Epidemiological Investigation Of PRRSV In Guangdong Province Of 2017 And Construction Of Reverse Genetics System For JXA1 Strains

To investigate the prevalence of PRRSV in Guangdong,4456 serum samples for antibody detection and 305 tissue samples for antigen detection were collected among large scale pig farms in Guangdong province during 2017.Finally,3263 sera were detected positive and the positive rate was 73.23%,114 antigen samples were detected positive and the positive rate was 37.38%.The positive rate of antibody was at a higher level,while the antigen positive rate was increased compared to the past five years.In order to reserch the relationship between PRRSV antigen-antibody levels with quarters,the detection results were classified by sersons.The results showed that the antibody positive rates were 69.45%,70.73%,77.66% and 79.46,from the first quarter to the fourth quarter,respectively,and the antigen positive rates were 62.16%,38.60%,33.33% and 29.57%,respectively.These data showed that from the first quarter to the fourth quarter,the antibody level increased and the antigens positive rate decreased.In order to reserch the relationship between PRRSV antigen-antibody levels with regions,the results were classified according to area.The results showed that the highest antibody positive rate was 91.60% in northern Guangdong,but the lowest antigen detection rate was 29.19% in Pearl River Delta region.In eastern Guangdong,more serum samples were sent for antibody detection,but the antibody level was the lowest in the four regions,which was 68.66%.However,the western Guangdong sent more tissue samples for pathogen detection,but the detection rate of the pathogen was the highest in the four regions,which up to 50.53%.These data indicate that there is no significant correlation between antibody level and antigen positive rate among different regions.To get more information of the prevalent PRRSV strains in Guangdong,ORF5 Gene were try to amplify and sequenced from the positive antigen samples.Finally,25 ORF5 sequences were obtained.Then,genetic evolution analysis and amino acid mutation analysis were executed based on the 25 ORF5 sequences.Genetic evolution analysis showed that 14 of the isolated sequences belonged to Sublineage 8.7 in Lineage 8,9 of the isolated sequences belonged to Lineage 3 and 2 of the isolated sequences belonged to Lineage 1.Genetic analysis based on ORF5 showed that the most prevalent PRRSV strain in Guangdong was Lineage 8.Amino acid analysis showed that the ORF5 isolated in Guangdong had wide range of amino acid site mutations and obvious subgroup characteristics.In order to further study the molecular epidemiology of PRRSV in Guangdong Province,three PRRSV strains were isolated from the corresponding samples of three Lineage.The three PRRSV strains belong to lineage 8 lineage 3 and lineage 1,respectively by sequenced the whole genome of the virus,indirect immunofluorescence assay and genetic analysis based on the whole virus genome.In order to better reserch the function,structure and pathogenesis of HP-PRRSV,The full-length genome of JXA1 strain was cloned to the pOKq vector and the CMV promoter,the BGH termination signal peptide were added to the two ends of the virus gene,respectively.Finally,the full-length infectious clone of JXA1 strain was successfully constructed.The result of virus rescue showed that genetic marker was inserted successfully and the rescued virus could produce similar CPE and growth curves to the parent virus.The construction of JXA1 reverse genetic platform will provide technical support for reserching pathogenesis and gene function of PRRSV.