Enteropathogenic Escherichia coli (EPEC) infection disrupts tight junctions (TJs) and perturbs intestinal barrier function in vitro. E. coli secreted protein F (EspF) is, in large part, responsible for these physiological and morphological alterations. We recently reported that the C57BL/6J mouse is a valid in vivo model of EPEC infection as EPEC colonizes the intestinal epithelium and effaces microvilli. Our current aim was to examine the effects of EPEC on TJ structure and barrier function of the mouse intestine and to determine the role of EspF in vivo. C57BL/6J mice were gavaged with approximately 2 x 10(8) EPEC organisms or PBS. At 1 or 5 days postinfection, mice were killed and ileal and colonic tissue was mounted in Ussing chambers to determine barrier function (measured as transepithelial resistance) and short circuit current. TJ structure was analyzed by immunofluorescence microscopy. Wild-type (WT) EPEC significantly diminished the barrier function of ileal and colonic mucosa at 1 and 5 days postinfection. Deficits in barrier function correlated with redistribution of occludin in both tissues. Infection with an EPEC strain deficient of EspF (delta espF) had no effect on barrier function at 1 day postinfection. Furthermore, delta espF had no effect on ileal TJ morphology and minor alterations of colonic TJ morphology at 1 day postinfection. In contrast, at 5 days postinfection, WT EPEC and delta espF had similar effects on barrier function and occludin localization. In both cases this was associated with immune activation, as demonstrated by increased mucosal tumor necrosis factor-alpha levels 5 days postinfection. In conclusion, these data demonstrate that WT EPEC infection of 6-8-week-old C57BL/6J mice (1) significantly decreases barrier function in the ileum and colon (2) redistributes occludin in the ileum and colon and (3) is dependent upon EspF to induce TJ barrier defects at early, but not late, times postinfection.