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Title: A Full-length cDNA Infectious Clone of North American Type 1 Porcine Reproductive and Respiratory Syndrome Virus: Expression of Green Fluorescent Protein in the Nsp2 Region

Author
item FANG, YING - SO DAKOTA STATE UNIV
item ROWLAND, RAYMOND - KANSAS STATE UNIV
item ROOF, MICHAEL - BOEHRINGER, INC
item Lunney, Joan
item CHRISTOPHER, JANE - SO DAKOTA STATE UNIV
item NELSON, ERIC - SO DAKOTA STATE UNIV

Submitted to: Journal of Virology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/13/2006
Publication Date: 12/1/2006
Citation: Fang, Y., Rowland, R.R., Roof, M., Lunney, J.K., Christopher, J., Nelson, E.A. 2006. A full-length cdna infectious clone of North American type 1 porcine reproductive and respiratory syndrome virus: expression of green fluorescent protein in the nsp2 region. Journal of Virology. 80:11447-11455.

Interpretive Summary: Porcine reproductive and respiratory syndrome (PRRS) is the most economically important pig disease in the United States. The disease was first described in 1987 in the US and in Europe in 1990. The PRRS virus (PRRSV) was first isolated in The Netherlands in 1991, and is represented by the European prototypic strain, Lelystad or genotype (type 1) PRRSV. In the US, the North American genotype, or type 2 PRRSV, was first isolated in 1992. Recently, a US Type 1 PRRSV isolates, termed North American Type 1, was identified in U.S. swineherds. This manuscript describes the characterization of an infectious clone for North American Type 1 PRRSV, SD01-08. In vitro studies showed that cloned virus maintained similar growth properties as that of the parental virus. In vivo studies showed that virological, pathological and immunological observations from animals challenged with cloned viruses were similar to those challenged with the parental virus and a modified live virus vaccine. Thus the infectious clone can now be used as a research tool to analyze type 1 PRRS and to develop improved vaccines for this important pig viral infection. To further explore the potential use of this clone as a viral backbone for expressing foreign genes, the green fluorescent protein (GFP) gene was inserted and proven to be expressed in tissue culture cells. The availability of this North American Type 1 infectious clone provides an important research tool for further study of the basic viral biology and pathogenic mechanisms of this group of Type 1 PRRSV in the US. In conjunction with the North American Type 2 infectious clones, a new generation of genetically engineered PRRSV vaccines could be constructed.

Technical Abstract: The recent emergence of a unique group of North American Type 1 porcine reproductive and respiratory syndrome virus (PRRSV) in the U.S. presents new disease control problems for a swine industry that has already been seriously impacted by North American Type 2 PRRSV. In this study, a full-length cDNA infectious clone was generated from a low virulent North American Type 1 PRRSV isolate, SD01-08. In vitro studies showed that cloned virus maintained similar growth properties as that of the parental virus. Virological, pathological and immunological observations from animals challenged with cloned viruses were similar to those challenged with the parental virus and a modified live virus vaccine. To further explore the potential use as a viral backbone for expressing foreign genes, the green fluorescent protein (GFP) was inserted into a unique deletion site located at amino acid positions 348/349 of the Nsp2 region in the virus, and expression of the Nsp2-GFP fusion protein visualized by fluorescent microscopy. The availability of this North American Type 1 infectious clone provides an important research tool for further study of the basic viral biology and pathogenic mechanisms of this group of Type 1 PRRSV in the US. In conjunction with the North American Type 2 infectious clones, a new generation of genetically engineered PRRSV vaccines could be constructed.