Investigation of selected determinants of Culicoides vectorial capacity of African horse sickness at Onderstepoort, South Africa

Culicoides biting midges (Diptera: Ceratopogonidae) are vectors of a variety of pathogens including African horse sickness virus (AHSV), a member of the Orbivirus genus in the Reoviridae family. AHSV causes African horse sickness (AHS), a disease of equids endemic in sub-Saharan Africa with an extremely high mortality rate. Culicoides (Avaritia) imicola Kieffer is considered to be the principal vector for AHSV and is the dominant Culicoides species in South Africa. Due to the global distribution of the vector species, the disease is at risk of spreading outside its traditional boundaries, which could have a severe economical impact on the equine industry. As part of the risk assessment it is essential to monitor known vectors as well as potential vector species. The present study compared two trapping methods for Culicoides midges. The conventional Onderstepoort light trap that was operated overnight was compared to mechanical aspiration from bait horses at sunset. Culicoides imicola was confirmed as the predominant species by both trapping methods. Other species, mainly Culicoides (Avaritia) bolitinos Meiswinkel and Culicoides (Avaritia) gulbenkiani Caeiro, were highly underrepresented in the light trap collections, but made a significant contribution to the mechanical aspiration catches. The time for optimal collection also differed between both trapping devices, leading to the conclusion that mechanical aspiration is a useful addition to conventional light trap collection and possibly the better choice when investigating insect vectors. Real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) of collected Culicoides midges showed an infection rate of 1.14%, exceeding previous estimates. Virus was present in midge pools collected from the light trap as well as the mechanical aspiration. Seven of the positive pools consisted only of C. imicola, four contained mixed species and one pool contained no C. imicola, suggesting the presence of another vector species. In a separate part of the study, field-collected C. imicola were fed with AHSV-positive blood. Individual midges were dissected and real-time RT-qPCR conducted on heads/thoraxes and abdomens immediately after feeding and after 10 days of incubation. While the majority of Culicoides were AHSV-positive directly after feeding (95.7%), virus was still present in 51% of the midges after 10 days of incubation. Significantly more midges were AHSV-positive in the abdomen compared to heads/thoraxes, indicating that C. imicola – like other Culicoides species – express a so-called mesenteronal escape barrier (MEB), i. e. virus replicates, but is unable to disseminate from the midgut of the midge. The mean amount of virus in the midges increased after 10 days of incubation, most likely due to viral replication in the abdomen. Replication in the salivary glands could not be shown. The RT-qPCR assay proved useful for investigation of midge pools as well as individual Culicoides midges.