Monoclonal antibodies to the GP5 of porcine reproductive and respiratory syndrome virus are more effective in virus neutralization than monoclonal antibodies to the GP4

The arterivirus porcine reproductive and respiratory syndrome virus (PRRSV) contains six structural proteins the roles of which are not completely understood. In a preceding study, immunization with the dutch isolate I10 of PRRSV had led to the development of MAbs against four structural proteins [Wieczorek-Krohmer, M., 1994. Herstellung und Charakterisierung von monoklonalen Antikörpern gegen das Virus des Porzinen Reproduktiven und Respiratorischen Syndroms (PRRSV). Inaugural-Dissertation, Ludwig-Maximilians-Universität, München] here finally identified by reaction with individual plasmid-expressed PRRSV proteins as products of ORFs 3 (GP₃), 4 (GP₄), 5 (GP₅) and 7 (N). Surprisingly, the MAbs against GP₅ revealed the presence of two antigenically distinct virus populations in the isolate I10, the population PRRSV-`PPV', isolated from plaques and the PRRSV-`EPV', gained by end point dilution. MAbs against GP₃, GP₄ and N reacted with both I10 populations as well as with natural PRRSV isolates. However, the anti-GP₅ MAbs exclusively recognized PRRSV-`PPV'. In this study immunization of mice with both separated I10 populations confirmed that solely PRRSV-`PPV' possesses the property to induce an immune response ultimately leading to the establishment of MAbs against GP₅. Whereas the 15 anti-GP₅ MAbs (derived from four independent fusions) reacted exclusively with PRRSV-`PPV' of the isolate I10, anti-GP<sub>4</sub> MAbs detected their target antigen on various isolates of European origin and were able to neutralize them. As indicated by competition assays and selection of neutralization-resistant virus mutants, all GP<sub>5</sub> MAbs are directed against a single antigenic site on the ORF 5 protein. Both groups of neutralizing antibodies bound to the surface of purified virions demonstrating that the recognized epitopes represent surface structures of the virion envelope. However, anti-GP<sub>5</sub> MAbs mediated the binding of more gold granules than anti-GP<sub>4</sub> MAbs. Comparison of the neutralizing effect of anti-GP<sub>4</sub> and anti-GP<sub>5</sub> MAbs revealed the anti-GP<sub>5</sub> MAbs as the more efficient antibodies. For the complete neutralization of about 100 ID<sub>50</sub> of PRRSV-`PPV' anti-GP₅ culture supernatant was effective up to a dilution of 1 : 1280 whereas the most effective anti-GP₄ antibodies exhibited a comparable effect only up to 1 : 64. These results indicate that PRRSV GP₅ in principle is a major target for neutralizing antibodies, as is found for other arteriviruses, but that in nature `ORF 5 escape mutants' may develop as easily as in vitro.