Sequence Analyses Of The PRRSV (AH0701) Complete Gene Isolated In Anhui And Development Of The Inactived Vaccine

Porcine reproductive and respiratory syndrome virus (PRRSV) is the causative agent of porcine reproductive and respiratory syndrome (PRRS), causes reproductive failure in breeding swine and respiratory problems in young pigs, which is responsible for significant economic losses to the swine industry throughout China. In order to investigate biological differences and genetic variation and genetic relationship of the genomes of PRRSV strains, the whole viral genome of PRRSV-AH0701 strain isolated from Anhui province in 2007 was sequenced and analysis of phylogenetic tree was constructed based on PRRSV-AH0701 and the documented strains isolated from other regions. In this study, we also developed the oil emulsion inactivated vaccine against PRRS with PRRSV-AH0701 strain, which should provide technological support for prevention and control of PRRS in Anhui province.Fifteen pairs of primer were designed and synthesized according to PRRSV complete genome sequence published in GenBank. Fifteen overlapping fragments covering the whole viral genome of PRRSV-AH0701 isolate were amplified by RT-PCR and the 5'and 3'ends were amplified with RACE. The cDNAs were cloned into pMD18-T for sequencing and the sequences were published in GenBank (GU461292). Sequences were aligned and phylogenetic trees were drawn as previously described. The results of sequence analysis demonstrated that PRRSV-AH0701 has a discontinuous deletion of 30 amino acids in Nsp2 gene, the same as PRRSV isolated from other regions of China after 2006, which was consistent with the characteristic of high pathogentic porcine reproductive and respiratory syndrome virus. The phylogenetic tree revealed that PRRSV-AH0701 strain had closer relationship with strains isolated in Shangdong and Hubei. Our results enriched information data of PRRSV genome and provided a valuable basis for further study on biological characteristics and molecular genetic variation of PRRSV.Oil emulsion inactivated vaccine against PRRSV was developed due to PRRSV-AH0701 strain. First, PRRSV-AH0701 was proliferated on Marc-145 cell with titer of 10^5.5 TCID50/0.1mL, the virus was collected and inactivated for developing the oil emulsion inactivated vaccine. The immune protection test of inactivated vaccince was conducted and the rate of protection is 80% (4/5), also immune efficiency was evaluated with ELISA antibody against PRRSV in experimental animals. Results showed that the immune dose of the inactivated vaccine was optimized at a dose of 2 mL, and the peak of antibody titer was observed at 14d post immunization. The antibody level could last eleven weeks. It was adviseded that the first vaccination should be introduced at 28d and revaccination at 56d with inactivated vaccine for the pig in PRRS -negative farm. For the in the PRRS-positive farm, first vaccination should be done at 28d and revaccination with attenuated vaccine at 56d was applied.As stated above, the satisfactory immune protection of the inactivated vaccince was achieved against porcine reproductive and respiratory syndrome due to preliminary experimental test and field trial. With the good safety and high immunoprotection, the developed inactivated vaccine is useful to prevent and control PRRS in Anhui province.