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Title: Mutations in classical swine fever virus NS4B affect virulence in swine

Author
item FERNANDEZ-SAINZ, IGNACIO - Oak Ridge Institute For Science And Education (ORISE)
item Gladue, Douglas
item Holinka-Patterson, Lauren
item O'DONNELL, VIVIAN - University Of Connecticut
item GUDMUNDSDOTTIR, INGIGERDUR - University Of Connecticut
item Prarat, Melanie
item Patch, Jared
item Golde, William
item LU, ZHIQIANG - Us Deparment Of Homeland Security
item RISATTI, GUILLERMO - University Of Connecticut
item Zhu, James
item Borca, Manuel

Submitted to: Journal of Virology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/2/2009
Publication Date: 2/1/2010
Citation: Fernandez-Sainz, I., Gladue, D.P., Holinka-Patterson, L.G., O'Donnell, V.K., Gudmundsdottir, I., Prarat, M.V., Patch, J.R., Golde, W.T., Lu, Z., Risatti, G.R., Zhu, J.J., Borca, M.V. 2010. Mutations in classical swine fever virus NS4B affect virulence in swine. Journal of Virology. 84(3):1536-1549.

Interpretive Summary: NS4B is one of the non-structural proteins of Classical Swine Fever Virus (CSFV), a virus causing a severe disease in swine. Knowledge of NS4B function is very limited. Genetic comparison of NS4B gene with gene database allowed the discovery of a particular amino acid sequence (called TIR) that is present in Toll-like-receptor (TLR) proteins. TLRs are host cell proteins that play a critical role in the induction of early immune response. CSFV harboring alteration of the TIR sequence in NS4B resulted in complete attenuation in swine. This attenuated virus could, however, behave as an experimental vaccine protecting pigs against the challenge with virulent wild type virus. Additional results indicated that CSFV NS4B is able to down regulate the host production of important cytokines, which regulates early events of the anti-viral immune response, or chemokines, which regulate cell migration during the inflammatory process. This mechanism may facilitate the spread of the virus during the infection in swine.

Technical Abstract: NS4B is one of the non-structural proteins of Classical Swine Fever Virus (CSFV), a virus causing a severe disease in swine. Protein domain analysis of the predicted amino acid sequence of NS4B in highly pathogenic CSFV strain Brescia (BICv) identified a Toll/Interleukin-1 receptor like domain (TIR). This TIR-like motif harbors two conserved domains named box 1 and box 2, also observed in other members of the TIR superfamily, including Toll-like receptors (TLRs). Mutations of NS4B Box 2 domain in BICv, that yield virus NS4B-VGIv, affected transcriptional activation of TLR-7 induced genes in swine macrophages including a sustained increase of IL-6 mRNA accumulation. Transfection of swine macrophages with wild type NS4B protein partially blocked the TLR-7 activating effect of Imiquimod R837. In vivo, NS4B-VGIv was completely attenuated in swine showing a reduced replication in the oronasal cavity and limited spreading from the site of virus entry to secondary target organs. Furthermore, the level and duration of IL-6 production in the tonsils of pigs intranasally inoculated with NS4B/VGIv was significantly higher than in animals infected with BICv. The peak of induction of IL-6 production in infected animals paralleled the ability of NS4B-VGIv infected animals to resist challenge with virulent BICv. Interestingly, treatment of peripheral blood monocuclear cell cultures with swine IL-6 results in significant decrease of BICv replication.