Location: Arthropod-Borne Animal Diseases Research
2013 Annual Report
Sub-objective A. Develop means to detect and characterize emergent viruses.
Objective 2: Assess the risk of North American ruminants to emergent, re-emergent, and previously uncharacterized bluetongue virus isolates.
Sub-objective A. Develop BTV “vector-transmitted” infectious models in target ruminant species to facilitate disease pathogenesis, disease transmission and vaccine efficacy studies.
Sub-objective B. Identify mammalian host innate and adaptive responses to insect transmitted BTV.
The bluetongue virus type 17 (BTV-17) core protein (VP7) genome sequence has been used to design X-TAG primers for polymerase chain reaction (PCR) amplification and oligonucleotide capture for an orbivirus fluorescent X-TAG microsphere assay.
The BTV-17 and epizootic hemorrhagic disease virus type 1 (EHDV-1) non-structural protein 1 (NS1) genome sequences have been used to design X-TAG primers for PCR amplification and oligonucleotide capture for a BTV/EHDV differential fluorescent X-TAG microsphere assay.
The outer capsid (VP2) gene sequences of BTV types 2, 10, 11, 13, and 17 have been used to design X-TAG primers for PCR amplification and oligo capture for a BTV serotype specific fluorescent X-TAG microsphere assay.
The VP7 and NS1 genes of BTV-17 were cloned into a cloning vector (pCR2) and an expression vector (pET).
The VP2 genes of BTV-2, 10, 11, 13, and 17 were cloned into a cloning vector (pCR2) and an expression vector (pET).
The VP7 and NS1 genes of EHDV were cloned into a cloning vector (pCR2).
Relative to Objective 2 (Develop BTV “vector-transmitted” infectious models in target ruminant species to facilitate disease pathogenesis, disease transmission and vaccine efficacy studies) the following progress was made:
A needle-free inoculation system was tested on sheep carcasses and live sheep with various settings to achieve optimal transdermal inoculum delivery. Injections of blue dye in the carcasses were used to evaluate depth and consistency of delivery.
Biting midge (Culicoides sonorensis) salivary proteins were collected on membranes, purified and concentrated. Proteins were identified by mass spectrometry and compared to known salivary proteins of other arthropods. Mice were injected intradermally with salivary proteins. At one, two and three days post inoculation, mice were euthanized. Blood, lymph nodes and injection site skin were harvested for analysis by flow cytometry, CBC chemistry panel, and histology. Specific mouse cell populations related to immune responses were up-regulated by the salivary proteins.
Irie, T., Liu, Y., Drolet, B.S., Carnero, E., Garcia-Sastre, A., Harty, R.N. 2012. Cytopathogenesis of Vesicular Stomatitis virus is regulated by the PSAP motif of M protein in a species-dependent manner. Viruses. 4: 1605-1618.
Mcvey, D.S., Wilson, W.C., Drolet, B.S. 2013. Reoviridae. McVey, D.S., Kennedy, M., Chengappa, M.M., Editors. Veterinary Microbiology, 3rd Edition. New Jersey: Wiley-Blackwell. p. 491-500.
Ruder, M.G., Howerth, E.W., Stallknecht, D.E., Allison, A.B., Carter, D.L., Drolet, B.S., Klement, E., Mead, D.G. 2012. Vector competence of Culicoides sonorensis for epizootic hemorrhagic disease virus serotype 7. Parasites & Vectors. 5:248.