Molecular and phylogenetic insights into Polychromophilus diversity in bat flies (Diptera: Nycteribiidae) from...

Haemosporidian parasites of the genus Polychromophilus are blood parasites infecting bats worldwide and transmitted by obligate hematophagous flies of the family Nycteribiidae. Despite increasing molecular data in Europe, their diversity, distribution, and host-vector associations remain poorly documented in parts of Western Europe. This study provides molecular evidence of Polychromophilus lineages in the nycteribiid bat fly Penicillidia conspicua parasitizing Miniopterus schreibersii (Miniopteridae) in Portugal. A total of 60 flies were collected from bats in two caves, and 25% (15/60) tested positive for Polychromophilus spp. based on amplification of the cytochrome b (cytB) gene. High-quality sequences of cytB and clpC genes were obtained and analyzed using Maximum-Likelihood phylogenetic inference. The resulting topologies revealed two well-supported clades corresponding to Polychromophilus melanipherus and Polychromophilus murinus. Our samples clustered within both clades, showing high genetic similarity to African and Asian Miniopterus-associated Polychromophilus lineages and to European and Neotropical Myotis-associated lineages. These findings indicate that multiple Polychromophilus lineages may coexist within the same host-vector system and suggest low geographic structuring across the Old World. The results reinforce the role of nycteribiid flies as a potential vector facilitating parasite dispersal across bat populations and regions. This study provides new and important vector-based molecular data, extending existing host-parasite knowledge to nycteribiid bat flies and strengthening evidence for established transmission cycles in the region, adding new insights into the evolutionary and biogeographic patterns of haemosporidian parasites in Europe.