Cloning and Expression of the Nucleoprotein of Peste des Petits Ruminants Virus in Baculovirus for Use in Serological Diagnosis

Abstract

Peste des petits ruminants (PPR) is a viral disease of goats and sheep characterized by erosive stomatitis, enteritis, and pneumonia. The virus is a member of the family Paramyxoviridae and the genus Morbillivirus. The disease has high morbidity and mortality rates and has a substantial economic impact in developing countries. We have cloned and sequenced the cDNA of the nucleocapsid (N) gene of the Nigeria 75/1 strain of PPR virus (PPRV). A comparison of its nucleotide and deduced amino acid sequence with those of the N gene of the tissue culture-attenuated strain of PPRV was performed. A divergence of 8.9 and 5.0% was found at the nucleotide and amino acid level, respectively. A recombinant baculovirus that expresses the N protein in insect cells and larvae (Spodoptera frugiperda) was generated. The recombinant protein, characterized by Western blot analysis, was shown to have a molecular weight of 58 kDa and was recognized by anti-PPRV antibodies. The recombinant protein was used successfully as a coating antigen in an enzyme-linked immunosorbent assay for the serological diagnosis of PPRV.

Global distribution of peste des petits ruminants virus and prospects for improved diagnosis and control

“…Currently, the OIE recommend the use of the competitive PPRVspecific anti-H monoclonal based ELISA (cH-ELISA) (Anderson & McKay, 1994) and virus neutralization tests (FAO, 1996). However, several alternatives exist (Choi et al, 2005;Libeau et al, 1995) including the indirect N ELISA (Ismail et al, 1995), immunofiltration (Dhinakar Raj et al, 2008), a novel sandwich ELISA (Saravanan et al, 2008), haemagglutination tests (Dhinakar Raj et al, 2000;Ezeibe et al, 2008) and latex agglutination tests (Keerti et al, 2009).…”

Diagnosis and control of peste des petits ruminants: a comprehensive review

“…Several attempts have been made since long to express various viral proteins in different expression systems viz., bacterial, mammalian, yeast and insect cells and to assess potential use of recombinant proteins in various diagnostic assays. In this direction, Ismail et al [65] cloned and sequenced the cDNA of the N gene of the Nigeria 75/1 strain of PPRV. The recombinant baculovirus expressed N protein in insect cell system was used successfully as a coating antigen in ELISA for serodiagnosis of PPRV.…”

Outbreaks of Peste Des Petits Ruminants (PPR) in Goats in Punjab, India

“…lacrimal, nasal, saliva, feces (Rossiter and Taylor, 1994). The first clinical description of PPR was made in 1942 in West Africa and characterized by necrotic and erosive stomatitis, enteritis and pneumonia (Ismail et al, 1995). In India, first PPR outbreak was recorded in Tamil Nadu in the year1987 (Shaila et al, 1989).…”