West Nile virus (WNV) is a serious human pathogen with a mortality rate of 0.1%. The National Institute forAllergy and Infectious Disease (NIAID) Biodefense Research Agenda classifies WNV as a Category BPriority Pathogen on the basis of its transmission by an arthropod host, its infectivity on aerosolizedexposure, and its potential threat to public health. Of particular concern is the possibility that West Nile virusmight be used as an agent of bioterrorism. There is currently no vaccine for WNV available for use inhumans. Thus, development of a WNV vaccine is a national biodefense and public health priority that isconsistent with the overall goals of NIH and MBRS. The goals of this Pilot Research Project are todevelop a candidate vaccine for West Nile virus and to test its immunogenicity in mice. This proposalincorporates two novel concepts: the use of Nodamura virus (NoV) RNA replicons to amplify WNV mRNAs inthe yeast Saccharomyces cerevisiae and inoculation of animals with purified total yeast RNA containingamplified NoV-WNV RNAs. The advantages of the nodavirus expression system as a source of vaccinematerial include exponential amplification of the chimeric mRNA; its ability to initiate RNA replication in awide variety of host cells, including those from mammals, insects, plants, and yeast, when the RNA isintroduced by transfection; and the lack of pathogenicity of NoV for humans. Yeast cells provide a safe andinexpensive source of vaccine material, and are currently used to produce Hepatitis B vaccines.
The specific aims of this proposal are:1. Construct NoV RNA2-based replicons that contain WNV structural (glycoprotein E, gpE) or nonstructural(NS1) proteins and determine the extent to which RNA replication amplifies NoV2-gpE and NoV-NS1mRNA and protein levels in yeast and mammalian cells2. Evaluate the WNV structural (gpE) and nonstructural (NS1) proteins as potential vaccine candidates bydefining the humoral and cell-mediated immune responses to these proteins elicited in inoculated mice.Relevance to public health: West Nile virus is a severe human pathogen that poses a threat to the publichealth and biosafety of the United States. Our goal is to develop safe, effective candidate vaccines that willprevent the spread of West Nile virus-associated diseases.
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