Boosting systemic and secreted antibody responses in mice orally immunized with recombinant Bacillus subtilis strains following parenteral priming with a DNA vaccine encoding the enterotoxigenic Escherichia coli (ETEC) CFA/I fimbriae B subunit

Recombinant Bacillus subtilis strains, either spores or vegetative cells, may be employed as safe and low cost orally delivered live vaccine vehicles. In this study, we report the use of an orally delivered B. subtilis vaccine strain to boost systemic and secreted antibody responses in mice i.m. primed with a DNA vaccine encoding the structural subunit (CfaB) of the CFA/I fimbriae encoded by enterotoxigenic Escherichia coli (ETEC), an important etiological agent of diarrhea among travelers and children living in endemic regions. DBA/2 female mice submitted to the prime-boost immunization regimen developed synergic serum (IgG) and mucosal (IgA) antibody responses to the target CfaB antigen. Moreover, in contrast to mice immunized only with one vaccine formulation, sera harvested from prime-boosted vaccinated individuals inhibited adhesion of ETEC cells to human red blood cells. Additionally, vaccinated dams conferred full passive protection to suckling newborn mice challenged with a virulent ETEC strain. Taken together the present results further demonstrate the potential use of recombinant B. subtilis strains as an alternative live vaccine vehicle.

Protection induced by pneumococcal surface protein A (PspA) is enhanced by conjugation to a Streptococcus pneumoniae capsular polysaccharide

The currently available anti-pneumococcal vaccines are based on capsular polysaccharide (PS), plain or conjugated to a carrier protein. Conjugated vaccines are expensive products, especially in the case of pneumococcus, in which reasonable coverage requires from 7 to 13 serotypes. To obtain increased coverage with fewer components, we evaluated the immunogenicity of the pneumococcal surface protein A (PspA), conjugated to capsular polysaccharide serotype 23F, aiming at induction of an immune response against both components. Mice immunized with PS23F-rPspA1 conjugate produced antibodies against both PS and rPspA1, comparable or slightly higher than that obtained by free PS. The immunized animals challenged with a lethal dose of a virulent strain bearing a homologous PspA, showed that the PS23F-rPspA1 conjugate induced higher survival than rPspA1 alone or in combination with PS. This increased protection was shown to correlate with the enhanced capacity of the antibodies to bind to the pneumococcal surface and to induce complement deposition. Our results indicate that the use of PS-PspA conjugates may be a way to increase coverage against pneumococci with fewer components.

Stable episomal expression system under control of a stress inducible promoter enhances the immunogenicity of Bacillus subtilis as a vector for antigen delivery

Bacillus subtilis has been successfully engineered to express heterologous antigens genetically fused to surface-exposed spore coat proteins asa vaccine vehicle endowed with remarkable heat resistance and probiotic effects for both humans and animals. Nonetheless, the immunogenicityof passenger antigens expressed by B. subtilis spores is low particularly following oral delivery. In this work, we describe a new episomalexpression system promoting enhanced immunogenicity of heterologous antigens carried by B. subtilis strains, either in the form of sporesor vegetative cells, following oral or parenteral delivery to mice. Based on a bi-directional replicating multicopy plasmid, the gene encodingthe B subunit of the heat-labile toxin (LTB), produced by enterotoxigenic Escherichia coli (ETEC) strains, was cloned under the controlof the B. subtilis glucose starvation inducible (gsiB) gene promoter, active in vegetative cells submitted to heat and other stress conditions.The recombinant plasmid proved to be structurally and segregationally stable in both cells and spores under in vitro and in vivo conditions.Moreover, BALB/c mice orally immunized with B. subtilis cells or spores elicited enhanced anti-LTB systemic (serum IgG) and secreted(fecal IgA) antibody responses, thus, suggesting that antigen expression occurred during in vivo transit. These results indicate that the newepisomal expression system may improve the performance of B. subtilis as a live orally-delivered vaccine carrier.