The Significance of the 98th Amino Acid in GP2a for Porcine Reproductive and Respiratory Syndrome Virus Adaptation in Marc-145 Cells

Abstract

Porcine reproductive and respiratory syndrome virus (PRRSV) is one of the most important pathogens in the pig industry. Marc-145 cells are widely used for PRRSV isolation, vaccine production, and investigations into virus biological characteristics. Despite their significance in PRRSV research, Marc-145 cells struggle to isolate specific strains of the North American virus genotype (PRRSV-2). The involvement of viral GP2a, GP2b, and GP3 in this phenomenon has been noted. However, the vital amino acids have not yet been identified. In this study, we increased the number of blind passages and successfully isolated two strains that were previously difficult to isolate with Marc-145 cells. Both strains carried an amino acid substitution in GP2a, specifically phenylalanine to leucine at the 98th amino acid position. Through a phylogenetic and epidemiologic analysis of 32 strains, those that were not amenable to isolation widely exhibited this mutation. Then, by using the PRRSV reverse genetics system, IFA, and Western blotting, we identified the mutation that could affect the tropism of PRRSV-2 for Marc-145 cells. Furthermore, an animal experiment was conducted. Through comparisons of clinical signs, mortality rates, and viral load in the organs and sera, we found that mutation did not affect the pathogenicity of PRRSV-2. In conclusion, our study firmly establishes the 98th amino acid in GP2a as a key determinant of PRRSV-2 tropism for Marc-145 cells.

Reverse genetics construction and pathogenicity of a novel recombinant NADC30-like PRRSV isolated in China

“…The K160 residue in GP2a is associated with PRRSV infectivity in MARC-145 cells and PAMs ( 38 ). Recently, the 98th amino acid of GP2a was found to play a key role in PRRSV adaptation to MARC-145 cells, and PRRSV strains with phenylalanine at this residue fail to infect MARC-145 cells ( 39 ). In line with their finding, GS2022 carrying phenylalanine at the 98th residue of GP2a also can infect PAMs but not MARC-145 cells ( Figure 1B ).…”