Dynamics of epizootic hemorrhagic disease virus infection within the vector, Culicoides sonorensis (Diptera: Ceratopogonidae)

Abstract

Culicoides sonorensis biting midges are confirmed vectors of epizootic hemorrhagic disease virus (EHDV), which causes mortality in white-tailed deer and ruminant populations. Currently, of the seven EHDV serotypes, only 1, 2, and 6 are detected in the USA, and very few studies have focused on the infection time course of these serotypes within the midge. The objective of this current research was to characterize EHDV-2 infection within the midge by measuring infection prevalence, virus dissemination, and viral load over the course of infection. Midges were fed a blood meal containing 106.9 PFU/ml EHDV-2, collected every 12 h from 0–2 days post feeding (dpf) and daily from 3–10 dpf, and cohorts of 20 C. sonorensis were processed using techniques that assessed EHDV infection and dissemination. Cytopathic effect assays and quantitative (q)PCR were used to determine infection prevalence, revealing a 50% infection rate by 10 dpf using both methods. Using immunohistochemistry, EHDV-2 infection was detectable at 5 dpf, and shown to disseminate from the midgut to other tissues, including fat body, eyes, and salivary glands by 5 dpf. Stain intensity increased from 5–8 dpf, indicating replication of EHDV-2 in secondary infection sites after dissemination. This finding is also supported by trends in viral load over time as determined by plaque assays and qPCR. An increase in titer between 4–5 dpf correlated with viral replication in the midgut as seen with staining at day 5, while the subsequent gradual increase in viral load from 8–10 dpf suggested viral replication in midges with disseminated infection. Overall, the data presented herein suggest that EHDV-2 disseminates via the hemolymph to secondary infection sites throughout the midge and demonstrate a high potential for transmission at five days at 25°C after an infective blood-meal.

Evaluation of A Baculovirus-Expressed VP2 Subunit Vaccine for the Protection of White-Tailed Deer (Odocoileus virginianus) from Epizootic Hemorrhagic Disease

“…EHDV-2 Alberta (SV-124-Canada 1962; ABADRU's virus collection) and EHDV-6 Indiana (12-38993; provided by SCWDS) were used for VP2 cloning and virus neutralization tests. EHDV-2 strain cc12-304, prepared from the spleen of an infected white-tailed deer from Kansas in 2012 [28], was kindly provided by Mark Ruder, SCWDS, propagated in a Culicoides sonorensis cell line (CuVa W8 cells), and used for the WTD challenge [29].…”

Analysis of Conserved, Computationally Predicted Epitope Regions for VP5 and VP7 Across three Orbiviruses

“… 4 – 6 Both BTV and EHDV infect wild and domestic ruminants, causing varying severity of disease depending on different species. 7 – 11 Epizootic hemorrhagic disease virus affects white-tailed deer the most severely. 12 Bluetongue virus is symptomatically seen most often in sheep.…”

Continuous Cell Lines from the European Biting Midge Culicoides nubeculosus (Meigen, 1830)

“…However, our results suggest that the situation within C. nubeculosus may be more complex if the apparent low ability of CNE/LULS44 cells in vitro to release mature BTV-1 particles reflects the conditions in intact midges. Despite broad assessments of susceptibility to infection and studies of barriers to arbovirus dissemination in Culicoides [ 52 , 53 , 54 ], BTV infection and release at the cellular level remains poorly explored. Further studies are needed to determine whether our results with BTV-1 are broadly representative of multiple BTV serotypes, whether the phenotypes represented in the CNE/LULS44 cell line include midgut cells (potentially limiting or blocking virion release), and whether manipulation of the culture conditions (temperature, pH, osmotic pressure) could facilitate the release of virus particles.…”