2, a gradual decrease in bacterial motility was clearly observed in the presence of increasing concentrations of BE. This result further verifies that BE specifically targets AI-2-mediated bacterial virulence pathways in E. coli O157:H7. To elucidate the effect of BE on an AI-3-mediated QS system, we examined whether the activation of ler promoter

by norepinephrine was also compromised by addition of BE. To address this question, mTOR inhibitor we created a green fluorescent protein (GFP) reporter strain, in which the gfp gene was transcribed by the ler promoter. As shown in Fig. 3, green fluorescence intensity was increased ∼1.37 fold by the addition of norepinephrine (second vs. third bar). The addition of BE, however, decreased the norepinephrine-stimulated production of GFP significantly (fourth vs. third bar). This result suggests

that BE can prevent the transcription of ler, regulated by AI-3-mediated QS system, from being activated and therefore may block a complex signaling cascade that regulates the expression of genes encoding proteins necessary click here for full virulence of E. coli O157:H7. Next, we tried to determine whether BE could attenuate the virulence of E. coli O157:H7 in vivo using C. elegans as a host. Caenorhabditis elegans is used as a simple and economic invertebrate animal model for the study of mechanisms of microbial pathogenesis (Nicholas & Hodgkin, 2004; Sifri et al., Morin Hydrate 2005). In particular, it was reported that C. elegans is a good model organism

to evaluate the virulence of E. coli O157:H7 and the antibacterial efficacy of many types of chemical compounds (Breger et al., 2007; Lee et al., 2008). As shown in Fig. 4, there were no significant differences in the survival rate of C. elegans for 2 days, but the survival rate of the nematodes fed on E. coli O157:H7 in the presence of 0.5% (v/v) of BE were significantly higher than those fed only on the pathogen for 3 days or more (Fig. 4). Notably, the survival rates of C. elegans fed on E. coli O157:H7 with 0% and 0.5% of BE after 8 days were 21.5% and 50%, respectively (Fig. 4). However, the survival rate of the nematodes fed on E. coli OP50, an avirulent strain routinely used as a nutrient source for C. elegans, was not affected by the presence of 0.5% BE (Fig. 4). These results suggest that BE can considerably protect the nematodes against a pathogenic attack by E. coli O157:H7, and thus, BE treatment can be developed as an agent to attenuate bacterial virulence in vivo. We then examined the effects of BE on the expression of virulence-associated genes by qRT-PCR. We analyzed the transcript levels of luxS and pfS, because these two genes are critically involved in AI-2 synthesis (Gonzalez Barrios et al., 2006). We also tested flhD and eae, which are involved in flagella regulation and type III secretion, respectively (Hughes et al., 2009). As shown in Fig.