Identity and diversity of blood meal hosts of biting midges (Diptera: Ceratopogonidae: Culicoides Latreille) in Denmark

Background: Host preference studies in haematophagous insects e. g. Culicoides biting midges are pivotal to assess transmission routes of vector-borne diseases and critical for the development of veterinary contingency plans to identify which species should be included due to their risk potential. Species of Culicoides have been found in almost all parts of the world and known to live in a variety of habitats. Several parasites and viruses are transmitted by Culicoides biting midges including Bluetongue virus and Schmallenberg virus. The aim of the present study was to determine the identity and diversity of blood meals taken from vertebrate hosts in wild-caught Culicoides biting midges near livestock farms. Methods: Biting midges were collected at weekly intervals for 20 weeks from May to October 2009 using light traps at four collection sites on the island Sealand, Denmark. Blood-fed female biting midges were sorted and head and wings were removed for morphological species identification. The thoraxes and abdomens including the blood meals of the individual females were subsequently subjected to DNA isolation. The molecular marker cytochrome oxidase I (COI barcode) was applied to identify the species of the collected biting midges (GenBank accessions JQ683259-JQ683374). The blood meals were first screened with a species-specific cytochrome b primer pair for cow and if negative with a universal cytochrome b primer pair followed by sequencing to identify mammal or avian blood meal hosts. Results: Twenty-four species of biting midges were identified from the four study sites. A total of 111,356 Culicoides biting midges were collected, of which 2,164 were blood-fed. Specimens of twenty species were identified with blood in their abdomens. Blood meal sources were successfully identified by DNA sequencing from 242 (76%) out of 320 Culicoides specimens. Eight species of mammals and seven species of birds were identified as blood meal hosts. The most common host species was the cow, which constituted 77% of the identified blood meals. The second most numerous host species was the common wood pigeon, which constituted 6% of the identified blood meals. Conclusions: Our results suggest that some Culicoides species are opportunistic and readily feed on a variety of mammals and birds, while others seems to be strictly mammalophilic or ornithophilic. Based on their number, dispersal potential and blood feeding behaviour, we conclude that Culicoides biting midges are potential vectors for many pathogens not yet introduced to Denmark.