Abstract
Effective vaccination against influenza virus infection is a serious problem mainly due to antigenic variability of the virus. Among many of investigated antigens, the extracellular domain of the M2 protein (M2e) features high homology in all strains of influenza A viruses and antibodies against M2e and is protective in animal models; this makes it a potential candidate for generation of a universal influenza vaccine. However, due to the low immunogenicity of the M2e, formulation of a vaccine based on this antigen requires some modification to induce effective immune responses. In this work we evaluated the possible use of Bacillus subtilis spores as a carrier of the Influenza A M2e antigen in mucosal vaccination. A tandem repeat of 4 consensus sequences coding for human—avian—swine—human M2e (M2eH-A-S-H) peptide was fused to spore coat proteins and stably exposed on the spore surface, as demonstrated by the immunostaining of intact, recombinant spores. Oral immunization of mice with recombinant endospores carrying M2eH-A-S-H elicited specific antibody production without the addition of adjuvants. Bacillus subtilis endospores can serve as influenza antigen carriers. Recombinant spores constructed in this work showed low immunogenicity although were able to induce antibody production. The System of influenza antigen administration presented in this work is attractive mainly due to the omitting time-consuming and cost-intensive immunogen production and purification. Therefore modification should be made to increase the immunogenicity of the presented system.
Highlights
Influenza virus belongs to the Orthomyxoviridae family and because of its high virulence and adaptive abilities it is one of the world’s most dangerous pathogens of warm-blooded vertebrates
Construction and chromosomal integration of gene fusions To obtain recombinant B. subtilis spores expressing influenza matrix 2 (M2) protein (M2e) antigen the synthetic M2eH-A-S-H sequence coding tandem repeat of 4 consensus sequence of human—avian— swine—human M2e was fused in frame with the spore coat protein genes cotB, cotC, cgeA and cotZ
All gene fusions were integrated into the B. subtilis chromosome and individual clones for each transformation were tested by PCR, and named BTL20 (CotC-M2eH-A-S-H), BTL21 (CotB-M2eH-A-S-H), BTL23 (CotB-LINKER-M2eH-A-S-H), BTL22 (CotZ-M2eH-A-S-H), BTL24 (CotZ-LINKER-M2eH-A-S-H), BTL25 (CgeA-M2eH-A-S-H), BTL26 (CgeA-LINKER-M2eH-A-S-H) and used for further analysis
Summary
Influenza virus belongs to the Orthomyxoviridae family and because of its high virulence and adaptive abilities it is one of the world’s most dangerous pathogens of warm-blooded vertebrates. Influenza Antigen Spore Display responsible for epidemics and pandemics. Due to the frequent antigenic drifts and antigenic shifts of the circulating virus there is a need for annual vaccine re-formulation and vaccination. This is a serious problem for influenza vaccination, which could be solved by generation of a universal vaccine able to confer cross protection against different influenza variants and subtypes. Studied antigens that could induce cross-reactive immune response are the conserved stalk domain of the hemagglutinin (HA2) [3,4], influenza A nucleoprotein (NP) [5,6] and the matrix 2 (M2) protein [7]
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