Haemosporidian Parasite Infections and Host‐Feeding Preferences of Hematophagous Insects Collected at the Birds of Prey Nests with Confirmation of Vectors for <i>Haemoproteus</i> spp. and <i>Leucocytozoon</i> sp.

Avian haemosporidian parasites, such as Haemoproteus, Leucocytozoon, and Plasmodium are transmitted by blood-sucking insects and can negatively affect their host. These parasites have specific life cycles that allow them to survive and spread efficiently through their vectors. However, the research on vectors and their feeding preferences are scarce. Therefore, in this study, we investigated haemosporidian parasite vectors and their feeding preferences in insects collected at birds of prey nests. To catch the vectors, UV-light traps were hung at the nests of birds of prey between June and July 2020 and 2021. Parous and nulliparous biting midge females were separated. Parous females were dissected for salivary gland preparations, and their remnants were stored in 96% EtOH for PCR-based analysis. We screened parous females for the presence of parasites amplifying a fragment of the cytochrome b gene of haemosporidian parasites using specific primers (Plas1F/HaemNR3 and 3760F/HaemJR4). To identify the feeding preference, we analyzed 73 engorged biting midge females using vertebrate specific primers L14841/H15149. In total, we dissected and analyzed 1262 parous biting midges (Culicoides spp.) and three blackfly (Simulium spp.) females. We found that Simulium vernum is a natural vector of Leucocytozoon sp. and identified Haemoproteus spp. sporozoites in Culicoides kibunensis, Culicoides pictipennis, and Culicoides segnis. DNA of three birds and two mammals was detected in engorged biting midges. In conclusion, our study provides valuable insights into the identification of natural vectors of avian haemosporidian parasites and their feeding preferences, contributing to a better understanding of the ecological dynamics that influence parasite transmission.