Epitope mapping porcine reproductive and respiratory syndrome virus by phage display: the nsp2 fragment of the replicase polyprotein contains a cluster of B-cell epitopes

We screened phage display libraries of porcine reproductive and respiratory syndrome virus (PRRSV) protein fragments with sera from experimentally inf ected pigs to identify linear B-cell epitopes that are commonly recognized during infection in vivo. We identified 10 linear epitope sites (ES) 11 to 53 amino acids in length. In the replicase polyprotein, a total of eight ES were identified, six of which localized to the Nsp2 replicase polyprotein processing end product, In the structural proteins, a total of two ES were identified, in the ORF3 and ORF4 minor envelope glycoproteins, The ORF4 ES was previously identified by monoclonal antibody mapping (J. J, M. Meulenbe rg, A. P. van Nieuwstadt, A, van Essen-Zandenbergen, and J, P, M, Langeveld , J. Virol, 71:6061-6067, 1997), but its immunogenicity had not been examin ed in pigs, We found that six experimentally PRRSV-infected pigs consistent ly had very high antibody titers against the ORF4 ES, In some animals, sera diluted 1:62,500 still gave weak positive enzyme immunoassay reactivity ag ainst the ORF4 ES, This hitherto unrecognized immunodominance likely caused phages displaying the ORF4 ES to outcompete phages displaying other ES dur ing library screening with porcine sera and accounted for our failure to id entify more than two ES in the structural genes of PRRSV, Genetic analysis showed that variable ES were also the most immunogenic in vivo. Serological analysis indicated differences in the immunoglobulin A responses between s hort-term and longer-term viremic pigs towards some ES, The implications of these findings for PRRSV diagnostics and immunopathogenesis are discussed.