Abstract：S-layer protein CTC surface display system of Bacillus thuringiensis was used to test the possibility of displaying H5N1 avian influenza virus haemagglutinin HA1 on cell surface. Two fusion genes were constructed by replacing the central part below the surface anchor sequence slh of S-layer protein gene ctc with part of ha1 gene (ha1p). Two recombinant B. thuringiensis strains were constructed by transferring recombinant plasmids to B. thuringiensis plasmid-free derivative strain BMB171, such as BCCH (harboring ctc-ha1p as well as a plasmid carrying surface display helper csaAB operon) and CH (harboring csa-ctc-ha1p). Hemagglutination assay showed recombinant HA1 proteins were displayed on the surface of two recombinant strains. Hemagglutination inhibition assay showed recombinant HA1 proteins were specific to standard HI (Hemagglutination Inhibition test) serum of type H5 AIV. After immunizing mice with recombinant strains, the recombinant HA1 proteins elicited a humoral respone to HA1 and exhibited immunogenicity as assayed by enzyme-linked immunosorbent assay. Meanwhile these assay showed recombinant strain CH exhibited the higher activity than BCCH. This demonstrated that the construction way of fusion gene csa-ctc-ha1p was more appropriate to S-layer protein displaying heterologous antigen on cell surface. The strategy developed in this study gives a possibility to generate heat stable, oral, veterinary vaccine against AIV with B. thuringiensis S-layer protein CTC surface display system.
