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Display of H5N1 Avian influenza virus haemagglutinin HA1 on Bacillus thuringiensis cell surface and its immunogenicity for mice

  • Liu Mei (a1) (a2), Lu Lin-Jing (a1), Huang Jun-Yan (a2), Zhang Shu-Huan (a1), Bi Ding-Ren (a1) (a2) and Sun Ming (a2)...


The S-layer protein CTC surface display system of Bacillus thuringiensis was used to test the possibility of displaying H5N1 Avian influenza virus (AIV) haemagglutinin HA1 on the cell surface of B. thuringiensis. Two recombinant plasmids, pCTC-HA1P and pCSHA1P, were constructed by replacing the central part below the surface anchor sequence slh of S-layer protein gene ctc with part ha1 gene (ha1p). pCTC-HA1P harboured the fusion gene ctc-ha1p and pCSHA1P the fusion gene csa-ctc-ha1p, csa representing the csaAB operon (very important in anchoring S-layer protein on the bacterial cell surface). Two recombinant B. thuringiensis strains were constructed by electrotransferring recombinant plasmids to B. thuringiensis plasmid-free derivative strain BMB171. Strains obtained were CH (bearing pCSHA1P) and BCCH (bearing pCTC-HA1P as well as the csaAB operon-carrying plasmid pMIL-CSA). The vegetative cells of CH and BCCH were used as antigens in haemagglutination (HA) and haemagglutination inhibition (HI) assays. HA assay showed recombinant HA1 proteins successfully displayed on the cell surface of CH and BCCH. HI assay showed that these recombinant HA1 proteins were specific to standard positive HI (haemagglutination inhibition test) serum of subtype H5 AIV. After immunization of mice with vegetative cells, both CH and BCCH elicited a humoral response to HA1 and exhibited immunogenicity as indicated by enzyme-linked immunosorbent assay (ELISA). ELISA also showed that CH exhibited a higher immunogenicity than BCCH. The strategy developed in this study suggests the possibility of generating a heat-stable and oral veterinary vaccine against AIV with the B. thuringiensis S-layer protein CTC surface display system.


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