Molecular epidemiology of epizootic hemorrhagic disease viruses in the United States
Murphy, Molly Danielle
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The epizootic hemorrhagic disease viruses (EHDV), the causative agents of epizootic hemorrhagic disease in wild and domestic ruminants, are endemic to the southeastern United States, with outbreaks of severe clinical disease occurring in white-tailed deer periodically. Outbreaks occur during late summer and early fall, concordant with the seasonal abundance pattern of Culicoides midges, some species of which have been demonstrated to be competent biological vectors of EHDV. A member of the Orbivirus genus in the family Reoviridae, the EHDV genome is comprised of ten double-stranded RNA fragments, encoding three non-structural and seven structural proteins. The high error rate of the RNA polymerase, in concert with the potential for segment reassortment, suggests that a high amount of genetic variation in EHDV is possible. The contributing factors to genetic variation among EHDV isolates are currently unknown. The objectives of this study were to (1) determine the association of time and geographic space on genetic variation among field isolates of EHDV-2 from the southeastern U.S., (2) characterize the viruses present during a 2001 outbreak of EHDV-2 -related hemorrhagic disease in the western U.S., and (3) characterize the EHDV-1 viruses present during a 1999 oubreak in the eastern U.S. To this end, field isolates of EHDV-1 and -2 were sequenced at two loci; the S10 locus, which encodes the viral egress protein NS3, and the L2 locus, which encodes the surface epitope protein L2. The genetic relationships among the sequences were determined through phylogenetic analyses. The results of the studies addressing the first two objectives demonstrated that the EHDV-2 viruses appear to be evolving very slowly and underwent very little nucleotide change during intra-epizootic periods. Additionally, these studies indicated that as of 1972, the EHDV-2 viruses likely existed in geographically distinct western and eastern populations, whereas between 1990 and 2001, they appeared to exist as a single population spanning both regions. The EHDV-1 isolates collected during the 1999 epizootic also demonstrated very little genetic variation; many of the isolates were identical at one or both loci. However, in contrast to EHDV-2 isolates collected between 1990-2001, EHDV-1 isolates collected between 1999-2001 appeared to exist in distinct eastern and western populations. Additionally, among four EHDV-1 isolates collected from an enzootic focus in 2001, none were identical at the S10 locus and only two were identical at the L2 locus. The enzootic isolates demonstrated a higher level of genetic variability than that which was described for the 1999 eastern epizootic isolates. This indicates that EHDV-1 viruses existing in enzootic and epizootic cycles may be evolving at different rates, or via different mechanisms.