Potential for colonization of O111:H25 atypical enteropathogenic E. coli.

Using clonal phylogenetic methods, it has been demonstrated that O111:H25 atypical enteropathogenic E. coli (aEPEC) strains belong to distinct clones, suggesting the possibility that their ability to interact with different hosts and abiotic surfaces can vary from one clone to another. Accordingly, the ability of O111:H25 aEPEC strains derived from human, cat and dogs to adhere to epithelial cells has been investigated, along with their ability to interact with macrophages and to form biofilms on polystyrene, a polymer used to make biomedical devices. The results demonstrated that all the strains analyzed were able to adhere to, and to form pedestals on, epithelial cells, mechanisms used by E. coli to become strongly attached to the host. The strains also show a Localized-Adherence-Like (LAL) pattern of adhesion on HEp-2 cells, a behavior associated with acute infantile diarrhea. In addition, the O111:H25 aEPEC strains derived either from human or domestic animals were able to form long filaments, a phenomenon used by some bacteria to avoid phagocytosis. O111:H25 aEPEC strains were also encountered inside vacuoles, a characteristic described for several bacterial strains as a way of protecting themselves against the environment. They were also able to induce TNF-α release via two routes, one dependent on TLR-4 and the other dependent on binding of Type I fimbriae. These O111:H25 strains were also able to form biofilms on polystyrene. In summary the results suggest that, regardless of their source (i.e. linked to human origin or otherwise), O111:H25 aEPEC strains carry the potential to cause human disease.

A universal polysaccharide conjugated vaccine against O111 E. coli.

E. coli O111 strains are responsible for outbreaks of blood diarrhea and hemolytic uremic syndrome throughout the world. Because of their phenotypic variability, the development of a vaccine against these strains which targets an antigen that is common to all of them is quite a challenge. Previous results have indicated, however, that O111 LPS is such a candidate, but its toxicity makes LPS forbidden for human use. To overcome this problem, O111 polysaccharides were conjugated either to cytochrome C or to EtxB (a recombinant B subunit of LT) as carrier proteins. The O111-cytochrome C conjugate was incorporated in silica SBA-15 nanoparticles and administered subcutaneously in rabbits, while the O111-EtxB conjugate was incorporated in Vaxcine(TM), an oil-based delivery system, and administered orally in mice. The results showed that one year post-vaccination, the conjugate incorporated in silica SBA-15 generated antibodies in rabbits able to inhibit the adhesion of all categories of O111 E. coli to epithelial cells. Importantly, mice immunized orally with the O111-EtxB conjugate in Vaxcine(TM) generated systemic and mucosal humoral responses against all categories of O111 E. coli as well as antibodies able to inhibit the toxic effect of LT in vitro. In summary, the results obtained by using 2 different approaches indicate that a vaccine that targets the O111 antigen has the potential to prevent diarrhea induced by O111 E. coli strains regardless their mechanism of virulence. They also suggest that a conjugated vaccine that uses EtxB as a carrier protein has potential to combat diarrhea induced by ETEC.

The influence of environmental bacteria in freshwater stingray wound-healing.

Invasion by bacteria can influence the course of healing of wounds acquired in aquatic environment. In this study, the bacteria present in Potamotrygon motoro stingray mucus and in the Alto Paraná river water were identified, and their ability to induce tissue injury and resist antibiotics was determined. Biochemical identification analysis showed that 97% of all bacterial isolates were Gram negative, Aeromonas spp., Enterobacter cloacae and Citrobacter freundii being the species most prevalent. Gelatinase and caseinase were produced by Aeromonas hydrophila, Aeromonas sobria and Pseudomonas aeruginosa strains. Erythrocyte hemolysis assay showed that A. sobria, A. hydrophila and to a lesser extent, other Gram-negative bacteria produced hemolysin. It was also observed that molecules released in culture by these bacteria were toxic to human epithelial cells. Antibiogram results showed that 68% of all bacterial isolates were resistant to at least one type of antibiotic, mainly B-lactams. Finally, it was demonstrated that although P. motoro venom was toxic to epithelial cells it did not influence bacterial proliferation. In summary, the results obtained in this work indicate that during the accident, the mucus of P. motoro and the environmental water may transfer into the wound pathogenic multi-resistant bacteria with the potential to cause severe secondary infections.

