Adherence of enterohemorrhagic Escherichia coli (EHEC) to the intestinal epithelium is critical for initiation of a bacterial infection. An in vitro infection study previously indicated that EHEC bacteria initially adhere diffusely and then proliferate to develop MC, a process that is mediated by various secreted proteins, such as EspA, EspB, EspD, Tir, and intimin, as well as other putative adherence factors. In the present study, we investigated the role of a large 93-kb plasmid (pO157) in the adherence of O157:H7 (O157Sakai) and found the toxB gene to be involved in the full adherence phenotype. A pO157-cured strain of O157Sakai (O157Cu) developed microcolonies on Caco-2 cells; however, the number of microcolonies was lower than that of O157Sakai, as were the production and secretion levels of EspA, EspB, and Tir. Introduction of a mini-pO157 plasmid (pIC37) composed of the toxB and ori regions restored full adherence capacity to O157Cu, including production and secretion of the proteins. In contrast, introduction of a pO157 mutant possessing toxB::Km into O157Cu could not restore the full adherence phenotype. Expression of truncated versions of His-tagged ToxB also promoted EspB production and/or secretion by O157Cu. These results suggest that ToxB contributes to the adherence of EHEC to epithelial cells through promotion of the production and/or secretion of type III secreted proteins.

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Helicobacter pylori is associated with gastric disorders in humans and some experimental animals, and possesses the luxS/type 2 autoinducer (AI-2) system. The effects of a specific luxS mutation on the characteristics of H. pylori were examined. On 0?3 % agar medium, motility of H. pylori HPKY08 (luxS : : cat) was significantly lower than that of wild-type H. pylori TK1402. The luxS-complemented strain HPKY21 exhibited motility comparable to that of H. pylori TK1402. It was shown that the luxS/AI-2 system plays an important role in H. pylori motility. The luxS mutant exhibited a reduced infection rate relative to the wild-type parent strain TK1402 in a Mongolian gerbil model. At 3 months after oral inoculation, lower numbers of H. pylori were detected by semi-quantitative real-time reverse transcription PCR (qRT-PCR) in luxS " mutant-infected gerbils than in TK1402-infected gerbils. Gastric inflammation and increased antibody titre for H. pylori were observed in TK1402-infected gerbils only. INTRODUCTIONQuorum sensing (QS) is a cell-to-cell communication system that regulates bacterial phenotypes, including the expression of virulence factor genes. The signalling molecules are known as autoinducers (AIs), and when these molecules reach a critical threshold concentration within a bacterial population, a signal transduction cascade is triggered, and this forms the basis for alterations in gene expression (Fuqua et al., 1994). Many Gram-negative bacteria utilize N-acylhomoserine lactone molecules, AI-1, as signals, while Gram-positive bacteria actively export peptides as signalling molecules. There is a second signalling system involved in a wide range of bacterial species (Schauder et al., 2001), and this system is utilized by both Gram-positive and Gram-negative bacteria. The signalling molecule known as type 2 autoinducer (AI-2) is a furanosyl borate diester (Chen et al., 2002), and the enzyme responsible for its synthesis is encoded by the luxS gene (Surette & Bassler, 1999). The genomes of many bacterial species, notably Escherichia coli, Salmonella enterica serovar Typhimurium, Shigella flexneri, Proteus mirabilis, Vibrio cholerae, Vibrio vulnificus, Campylobacter jejuni, Porphyromonas gingivalis, Bacillus subtilis, Streptococcus pyogenes, Streptococcus mutans, Clostridium perfringens and Clostridium difficile, include luxS homologues. In several of these, luxS-related AI-2 signals are involved in bacterial characteristics such as biofilm formation (Balestrino et al., 2005;Blehert et al., 2003;Fong et al., 2001;Wen & Burne 2004), flagella and motility (Jeon et al., 2003;Schneider et al., 2002;Stroeher et al., 2003), type III secretion systems (Sperandio et al., 1999), toxin production (Ohtani et al., 2002) and virulence (Lyon et al., 2001; Parsonnet et al., 1991;Stroeher et al., 2003).Helicobacter pylori has been identified as the aetiological agent of chronic active gastritis, peptic ulcer disease (Blaser, 1992;Graham, 1989), gastric adenocarcinoma (Parsonnet et al., 1991) and mucosal-associated lymphoid t...

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 has been considered as an agent responsible for outbreak of hemorrhagic colitis and the hemolytic uremic syndrome. We examined the effect of the probiotic agent Clostridium butyricum MIYAIRI strain 588 on EHEC O157:H7 infections in vitro and in vivo using gnotobiotic mice. The growth of EHEC O157:H7 and the production of Shiga-like toxins in broth cultures were inhibited by co-incubation with C. butyricum. The antibacterial effects of butyric and lactic acid were demonstrated in a dose-dependent manner. In addition, the inhibitory effect of butyric acid on the viability of EHEC was demonstrated not only at low pH, but also at neutral pH adjusted to 7.0. Flowcytometric analysis showed that pre-incubation of Caco-2 cells with C. butyricum and E. coli K12 inhibited the adhesion of EHEC O157:H7. However, the effect of C. butyricum on adhesion of EHEC to Caco-2 cells was more inhibitory than that of E. coli K12. Gnotobiotic mice mono-associated with EHEC O157:H7 died within 4-7 days after the infection. On the other hand, all gnotobiotic mice prophylactically pre-treated with C. butyricum survived exposure to EHEC O157:H7 and of the gnotobiotic mice therapeutically post-treated with C. butyricum, 50% survived. Both counts of EHEC O157:H7 and the amounts of shiga-like toxins (Stx1 and Stx2) in fecal contents of gnotobiotic mice di-associated with EHEC O157:H7 and C. butyricum were less than those of gnotobiotic mice mono-associated with EHEC O157:H7. These results indicated that the probiotic bacterium C. butyricum MIYAIRI strain 588 has preventive and therapeutic effects on EHEC O157:H7 infection in gnotobiotic mice.

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