Many studies have linked the antimicrobial properties of kefir with the presence of bacteriocins and organic acids. In the present work, results obtained from bacteriostatic and bactericidal studies, and from RP-HPLC, Mass Spectrometry and proton NMR analysis, show that a sample of milk kefir grains is able to produce an antimicrobial fraction, denoted FK-1000, composed of sugars and amino acids, predominantly polymers of alanine, doublets of tyrosine and phenylalanine. Since this fraction is a lyophilized product whose molecular profile is different from bacteriocins and simple carboxylic acids, its antimicrobial effect cannot be attributed to these molecules, or to alcohols or hydrogen peroxide. The fraction is bactericidal against weak-acid-resistant MRSA and weak-acid resistant P. aeruginosa at pH 5, and is bacteriostatic against both pathogens at pH 7. In combination formulation, the FK-1000 fraction is able to increase fivefold the effect of streptomycin against P. aeruginosa and it is not toxic to human epithelial cells at antimicrobial concentrations. 16 S rRNA microbiota analysis of antimicrobial-producing and non-producing kefir grains demonstrated that they are distinct. In summary, the results indicate that milk kefir grains can produce different classes of molecules with potent antibiotic activity against resistant bacteria.
The ability of an oil-based carrier vehicle to act as an antigen delivery system via the oral and/or parenteral routes was investigated. The formulation consists of hydrophilic macromolecules (antigens) solubilised in oil phase, in the absence of water, by virtue of being wrapped in a sheath of phospholipid amphiphile. Results obtained demonstrate that the level of mucosal IgA antibodies detected in the stools of mice immunised orally with cholera toxin B fragment (CTB) or E. coli heat-labile toxin (LT) in oil is much higher than the level of IgA produced by mice immunised with CTB or LT alone. In addition, mice immunised orally with Y. pestis antigens (F1 and V) and CTB as immunostimulant in oil produce a significantly increased (p<0.02) systemic IgG response against both antigens (F1 and V) than mice orally immunised with F1, V and CTB without oil. Six out of ten mice immunised with F1 and V antigens in oil survived an aerosol challenge of 100 LD 50 doses of virulent Y. pestis. Furthermore, animals immunised sub-cutaneously with the HIV antigen (HGP-30) in oil induced much stronger humoral and cellular responses against the antigen than mice immunised with the antigen alone. Taken together, these findings indicate that oil can be used successfully as an antigen delivery system in vaccine formulations without the necessity of an aqueous phase or an emulsification process. This greatly enhances the stability and ease of production of a formulation manufactured for commercial use.
The serogroup O55 of E. coli is composed of strains whose mechanisms of virulence are different from each other. Since the O55 polysaccharides are present in all E. coli O55 strains, and so are the polymers that compose the capsule of O55 atypical enteropathogenic E. coli (aEPEC), it was investigated whether anti-O55 antibodies were able to help the innate immune system to eliminate capsulated aEPEC and Shiga toxin-producing E. coli (STEC) belonging to the serogroup O55. The results demonstrate that the capsule of EPEC was able to inhibit the deposition of C3b on the bacterial surface and, as a consequence, their lysis by the alternative pathway of the complement system. However, in the presence of antibodies, the ability of the complement to lyse these pathogens was restored. It was also observed that macrophages were able to ingest EPEC and STEC, but they were only able to kill the ingested pathogens in the presence of antibodies. Anti-O55 antibodies were also able to inhibit aEPEC and STEC O55 adherence to human epithelial cells. In summary, the results demonstrated that the O55 polysaccharides have the potential to induce an effective humoral immune response against STEC and EPEC, indicating that they are good antigen targets to be used in vaccine formulations against these pathogens.
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