Comparative Phylogeography of and (Plantaginaceae) Across Europe: Integrating Hybridization and Polyploidy in Phylogeography.

Climatic fluctuations in the Pleistocene caused glacial expansion-contraction cycles in Eurasia and other parts of the world. Consequences of these cycles, such as population expansion and subsequent subdivision, have been studied in many taxa at intraspecific population level across much of the Northern Hemisphere. However, the consequences for the potential of hybridization and polyploidization are poorly understood. Here, we investigated the phylogeographic structure of two widespread, closely related species, and , across their European distribution ranges. We assessed the extent and the geographic pattern of polyploidization in both species and hybridization between them. We used genome-scale SNP data to clarify phylogenetic relationships and detect possible hybridization/introgression events. In addition, crossing experiments were performed in different combination between and individuals of two ploidy levels and of different geographic origins. Finally, we employed ecological niche modeling to infer macroclimatic differences between both species and both ploidy levels. We found a clear genetic structure reflecting the geographical distribution patterns in both species, with showing higher genetic differentiation than . . We retrieved significant signals of hybridization and introgression in natural populations from the genetic data and corroborated this with crossing experiments. However, there were no clear phylogeographic patterns and unequivocal macroclimatic niche differences between diploid and tetraploid lineages. This favors the hypothesis, that autopolyploidization has happened frequently and in different regions. The crossing experiments produced viable hybrids when the crosses were made between plants of the same ploidy levels but not in the interploidy crosses. The results suggest that hybridization occurs across the overlapping areas of natural distribution ranges of both species, with apparently directional introgression from to . Nevertheless, the two species maintain their species-level separation due to their adaptation to different habitats and spatial isolation rather than reproductive isolation.