The influence of environmental bacteria in freshwater stingray wound-healing

Invasion by bacteria can influence the course of healing of wounds acquired in aquatic environment. In this study, the bacteria present in Potamotrygon motoro stingray mucus and in the Alto Paraná river water were identified, and their ability to induce tissue injury and resist antibiotics was determined. Biochemical identification analysis showed that 97% of all bacterial isolates were Gram negative, Aeromonas spp., Enterobacter cloacae and Citrobacter freundii being the species most prevalent. Gelatinase and caseinase were produced by Aeromonas hydrophila, Aeromonas sobria and Pseudomonas aeruginosa strains. Erythrocyte hemolysis assay showed that A. sobria, A. hydrophila and to a lesser extent, other Gram-negative bacteria produced hemolysin. It was also observed that molecules released in culture by these bacteria were toxic to human epithelial cells. Antibiogram results showed that 68% of all bacterial isolates were resistant to at least one type of antibiotic, mainly B-lactams. Finally, it was demonstrated that although P. motoro venom was toxic to epithelial cells it did not influence bacterial proliferation. In summary, the results obtained in this work indicate that during the accident, the mucus of P. motoro and the environmental water may transfer into the wound pathogenic multi-resistant bacteria with the potential to cause severe secondary infections.

Lipopolysaccharide as an Antigen Target for the Formulation of a Universal Vaccine against Escherichia coli O111 Strains

A promising approach to developing a vaccine against O111 strains of diarrheagenic Escherichia coli thatexhibit different mechanisms of virulence is to target either the core or the polysaccharide chain (O antigen)of their lipopolysaccharide (LPS). However, due to structural variations found in both these LPS components,to use them as antigen targets for vaccination, it is necessary to formulate a vaccine able to induce a humoralimmune response that can recognize all different variants found in E. coli O111 strains. In this study, it was demonstrated that, despite differences in composition of oligosaccharide repeat units between O111ab and O111ac LPS subtypes, antibodies against one O111 subtype can recognize and inhibit the adhesion to human epithelial cells of all categories of O111 E. coli(enteropathogenic E. coli [EPEC], enterohemorrhagic E. coli [EHEC], and enteroaggregative E. coli [EAEC]) strains regardless of the nature of their flagellar antigens, mechanisms of virulence, or O111 polysaccharide subtypes. These antibodies were also able to increase the clearance of different strains of O111 E. coli by macrophages. PCR analyses of the pathways involved in O111 LPS core biosynthesis showed that all EAEC strains have core type R2, whereas typical EPEC and EHEC havecore type R3. In contrast, atypical EPEC strains have core types R2 and R3. In summary, the results presentedherein indicate that the O111 polysaccharide and LPS core types R2 and R3 are antigen targets for panspecific immunotherapy against all categories of O111 E. coli.

Lipopolysaccharide as an antigen target for the formulation of a universal vaccine against Escherichia coli O111 strains.

A promising approach to developing a vaccine against O111 strains of diarrheagenic Escherichia coli that exhibit different mechanisms of virulence is to target either the core or the polysaccharide chain (O antigen) of their lipopolysaccharide (LPS). However, due to structural variations found in both these LPS components, to use them as antigen targets for vaccination, it is necessary to formulate a vaccine able to induce a humoral immune response that can recognize all different variants found in E. coli O111 strains. In this study, it was demonstrated that, despite differences in composition of oligosaccharide repeat units between O111ab and O111ac LPS subtypes, antibodies against one O111 subtype can recognize and inhibit the adhesion to human epithelial cells of all categories of O111 E. coli (enteropathogenic E. coli [EPEC], enterohemorrhagic E. coli [EHEC], and enteroaggregative E. coli [EAEC]) strains regardless of the nature of their flagellar antigens, mechanisms of virulence, or O111 polysaccharide subtypes. These antibodies were also able to increase the clearance of different strains of O111 E. coli by macrophages. PCR analyses of the pathways involved in O111 LPS core biosynthesis showed that all EAEC strains have core type R2, whereas typical EPEC and EHEC have core type R3. In contrast, atypical EPEC strains have core types R2 and R3. In summary, the results presented herein indicate that the O111 polysaccharide and LPS core types R2 and R3 are antigen targets for panspecific immunotherapy against all categories of O111 E. coli.