Abstract
To develop a safe and effective mucosal vaccine against pathogenic influenza viruses, we constructed recombinant Lactobacillus casei strains that express conserved matrix protein 2 with (pgsA-CTA1-sM2/L. casei) or without (pgsA-sM2/L. casei) cholera toxin subunit A1 (CTA1) on the surface. The surface localization of the fusion protein was verified by cellular fractionation analyses, flow cytometry and immunofluorescence microscopy. Oral and nasal inoculations of recombinant L. casei into mice resulted in high levels of serum immunoglobulin G (IgG) and mucosal IgA. However, the conjugation of cholera toxin subunit A1 induced more potent mucosal, humoral and cell-mediated immune responses. In a challenge test with 10 MLD50 of A/EM/Korea/W149/06(H5N1), A/Puerto Rico/8/34(H1N1), A/Aquatic bird /Korea/W81/2005(H5N2), A/Aquatic bird/Korea/W44/2005(H7N3), and A/Chicken/Korea/116/2004(H9N2) viruses, the recombinant pgsA-CTA1-sM2/L. casei provided better protection against lethal challenges than pgsA-sM2/L. casei, pgsA/L. casei and PBS in mice. These results indicate that mucosal immunization with recombinant L. casei expressing CTA1-conjugated sM2 protein on its surface is an effective means of eliciting protective immune responses against diverse influenza subtypes.
Highlights
Vaccination remains most economical and effective means against respiratory diseases caused by influenza viruses [1]
Since sM2 is a highly conserved and promising target for a universal vaccine and cholera toxin subunit A1 (CTA1) is strong mucosal adjuvant, in this study, we developed constructs using a consensus sM2 gene reconstituted from the analysis of H1N1, H5N1 and H9N2 influenza viruses with or without the fusion of CTA1
To determine the cellular localization of the sM2 and CTA1 proteins expressed on the surface of L. casei via the cell wall anchor protein pgsA, membrane and cytoplasmic fractions were subjected to western blot analysis
Summary
Vaccination remains most economical and effective means against respiratory diseases caused by influenza viruses [1]. Based on the circulating viruses in the population, trivalent vaccine strains have been developed and are used for the influenza virus protection [2]. The most acceptable current available strategy is the intramuscular administration of inactivated vaccines produced by egg-based manufacturing systems which while effective, are hampered by limited capacity and flexibility [3]. Vaccine strains must be frequently adapted to match the circulating viruses throughout the world [4]. The levels of antibody induced by the inactivated vaccine have been observed to decrease by 75% over an 8-month period [2,5]. Alternative strategies for developing broadly cross-protective, safe and effective vaccines against influenza viral infections are of prominent importance
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