A Single Amino Acid Substitution in Porcine Reproductive and Respiratory Syndrome Virus Glycoprotein 2 Significantly Impairs Its Infectivity in Macrophages.

Porcine reproductive and respiratory syndrome virus (PRRSV) has a restricted tropism for macrophages and CD163 is a key receptor for infection. In this study, the PRRSV strain NCV1 was passaged on MARC-145 cells for 95 passages, and two plaque-clones (C1 and C2) were randomly selected for further analysis. The C1 virus nearly lost the ability to infect porcine alveolar macrophages (PAMs), as well as porcine kidney cells expressing porcine CD163 (PK15-pCD163), while the C2 virus replicates well in these two cell types. Pretreatment of MARC-145 cells with an anti-CD163 antibody nearly blocked C1 virus infection, indicating that the virus still required CD163 to infect cells. The C1 virus carried four unique amino acid substitutions: three in the nonstructural proteins and a K160I in GP2. The introduction of an I160K substitution in GP2 of the C1 virus restored its infectivity in PAMs and PK15-pCD163 cells, while the introduction of a K160I substitution in GP2 of the low-passaged, virulent PRRSV strain NCV13 significantly impaired its infectivity. Importantly, pigs inoculated with the rNCV13-K160I mutant exhibited lower viremia levels and lung lesions than those infected with the parental rNCV13. These results demonstrated that the K160 residue in GP2 is one of the key determinants of PRRSV tropism.