Molecular Evolution And Variation Of Porcine Reproductive And Respiratory Syndrome Viruses Isolated In Chongqing

Porcine reproductive and respiratory syndrome(PRRS),caused by porcine reproductive and respiratory syndrome virus(PRRSV),is a serious infectious disease of swine.It is characterized by severe reproductive failure in sows and respiratory distress in piglets and growing pigs.Since it's first reported in the United States in 1987,it has been spread to most countries and caused large economically lost.A highly pathogenic disease which is characterized by high-fever,high morbidity and mortality rate was outbreaked in the south of China in 2006,and then spread quickly to most provinces. Most experts believed that PRRSV mutant was one of the main etiologies.Tissue samples of pigs were collected in Chongqing area in 2008,and analyzed by RT-PCR.The positive samples were inoculated on Marc-145 cell monolayer,and 1 PRRSV strain named CSY-1 was successfully isolated.The isolate was identified as variat by Real-time FQ-PCR.The cDNA fragments covering full-length genome of strain CBB-1 and CWZ-1 isolated in 2006 and 2007 were amplified by RT-PCR and 3' -Full RACE,respectively. The complete genomes were obtained by assembling these sequence.Results of sequence comparison showed that the length of complete nucleotide sequence was 15353bp and 15346bp,respectively.CWZ-1 and CBB-1 had the same genomic structure and shared no more than 62.0%of nucleotide homology with LV.But they shared 89.3%～99.4%of nucleotide homology with American type isolates,shared 98.9%～99.4%of nucleotide homology with variation strain,indicating that CWZ-1 and CBB-1 belonged to American type.Nucleotide analysis showed that these strains had 30 amino acid deletions in Nsp2 protein and several amino acid mutations in some structural protein.The virulence of PRRSV weakened or enhancement was probably due to the effect of several mutations.The Nsp2 and ORF5 genes of PRRSV Chongqing isolates were amplified by RT-PCR,isolated and sequenced.Twelve sequences of Nsp2 gene and fourteen sequences of ORF5 gene were obtained.Compared with PRRSV mutant,sequence analysis showed Nsp2 gene shared 97.3%～99.5%of nucleotide homology and 95.7%～99.2%of amino acid identity,ORF5 gene shared 98.2%～99.7%of nucleotide homology and 98.0%～99.5%of amino acid identity.Compared with traditional strains, the Nsp2 and ORF5 genes and deduced amino acid of all isolates existed many point mutations.The phylogenetic tree indicated that all Chongqing isolates were the variants, and belonged to JXA1 subgroup.The results demonstrated that Nsp2 of gene of strains of Chongqing had 30 amino acid deletions in 481 and 532～560.This proved the variant of PRRSV was the dominant strain in Chongqing.