Dynamic Investigation And Genetic Evolution Analysis Of PRRSV In Partial Regions Of China In2011-2012

PRRS(Porcine Reproductive and Respiratory Syndeom)was found in south of America in1987, the main symptoms of PRRS is breeding disorder of pregnant sows and respiratory distressof piglets and growing pigs. The disease is caused by the PRRSV(Porcine Reproductive andRespiratory Syndrom virus), PRRSV is belong to Arteriviridae, and PRRSV is a small envelopedvirus with a positive-sense, single-stranded RNA,the characteristic of PRRSV is extensive geneticvariation and multiple strains.In this study,we collected1707samples in119ordinary surveillance slaughterhouses,1255suspected cases in164pig farms from12different provinces in2011,and in2012we clllected1485samples in105oridinary surveillance slaughterhouses and588suspected cases in152pigfarms from12different provinces. Test results showed that the positive rate of ordinarysurveillance slaughterhouses is16.75%, the positive rate of suspected cases is42.8%in2011; andin2012the positive rate of ordinary surveillance slaughterhouses is15.26%the positive rate ofsuspected cases is21.09%. Through the analysis and comparison of results, we found that thepositive rate of ordinary surveillance slaughterhouses and suspected cases were all decreased,andthe positive rate of pig farms that had suspected cases had a substantial decline.Isolate virus in positive samples used Marc-145cells. Using Primer Premier version5.0software to design two pairs of primers T1,T2and P1,P2for the hypervariable region of NSP2gene sequence and ORF5gene sequences based on the domestic and foreign gene of PRRSVreference strains sequence in GenBank. Amplify the NSP2gene hypervariable region fragment andORF5gene sequence by RT-PCR. Connected and transformed the amplified target fragment andpGEM-T easy vector. Obtained the recombinant plasmids containing the target fragment of420bpand603bp. Identified the bacterium by PCR and sent the positive bacterium to sequencing.Theresult showed that we totally obtained57the hypervariable region gene sequence of NSP2geneand59ORF5gene sequences. The nucleotide sequence homology of NSP2gene was89.6%~100%and the nucleotide sequence homology of ORF5gene was90.5%~100%. The homology of theamino acid sequence deduced by ORF5gene was82.1%~99.5%.Analyse the phylogenetic treesconstructd with the sequence of classical strains JXA1, CH-1a, VR-2332, we found that the strainswe obtained were all American type strain.Compared and analysed the sequence of the gene obtained using the MegAlign software inDNAstar software package, and compared to domestic strains CH-1a, HB-1(sh)2002, JXA1andabroad standard strain VR-2332. The result show that the nucleotide variation of NSP2gene mainly occurred in discontinuous90nucleotide deletion and the variation of ORF5gene mainlyoccurred in amino acid point mutations.Aalysed the phylogenetic trees based on reference strains and isolated strains, the PRRSVstrains obtained in this research were closer with the domestic isolates JXA1in geneticrelationship. In2011-2012the dominant strains were still mutant strains represented by JXA1inChina, belong to the subgroup4.