The Locus of Enterocyte Effacement Pathogenicity Island of Shiga Toxin-Producing Escherichia coli O157: H7 and Other Attaching and Effacing E. coli

Escherichia coli O157:H7 is responsible for a human toxico-infection that can lead to severe complications such as hemolytic uremic syndrome. Inside the intestine, E. coli O157:H7 forms typical attaching-effacing lesions and produces Shiga toxins. The genes that are responsible for these lesions are located in a pathogenicity island called the locus of enterocyte effacement (LEE). LEE gene expression is influenced by quorum sensing through the luxS system. In this study, the effect of glucose on the expression of several genes from LEE, on the expression of Shiga toxin genes, and on the expression of luxS was assessed with real-time, reverse transcription PCR. All concentrations of glucose (from 0.1 to 1%) were able to down-regulate genes from the LEE operon. A slight down-regulation of genes implicated in Shiga toxin expression was also observed but was significant for low doses of glucose (0.1 to 0.5%) only. A slight but significant increase in luxS expression was observed with 1% glucose. This confirms that in addition to quorum sensing, the presence or absence of nutrients such as glucose has an impact on the down- or upregulation of LEE-encoded virulence genes by the bacterium. The influence of glucose on the virulence of E. coli O157:H7 has received little attention, and these results suggest that glucose can have an important effect on the virulence of E. coli O157:H7.

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The Gene tia, Harbored by the Subtilase-Encoding Pathogenicity Island, Is Involved in the Ability of Locus of Enterocyte Effacement-Negative Shiga Toxin-Producing Escherichia coli Strains To Invade Monolayers of Epithelial Cells

Infection and Immunity ◽

10.1128/iai.00613-17 ◽

2017 ◽

Vol 85 (12) ◽

Cited By ~ 6

Author(s):

Roslen Bondì ◽

Paola Chiani ◽

Valeria Michelacci ◽

Fabio Minelli ◽

Alfredo Caprioli ◽

...

Keyword(s):

Escherichia Coli ◽

Epithelial Cells ◽

Shiga Toxin ◽

Cultured Cells ◽

Pathogenicity Island ◽

Content Type ◽

Cell Monolayers ◽

E Coli ◽

Enterocyte Effacement ◽

K 12

ABSTRACT Locus of enterocyte effacement (LEE)-negative Shiga toxin (Stx)-producing Escherichia coli (STEC) strains are human pathogens that lack the LEE locus, a pathogenicity island (PAI) involved in the intimate adhesion of LEE-positive strains to the host gut epithelium. The mechanism used by LEE-negative STEC strains to colonize the host intestinal mucosa is still not clear. The cell invasion determinant tia, previously described in enterotoxigenic E. coli strains, has been identified in LEE-negative STEC strains that possess the subtilase-encoding pathogenicity island (SE-PAI). We evaluated the role of the gene tia, present in these LEE-negative STEC strains, in the invasion of monolayers of cultured cells. We observed that these strains were able to invade Caco-2 and HEp-2 cell monolayers and compared their invasion ability with that of a mutant strain in which the gene tia had been inactivated. Mutation of the gene tia resulted in a strong reduction of the invasive phenotype, and complementation of the tia mutation with a functional copy of the gene restored the invasion activity. Moreover, we show that the gene tia is overexpressed in bacteria actively invading cell monolayers, demonstrating that tia is involved in the ability to invade cultured monolayers of epithelial cells shown by SE-PAI-positive E. coli, including STEC, strains. However, the expression of the tia gene in the E. coli K-12 strain JM109 was not sufficient, in its own right, to confer to this strain the ability to invade cell monolayers, suggesting that at least another factor must be involved in the invasion ability displayed by the SE-PAI-positive strains.

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STEC-EPEC Oligonucleotide Microarray: A New Tool for Typing Genetic Variants of the LEE Pathogenicity Island of Human and Animal Shiga Toxin–Producing Escherichia coli (STEC) and Enteropathogenic E. coli (EPEC) Strains

Clinical Chemistry ◽

10.1373/clinchem.2005.059766 ◽

2006 ◽

Vol 52 (2) ◽

pp. 192-201 ◽

Cited By ~ 46

Author(s):

Patricia Garrido ◽

Miguel Blanco ◽

Mercedes Moreno-Paz ◽

Carlos Briones ◽

Ghizlane Dahbi ◽

...

