Isolation And Identification Of NADC30-like PRRSV And Construction Of Its Reverse Genetic Platform

Porcine reproductive and respiratory syndrome(PRRS)is an acute infectious disease caused by porcine reproductive and respiratory syndrome virus(PRRSV),which can cause reproductive and respiratory disorders in pigs.PRRSV is highly infectious to pigs of different breeds,ages and genders,and has had a huge impact on the global pig industry.The NADC30-like PRRSV strain was first reported in Henan in 2013.In recent years,this subtype strain has broken out in a large area in my country,and has gradually developed into a dominant strain in various places.This subtype strain has many new features,such as high recombination variability,relatively weak pathogenicity,and strong immunosuppressive ability,making it difficult for commercial vaccines to effectively prevent and control it.Therefore,in-depth study of the characteristics of NADC30-like PRRSV strains is very important for the prevention and control of this disease.Based on the analysis of clinical cases,this study conducted in-depth research on the genome and pathogenic characteristics of NADC30-like PRRSV strains,and constructed a NADC30-like PRRSV reverse genetic manipulation platform.The specific contents and results of this study are as follow :1.Isolation and sequence analysis of NADC30-like PRRSV strainA strain of NADC30-like PRRSV was successfully isolated from the disease materials submitted for inspection and named as XM-2020.The full sequence of the strain was subsequently determined,and its genomic characteristics were analyzed using bioinformatics software.The comparison results showed that there were 131 discontinuous amino acid deletions in the Nsp2 region of the XM-2020 strain,which was consistent with the genomic characteristics of the NADC30-like PRRSV strain.There are two recombination breakpoints in its sequence,indicating that this strain recombines with HP-RRRSV.2.Pathogenicity assessment of XM-2020 strainThe pathogenicity comparison test with the HP-PRRSV strain showed that the pathogenicity of the XM-2020 strain was lower than that of the HP-PRRSV strain,specifically,the piglets inoculated with the XM-2020 strain showed milder clinical symptoms and lower Serum antibody levels.Interestingly,the piglets infected with the XM-2020 strain developed diarrhea symptoms during the disease period,and there were obvious lesions in the intestine.3.Reverse genetic platform of XM-2020 was establishedThe entire genome of XM-2020 was successfully amplified and spliced to construct an infectious clone.The r XM-2020 strain and the r XM-2020-BFP strain were successfully rescued by transfecting BHK-21 cells,and it was determined that the rescued strains had similar growth curves to the parental strains.In conclusion,this study successfully isolated and identified a NADC30-like PRRSV strain,and analyzed the genomic characteristics of this strain,which provided reference data for determining the local PRRSV prevalence.This strain is less pathogenic than HP-PRRSV,but can cause lesions in the intestine,which is of great significance for the prevention and control of NADC30-like PRRSV subsets.In addition,the successful construction of the reverse genetic platform provides technical support for the next step in the related research on the enhancement of the ability of NADC30-like PRRSV subsets to infect various organs.