The mechanisms used by Shiga toxin (Stx)-producing Escherichia coli to adhere to epithelial cells are incompletely understood. Two cosmids from an E. coli O157:H7 DNA library contain an adherence-conferring chromosomal gene encoding a protein similar to iron-regulated gene A (IrgA) of Vibrio cholerae (M. B. Goldberg, S. A. Boyko, J. R. Butterton, J. A. Stoebner, S. M. Payne, and S. B. Calderwood, Mol. Microbiol. 6:2407-2418, 1992). We have termed the product of this gene the IrgA homologue adhesin (Iha), which is encoded by iha. Iha is 67 kDa in E. coli O157:H7 and 78 kDa in laboratory E. coli and is structurally unlike other known adhesins. DNA adjacent to iha contains tellurite resistance loci and is conserved in structure in distantly related pathogenic E. coli, but it is absent from nontoxigenic E. coli O55:H7, sorbitol-fermenting Stx-producing E. coli O157:H؊, and laboratory E. coli. We have termed this region the tellurite resistance-and adherence-conferring island. We conclude that Iha is a novel bacterial adherence-conferring protein and is contained within an E. coli chromosomal island of conserved structure. Pathogenic E. coli O157:H7 has only recently acquired this island.Escherichia coli O157:H7 and other Shiga toxin (Stx)-producing E. coli (STEC) strains cause diarrhea, hemorrhagic colitis, and the hemolytic uremic syndrome. The mechanisms underlying the adherence of STEC to epithelial cells are only partly understood (35). The ability to adhere to epithelial cells is an important virulence trait, because adherence presumably enables enteric pathogens to deliver toxins efficiently to host organs, overcome peristaltic clearance, and gain access to hostderived nutrients.Intimin is the best-characterized E. coli O157:H7 adherence molecule. Encoded by eae, intimin mediates the attaching and effacing lesion caused by enteropathogenic E. coli (EPEC) and many STEC serotypes (21) and is an important component of pathogenicity. However, cloned eae from EPEC and STEC do not confer the adherent phenotype upon laboratory E. coli (18,25,28). Moreover, though the cloned EPEC locus of enterocyte effacement, which includes eae, does confer the adherence phenotype on E. coli K-12 (27), the cloned E. coli O157:H7 locus of enterocyte effacement does not (12).We describe an E. coli O157:H7 gene that renders laboratory E. coli adherent to epithelial cells and explore evolutionary aspects of its acquisition.(These data were presented in part at the 3rd International Symposium and Workshop on Shiga Toxin-Producing Escherichia coli Infections, Baltimore, Md., 22 to 26 June 1997.) MATERIALS AND METHODSBacteria. The bacteria analyzed in this study are described in Table 1. The bacteria were inoculated directly from frozen stock (in Luria-Bertani [LB] broth-15% glycerol, maintained at Ϫ70°C) into LB broth (26). The cultures were grown overnight under standardized conditions (37°C; 14 to 16 h; stationary cultures) for adherence assays and protein preparations. The bacteria were grown in a shaking incubator (37°C; 14 to 24 h) f...

A fimbrial adhesin, designated F1845, was found to be responsible for the diffuse HEp-2 cell adherence of a diarrheal Escherichia coli isolate. The genetic determinant of F1845 was cloned, and the order of the genes necessary for production of F1845 was determined by maxicell analysis. Five polypeptides with apparent sizes of 10, 95, 27, 15.5, and 14.3 kilodaltons (kDa) were found to be encoded in that order by the F1845 determinant. The nucleotide sequence of the 14.3-kDa subunit gene was determined and found to share extensive homology in its signal sequence with the gene encoding the structural subunit of the AFA-1 hemagglutinin of a uropathogenic E. coli strain (A. Labigne-Roussel, M.A. Schmidt, W. Walz, and S. Falkow, J. Bacteriol. 162:1285-1292, 1985) but not in the region encoding the mature protein. Southern blot hybridizations indicated that the F1845 determinants are of chromosomal origin. Hybridization studies using a probe from the region encoding the 95-kDa polypeptide indicated that related sequences may be plasmid associated in some strains and chromosomal in others. Additional hybridization studies of E. coli isolates possessing sequence homology to the F1845 determinant suggest that the sequences in the 5' region of the F1845 structural subunit gene are more highly conserved than sequences in the 3' region.

