Blood Relatives: Linking Evolutionary History and Conservation of Medicinal Leeches (Hirudo spp.).

Medicinal leeches of the genus Hirudo are both historically important in medicine and increasingly studied as models in ecology and biomedicine. However, the taxonomy, population structure and evolutionary history of this genus remain partially unresolved. Here, we present the most comprehensive phylogenetic analysis of the Hirudo complex to date, incorporating 350 COX1 sequences (103 newly generated) and 247 complete mitogenomes from across the distribution range. In Hirudo verbana, three major clades corresponding to Iberian (A), Western (B) and Eastern (C) haplogroups were found, with sub-haplogroups reflecting glacial refugia and Pleistocene expansions. Diversity was highest in Eastern populations, consistent with Anatolia and the Black Sea basin acting as long-term refugia. Bayesian divergence dating indicates that the genus originated in the mid-Miocene (~15 Mya), with an early split of H. nipponia in East Asia. Subsequent radiations shaped the Western Eurasian clade, with H. sulukii diverging ~9.9 Mya, H. orientalis splitting from European lineages ~6.4 Mya and differentiating ~440 kya, and recent radiations of H. verbana, H. medicinalis and H. troctina occurring during the late Pleistocene. Notably, nucleotide diversity within H. nipponia (~8.8%) and deep divergence among its haplogroups suggest the presence of multiple cryptic species, with H. tianjinensis clustering within these lineages. We also molecularly identified several Italian specimens as Haemopis sanguisuga or a potential new Dina species. These findings emphasise the importance of taxonomic clarity and genetic monitoring to guide conservation and prevent overexploitation of Hirudo leeches, ensuring the long-term sustainability and biomedical utility of these ecologically significant annelids.