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Research Project: Development of Deoptimized Live Attenuated Foot-Mouth-Disease Virus Vaccine Candidates

Location: Foreign Animal Disease Research

Project Number: 3022-32000-064-003-I
Project Type: Interagency Reimbursable Agreement

Start Date: Aug 9, 2018
End Date: Sep 30, 2022

The main goal of this project is the development of weakened Foot-and-Mouth Disease Virus (FMDV) vaccine candidates that are partially impaired for growing in the animal host. These weakened viruses will be constructed by using recently developed techniques in which the viral genomic coding sequences are changed without altering the overall architecture of the virus. These viruses grow in the animals and induce immunity without causing or transmitting disease to other animals. Specific objectives include: 1. Construction of FMDV strains containing deoptimized genome coding regions P1, P2, and/or P3 sequences with DIVA markers in FMDV A24 infectious clone backbone. 2. Conduct in vitro characterization of deoptimized FMDV strains. 3. Conduct in vivo testing of mutant virus 1 in swine. 4. Conduct in vivo testing of mutant 2 virus 2 in swine.

1. Previously designed sequences will be used to generate new deoptimized FMDV strains by reverse genetics. Infectious clones will be transcribed to generate RNA followed by electroporation in baby hamster kidney (BHK)-21 cells to derive infectious virus. Viruses will be isolated, propagated and sequenced to confirm genetic stability. High titer stocks of at least 2 mutant viruses will be produced for further in vitro and in vivo characterization. 2. Strains derived in Objective 1 will be characterized for their ability to grow in multiple cell lines in comparison to wild type A24 FMDV. Two recombinant viruses showing acceptable replication rates in vaccine producing cell lines, and a clear attenuated phenotype in primary cells will be selected for further characterization in vivo in swine. 3. The degree of attenuation and the determination of minimal protective dose of mutant 1 will be evaluated in swine. After challenge all animals will be monitored for additional 21 days and evaluated for the presence of viral RNA, virus, cytokines, and neutralizing antibodies in serum as well as for cell mediated immunity. All animals will be assayed pre and post challenge for the presence of antibodies against nonstructural proteins to confirm DIVA features on live vaccine candidates.