title: Subtractive hybridization and identification of putative adhesins in a Shiga toxin-producing eae-negative Escherichia coli.
abstract: Adherence to epithelial cells by specific adhesins is a characteristic of Shiga toxin-producing Escherichia coli (STEC) strains. The eae-encoded protein intimin is the main adhesin implicated in intestinal colonization in vivo. We recently showed that STEC strains isolated in Chile displayed a wide variety of adhesins; here we demonstrate that some of these STEC strains are eae-negative and still adhere to epithelial cells at a level 100-fold higher than enterohaemorrhagic E. coli (EHEC) O157 : H7 prototype strain EDL933. This phenotype is associated with the presence of adherence factors different from the intimin protein. Subtractive hybridization between EHEC EDL933 and STEC eae-negative strain 472-1 was used to identify regions implicated in adhesion. In addition to the saa gene, we identified 18 specific genes in STEC 472-1, 16 of which had nucleotide identity to Salmonella ST46 phage genes; the two remaining ones shared identity to a gene encoding a hypothetical protein of uropathogenic E. coli. The DNA sequence of the STEC 472-1 psu-int region identified five open reading frames with homology to phage genes. We constructed mutant strains in the saa gene and the psu-int region to study the participation of these genes in the adherence to epithelial cells and our results demonstrated that STECDeltasaa and STECDeltapsu-int mutants displayed a 10-fold decrease in adherence as compared to the STEC 472-1 wild-type strain. Overall, our results suggest that STEC strain 472-1 adheres to epithelial cells in an eae-independent matter and that saa and psu-int participate in this adhesion process.
authors.affiliation: Programa de Microbiología y Micologia, ICBM, Facultad de Medicina, Universidad de Chile, Santiago, Chile.
authors.forename: Maricel
authors.initials: M
authors.lastname: Vidal
authors.affiliation: Programa de Microbiología y Micologia, ICBM, Facultad de Medicina, Universidad de Chile, Santiago, Chile.
authors.forename: Valeria
authors.initials: V
authors.lastname: Prado
authors.affiliation: Department of Clinical Laboratory Sciences, University of Texas Medical Branch, Galveston, TX 77555-1070, USA.|Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX 77555-1070, USA.
authors.forename: Gregory C
authors.initials: GC
authors.lastname: Whitlock
authors.affiliation: Programa de Biologia Celular y Molecular, ICBM, Facultad de Medicina, Universidad de Chile, Santiago, Chile.
authors.forename: Aldo
authors.initials: A
authors.lastname: Solari
authors.affiliation: Department of Pathology and Sealy Center for Vaccine Development, University of Texas Medical Branch, Galveston, TX 77555-1070, USA.|Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX 77555-1070, USA.
authors.forename: Alfredo G
authors.initials: AG
authors.lastname: Torres
authors.affiliation: Programa de Microbiología y Micologia, ICBM, Facultad de Medicina, Universidad de Chile, Santiago, Chile.
authors.forename: Roberto M
authors.initials: RM
authors.lastname: Vidal
mesh_terms: D018829:Adhesins, Bacterial / Q000235:genetics / Q000378:metabolism*; D018830:Adhesins, Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001422:Bacterial Adhesion; D036022:Capsid Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D004847:Epithelial Cells / Q000382:microbiology; D019453:Escherichia coli O157 / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D009154:Mutation; D009693:Nucleic Acid Hybridization / Q000379:methods*; D054323:Shiga-Toxigenic Escherichia coli / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
chemical_list: D018829:Adhesins, Bacterial; D018830:Adhesins, Escherichia coli; D036022:Capsid Proteins; D029968:Escherichia coli Proteins; C439650:SAA protein, E coli; C074513:polarity suppression factor, bacteriophage P4; C073819:eaeA protein, E coli