Isolation of Porcine Reproductive and Respiratory Syndrome Virus GP5-Specific, Neutralizing Monoclonal Antibodies From Hyperimmune Sows

Abstract

Porcine reproductive and respiratory syndrome (PRRS) is a devastating disease which impacts the pig industry worldwide. The disease is caused by PRRS viruses (PRRSV-1 and -2) which leads to abortions and other forms of reproductive failure in sows and severe respiratory disease in growing pigs. Current PRRSV vaccines provide limited protection; only providing complete protection against closely related strains. The development of improved PRRSV vaccines would benefit from an increased understanding of epitopes relevant to protection, including those recognized by antibodies which possess the ability to neutralize distantly related strains. In this work, a reverse vaccinology approach was taken; starting first with pigs known to have a broadly neutralizing antibody response and then investigating the responsible B cells/antibodies through the isolation of PRRSV neutralizing monoclonal antibodies (mAbs). PBMCs were harvested from pigs sequentially exposed to a modified-live PRRSV-2 vaccine as well as divergent PRRSV-2 field isolates. Memory B cells were immortalized and a total of 5 PRRSV-specific B-cell populations were isolated. All identified PRRSV-specific antibodies were found to be broadly binding to all PRRSV-2 isolates tested, but not PRRSV-1 isolates. Antibodies against GP5 protein, commonly thought to possess a dominant PRRSV neutralizing epitope, were found to be highly abundant, as four out of five B cells populations were GP5 specific. One of the GP5-specific mAbs was shown to be neutralizing but this was only observed against homologous and not heterologous PRRSV strains. Further investigation of these antibodies, and others, may lead to the elucidation of conserved neutralizing epitopes that can be exploited for improved vaccine design and lays the groundwork for the study of broadly neutralizing antibodies against other porcine pathogens.

Reappraisal of PRRS Immune Control Strategies: The Way Forward

“…The issue of target proteins of the neutralizing antibody (NA) and relevant neutralizing epitopes has stimulated several studies. Thus, neutralizing GP5-specific monoclonal Abs have been isolated from the sera of hyperimmune sows [ 65 ], confirming the presence of neutralizing epitopes on GP5 glycoprotein [ 66 , 67 ]. Neutralizing epitopes have also been observed on the minor surface glycoproteins GP2, GP3, and GP4 [ 68 ].…”

“…Neutralizing epitopes have also been observed on the minor surface glycoproteins GP2, GP3, and GP4 [ 68 ]. The isolation of specific monoclonal Abs against these proteins would likely uncover additional neutralizing or potentially broadly neutralizing antibodies [ 65 ]. It has been observed that the nsp2 protein also contributes to the neutralizing activity of the structural proteins GP5-M in vitro, suggesting that there is at least one neutralizing epitope in nsp2, or in the spatial structure formed by nsp2 and the structural proteins together [ 69 ].…”

Reappraisal of PRRS Control Strategies: The Way forward

“…The issue of target proteins of neutralizing antibody (NA) and relevant neutralizing epitopes has stimulated several studies. Thus, neutralizing GP5-specific monoclonal Abs have been isolated from sera of hyperimmune sows [64], confirming the presence of neutralizing epitopes on GP5 glycoprotein [65,66] . Neutralizing epitopes has been observed also on the minor surface glycoproteins GP2, GP3, and GP4 [67] .…”

“…Neutralizing epitopes has been observed also on the minor surface glycoproteins GP2, GP3, and GP4 [67] . The isolation of specific monoclonal Abs against these proteins will likely uncover additional neutralizing or potentially broadly neutralizing antibodies [64] . It has been observed that also the nsp2 protein contributes to the neutralizing activity of the structural proteins GP5-M in vitro, suggesting that there is at least one neutralizing epitope in nsp2 or in the spatial structure formed by nsp2 and structural proteins together [68] .…”

Combined Subcutaneous-Intranasal Immunization With Epitope-Based Antigens Elicits Binding and Neutralizing Antibody Responses in Serum and Mucosae Against PRRSV-2 and SARS-CoV-2

“…Concerning PRRSV-2, difficulties include its high mutational rate, the immunomodulatory disease provoked, the virus persistence in secondary lymphoid tissues, and the documented reversion to virulence of live-attenuated virus vaccine strains ( 33 – 35 ). Additionally, glycosylation of PRRSV-2 proteins and the presence of immunodominant hypervariable decoy epitopes, as occurs in the GP5 protein, results in humoral immune responses that do not confer protection against heterologous strains ( 36 , 37 ). Concerning SARS-CoV-2, its high transmissibility and widespread infection in the human population have led to the emergence of variants of concern (VOCs) ( 38 ).…”