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Genetic Variants ◽

Pathogenicity Island ◽

Oligonucleotide Microarray ◽

Chromosomal Dna ◽

Emerging Pathogens ◽

E Coli ◽

Enterocyte Effacement ◽

Gene 4

Abstract Background: Shiga toxin–producing Escherichia coli (STEC) and enteropathogenic E. coli (EPEC) are important emerging pathogens that can cause a severe and sometimes fatal illness. Differentiation of eae, tir, espA, espD, and espB gene variants of the locus of enterocyte effacement (LEE) pathogenicity island represents an important tool for typing in routine diagnostics as well as in pathogenesis, epidemiologic, clonal, and immunologic studies. Methods: Type-specific oligonucleotide microarrays and a PCR scheme were designed and constructed for the detection and typing of genetic variants of the LEE genes. Oligonucleotide probes were tested for their specificity against the corresponding type strain by microarray hybridization using fluorescent DNA, either PCR-amplified (single, multiplex, long-range), chromosomal, or amplified chromosomal DNA. Results: The PCR scheme and the oligonucleotide microarray allowed us to distinguish 16 variants (α1, α2, β1, β2, γ1, γ2/θ, δ/κ, ε, ζ, η, ι, λ, μ, ν, ξ, ο) of the eae gene, 4 variants (α1, β1, γ1, γ2/θ) of the tir gene, 4 variants (α1, β1, β2, γ1) of the espA gene, 3 variants (α1, β1, γ1) of the espB gene, and 3 variants (α1, β1, γ1) of the espD gene. We found a total of 12 different combinations of tir, espA, espB, and espD genes among the 25 typed strains. Conclusions: The PCR scheme and the oligonucleotide microarray described are effective tools to rapidly screen multiple virulence genes and their variants in E. coli strains isolated from human and animal infections. The results demonstrate the great genetic diversity among LEE genes of human and animal STEC and EPEC strains.

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Molecular Evolution of a Pathogenicity Island from Enterohemorrhagic Escherichia coli O157:H7

Infection and Immunity ◽

10.1128/iai.66.8.3810-3817.1998 ◽

1998 ◽

Vol 66 (8) ◽

pp. 3810-3817 ◽

Cited By ~ 279

Author(s):

Nicole T. Perna ◽

George F. Mayhew ◽

György Pósfai ◽

Simon Elliott ◽

Michael S. Donnenberg ◽

...

Keyword(s):

Escherichia Coli ◽

Pathogenicity Island ◽

Enterohemorrhagic Escherichia Coli ◽

Open Reading Frames ◽

Escherichia Coli O157 ◽

E Coli ◽

History Of ◽

Enterocyte Effacement ◽

Targeting Vector ◽

Reading Frames

ABSTRACT We report the complete 43,359-bp sequence of the locus of enterocyte effacement (LEE) from EDL933, an enterohemorrhagicEscherichia coli O157:H7 serovar originally isolated from contaminated hamburger implicated in an outbreak of hemorrhagic colitis. The locus was isolated from the EDL933 chromosome with a homologous-recombination-driven targeting vector. Recent completion of the LEE sequence from enteropathogenic E. coli (EPEC) E2348/69 afforded the opportunity for a comparative analysis of the entire pathogenicity island. We have identified a total of 54 open reading frames in the EDL933 LEE. Of these, 13 fall within a putative P4 family prophage designated 933L. The prophage is not present in E2348/69 but is found in a closely related EPEC O55:H7 serovar and other O157:H7 isolates. The remaining 41 genes are shared by the two complete LEEs, and we describe the nature and extent of variation among the two strains for each gene. The rate of divergence is heterogeneous along the locus. Most genes show greater than 95% identity between the two strains, but other genes vary more than expected for clonal divergence among E. coli strains. Several of these highly divergent genes encode proteins that are known to be involved in interactions with the host cell. This pattern suggests recombinational divergence coupled with natural selection and has implications for our understanding of the interaction of both pathogens with their host, for the emergence of O157:H7, and for the evolutionary history of pathogens in general.

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Characterization of Escherichia coli O157:H7 and other Shiga toxin-producing E. coli serotypes isolated from sheep.

Journal of Clinical Microbiology ◽

10.1128/jcm.35.4.892-899.1997 ◽

1997 ◽

Vol 35 (4) ◽

pp. 892-899 ◽

Cited By ~ 90

Author(s):

I T Kudva ◽

P G Hatfield ◽

C J Hovde

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Escherichia Coli O157 ◽

E Coli

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Comparison between human and animal isolates of Shiga toxin-producing Escherichia coli O157 from Australia

Epidemiology and Infection ◽

10.1017/s0950268801006689 ◽

2002 ◽

Vol 128 (3) ◽

pp. 357-362 ◽

Cited By ~ 17

Author(s):

