Enterohaemorrhagic Escherichia coli (EHEC) O157 : H7 inhibits interferon (IFN)-c-stimulated tyrosine phosphorylation of signal transducer and activator of transcription (STAT)-1 in epithelial cells. We determined the effects of probiotics on EHEC-mediated disruption of IFN-c-stimulated STAT-1 activation in epithelial cell lines. Confluent Intestine 407, HEp-2 and Caco-2 epithelial cells were pre-treated (3 h) with either probiotics or surface-layer proteins derived from Lactobacillus helveticus R0052 prior to infection with EHEC O157 : H7 strain CL56 (m.o.i. 100 : 1, 6 h, 37 6C in 5 % CO 2 ). Subsequently, cells were washed and stimulated with human recombinant IFN-c (50 ng ml "1 , 0.5 h, 37 6C) followed by whole-cell protein extraction and immunoblotting for tyrosine-phosphorylated STAT-1. Relative to uninfected cells, STAT-1-activation was reduced after EHEC O157 : H7 infection. Pre-incubation with the probiotic L. helveticus R0052 followed by EHEC infection abrogated pathogen-mediated disruption of IFNc-STAT-1 signalling. As determined using Transwell inserts, probiotic-mediated protection was independent of epithelial cell contact. In contrast, pre-incubation with boiled L. helveticus R0052, an equal concentration of viable Lactobacillus rhamnosus R0011, or surface-layer proteins (0.14 mg ml "1 ) did not restore STAT-1 signalling in EHEC-infected cells. The viable probiotic agent L. helveticus R0052 prevented EHEC O157 : H7-mediated subversion of epithelial cell signal transduction responses. INTRODUCTIONOutbreaks of enterohaemorrhagic Escherichia coli (EHEC) serotype O157 : H7 infection occur frequently across the developed world (Serna & Boedeker, 2008). This foodborne enteric pathogen is acquired from contaminated foodstuffs and non-chlorinated drinking water (Karch et al., 2005;Maki, 2006;Rangel et al., 2005). In the severest cases of infection (~10-15 % of cases), the haemolytic uraemic syndrome develops as a systemic complication, leading to patient hospitalization, kidney failure and death (Serna & Boedeker, 2008;Tarr et al., 2005). Renal disease results from bacterial cytotoxin [Shiga-like toxin (Stx)-1 and -2] production in the gut, release into the systemic circulation and damage to vascular endothelial cells in the renal glomerulus (Blackall & Marques, 2004). EHEC pathogenesis is also attributed to intimate bacterial adhesion conferred by the locus of enterocyte effacement (LEE) pathogenicity island, the activity of effector proteins secreted through a type three secretion system (T3SS), and pathogen modulation of host cell signal transduction cascades (Bhavsar et al., 2007;Kaper et al., 2004).EHEC disease pathogenesis includes subversion of epithelial cell signal transduction cascades involved in host innate immune responses to infection (Bhavsar et al., 2007). For instance, EHEC inhibits interferon (IFN)-c-stimulated tyrosine phosphorylation of signal transducer and activator of transcription-1 (STAT-1) in multiple cell lines (Ceponis et al., 2003), which is mediated by a bacterially encoded fac...