Development of an epitope-based marker system for use in categorizing porcine reproductive and respiratory syndrome viruses

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Yang, Liuzhan
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Kenneth B. Platt
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Veterinary Microbiology and Preventive Medicine
Our faculty promote the understanding of causes of infectious disease in animals and the mechanisms by which diseases develop at the organismal, cellular and molecular levels. Veterinary microbiology also includes research on the interaction of pathogenic and symbiotic microbes with their hosts and the host response to infection.
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A marker system based on the presence or absence epitopes of the N, M, GP5 and GP3 proteins of porcine reproductive and respiratory syndrome (PRRS) virus was developed for use in epidemiological studies. Murine monoclonal antibodies (MAbs) were generated that represented: 7 continuous and 3 discontinuous epitopes of the N protein and were designated EpORF7-A through Hd; 3 discontinuous epitopes of the M protein designated EpORF6-Ad, Bd and Cd; 3 continuous epitopes of GP5 designated EpORF5-A, B and C; and 2 continuous epitopes of GP3 designated EpORF3-A and B. Monoclonal antibodies representing EpORF5-C and EpORF6-Ad and Bd had neutralizing activity. Epitope profiles were determined for 69 North American (NA) PRRS viruses and the Lelystad virus using a panel of MAbs that represented the 18 epitopes. The 69 NA viruses were categorized first into 5 major antigenic groups designated 115 through IV 15 and VI 15 based on the N protein. These 5 groups represented 82.6, 11.6, 2.9, 1.4 and 1.4% of all NA viruses respectively. The European Lelystad virus constituted a separate antigenic group designated V15. Groups 115, 1115, and 11115 were subsequently categorized into 9, 4 and 2 subgroups based on seroreactivity with MAbs representing specific epitopes of the M, GP3 and GP5 proteins. Groups IV15 and V15 each represented a single virus. The stability of each of the above 18 epitopes was evaluated for stability by serially passaging and maintaining 5 NA isolates in pigs in 3 separate experiments for periods ranging from 16 to 22 weeks. No differences were observed in the epitope profile of any of these 5 viruses following passage. Preliminary studies were also undertaken to identify the region of ORF 7 that encode specific epitopes. Analysis of the amino acid sequence of the N protein using the GCG package and blocking assays suggested that EpORF7-B, C, and D are located close to one another between position 70 and 122 and that EpORF7-A and E are spatially distant from each other and EpORF7-B, C and D.

Sat Jan 01 00:00:00 UTC 2000