Construction and the immunogenicity of the recombinant Modified Vaccinia Virus Ankala co-expressing ORF4、ORF5 and ORF6 genes of Porcine Reproductive and Respiratory Syndrome Virus NJ-a strain

Abstract:

To develop investigate the recombinant MVA(rMVA) vaccines against PRRSV infection, the ORF4, ORF5 and ORF6 of PRRSV NJ-a strain were subcloned into the MVA transfer vector pⅡLR and the resultant recombinant vector was called pⅡLR-ORF4/ORF5/ORF6. The rMVA was generated by transfecting MVA-infected BHK-21 cells with the recombinant vector and screened by plaque purification after X-gal staining. After six rounds of purification, insertion of PRRSV GP4, GP5 and M genes into the MVA genome was confirmed by PCR analysis and expression of the three proteins was identified by Western-blot and IFA. Each of the tested mice was inoculated with 5×10⁵TCID₅₀/mouse of the rMVA-GP4/GP5/M and boosted 3 weeks later. Neutralization assay showed that PRRSV-specific neutralizing antibodies were detectable at 3 weeks and reached the highest titer (2⁵) by 8 weeks after the primary vaccination, which maintained stable until the end of the experiment. The significant lymphocyte proliferation responses were also observed in mice immunized with rMVA-GP4/GP5/M. These results indicate the rMVA co-expressing PRRSV ORF4, ORF5 andORF6 genes may be an attractive candidate vaccine for preventing PRRSV infection.