Characterization of Broadly Neutralizing Antibodies to Porcine Reproductive and Respiratory Syndrome Virus (PRRSV)
2020-01
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Characterization of Broadly Neutralizing Antibodies to Porcine Reproductive and Respiratory Syndrome Virus (PRRSV)
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2020-01
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Porcine reproductive and respiratory syndrome (PRRS) is an all too common disease with a devastating impact on pork producers in the US and worldwide. The disease is caused by the PRRS virus (PRRSV) which leads to abortions and other forms of reproductive failure in sows and severe respiratory disease in adults and neonates. Unfortunately, current vaccines against PRRSV provide limited protection and often only protects against closely related viruses. Efficacious vaccine-based prevention of infectious disease is based on antigen-specific long-lived memory B and T cells. Protective, or neutralizing, antibodies produced by memory B cells, that are activated in response to virulent pathogen challenge, are critical to this process. Thus, it is of utmost importance to investigate antibodies with neutralizing potential especially broadly neutralizing antibodies (bnAbs), which possess the ability to neutralize distantly related strains. In this manuscript, I take a reverse approach to the classic neutralization study, starting first with pigs we know are broadly neutralizing and then investigating the identity of the responsible B cells, using a novel technique for the isolation of PRRSV neutralizing antibodies. PBMCs were harvested from pigs sequentially exposed to divergent PRRSV isolates and vaccine. Memory B cells were then transduced, with a proprietary retroviral vector developed by AIMM Therapeutics, containing genes highly expressed by germinal center B cells, creating an immortalized B-cell population. B-cells were then sorted by FACS and five PRRSV-specific B cells were isolated. All identified PRRSV-specific antibodies were found to be broadly binding to all PRRSV-2 isolates tested, but not PRRSV-1. Antibodies against GP5 protein, commonly thought to be the PRRSV neutralizing epitope, were found to be highly abundant, as four out five clonal B cells were GP5 specific. Next, an isolated GP5 clone was discovered to be neutralizing against homologous, but not heterologous PRRSV. Sequencing of this clone’s heavy chain variable region and CDR3 revealed a gene that was heavily mutated compared to germline sequence, suggesting somatic hypermutation playing an essential role in generation of broadly neutralizing antibodies. 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 bnAbs against other porcine pathogens.
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University of Minnesota M.S. thesis. January 2020. Major: Comparative and Molecular Biosciences. Advisors: Michael Murtaugh, Cheryl Dvorak. 1 computer file (PDF); x, 105 pages.
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Young, Jordan. (2020). Characterization of Broadly Neutralizing Antibodies to Porcine Reproductive and Respiratory Syndrome Virus (PRRSV). Retrieved from the University Digital Conservancy, https://hdl.handle.net/11299/213054.
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