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. 2012 Jul;1(1):77-82.
doi: 10.7774/cevr.2012.1.1.77. Epub 2012 Jul 31.

Recombinant influenza viruses as delivery vectors for hepatis B virus epitopes

Jae-Min Song  1 , Kwang-Hee Lee, Baik-Lin Seong
Affiliations

Recombinant influenza viruses as delivery vectors for hepatis B virus epitopes

Jae-Min Song et al. Clin Exp Vaccine Res. 2012 Jul.

Abstract

Purpose: Neuraminidase (NA) of influenza virus contains stalk region that shows a great deal of variability in both amino acid sequence and length. In this paper, we investigated generation of recombinant influenza viruses that had hepatitis B virus (HBV) B cell epitopes in the NA stalk region as a dual vaccine candidate.

Materials and methods: We used the WSH-HK reassortant helper virus for rescue of recombinant influenza virus containing HBV epitopes and reverse genetic protocol based on the use of micrococcal nuclease-treated virus cores for reconstitution of ribonucleoproteins.

Results: We successfully generated a chimeric influenza viruses which contained 22 amino acid peptides in the stalk region derived from the surface and pre-surface protein HBV. The growth kinetics of the recombinant viruses was investigated after infection of Madin-Darby canine kidney (MDCK) and Madin-Darby bovine kidney (MDBK) cells and the rIV-BVPreS virus showed higher titer than other viruses in MDCK cells. We also confirmed the presence of HBV epitopes in the chimeric viruses by enzyme-linked immunosorbent assay (ELISA) using anti-HBV polyclonal antibody. When the ratio of recombinant virus verse wild type virus was calculated by ELISA, recombinant viruses exhibited 2 fold higher values than the wild type virus.

Conclusion: These results suggest that chimeric influenza virus which contained foreign antigens can be used as dual vaccine against both HBV and influenza viruses.

Keywords: Chimeric virus; Dual vaccine; NA stalk.

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Conflict of interest statement

No potential conflict of interest relevant to this article was reported.

Figures

Fig. 1
Fig. 1
Generation of recombinant influenza A virus. Reconstituted ribonucleoprotein (RNP) complex which has RNA in vitro transcribed with foreign epitopes and functional viral core proteins was transfected into Madin-Darby bovine kidney (MDBK) cells that had been infected with WSN-HK helper virus. Recombinant influenza A virus was selected by plaque assay on MDBK cells in the absence of protease and further amplified in MDBK cells. NA, neuraminidase; NP, nucleoprotein.
Fig. 2
Fig. 2
Amino acid sequences of the neuraminidase (NA) stalk mutants from residues 36 to 80 of WSN NA. The B cell epitope sequences derived from HBV surface (rIV-BVS) and PreS2 (rIV-BVPreS2) protein are italicized. NAmut is the partial amino acid sequences of mutant NA on the intermediate cloning vector, pT3WSN-NAmut, for introducing foreign epitopes.
Fig. 3
Fig. 3
Growth properties of recombinant influenza A viruses. rIV-BVS and rIV-BVPreS2 were cultured in Madin-Darby bovine kidney (MDBK) (A) and Madin-Darby canine kidney (MDCK) (B) cells for 72 hours then the virus titer of the sample was determined by plaque assay. PFU, plaque-forming unit.
Fig. 4
Fig. 4
Binding properties between recombinant influenza A viruses and hepatitis B virus (HBV) antibody. rIV-BVS and rIV-BVPreS2 were coated on the microtiter plates and tested the binding activity of the anti-HBV polyclonal antibody by enzyme-linked immunosorbent assay.
See this image and copyright information in PMC

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