New data on the distribution of the Vipera ammodytes (Linnaeus, 1758) mitochondrial lineages place their contact zone in western Bulgaria

Molecular studies have outlined several mitochondrial lineages of Vipera ammodytes, but the limits of their distribution ranges remain unclear due to limited sampling throughout the vast range of the species. One particularly understudied area is the Eastern Balkan Peninsula where at least three lineages occur, showing ranges that may be partly overlapping. We used two mitochondrial markers (cytb and ND2) to unveil mtDNA distribution patterns based on vipers from 31 localities across Bulgaria. Our results confirmed the presence of the north-eastern (NE) and the south-eastern (SE) mitochondrial clade in Bulgaria, the latter being represented by its southern (S) and eastern (E) subclades. Among the sampled localities, two were suspected to be potential contact zones between these mtDNA lineages based on old morphology-derived distributional records. The NE clade was absent from both potential contact zones. However, our data showed that in western Bulgaria, populations of this clade establish contact with populations of the E subclade in at least one area, and also come close to contact with populations of the S subclade. These results indicate the need for more detailed research in the potential areas of contact in western Bulgaria, integrating morphological data with extensive mitochondrial and nuclear DNA-sampling to better understand the phylogeographic patterns of intraspecific differentiation in V. ammodytes.