Characterization of Shiga Toxin-Producing Escherichia coli Strains Isolated from Human Patients in Germany over a 3-Year Period

ABSTRACT We have investigated 677 Shiga toxin-producing Escherichia coli (STEC) strains from humans to determine their serotypes, virulence genes, and clinical signs in patients. Six different Shiga toxin types (1, 1c, 2, 2c, 2d, and 2e) were distributed in the STEC strains. Intimin (eae) genes were present in 62.6% of the strains and subtyped into intimins α1, β1, γ1, ε, θ, and η. Shiga toxin types 1c and 2d were present only in eae-negative STEC strains, and type 2 was significantly (P < 0.001) more frequent in eae-positive STEC strains. Enterohemorrhagic E. coli hemolysin was associated with 96.2% of the eae-positive strains and with 65.2% of the eae-negative strains. Clinical signs in the patients were abdominal pain (8.7%), nonbloody diarrhea (59.2%), bloody diarrhea (14.3%), and hemolytic-uremic syndrome (HUS) (3.5%), and 14.3% of the patients had no signs of gastrointestinal disease or HUS. Infections with eae-positive STEC were significantly (P < 0.001) more frequent in children under 6 years of age than in other age groups, whereas eae-negative STEC infections dominated in adults. The STEC strains were grouped into 74 O:H types by serotyping and by PCR typing of the flagellar (fliC) genes in 221 nonmotile STEC strains. Eleven serotypes (O157:[H7], O26:[H11], O103:H2, O91:[H14], O111:[H8], O145:[H28], O128:H2, O113:[H4], O146:H21, O118:H16, and O76:[H19]) accounted for 69% of all STEC strains. We identified 41 STEC strains belonging to 31 serotypes which had not previously been described as human STEC. Twenty-six of these were positive for intimins α1 (one serotype), β1 (eight serotypes), ε (two serotypes), and η (three serotypes). Our study indicates that different types of STEC strains predominate in infant and adult patients and that new types of STEC strains are present among human isolates.

[1]  G. Duffy,et al.  Verocytotoxigenic E. coli. , 2004 .

[2]  Hideki Kobayashi,et al.  Prevalence and Characteristics of eae-Positive Escherichia coli from Healthy Cattle in Japan , 2003, Applied and Environmental Microbiology.

[3]  Gilbert S Chen,et al.  Shiga toxin-producing Escherichia coli in Montana: bacterial genotypes and clinical profiles. , 2003, The Journal of infectious diseases.

[4]  G. Dougan,et al.  Subtyping intimin genes from enteropathogenic Escherichia coli associated with outbreaks and sporadic cases in the United Kingdom and Eire. , 2003, Molecular and cellular probes.

[5]  L. Beutin,et al.  Animal host associated differences in Shiga toxin‐producing Escherichia coli isolated from sheep and cattle on the same farm , 2003, Journal of applied microbiology.

[6]  L. Beutin,et al.  HEp-2 Cell Adherence, Actin Aggregation, and Intimin Types of Attaching and Effacing Escherichia coli Strains Isolated from Healthy Infants in Germany and Australia , 2003, Infection and Immunity.

[7]  A. Friedrich,et al.  Shiga Toxin 1c-Producing Escherichia coli Strains: Phenotypic and Genetic Characterization and Association with Human Disease , 2003, Journal of Clinical Microbiology.

[8]  M. Blanco,et al.  Prevalence and characterization of Vero cytotoxin-producing Escherichia coli isolated from diarrhoeic and healthy sheep and goats , 2003, Epidemiology and Infection.

[9]  J. Blanco,et al.  Verotoxin-Producing Escherichia coli in Spain: Prevalence, Serotypes, and Virulence Genes of O157:H7 and Non-O157 VTEC in Ruminants, Raw Beef Products, and Humans , 2003, Experimental biology and medicine.

[10]  J. Blanco,et al.  Serotypes, Virulence Genes, and Intimin Types of Shiga Toxin (Verotoxin)-Producing Escherichia coli Isolates from Healthy Sheep in Spain , 2003, Journal of Clinical Microbiology.

[11]  L. Wieler,et al.  Description of a Novel Intimin Variant (Type ζ) in the Bovine O84:NM Verotoxin-Producing Escherichia coli Strain 537/89 and the Diagnostic Value of Intimin Typing , 2003, Experimental biology and medicine.

[12]  S. Djordjevic,et al.  stx1c Is the Most Common Shiga Toxin 1 Subtype among Shiga Toxin-Producing Escherichia coli Isolates from Sheep but Not among Isolates from Cattle , 2003, Journal of Clinical Microbiology.

