Molecular and Phenotypic Characterization of Escherichia coli O26:H8 among Diarrheagenic E. coli O26 Strains Isolated in Brazil

ABSTRACT Escherichia coli strains of serogroup O26 comprise two distinct groups of pathogens, characterized as enteropathogenic E. coli (EPEC) and enterohemorrhagic E. coli (EHEC). Among the several genes related to type III secretion system-secreted effector proteins, espK was found to be highly specific for EHEC O26:H11 and its stx-negative derivative strains isolated in European countries. E. coli O26 strains isolated in Brazil from infant diarrhea, foods, and the environment have consistently been shown to lack stx genes and are thus considered atypical EPEC. However, no further information related to their genetic background is known. Therefore, in this study, we aimed to discriminate and characterize these Brazilian O26 stx-negative strains by phenotypic, genetic, and biochemical approaches. Among 44 isolates confirmed to be O26 isolates, most displayed flagellar antigen H11 or H32. Out of the 13 nonmotile isolates, 2 tested positive for fliC H11, and 11 were fliC H8 positive. The identification of genetic markers showed that several O26:H11 and all O26:H8 strains tested positive for espK and could therefore be discriminated as EHEC derivatives. The presence of H8 among EHEC O26 and its stx-negative derivative isolates is described for the first time. The interaction of three isolates with polarized Caco-2 cells and with intestinal biopsy specimen fragments ex vivo confirmed the ability of the O26 strains analyzed to cause attaching-and-effacing (A/E) lesions. The O26:H32 strains, isolated mostly from meat, were considered nonvirulent. Knowledge of the virulence content of stx-negative O26 isolates within the same serotype helped to avoid misclassification of isolates, which certainly has important implications for public health surveillance.

[1]  L. Beutin,et al.  Virulence gene profiling of enterohemorrhagic (EHEC) and enteropathogenic (EPEC) Escherichia coli strains: a basis for molecular risk assessment of typical and atypical EPEC strains , 2011, BMC Microbiology.

[2]  L. Huicho,et al.  Phylogenetic relationships of Shiga toxin-producing Escherichia coli isolated from Peruvian children. , 2011, Journal of medical microbiology.

[3]  W. Elias,et al.  The ability of haemolysins expressed by atypical enteropathogenic Escherichia coli to bind to extracellular matrix components. , 2011, Memorias do Instituto Oswaldo Cruz.

[4]  L. Beutin,et al.  Identification of Genetic Markers for Differentiation of Shiga Toxin-Producing, Enteropathogenic, and Avirulent Strains of Escherichia coli O26 , 2011, Applied and Environmental Microbiology.

[5]  T. A. Gomes,et al.  Prevalence and Characteristics of the O122 Pathogenicity Island in Typical and Atypical Enteropathogenic Escherichia coli Strains , 2010, Journal of Clinical Microbiology.

[6]  L. Beutin,et al.  Evaluation of multiple-locus variable number of tandem-repeats analysis (MLVA) as a method for identification of clonal groups among enteropathogenic, enterohaemorrhagic and avirulent Escherichia coli O26 strains. , 2010, FEMS microbiology letters.

[7]  D. S. Horton,et al.  Differentiation of typical and atypical enteropathogenic Escherichia coli using colony immunoblot for detection of bundle‐forming pilus expression , 2009, Journal of applied microbiology.

[8]  J. Blanco,et al.  HeLa-cell adherence patterns and actin aggregation of enteropathogenic Escherichia coli (EPEC) and Shiga-toxin-producing E. coli (STEC) strains carrying different eae and tir alleles. , 2009, International microbiology : the official journal of the Spanish Society for Microbiology.

[9]  F. B. Andrade,et al.  Atypical enteropathogenic Escherichia coli genomic background allows the acquisition of non-EPEC virulence factors. , 2009, FEMS microbiology letters.

[10]  M. Morais,et al.  Evidence of Pathogenic Subgroups among Atypical Enteropathogenic Escherichia coli Strains , 2009, Journal of Clinical Microbiology.

[11]  J. Blanco,et al.  Virulence features of atypical enteropathogenic Escherichia coli identified by the eae(+) EAF-negative stx(-) genetic profile. , 2009, Diagnostic microbiology and infectious disease.

[12]  W. Elias,et al.  An overview of atypical enteropathogenic Escherichia coli. , 2009, FEMS microbiology letters.

[13]  J. Mainil,et al.  Putative Adhesins of Enteropathogenic and Enterohemorrhagic Escherichia coli of Serogroup O26 Isolated from Humans and Cattle , 2009, Journal of Clinical Microbiology.

[14]  C. Jenkins,et al.  Escherichia coli serogroup O26 – a new look at an old adversary , 2007, Journal of applied microbiology.

[15]  A. Mellmann,et al.  Shiga Toxin-Mediated Hemolytic Uremic Syndrome: Time to Change the Diagnostic Paradigm? , 2007, PloS one.

