[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].
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 p II LR and the resultant recombinant vector was called p II LR-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 x 10(5) TCID(50)/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 (2(5)) 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.
Zheng
Qi-sheng
QS
Key Laboratory of Animal Disease Diagnosis and Immunology, Ministry of Agriculture, Nanjing Agricultural University, Nanjing 210095, China. zhengqisheng2004@yahoo.com.cn
Bi
Zhi-xiang
ZX
Li
Peng
P
Chen
De-sheng
DS
Chen
Pu-yan
PY
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood; D015153:Blotting, Western; D005260:Female; D005455:Fluorescent Antibody Technique; D008213:Lymphocyte Activation; D051379:Mice; D008807:Mice, Inbred BALB C; D016133:Polymerase Chain Reaction; D019316:Porcine respiratory and reproductive syndrome virus / Q000276:immunology*; D011994:Recombinant Proteins / Q000276:immunology; D013601:T-Lymphocytes / Q000276:immunology; D014614:Vaccines, Synthetic / Q000276:immunology*; D014616:Vaccinia virus / Q000235:genetics*; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000276:immunology; D014763:Viral Matrix Proteins / Q000235:genetics* / Q000276:immunology; D015678:Viral Structural Proteins / Q000235:genetics* / Q000276:immunology; D014765:Viral Vaccines / Q000276:immunology*
D000914:Antibodies, Viral; D011994:Recombinant Proteins; D014614:Vaccines, Synthetic; D014759:Viral Envelope Proteins; D014763:Viral Matrix Proteins; D015678:Viral Structural Proteins; D014765:Viral Vaccines; C114789:glycoprotein 5, PRRSV