Two new species of Haemoproteus Kruse, 1890 (Haemosporida, Haemoproteidae) from European birds, with emphasis on DNA barcoding for detection of haemosporidians in wildlife

Two new species of Haemoproteus Kruse, 1890 (Haemosporida, Haemoproteidae) are described: Haemoproteus (Parahaemoproteus) homovelans n. sp. from Grey-faced Woodpecker, Picus canus Gmelin, and Haemoproteus (Parahaemoproteus) concavocentralis n. sp. recorded in Hawfinch, Coccothraustes coccothraustes (Linnaeus), both sampled in Bulgaria. The morphology of the gametocytes and their host-cells are described and mitochondrial cytochrome b (cyt b) gene sequences are generated. Haemoproteus homovelans possesses circumnuclear gametocytes lacking volutin granules. This parasite is particularly similar to Haemoproteus velans Coatney & Roudabush, 1937 also possessing circumnuclear gametocytes that are, however, overfilled with volutin. Haemoproteus concavocentralis can be readily distinguished from all described avian haemoproteids due to the presence of an unfilled concave space between the central part of advanced gametocytes and erythrocyte nucleus. Bayesian phylogenetic analyses of 40 haemosporidian cyt b lineages showed close relationships of H. concavocentralis (hHAWF2) with a group of Haemoproteus spp. possessing gametocytes that are pale-stained with Giemsa. The lineage hPICAN02 of H. homovelans clustered with parasites infecting non-passerine birds. Phylogenetic analyses support the current subgeneric classification of the avian haemoproteids and suggest that cyt b lineage hPIPUB01 (GenBank EU254552) has been incorrectly assigned to Haemoproteus picae Coatney & Roudabush, 1937, a common parasite of corvid birds (Passeriformes). This study emphasises the importance of combining molecular techniques and light microscopy in the identification and field studies of avian haemosporidian parasites. Future development of barcodes for molecular identification of haemoproteids will allow better diagnostics of these infections, particularly in veterinary studies addressing insufficiently investigated tissue pathology caused by these parasites.