[16]  H. Karch,et al.  Presence of virulence and fitness gene modules of enterohemorrhagic Escherichia coli in atypical enteropathogenic Escherichia coli O26. , 2007, Microbes and infection.

[17]  A. Mellmann,et al.  Shiga Toxin Gene Loss and Transfer In Vitro and In Vivo during Enterohemorrhagic Escherichia coli O26 Infection in Humans , 2007, Applied and Environmental Microbiology.

[18]  C. Dozois,et al.  Autotransporter-Encoding Sequences Are Phylogenetically Distributed among Escherichia coli Clinical Isolates and Reference Strains , 2007, Applied and Environmental Microbiology.

[19]  A. Cravioto,et al.  An adhesive factor found in strains ofEscherichia coli belonging to the traditional infantile enteropathogenic serotypes , 1979, Current Microbiology.

[20]  J. Harel,et al.  Identification of Virulence Genes Linked with Diarrhea Due to Atypical Enteropathogenic Escherichia coli by DNA Microarray Analysis and PCR , 2006, Journal of Clinical Microbiology.

[21]  L. Beutin,et al.  Characterization of two major groups of diarrheagenic Escherichia coli O26 strains which are globally spread in human patients and domestic animals of different species. , 2005, FEMS microbiology letters.

[22]  A. Torres,et al.  Identification and Characterization of the Locus for Diffuse Adherence, Which Encodes a Novel Afimbrial Adhesin Found in Atypical Enteropathogenic Escherichia coli , 2005, Infection and Immunity.

[23]  H. Karch,et al.  Molecular Profiling and Phenotype Analysis of Escherichia coli O26:H11 and O26:NM: Secular and Geographic Consistency of Enterohemorrhagic and Enteropathogenic Isolates , 2005, Journal of Clinical Microbiology.

[24]  I. Bátai,et al.  Occurrence of hlyA and sheA Genes in Extraintestinal Escherichia coli Strains , 2005, Journal of Clinical Microbiology.

[25]  G. Frankel,et al.  Enteropathogenic Escherichia coli: unravelling pathogenesis. , 2005, FEMS microbiology reviews.

[26]  L. Beutin,et al.  Genetical and functional investigation of fliC genes encoding flagellar serotype H4 in wildtype strains of Escherichia coli and in a laboratory E. coli K-12 strain expressing flagellar antigen type H48 , 2005, BMC Microbiology.

[27]  M. Sinclair,et al.  Escherichia coli and Community-acquired Gastroenteritis, Melbourne, Australia , 2004, Emerging infectious diseases.

[28]  T. A. Gomes,et al.  Emerging Enteropathogenic Escherichia coli Strains? , 2004, Emerging infectious diseases.

[29]  Hung Bui,et al.  A specific genetic background is required for acquisition and expression of virulence factors in Escherichia coli. , 2004, Molecular biology and evolution.

[30]  T. A. Gomes,et al.  Virulence Properties and Characteristics of Shiga Toxin-Producing Escherichia coli in São Paulo, Brazil, from 1976 through 1999 , 2004, Journal of Clinical Microbiology.

[31]  A. Torres,et al.  Characterization of the second long polar (LP) fimbriae of Escherichia coli O157:H7 and distribution of LP fimbriae in other pathogenic E. coli strains. , 2004, FEMS microbiology letters.

[32]  C. Moreira-Filho,et al.  Atypical enteropathogenic Escherichia coli strains: phenotypic and genetic profiling reveals a strong association between enteroaggregative E. coli heat-stable enterotoxin and diarrhea. , 2003, The Journal of infectious diseases.

[33]  É. Oswald,et al.  Characterization of the Novel Factor Paa Involved in the Early Steps of the Adhesion Mechanism of Attaching and Effacing Escherichia coli , 2003, Infection and Immunity.

[34]  R. Robins-Browne,et al.  Identification of a Novel Fimbrial Gene Cluster Related to Long Polar Fimbriae in Locus of Enterocyte Effacement-Negative Strains of Enterohemorrhagic Escherichia coli , 2002, Infection and Immunity.

[35]  K. Irino,et al.  First Shiga Toxin-Producing Escherichia coli Isolate from a Patient with Hemolytic Uremic Syndrome, Brazil , 2002, Emerging infectious diseases.

[36]  Tetsuya Hayashi,et al.  toxB Gene on pO157 of Enterohemorrhagic EscherichiacoliO157:H7 Is Required for Full Epithelial Cell Adherence Phenotype , 2001, Infection and Immunity.

[37]  H. Karch,et al.  Identification and Characterization of a Novel Genomic Island Integrated at selC in Locus of Enterocyte Effacement-Negative, Shiga Toxin-Producing Escherichia coli , 2001, Infection and Immunity.

[38]  P. Srimanote,et al.  Characterization of Saa, a Novel Autoagglutinating Adhesin Produced by Locus of Enterocyte Effacement-Negative Shiga-ToxigenicEscherichia coli Strains That Are Virulent for Humans , 2001, Infection and Immunity.

