The attaching and effacing (A/E) pathogens enterohemorrhagic Escherichia coli (EHEC) and enteropathogenic E. coli (EPEC) cause serious disease in humans resulting in profuse diarrhea and occasional death. These pathogens attach to the surfaces of intestinal epithelial cells and use a type-three secretion system (TTSS) to inject effector proteins which collapse (efface) the microvilli of the host cells and hijack normal host cellular processes. In vitro (cell culture) work has demonstrated that tight junctions (TJs) are one of the structures targeted by these pathogens. In order to test whether TJs are disrupted in vivo, the A/E pathogen, Citrobacter rodentium (CR), was used to infect mice as a small animal model. CR contains the major pathogenic effector proteins found in all A/E pathogens and colonizes the colon at high levels. Following infection, the TJ proteins claudins-1, -3 and -5 are redistributed from the cell membrane to the cell cytoplasm. In addition, molecular tracers injected into the lumen of murine colons following 7-day infection by wild-type (WT) CR penetrate the epithelium. These effects are dependent on the effector protein EspF but not Map. Morphologically, TJ regions appear smaller following WT CR infection when compared with TTSS or espF mutant bacterial infections. The effects on TJs require the intimate attachment of the bacteria to the host cells. Bacterially induced inflammation does not affect the localization of claudins-1, -3 or -5. This work was funded by the following agencies: HHMI, CAG/Aztra Zeneca/CIHR, MSFHR and The Killam Foundation.