Two novel putative Escherichia coli virulence genes, iha and iroN from E. coli (iroN E. coli ), were detected in 55 and 39%, respectively, of 67 E. coli isolates from patients with urosepsis. iha and iroN E. coli exhibited divergent associations with other putative virulence genes, phylogenetic markers, host characteristics, and antimicrobial resistance.The virulent strains of Escherichia coli that cause urinary tract infections (UTIs) and other extraintestinal infections in humans (i.e., extraintestinal pathogenic E. coli [ExPEC]) owe their pathogenic potential largely to the presence of specialized virulence factors (VFs) which are absent from commensal members of the species and which allow ExPEC strains to colonize host mucosal surfaces, injure and invade host tissues, foil host defense mechanisms, and incite an injurious host inflammatory response (4,5,9,20). Currently recognized putative VFs (PVFs) of ExPEC include adhesins, siderophores, toxins, protectins, and invasins, some of which are encoded on pathogenicity-associated islands (PAIs) (2,7,8,17,24,26). PAIs are large blocks of established or suspected virulence genes that are inserted into the E. coli genome (often at tRNA loci) and which may provide a mechanisms for coordinate horizontal transfer of virulence genes between lineages within E. coli and even between species (3,7,21,24,25).Two recently described PAI-linked PVF genes of ExPEC, iha and iroN from E. coli (iroN E. coli ), are of interest both in their own right and because of their potential utility as markers for their respective PAIs of origin (23,27). iha, an novel nonhemagglutinating adhesin which in vitro confers HeLa cell adherence capability to (nonadhering) E. coli K-12, was first identified as part of a tellurite resistance-associated PAI (termed TAI, the tellurite resistance-adherence-conferring island) from an E. coli O157:H7 isolate from a patient with hemorrhagic colitis (27). iha exhibits nearly perfect sequence identity with the sequenced portion of an open reading frame (ORF) of unknown function (ORF "R4") from a PAI in archetypal ExPEC strain CFT073 (8,17). In addition to the iha homologue and multiple other ORFs of unknown significance, this PAI from strain CFT073 also contains an hly (hemolysin) operon and one of the strain's two pap operons (named for pilus associated with pyelonephritis; P fimbriae), which includes the F7-2 allele of papA (the P fimbrial structural subunit gene) and allele II of papG (the P fimbrial digalactoside-specific adhesin gene) (8,16,17). Consistent with the hypothesis that iha is a PVF in ExPEC, probes derived from PAI regions immediately adjacent to iha as it occurs in strain CFT073 hybridized significantly more frequent with UTI or bacteremia isolates of E. coli than with commensal E. coli (8).iroN E. coli , a novel catechole siderophore receptor which exhibits increased expression in urine, was recently identified in archetypal ExPEC strain CP9 as part of a PAI which also includes one of this strain's two pap operons (i.e., the pap operon con...

The transcriptional organization of the gene cluster encoding the F1845 fimbrial adhesin of a diarrhoea-associated Escherichia coli was investigated. Genes daaA to daaE were determined to constitute a single transcriptional unit under the control of the daaA promoter. The nucleotide sequence of daaA and that of an upstream open reading frame encoded on the opposite strand, designated daaF, were determined to share limited homology with the papB and papI genes of the P fimbrial adhesin, respectively. The 5' termini of the daaF and daaABCDE transcripts were mapped by primer extension and nuclease protection analyses. The promoters for these transcripts were associated with potential regulatory sequences including two consensus leucine-responsive regulatory protein (Lrp)-binding sites which contained differentially methylated GATC sequences, a cAMP-CRP-binding site, and an integration host factor (IHF)-binding site. Expression of the daa locus was determined to be dependent on Lrp, subject to catabolite repression, and dependent on IHF.