Location: Foreign Animal Disease Research
Project Number: 8064-32000-057-24
Start Date: Jun 01, 2009
End Date: Dec 31, 2013
Objective 1a, host-pathogen interaction of the huAd5 vector will be studied by inoculating cattle with huAd5 vectors containing reporter genes or FMDV capsid and tracking its distribution and transgene expression in target tissues such as muscle and regional lymph nodes. The identification and tracking tissue distribution of AdFMD vaccines will be useful for future design and assessment of AdFMD vaccine with increased potency. Objective 1b. ARS, PIADC will utilize the experimental closed Holstein herd from the University of Vermont, to conduct tissue typing analysis of Class I major histocompatibility complex (MHCI) among herd members using PCR developed in collaboration with the University of Copenhagen, Denmark. Relevant Class I genes corresponding to animals in the experimental herd will be expressed in E.coli for analysis of peptide binding in vitro. These data will be used to predict peptides of FMDV capsids from strains A24, A22, O1 Manisa and O1 Campos. Amendment 3, Objective 1b Enhanced: To enhance the fidelity of data generated, a robust system to check and confirm the predicting algorithm is warranted to generate a peptide library for the viral protein, P1 region of the FMDV. This library will allow for systematic analysis of the bovine immune responses to confirm the predictions fo the computer and identify peptides that the algorithm might miss. These data will be used to improve the functionality of the predictor program, identifying T cell epitopes which induce immune response. Objective 2a, will be accomplished by sequence analysis and determination of antigenic profiles of FMDV and bovine rhinovirus polymerase proteins and the determination of cross-reactivity between them. Identification and expression of FMDV-specific 3D epitopes by recombinant technologies will be conducted. Proof of concept for specific and sensitivity of ELISA tests will be developed. Objective 2b will be accomplished by cloning and expressing bovine IFNA. The expressed IFNA will be tested for antiviral activity. The bIFNA with the highest activity will be identified. Objective 3a, will be accomplished through identification and cloning of all members of the type III IFN gene family of cattle using Ad5-vectors. Examination of antiviral properties against FMDV will be conducted. Dose-response experiments will be conducted in-vitro. A study of IFN stimulated gene induction and specific type III receptor gene expression by real time RT-PCR and/or microarray analysis, will be conducted to examine gene induction after treatment with type III IFN alone or in combination with other Biotherapeutics. 3.b. will be accomplished by conducting vaccine challenge studies on swine and cattle utilizing different doses of polyICLC in combination with the optimal dose of Ad5-IFNa, and a suboptimal dose of AD5-IFNa. The cellular components of the innate immune response including natural killer cells, dendritic cells will be examined. Based on these studies, the optimal amounts of polyICLC and Ad5-IFNa will be administered to swine and cattle and challenged with FMDV. Amend 3: Obj 2a: include comprehnesive bench standardization of competiive ELISA 3D test.