V. austriaca - V. orbiculata diploid-polyploid complex, including populations showing intermediate morphologies. A combination of nuclear markers (microsatellites), plastid DNA regions (trnH-psbA, ycf6-psbM) and ploidy level estimations using flow cytometry are employed to assess the genetic structure and evolutionary dynamics of this polyploid complex. To reconstruct the evolutionary history, an approximate Bayesian computation approach is combined with projections of the species distribution models onto the climatic scenarios of the Mid-Holocene (6 ka BP) and Last Glacial Maximum (22 ka BP). Four main groups were found: one well-established entity within the diploid level, V. dalmatica, a second diploid-tetraploid group which corresponds to V. orbiculata, a hexaploid cluster harboring V. austriaca subsp. jacquinii individuals, and an enigmatic tetraploid group. According to the molecular data obtained, this latter cluster represents an allopolyploid cryptic lineage −with V. orbiculata and V. dalmatica as putative parents− morphologically similar to V. orbiculata, but genetically more related to V. austriaca subsp. jacquinii. Veronica dalmatica and this “uncertain tetraploid” group are involved in the formation of the hexaploid taxon V. austriaca subsp. jacquinii, with the possibility of recent gene flow among different cytotypes. The present study supports a scenario of diversification from a diploid common ancestor leading to two different but interrelated lineages. The first one would correspond with the diploid V. orbiculata plus tetraploid individuals of this species arising through allo- and autopolyploidization, and the second one would involve all ploidy levels with allopolyploidization being prevalent.
Keywords: Allopolyploidy; Autopolyploidy; Balkan Peninsula; Last Glacial Maximum; Microsatellites; Veronica.