[13]  T. Pysher,et al.  Response to Shiga toxin 1 and 2 in a baboon model of hemolytic uremic syndrome , 2003, Pediatric Nephrology.

[14]  Teresa M. Large,et al.  Molecular Characterization of a Serotype O121:H19 Clone, a Distinct Shiga Toxin-Producing Clone of Pathogenic Escherichia coli , 2002, Infection and Immunity.

[15]  G. Dougan,et al.  An eight‐month study of a population of verocytotoxigenic Escherichia coli (VTEC) in a Scottish cattle herd , 2002, Journal of applied microbiology.

[16]  L. Beutin,et al.  Serotypes and virulence factors of Shiga toxin‐producing Escherichia coli isolated from healthy Norwegian sheep , 2002, Journal of applied microbiology.

[17]  S. Djordjevic,et al.  Virulence Properties and Serotypes of Shiga Toxin-Producing Escherichia coli from Healthy Australian Cattle , 2002, Applied and Environmental Microbiology.

[18]  A. Siitonen,et al.  Clinical Escherichia coli Strains Carrying stx Genes: stx Variants and stx-Positive Virulence Profiles , 2002, Journal of Clinical Microbiology.

[19]  L. Beutin,et al.  Genetic Diversity of Intimin Genes of Attaching and Effacing Escherichia coli Strains , 2002, Journal of Clinical Microbiology.

[20]  L. Beutin,et al.  A Multiresistant Clone of Shiga Toxin-Producing Escherichia coli O118:[H16] Is Spread in Cattle and Humans over Different European Countries , 2002, Applied and Environmental Microbiology.

[21]  P. Srimanote,et al.  Characterization of a Novel Type IV Pilus Locus Encoded on the Large Plasmid of Locus of Enterocyte Effacement-Negative Shiga-Toxigenic Escherichia coli Strains That Are Virulent for Humans , 2002, Infection and Immunity.

[22]  H. Karch,et al.  Identification, Characterization, and Distribution of a Shiga Toxin 1 Gene Variant (stx1c) in Escherichia coli Strains Isolated from Humans , 2002, Journal of Clinical Microbiology.

[23]  K. Frydendahl Prevalence of serogroups and virulence genes in Escherichia coli associated with postweaning diarrhoea and edema disease in pigs and a comparison of diagnostic approaches. , 2002, Veterinary microbiology.

[24]  T. Whittam,et al.  Molecular Evolution of the Intimin Gene in O111 Clones of Pathogenic Escherichia coli , 2002, Journal of bacteriology.

[25]  A. Friedrich,et al.  Escherichia coli harboring Shiga toxin 2 gene variants: frequency and association with clinical symptoms. , 2002, The Journal of infectious diseases.

[26]  L. Beutin,et al.  Evaluation of the VTEC-Screen "Seiken" test for detection of different types of Shiga toxin (verotoxin)-producing Escherichia coli (STEC) in human stool samples. , 2002, Diagnostic microbiology and infectious disease.

[27]  L. Beutin,et al.  Isolation of a Lysogenic Bacteriophage Carrying thestx1OX3 Gene, Which Is Closely Associated with Shiga Toxin-Producing Escherichia coli Strains from Sheep and Humans , 2001, Journal of Clinical Microbiology.

[28]  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.

[29]  R. Amils,et al.  Association between intimin (eae) and EspB gene subtypes in attaching and effacing Escherichia coli strains isolated from diarrhoeic lambs and goat kids. , 2001, Microbiology.

[30]  N. Pradel,et al.  Heterogeneity of Shiga Toxin-Producing Escherichia coli Strains Isolated from Hemolytic-Uremic Syndrome Patients, Cattle, and Food Samples in Central France , 2001, Applied and Environmental Microbiology.

[31]  S. Djordjevic,et al.  Virulence Properties and Serotypes of Shiga Toxin-Producing Escherichia coli from Healthy Australian Slaughter-Age Sheep , 2001, Journal of Clinical Microbiology.

[32]  S. Djordjevic,et al.  The Common Ovine Shiga Toxin 2-ContainingEscherichia coli Serotypes and Human Isolates of the Same Serotypes Possess a Stx2d Toxin Type , 2001, Journal of Clinical Microbiology.

[33]  E. Boedeker,et al.  Complete Nucleotide Sequence and Analysis of the Locus of Enterocyte Effacement from Rabbit Diarrheagenic Escherichia coli RDEC-1 , 2001, Infection and Immunity.

[34]  T. Whittam,et al.  Pathogenesis and evolution of virulence in enteropathogenic and enterohemorrhagic Escherichia coli. , 2001, The Journal of clinical investigation.

