Increased genetic structuring of isolated Salamandra salamandra populations (Caudata: Salamandridae) at the margins of the Carpathian Mountains

The traditional southern Pleistocene refugia hypothesis in Europe has lately been challenged for several animal and plant species, and importance of more northern areas has been pointed out. The Carpathian Basin is one of the recently recognized biodiversity hotspots in Europe, representing rich fauna especially at the marginal regions. Marginal populations are prone to have lower genetic diversity and higher genetic differentiation than central populations. Here we examined one mitochondrial DNA fragment and nine nuclear (microsatellite) loci to describe the genetic diversity and phylogeographical pattern of fire salamander ( Salamandra salamandra ) populations in the Carpathian Basin with focusing on the southern margins of the Western Carpathians, where the species is present with isolated populations. Analyses of microsatellites indicated reduced genetic diversity for most of the isolated populations. While the analyses of the mitochondrial DNA showed the homogeneity of Carpathian Basin populations, the analyses with the nuclear markers revealed a more recent genetic split between western (Alpine) and eastern (Carpathian) populations, and separated the Apuseni Mountains population. Using Approximate Bayesian Computation analyses we identified the most probable colonization scenario for the isolated North Hungarian populations. Split between isolated salamander populations from the central populations in the Carpathian Mountains dates back to the beginning of the Late Pleistocene, while split between most of the isolated Hungarian populations can be associated with the Last Glacial Maximum. We found evidence for long-time isolation between the marginal Carpathian Basin and central fire salamander populations. Our results also proved the glacial survival of S. salamandra in the temperate forests of northeast Pannonia (North Hungarian Mountains), confirming that the Carpathian Basin served as important northerly refugia during the Pleistocene climatic oscillations.