N. FEGAN ◽

P. DESMARCHELIER

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Human Infection ◽

Pulsed Field Gel Electrophoresis ◽

Toxin Gene ◽

Escherichia Coli O157 ◽

Animal Population ◽

E Coli ◽

Virulence Markers ◽

Animal Isolates

There is very little human disease associated with enterohaemorrhagic Escherichia coli O157 in Australia even though these organisms are present in the animal population. A group of Australian isolates of E. coli O157:H7 and O157:H- from human and animal sources were tested for the presence of virulence markers and compared by XbaI DNA macrorestriction analysis using pulsed-field gel electrophoresis (PFGE). Each of 102 isolates tested contained the gene eae which encodes the E. coli attaching and effacing factor and all but one carried the enterohaemolysin gene, ehxA, found on the EHEC plasmid. The most common Shiga toxin gene carried was stx2c, either alone (16%) or in combination with stx1 (74%) or stx2 (3%). PFGE grouped the isolates based on H serotype and some clusters were source specific. Australian E. coli O157:H7 and H- isolates from human, animal and meat sources carry all the virulence markers associated with EHEC disease in humans therefore other factors must be responsible for the low rates of human infection in Australia.

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Outbreak of Shiga Toxin–Producing Escherichia coli (STEC) O104:H4 Infection in Germany Causes a Paradigm Shift with Regard to Human Pathogenicity of STEC Strains

Journal of Food Protection ◽

10.4315/0362-028x.jfp-11-452 ◽

2012 ◽

Vol 75 (2) ◽

pp. 408-418 ◽

Cited By ~ 151

Author(s):

LOTHAR BEUTIN ◽

ANNETT MARTIN

Keyword(s):

Escherichia Coli ◽

Hemolytic Uremic Syndrome ◽

Shiga Toxin ◽

Outbreak Strain ◽

Fenugreek Seeds ◽

E Coli ◽

Aggregative Adherence ◽

Uremic Syndrome ◽

Enterocyte Effacement ◽

The Way

An outbreak that comprised 3,842 cases of human infections with enteroaggregative hemorrhagic Escherichia coli (EAHEC) O104:H4 occurred in Germany in May 2011. The high proportion of adults affected in this outbreak and the unusually high number of patients that developed hemolytic uremic syndrome makes this outbreak the most dramatic since enterohemorrhagic E. coli (EHEC) strains were first identified as agents of human disease. The characteristics of the outbreak strain, the way it spread among humans, and the clinical signs resulting from EAHEC infections have changed the way Shiga toxin–producing E. coli strains are regarded as human pathogens in general. EAHEC O104:H4 is an emerging E. coli pathotype that is endemic in Central Africa and has spread to Europe and Asia. EAHEC strains have evolved from enteroaggregative E. coli by uptake of a Shiga toxin 2a (Stx2a)–encoding bacteriophage. Except for Stx2a, no other EHEC-specific virulence markers including the locus of enterocyte effacement are present in EAHEC strains. EAHEC O104:H4 colonizes humans through aggregative adherence fimbrial pili encoded by the enteroaggregative E. coli plasmid. The aggregative adherence fimbrial colonization mechanism substitutes for the locus of enterocyte effacement functions for bacterial adherence and delivery of Stx2a into the human intestine, resulting clinically in hemolytic uremic syndrome. Humans are the only known natural reservoir known for EAHEC. In contrast, Shiga toxin–producing E. coli and EHEC are associated with animals as natural hosts. Contaminated sprouted fenugreek seeds were suspected as the primary vehicle of transmission of the EAHEC O104:H4 outbreak strain in Germany. During the outbreak, secondary transmission (human to human and human to food) was important. Epidemiological investigations revealed fenugreek seeds as the source of entry of EAHEC O104:H4 into the food chain; however, microbiological analysis of seeds for this pathogen produced negative results. The survival of EAHEC in seeds and the frequency of human carriers of EAHEC should be investigated for a better understanding of EAHEC transmission routes.

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Prevalence of shiga toxin-producing Escherichia coli O157, O26 and O111 in milk, meat and faeces of cattle, sheep and pigs slaughtered in Benin

Journal of Applied Biosciences ◽

10.35759/jabs.152.6 ◽

2020 ◽

Vol 152 ◽

pp. 15667-15675

Author(s):

Chakirath Folakè Arikè Salifou ◽

Cyrille Boko ◽

Isidore Houaga ◽

Raoul Agossa ◽

Isabelle Ogbankotan ◽

...