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

[36]  G. Dougan,et al.  Enterohaemorrhagic Escherichia coliO157:H7 target Peyer's patches in humans and cause attaching/effacing lesions in both human and bovine intestine , 2000, Gut.

[37]  L. Beutin,et al.  Investigation of human infections with verocytotoxin-producing strains of Escherichia coli (VTEC) belonging to serogroup O118 with evidence for zoonotic transmission , 2000, Epidemiology and Infection.

[38]  B. Finlay,et al.  Exploitation of host cells by enteropathogenic Escherichia coli. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[39]  F. Scheutz,et al.  Prevalence and Characterization of Shiga Toxin-Producing Escherichia coli Isolated from Cattle, Food, and Children during a One-Year Prospective Study in France , 2000, Journal of Clinical Microbiology.

[40]  A. Caprioli,et al.  A New Shiga Toxin 2 Variant (Stx2f) fromEscherichia coli Isolated from Pigeons , 2000, Applied and Environmental Microbiology.

[41]  A. O’Brien,et al.  Elastase in Intestinal Mucus Enhances the Cytotoxicity of Shiga Toxin Type 2d* , 2000, The Journal of Biological Chemistry.

[42]  A. Caprioli,et al.  Typing of Intimin Genes in Human and Animal Enterohemorrhagic and Enteropathogenic Escherichia coli: Characterization of a New Intimin Variant , 2000, Infection and Immunity.

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

[44]  S. McEwen,et al.  Associations between Virulence Factors of Shiga Toxin-ProducingEscherichia coli and Disease in Humans , 1999, Journal of Clinical Microbiology.

[45]  J. Meng,et al.  Virulence genes of Shiga toxin-producing Escherichia coli isolated from food, animals and humans. , 1998, International journal of food microbiology.

[46]  A. Siitonen,et al.  Shiga Toxin-Producing Escherichia coliin Finland from 1990 through 1997: Prevalence and Characteristics of Isolates , 1998, Journal of Clinical Microbiology.

[47]  S. Lauwers,et al.  Identification of New Verocytotoxin Type 2 Variant B-Subunit Genes in Human and Animal Escherichia coliIsolates , 1998, Journal of Clinical Microbiology.

[48]  Kim Ziebell,et al.  Association of Enterohemorrhagic Escherichia coliHemolysin with Serotypes of Shiga-Like-Toxin-ProducingEscherichia coli of Human and Bovine Origins , 1998, Applied and Environmental Microbiology.

[49]  L. Beutin,et al.  Human infections with Shiga toxin-producing Escherichia coli other than serogroup O157 in Germany. , 1998, Emerging infectious diseases.

[50]  Nicole T. Perna,et al.  Molecular Evolution of a Pathogenicity Island from Enterohemorrhagic Escherichia coli O157:H7 , 1998, Infection and Immunity.

[51]  I. Carle,et al.  Comparison of 14 PCR systems for the detection and subtyping of stx genes in Shiga-toxin-producing Escherichia coli. , 1998, Research in microbiology.

[52]  B. Souweine,et al.  Non-O157:H7 Stx2-Producing Escherichia coli Strains Associated with Sporadic Cases of Hemolytic-Uremic Syndrome in Adults , 1998, Journal of Clinical Microbiology.

[53]  G. Dougan,et al.  Detection of Intimins α, β, γ, and δ, Four Intimin Derivatives Expressed by Attaching and Effacing Microbial Pathogens , 1998, Journal of Clinical Microbiology.

[54]  A. Caprioli,et al.  Enteroaggregative, Shiga Toxin-ProducingEscherichia coli O111:H2 Associated with an Outbreak of Hemolytic-Uremic Syndrome , 1998, Journal of Clinical Microbiology.

[55]  G. Dougan,et al.  Effect of Prior Experimental Human EnteropathogenicEscherichia coli Infection on Illness following Homologous and Heterologous Rechallenge , 1998, Infection and Immunity.

[56]  K. Ludwig,et al.  Pathomechanisms in the haemolytic-uraemic syndrome. , 1998, Nephrology, dialysis, transplantation : official publication of the European Dialysis and Transplant Association - European Renal Association.

[57]  L. Beutin,et al.  Epidemiological relatedness and clonal types of natural populations of Escherichia coli strains producing Shiga toxins in separate populations of cattle and sheep , 1997, Applied and environmental microbiology.

[58]  A. Siitonen,et al.  Shiga toxin-producing Escherichia coli serotype OX3:H21 as a cause of hemolytic-uremic syndrome. , 1997, Clinical infectious diseases : an official publication of the Infectious Diseases Society of America.

