The Evolution And Epidemiological Characteristics Of Lineage 3 And 1 PRRSV In China And HP-PRRSV Fidelity Analysis

PRRSV is prone to genetic variation.Since the emergence of PRRSV in the 1980 s,PRRSV has been evolving continuously and developed many epidemic lineages and endemic lineages,and spread.worldwide.It has been almost more than 20 years in China since the first being reported.The prevalence and variation of endemic PRRSVs,combined with exotically introduced PRRSVs,make the situation of PRRSV in China become more complex.Lineage 3 PRRSV,belonging to the genotype II,has a long epidemic history in China.frequently appeared.The novel lineage 3 viruses constantly emerging in recent years in southern China are characterized by a high detection rate and significant pathogenicity.It increases the risk to threaten the pig industry in China.In order to investigate the prevalence of lineage 3 in South China,a cross-sectional epidemiology investigation was performed in PRRSV positive pig farms in South China from January 2016 to July 2017,and the genome and biological characteristics of the lineage 3 isolates were investigated.Results showed that the prevalence of lineage3 in South China was 35.68%(95% CI: 27.6-44.3%).The genome and phylogenetic analysis of GDZS2016 and FJNP2017 isolates indicate that the genomes of the two strains share 95.7%–96.2% homology with JXA1-like strains(HP-PRRSV)and located in JXA1-like group in the evolution tree.The recombination analysis showed HPPRRSV provides both strains with most of the ORF1 gene,encoding non-structural proteins,while most parts of the structural protein gene region of the two strains were provided by lineage 3 viruses.The results of the animal experiments suggested that FJNP2017 strain with the complete ORF1 gene of HP-PRRSVs showed higher virulence than the GDZS2016 strain with part of the ORF1 gene from HP-PRRSV.These results give a reason for the increased pathogenicity of lineage 3 viruses in pig farms.With the development of phylogeny,phylodynamics and phylogeography have been widely used in the animal husbandry industry as a real-time surveillance system for animal pathogens.In this study,the phylogeny,phylodynamics,and phylogeography combined with big data were used to analyze the origin,exotic introduction,evolution history and current epidemic status of lineage 3 and lineage 1 PRRSV in China.The results showed that the Lineage 3 strain originated in Taiwan.It has developed into two endemic clusters: mainland China cluster and Taiwan island cluster.The epidemic history and viral dispersal route analysis indicated that the epicenter of lineage 3 was southern China and lineage 3 viruses spread to interior China through this area.Since the lineage 1 PRRSV was first reported in China in 2013,these strains have been continuously imported from the United States and had three sub-lineage 1 appeared in China: lineage 1.5(NADC34-like),lineage 1.6 and New Intro cluster(NADC30-like).These viruses with high genetic diversity had widely distributed in China,especially in Henan Province.Lineage 1 viruses have developed an endemic cluster in China: New Intro 1.1(NADC30-like cluster).Our result demonstrated that the highfrequency recombination events in the Chinese lineage 1 viruses occurred among the endemic PRRSV strains in China.The recombination hotspots of lineage 1 viruses are mainly located in Nsp9 and ORF2a-ORF4 regions.For the lack of proofreading capability of PRRSV’s polymerase,the replication of PRRSV is an error-prone process that contributes to the mutations.Chinese HP-PRRSV is prone to genetic mutations.It is reported that the virulence return of the commercial HPPRRSV vaccine occurs.In this study,the nucleic acid mutagens were used as a selection pressure to act on the JXA1 strain,and the fidelity-determining sites on the RdRp of JXA1 strain were screened.Using the JXA1 infectious cloning platform,the selected Ala86 Thr and Val155 Ile sites were verified.The results showed that the rescued rJXA1-86 and rJXA1-86-155 were resistant to ribavirin and had a lower mutation frequency than the parental strain.In summary,lineage 1 and lineage 3 PRRSVs were systematically analyzed.Results reveal the exotic introduction,evolution history and current epidemic status of these circulating viruses in China.In addition,we screened and identified the fidelity rJXA1-86 and rJXA1-86-155 strains in vitro.This research provides a reference for the prevention and control of lineage 1 and lineage 3 viruses,and also lay the foundation for developing a safe and stable HP-PRRSV vaccine.