Enterohaemorrhagic Escherichia coli O157 : H7 is a human pathogen that causes no apparent disease in cattle, its primary reservoir host. Recent research has demonstrated that E. coli O157 : H7 predominately colonizes the distal few centimetres of the bovine rectum, and in this study, the LEE4 operon encoding a type III secretion system translocon and associated proteins was shown to be essential for colonization. A deletion mutant of LEE4 failed to colonize cattle, in contrast to a co-inoculated strain containing a chromosomal complement of the operon, therefore fulfilling 'molecular' Koch's postulates for this virulence determinant. In addition, attaching and effacing (A/E) lesions were detectable in E. coli O157 : H7 microcolonies from the terminal rectum of both naturally and experimentally colonized cattle when examined by transmission electron microscopy. This study proves that type III secretion is required for colonization of cattle by E. coli O157 : H7, and that A/E lesion formation occurs at the bovine terminal rectum within E. coli O157 : H7 microcolonies. The research confirms the value of using type III secreted proteins as vaccine candidates in cattle. INTRODUCTIONEnterohaemorrhagic Escherichia coli (EHEC) has emerged in developed countries over the past 20 years as an important cause of human intestinal disease. In addition to bloody diarrhoea, intestinal infection can lead to potentially fatal systemic sequelae resulting from the activity of Shiga toxins. The majority of these infections in the USA, Canada, UK and Japan are caused by E. coli O157 : H7 (Nataro & Kaper, 1998). This serotype has been frequently isolated from cattle faeces, and most human E. coli O157 : H7 infections originate, either directly or indirectly, from this source (Besser et al., 1999;Borczyk et al., 1987). It is widely acknowledged that controlling E. coli O157 : H7 within the bovine population would be an effective method of reducing transmission to humans (Stevens et al., 2002).In common with other EHEC and EPEC (enteropathogenic E. coli), E. coli O157 : H7 contains a pathogenicity island, known as the locus of enterocyte effacement (LEE), that confers the attaching and effacing (A/E) phenotype McDaniel & Kaper, 1997). The LEE encodes a type III secretion system (TTSS) (Hueck, 1998), various translocators and effectors, the outer-membrane protein intimin (Jerse et al., 1990) and its receptor, termed Tir (translocated intimin receptor) (Kenny et al., 1997). The LEE is arranged into several polycistronic operons termed LEE1 to LEE5 (Elliott et al., 1998). The LEE4 operon encodes several proteins essential for the A/E phenotype. These include SepL (Kresse et al., 2000) and EscF (Wilson et al., 2001), both essential components of the LEE TTSS, and Esps (EPEC secreted proteins) A, B, D and F (Knutton et al., 1998;Taylor et al., 1998;Wachter et al., 1999; and the non-LEE-encoded Cif, EspI/NleA and TccP (Marches et al., 2003;Mundy et al., 2004;Gruenheid et al., 2004;Garmendia et al., 2004). Other than EspF, these are all likel...

Recent research has established that the terminal rectum is the predominant colonization site of enterohemorrhagic Escherichia coli O157:H7 in cattle. The main aim of the present work was to investigate pathological changes and associated immune responses at this site in animals colonized with E. coli O157:H7. Tissue and gastrointestinal samples from a total of 22 weaned Holstein-cross calves challenged with E. coli O157:H7 were analyzed for bacterial colonization and pathology. Five unexposed age-matched calves were used as comparative negative controls. E. coli O157:H7 bacteria induced histopathological alterations of the rectal mucosa with enterocyte remodeling. This was often associated with removal of the colonized epithelial layer. Immunogold labeling and transmission electron microscopy (TEM) showed E. coli O157 bacteria on pedestals, as part of attaching and effacing lesions. These pathological changes induced a local infiltration of neutrophils that was quantified as larger in infected animals. Rectal mucosal immunoglobulin A responses were detected against the E. coli O157:H7 antigen. This work presents evidence that E. coli O157:H7 is not a commensal bacteria in the bovine host and that the mucosal damage produced by E. coli O157:H7 colonization of the terminal rectum induces a quantifiable innate immune response and production of specific mucosal antibodies.Enterohemorrhagic Escherichia coli (EHEC) infection has emerged in the last 20 years as a cause of diarrhea that can lead to the more serious consequence of hemolytic-uremic syndrome and thrombotic microangiopathy. The majority of EHEC infections are caused by E. coli O157:H7 (24), and this serotype has been isolated frequently from cattle feces. Many human EHEC O157 infections originate, either directly or indirectly, from exposure to cattle feces (17), and a key step in protecting humans from EHEC infection is to understand and control E. coli O157:H7 colonization of cattle.Experimental challenges have suggested a variety of colonization sites in cattle (4,5,12). However, more recently, the terminal rectal mucosa has been identified as the major site of E. coli O157:H7 colonization (25), and this finding has been confirmed in slaughter animals (20). From an understanding of where E. coli O157:H7 colonizes the bovine intestinal tract, there is an opportunity to examine pathological changes at the site and to determine whether these changes correlate with the development of immunological responses. The main aim of the research is to underpin methods to control this pathogen in its main animal reservoir.A feature of E. coli O157:H7 infection is the formation of attaching and effacing (A/E) lesions, characterized by the elimination of the microvilli and intimate enterocyte attachment (7, 16). In vivo, A/E lesions are present at the terminal rectum of naturally and experimentally infected cattle, and inactivation of the type III secretion apparatus that is essential for this phenotype prevents E. coli O157:H7 colonization of cattle (27). The profound...