[59]  P. Hatfield,et al.  Characterization of Escherichia coli O157:H7 and other Shiga toxin-producing E. coli serotypes isolated from sheep , 1997, Journal of clinical microbiology.

[60]  L. Beutin,et al.  Rapid detection and isolation of shiga-like toxin (verocytotoxin)-producing Escherichia coli by direct testing of individual enterohemolytic colonies from washed sheep blood agar plates in the VTEC-RPLA assay , 1996, Journal of clinical microbiology.

[61]  C. Lingwood,et al.  Interaction of verotoxin 2e with pig intestine , 1996, Infection and immunity.

[62]  A. O’Brien,et al.  Activation of Shiga-like toxins by mouse and human intestinal mucus correlates with virulence of enterohemorrhagic Escherichia coli O91:H21 isolates in orally infected, streptomycin-treated mice , 1996, Infection and immunity.

[63]  A. Caprioli,et al.  Genotyping of Shiga-like toxin genes in non-O157 Escherichia coli strains associated with haemolytic uraemic syndrome. , 1995, Journal of medical microbiology.

[64]  L. Beutin,et al.  Virulence markers of Shiga-like toxin-producing Escherichia coli strains originating from healthy domestic animals of different species , 1995, Journal of clinical microbiology.

[65]  L. Beutin,et al.  Molecular analysis of the plasmid-encoded hemolysin of Escherichia coli O157:H7 strain EDL 933 , 1995, Infection and immunity.

[66]  J. Heesemann,et al.  Development of PCR for screening of enteroaggregative Escherichia coli , 1995, Journal of clinical microbiology.

[67]  T. McDaniel,et al.  A genetic locus of enterocyte effacement conserved among diverse enterobacterial pathogens. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[68]  L. Beutin,et al.  Heterogeneity of the amino-acid sequences of Escherichia coli Shiga-like toxin type-I operons. , 1995, Gene.

[69]  W. L. Payne,et al.  Simultaneous identification of strains of Escherichia coli serotype O157:H7 and their Shiga-like toxin type by mismatch amplification mutation assay-multiplex PCR , 1995, Journal of clinical microbiology.

[70]  L. Beutin,et al.  The large-sized plasmids of enterohemorrhagic Escherichia coli O157 strains encode hemolysins which are presumably members of the E. coli alpha-hemolysin family. , 1994, FEMS microbiology letters.

[71]  P. Manning,et al.  Cloning and nucleotide sequence of a variant Shiga-like toxin II gene from Escherichia coli OX3:H21 isolated from a case of sudden infant death syndrome. , 1992, Microbial pathogenesis.

[72]  M. McKee,et al.  Two copies of Shiga-like toxin II-related genes common in enterohemorrhagic Escherichia coli strains are responsible for the antigenic heterogeneity of the O157:H- strain E32511 , 1991, Infection and immunity.

[73]  M. Levine,et al.  A sensitive and specific DNA probe to identify enteroaggregative Escherichia coli, a recently discovered diarrheal pathogen. , 1990, The Journal of infectious diseases.

[74]  V. Gannon,et al.  Molecular cloning and nucleotide sequence of another variant of the Escherichia coli Shiga-like toxin II family. , 1990, Journal of general microbiology.

[75]  Y. Takeda,et al.  Cloning and nucleotide sequencing of Vero toxin 2 variant genes from Escherichia coli O91:H21 isolated from a patient with the hemolytic uremic syndrome. , 1990, Microbial pathogenesis.

[76]  M. P. Jackson,et al.  Cloning and sequencing of a Shiga-like toxin type II variant from Escherichia coli strain responsible for edema disease of swine , 1988, Journal of bacteriology.

[77]  M. Levine,et al.  A DNA probe to identify enterohemorrhagic Escherichia coli of O157:H7 and other serotypes that cause hemorrhagic colitis and hemolytic uremic syndrome. , 1987, The Journal of infectious diseases.

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

[79]  B. Rowe,et al.  Cloning of genes determining the production of vero cytotoxin by Escherichia coli. , 1985, Journal of general microbiology.

[80]  B. Levin,et al.  Genetic diversity in relation to serotype in Escherichia coli , 1985, Infection and immunity.

[81]  I. Ørskov,et al.  2 Serotyping of Escherichia coli , 1984 .

[82]  J. Wells,et al.  Hemorrhagic colitis associated with a rare Escherichia coli serotype. , 1983, The New England journal of medicine.

[83]  K. Bettelheim,et al.  CYTOTOXIC ESCHERICHIA COLI SEROTYPES , 1980, The Lancet.

[84]  N. Day,et al.  CYTOTOXIC ENTEROPATHOGENIC ESCHERICHIA COLI , 1980, The Lancet.