Cloning ORF3 Gene And Molecular Epidemiology Of Porcine Reproductive And Respiratory Syndrome Virus In Guangxi Province

Porcine reproductive and respiratory syndrome virus(PRRSV)is the causative agent of PRRS,which caused reproductive failure in sows and respiratory problems in piglets.It has caused great damages for worldwide hog industry since being recognized.In the sumer and autumn of 2006,part regins in our country occured an infectious disease named ardent fever disease,which caused great damages.We found the variable strains of highly pathogenic PRRS and proved it caused this pestilence.In this study,we identified 56 sample collected from the pig farms in Guangxi from ??? 2006 to Feb.2008 by RT-PCR.The result showed that 30 samples were positive,the positive rate was 54%.According to the reported complete nucleotide sequences of ATCC VR-2332 and China type strain CH-1a, a pair of primers GP3-1/GP3-2 were designed to amplify ORF3 gene of the 30 PRRSV positive samples with Primer software.The fragments of complete ORF3 gene obtained from RT-PCR was approximate 898bp in length which was correspond to the objective fragments.The ORF3 gene was cloned into pMD18-T vector and sequenced.The results of sequencing showed that complete 756 bp ORF3 gene had a complete open reading flame which encoded 254 amino acids.The 30 strains ORF3 genes samples whose nucleotide homologies were 90.8%～99.9%compared with reference viruses VR-2332,FJ04A,JXA1,HuN,LV,RespPRRS MLV and CH-1a strain were 87.9～90.0%, 89.4%～90.7%,92.1%～99.7%,91.9%～99.4%,40.7%～41.2%,88.2%～89.6%, 93.1～96.7%.The homology of deduced animo acid sequences were 90.2～92.5%, 91.3%～93.8%,93.0%～99.7%,93.8%～99.6%,54.9%～56.8%,90.2%～92.5% and 95.9～97.6%respectively.The result indicated that the domestic PRRSV epidemic strains were significantly genetic divergent from each other,30 samples belonged to North American genotype.Phylogenetic tree among 30 strains and other 48 PRRSV strains published were obtained by analyzing nucleotide sequences of ORF3 gene.The ORF3 gene homologies of 30 strains which belonged to subgroupⅡof the three subgroups were closely to the demonic type strains isolated during the period of 'ardent fever disease' from 2006 to 2007,which was 97.7%～99.7%.Compared the deduced amino acids,we found out 7 potential glycosylation sites existed in the ORF3 encoded protain of different subgroups strains isolated from different regins of GuangXi.In this study,the genetic variability was study by comparing the PRRSV ORF3 gene nucleotide.We can understand PRRS epidimiology in south of China,and it should be a good foundation for controling this disease.