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United States Department of Agriculture

Agricultural Research Service

Research Project: SWINE VIRAL DISEASES PATHOGENESIS AND IMMUNOLOGY Title: Genetic stability of PRRSV VR-2332 nsp2 deletion mutants in swine

Authors
item Guo, Baoqing - IOWA STATE UNIVERSITY
item Faaberg, Kay
item Lager, Kelly
item Kehrli Jr, Marcus

Submitted to: American Society for Virology Meeting
Publication Type: Abstract Only
Publication Acceptance Date: June 18, 2009
Publication Date: July 11, 2009
Citation: Guo, B., Faaberg, K.S., Lager, K.M., Kehrli, Jr., M.E. 2009. Genetic stability of PRRSV VR-2332 nsp2 deletion mutants in swine [abstract]. 28th Annual Meeting of the American Society for Virology, Vancouver, Canada. Paper No. W33-12.

Technical Abstract: PRRSV nonstructural protein 2 (nsp2) is the largest putative cleavage product of this arterivirus replicase polyprotein, and is composed of a cysteine protease (PL2) domain at the N-terminus, a middle hypervariable region and four transmembrane domains near the C-terminus. Previous studies demonstrated that up to 403 amino acids could be deleted from the hypervariable region of nsp2 without loss of viral replication in cell culture (Han et al., J. Virol, 2007). Four PRRSV nsp2 deletion mutants along with wild-type (wt) recombinant virus were next evaluated in swine. Serum samples (up to day 35) were evaluated for acquisition of antibody, presence of interferon gamma (IFN-gamma, an antiviral that activates monocyte cytotoxicity and enhances NK cell activity). For this presentation, viral growth kinetics and nsp2 sequence analysis were also completed. Three mutant viruses were shown to replicate at a rate 10-1000 fold less than the wt virus. No viral titer was obtained from the mutant with the largest deletion, but qRT-PCR and ELISA results suggested some replication had occurred. The nsp2 genes of each mutant and wt virus at day 35 were amplified by RT-PCR, cloned and sequenced. The amino acid sequence analysis of the deletion mutants and wt virus indicated that the nsp2 genes of the deletion mutants were genetically stable. Virus obtained from serum from swine inoculated with the largest deletion mutant was further passaged in swine to substantiate viral replication. This deletion mutant showed an increased capacity to replicate, and the nsp2 sequence was again shown to be stable. Since the deleted nsp2 sequences contained likely antigenic epitopes, the variable effects of the individual nsp2 deletions on the immune responses were evaluated and will be presented. In addition, whole genome sequence analysis of the mutant viruses recovered from swine may identify compensatory effects for virus genome stability.

Last Modified: 10/22/2014
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