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United States Department of Agriculture

Agricultural Research Service


Location: Arthropod-borne Animal Diseases Research

2012 Annual Report

1a. Objectives (from AD-416):
Objective 1: Identify biological determinants of disease susceptibility associated with arboviral infections. Subobjective 1A. Assess the role of insect salivary proteins on the pathogenesis of bluetongue virus in relevant target vertebrate hosts. Subobjective 1B. Assess the role of insect salivary proteins on the pathogenesis of vesicular stomatitis virus in relevant target vertebrate hosts. Subobjective 1C. Identify and characterize the vertebrate host receptors for bluetongue virus. Subobjective 1D. Assess vesicular stomatitis virus-induced physiological variations and determine their affect on vector-host selection. Objective 2: Determine the host-range specificity of exotic bluetongue viruses. Subobjective 2A: Determine the susceptibility of U.S livestock to exotic bluetongue virus. Subobjective 2B: Determine the susceptibility of U.S wildlife to exotic bluetongue virus.

1b. Approach (from AD-416):
Arthropod-borne diseases pose significant concerns to the U.S. livestock industry. This project will investigate several biological relationships among host, vector and virus that will lead to improved disease control and risk assessment of emerging and re-emerging, domestic and exotic arboviruses. Biological determinants of arthropod-borne viral diseases of animals can be associated with the insect vector, the arbovirus, or the animal. One potential insect determinant is insect saliva, which may affect arbovirus transmission and subsequent infection. Culicoides sonorensis saliva will be used to examine interactions between saliva, arboviruses, and the immune response of susceptible animals. This may help to identify ways to interrupt disease transmission. A second important arboviral-animal determinant is virus attachment, mediated by cellular receptor(s) and allowing subsequent infection. Candidate receptor molecules for bluetongue virus (BTV) will be identified from sub-cellular fractions. This will provide a better understanding of BTV pathogenesis and may lead to more targeted vaccine strategies. The final biological determinant to be addressed is the effect of virus infection on host selection by insect vectors. The effect of vesicular stomatitis virus (VSV) infection on insect feeding and host defensive behaviors that could affect virus acquisition and transmission will be examined. This may help in the design and implementation of more efficient and cost-effective bite transmission control strategies. Introduction of exotic arboviruses is an ongoing risk and reality. The susceptibility of North American sheep and white-tailed deer to BTV-8, which is causing devastating disease in Europe, will be determined to provide valuable information for risk assessment.

3. Progress Report:
This is the final report for 5430-32000-002-00D, which terminated in FY2012 and was replaced by 5430-32000-006-00D. Relative to Objective 1 (Identify biological determinants of disease associated with arboviral infections) the following progress was made: To determine any differences in salivary proteins ABADRU's three Culicoides colonies, saliva was collected from over 600 midges from each colony, pooled and analyzed by electrophoresis. There appear to be no discernable differences between the three colonies. Protein sequencing analysis of Culicoides saliva showed at least 42 discernible components. The most abundant of these include homologs of maltase and late trypsin, as well as members of a family of D7 odorant binding proteins, and several kunitz-type protease inhibitor related proteins. Cloning and expression of abundant Culicoides salivary proteins has begun. This will facilitate functional studies of the individual proteins. Culicoides salivary proteins were shown to down regulate antiviral immune responses (IFN-gamma) of mouse fibroblast cells. This suggests that when virus is delivered into the skin of an animal by insect bite, the saliva may enable the virus to establish infection locally by immunosuppressing the host response to the virus. Culicoides salivary proteins were shown to increase mouse immune cell populations by 2-8 fold in regional lymph nodes demonstrating the immunogenicity of these proteins. Genetic diagnostic assays for emerging and re-emerging insect transmitted viruses affecting livestock and wildlife were developed. This includes: BTV, epizootic hemorrhagic disease virus (EHDV), and vesicular stomatitis virus (VSV). Several exotic serotypes have been isolated in Florida from samples submitted to the National Veterinary Services Laboratory for export purposes. One serotype, has been isolated multiple times. The ABADRU has a collection of virus strains from Central America and the Caribbean region including some of this serotype. These exotic strains related to the ones isolated in Florida are being propagated for viral RNA isolation. The viral RNA is then submitted to whole genome amplification and sequenced. This information will be analyzed using bioinformatics to test the hypothesis that the introduced virus serotype came from this southern regions perhaps during hurricane or from ship travel. Relative to Objective 2 (Determine the host–range specificity of exotic bluetongue viruses) the following progress was made: Final analysis of tissue and blood samples from BTV-8 infected white-tailed deer was completed and a manuscript is in preparation. Comparative sequence analysis between the original BTV-8 inoculum from The Netherlands and virus isolated from infected deer showed no significant genomic changes. North American sheep were inoculated with BTV-8 from The Netherlands to determine their susceptibility to this exotic BTV type. See accomplishment section below.

4. Accomplishments
1. Infection of North American sheep by the European Bluetongue serotype 8 (BTV-8) virus. To determine the risk of US sheep to BTV-8, ARS researchers at Manhattan, KS inoculated sheep with BTV-8 obtained from The Netherlands. The disease level observed was similar to domestic BTV virus types and less severe than what has been reported for European sheep species. Our results indicate that if BTV-8 were to be introduced into the U.S., our sheep populations would be highly susceptible to infection, but clinical disease would not be more severe than outbreaks resulting from many of our domestic BTV types.

Review Publications
Bowen, J., Mecham, J.0., Hamlin, M., Henderson, B., Kim, M., Mirjankar, N., Lavine, B.K. 2011. Development of field-deployable instrumentation based on “antigen–antibody” reactions for detection of hemorrhagic disease in ruminants. Microchemical Journal.

Ruder, M.G., Allison, A.B., Stallknecht, D.E., Mead, D.G., Mcgraw, S.M., Carter, D.L., Kubiski, S.V., Batten, C.A., Klement, E., Howerth, E.W. 2012. Susceptibility of white-tailed deer (Odocoileus virginianus) to experimental infection with epizootic hemorrhagic disease virus serotype 7. Journal of Wildlife Diseases. 48(3):676-685.

Reeves, W.K. 2011. Behavior of larval Culicoides Sonorensis (Diptera: Ceratopogonidae) in response to an invertebrate predator, Hydra Littoralis (Anthomedusae: Hydridae). Entomological News. 121: 298-301.

Last Modified: 06/26/2017
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