[39]  C. Moreira-Filho,et al.  Genetic differences between Escherichia coli O26 strains isolated in Brazil and in other countries. , 2001, FEMS microbiology letters.

[40]  J. R. Andrade,et al.  Phenotypic and genotypic characteristics of Escherichia coli strains of non-enteropathogenic E. coli (EPEC) serogroups that carry EAE and lack the EPEC adherence factor and Shiga toxin DNA probe sequences. , 2001, The Journal of infectious diseases.

[41]  L. Trabulsi,et al.  10.321/eid0805.Typical and Atypical Enteropathogenic Escherichia coli , 2002, Emerging infectious diseases.

[42]  O. Clermont,et al.  Rapid and Simple Determination of theEscherichia coli Phylogenetic Group , 2000, Applied and Environmental Microbiology.

[43]  F. Grimont,et al.  Identification of Escherichia coli flagellar types by restriction of the amplified fliC gene. , 2000, Research in microbiology.

[44]  H. Karch,et al.  Molecular Analysis of H Antigens Reveals that Human DiarrheagenicEscherichia coli O26 Strains That Carry the eaeGene Belong to the H11 Clonal Complex , 2000, Journal of Clinical Microbiology.

[45]  T. R. Ward,et al.  Iha: a Novel Escherichia coli O157:H7 Adherence-Conferring Molecule Encoded on a Recently Acquired Chromosomal Island of Conserved Structure , 2000, Infection and Immunity.

[46]  R. Robins-Browne,et al.  Identification of a novel genetic locus that is required for in vitro adhesion of a clinical isolate of enterohaemorrhagic Escherichia coli to epithelial cells , 2000, Molecular microbiology.

[47]  H. Karch,et al.  Molecular characteristics and epidemiological significance of Shiga toxin-producing Escherichia coli O26 strains. , 2000, Journal of clinical microbiology.

[48]  H. Karch,et al.  A Genomic Island, Termed High-Pathogenicity Island, Is Present in Certain Non-O157 Shiga Toxin-Producing Escherichia coli Clonal Lineages , 1999, Infection and Immunity.

[49]  H. Karch,et al.  The large plasmids of Shiga-toxin-producing Escherichia coli (STEC) are highly variable genetic elements. , 1999, Microbiology.

[50]  J. Harel,et al.  Distribution of a novel locus called Paa (porcine attaching and effacing associated) among enteric Escherichia coli. , 1999, Advances in experimental medicine and biology.

[51]  V. Sperandio,et al.  Virulence properties of atypical EPEC strains. , 1999, Journal of medical microbiology.

[52]  G. Dougan,et al.  Enteropathogenic and enterohaemorrhagic Escherichia coli : more subversive elements , 1998, Molecular microbiology.

[53]  M. Nakazawa,et al.  Detection and sequences of the enteroaggregative Escherichia coli heat-stable enterotoxin 1 gene in enterotoxigenic E. coli strains isolated from piglets and calves with diarrhea , 1997, Journal of clinical microbiology.

[54]  H. O. Saridakis.  Non production of shiga - like toxins by escherichia coli serogroup o26 , 1994 .

[55]  T. Whittam,et al.  Clonal relationships among Escherichia coli strains that cause hemorrhagic colitis and infantile diarrhea , 1993, Infection and immunity.

[56]  L. Trabulsi,et al.  Attaching-effacing lesions and intracellular penetration in HeLa cells and human duodenal mucosa by two Escherichia coli strains not belonging to the classical enteropathogenic E. coli serogroups , 1993, Infection and immunity.

[57]  G. Schoolnik,et al.  Cloning and characterization of the bundle‐forming pilin gene of enteropathogenic Escherichia coli and its distribution in Salmonella serotypes , 1993, Molecular microbiology.

[58]  J. Yu,et al.  A genetic locus of enteropathogenic Escherichia coli necessary for the production of attaching and effacing lesions on tissue culture cells. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[59]  S. Moseley,et al.  Molecular characterization of a fimbrial adhesin, F1845, mediating diffuse adherence of diarrhea-associated Escherichia coli to HEp-2 cells , 1989, Journal of bacteriology.

[60]  J. Saunders,et al.  Interaction of enteropathogenic Escherichia coli 0111 with rabbit intestinal mucosa in vitro. , 1989, Gastroenterology.

[61]  M. Levine.  Escherichia coli that cause diarrhea: enterotoxigenic, enteropathogenic, enteroinvasive, enterohemorrhagic, and enteroadherent. , 1987, The Journal of infectious diseases.

[62]  M. Levine,et al.  Detection of an adherence factor of enteropathogenic Escherichia coli with a DNA probe. , 1985, The Journal of infectious diseases.

[63]  I. Scaletsky,et al.  Non production of cytotoxin among enteropathogenic strains of Escherichia coli isolated in Sao Paulo, Brasil , 1983 .