The Key Amino Acid Sites Of PRRSV Affect Cell Tropism

Porcine reproductive and respiratory syndrome(PRRS),caused by porcine reproductive and respiratory syndrome virus(PRRSV),is a significant threat to the global pig industry.In recent years,the PRRSV strains in China has become more and more complicated,and the mutant strains HP-PRRSV,NADC30-Like and GM2-Like strains,which could not passage in the MARC-145 cells,have emerged successively.Currently,extensive fundamental research and most anttenuated vaccine strains were obtained from the MARC-145 cells.Therefore,it is significant to research against these PRRSVs for scientific prevention and control of PRRS.The prevalence of PRRS in 2018 in South China was studied and it was found that the prevalence of PRRS in South China was still dominated by HP-PRRSV.Whole genome sequencing of 7 strains showed that strains that did not infect Marc-145 cells were ubiquitous in the three subtypes.The PRRSV GM2-Like 1138 isolated by our laboratory can infect porcine pulmonary macrophages(PAM)but not the passaged cell line African green monkey kidney epithelial cells(Marc-145 cells).The HP-PRRSV JX-GD and the NADC30-like LYNA strain can infect both PAM cells and Marc-145 cells.The amino acids in the GP2a-GP3 region of PRRSV strains isolated from the laboratory and the known cell-adapted strains in Gen Bank were analyzed.It was found that the 98 th amino acid of GP2 a was mainly leucine and phenylalanine.The strain which GP2 a 98th amino acid is phenylalanine can not be infected with Marc-145 cell line,while 98 amino acid is leucine can infect PAM and Marc-145 cells.It inferred GP2 a 98th amino acid possible A key site of PRRSVs that could not infect Cells.In order to verify whether the 98 th amino acid of GP2 a is a key site affecting PRRSV infection of Marc-145,the laboratory constructed pok JXA1 infectious clone as the backbone.Mutated GP2 a 98 leucine to phenylalanine,The r JXA1-98 Phe virus was successfully saved.r JXA1-98 Phe can infect PAM but not Marc-145 cells,and its parental JXA1 strain can infect both cells.It indicates that the 98 th amino acid of GP2 a is the key to determine the adaptability of PRRSV to Marc-145 cells.The cell adaptability was reversely verified.The infectious clone pok1138 was constructed by using the strain 1138 which could not infect Marc-145 cells isolated in our laboratory,and the 98 th amino acid of GP2 a was leucine,which successfully rescued r1138-98 Leu.r1138-98 Leu can infect PAM but Marc-145 cells,indicating that the 98 th amino acid of GP2 a is a key site determining the adaptability of PRRSV to Marc-145 cells.In this study,we determined that the 98 th amino acid of GP2 a is a key site for PRRSVinfected Marc-145 cells by sequence analysis.The change of this site has no significant effect on the proliferation rate of PRRSV on PAM cells.influences.This study provides new research ideas and methods for the study of PRRSV cell adaptation in different cells.