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Virulence Genes ◽

Animal Origin ◽

Escherichia Coli O157 ◽

E Coli ◽

Milk Samples ◽

Food Of Animal Origin ◽

Cattle Faeces ◽

Risk Assessment And Management

Objectives: The study aimed to search for E. coli O157 and non-O157 in milk, meat and faeces of cattle, sheep and pigs slaughtered in Cotonou. Methodology and Results: One hundred and Seventy-Five (175) samples including 25 meat, 25 faeces per species and 25 milk from cattle were analysed for E. coli O157; O26 and O111 and the virulence genes were identified by PCR. The SAS software (1998) and the bilateral Z test were used to calculate and compare the identification frequencies. E. coli O157 was identified in 4% of cattle faeces, 4% of sheep faeces, and 20% of beef and, in 20% of milk samples. E. coli O26 was identified in 12% of cattle faeces and, in 8% of beef samples. E. coli O111 was identified at frequencies of 8%, and 12% in faeces of sheep and pigs, respectively. The eae gene was detected in 4% of beef, ovine meat, milk, pig faeces and in sheep faeces. stx1 was detected in 8% of milk, and in 4% of bovine and sheep faeces. The strains possessing the gene were all of E. coli O157 with the exception of one from pig faeces identified as O111. Conclusions and application of findings: The presence of these serogroups of E. coli with virulence genes poses a real food safety problem in Benin. This study findings must be taken into account for risk assessment and management related to the consumption of food of animal origin. Keywords: Benin, E. coli O157, O26, O111, faeces, meat, milk

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Simultaneous Enrichment of Shiga Toxin–Producing Escherichia coli O157 and O26 and Salmonella in Food Samples Using Universal Preenrichment Broth

Journal of Food Protection ◽

10.4315/0362-028x-72.10.2065 ◽

2009 ◽

Vol 72 (10) ◽

pp. 2065-2070 ◽

Cited By ~ 8

Author(s):

MASASHI KANKI ◽

KAZUKO SETO ◽

JUNKO SAKATA ◽

TETSUYA HARADA ◽

YUKO KUMEDA

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Food Samples ◽

Escherichia Coli O157 ◽

E Coli ◽

Significant Difference ◽

Peptone Water ◽

Radish Sprouts ◽

Pork Samples ◽

Better Than

Universal preenrichment broth (UPB) was compared with modified Escherichia coli broth with novobiocin (mEC+n) for enrichment of Shiga toxin–producing E. coli O157 and O26, and with buffered peptone water (BPW) for preenrichment of Salmonella enterica. Ten strains each of the three pathogens were inoculated into beef and radish sprouts following thermal, freezing, or no treatment. With regard to O157 and O26, UPB incubated at 42°C recovered significantly more cells from inoculated beef than UPB at 35°C and from radish sprout samples than UPB at 35°C and mEC+n. With regard to Salmonella, UPB incubated at 42°C was as effective as UPB at 35°C and BPW at recovering cells from beef and radish sprout samples. No significant difference was noted between the effectiveness of UPB at 42°C and UPB at 35°C or BPW in the recovery of Salmonella from 205 naturally contaminated poultry samples. By using UPB at 42°C, one O157:H7 strain was isolated from the mixed offal of 53 beef samples, 6 cattle offal samples, and 50 pork samples all contaminated naturally, with no pathogen inoculation. The present study found that UPB incubated at 42°C was as effective as, or better than, mEC+n for enrichment of O157 and O26 and comparable to BPW for preenrichment of Salmonella. These findings suggest that a great deal of labor, time, samples, and space may be saved if O157, O26, and Salmonella are enriched simultaneously with UPB at 42°C.

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Molecular analysis of Escherichia coli O157 strains isolated from cattle and pigs by the use of PCR and pulsed-field gel electrophoresis methods

Veterinární Medicína ◽

10.17221/5819-vetmed ◽

2012 ◽

Vol 47 (No. 6) ◽

pp. 149-158 ◽

Cited By ~ 15

Author(s):

J. Osek ◽

P. Gallien

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Genetic Relatedness ◽

Rflp Analysis ◽

Toxin Gene ◽

Escherichia Coli O157 ◽

Chromosomal Dna ◽

E Coli ◽

Shiga Toxin Genes ◽

Porcine Origin

Fourteen Escherichia coli O157 strains isolated from cattle and pigs in Poland and in Germany were investigated, using PCR, for the genetic markers associated with Shiga toxin-producing E. coli (STEC). Only two strains, both of cattle origin, were positive for the fliC (H7) gene and could be classified as O157 : H7. Nine isolates had stx shiga toxin genes, either stx1 (1 strain), stx2 (4 isolates) or both (4 strains). The stx2-carrying samples were further subtyped by PCR for the stx2c, stx2d, and stx2e toxin variants. It was shown that all but one stx2-positive bacteria possessed the stx2c Shiga toxin gene type and one stx2 STEC isolate had the stx2d virulence factor sub-type. The eaeA (intimin) gene was found in 9 strains (8 isolates from cattle and one strain from pigs); all of them harboured the genetic marker characteristic of the gamma intimin variant. The translocated intimin receptor (tir) gene was detected in 7 isolates tested and among them only one tir-positive strain was recovered from pigs. The ehly E. coli enterohemolysin gene was amplified in all but one strains obtained from cattle and only in one isolate of porcine origin. The genetic relatedness of the analysed E. coli O157 strains was examined by restriction fragment length polymorphism (RFLP) of chromosomal DNA digested with XbaI. Two distinct but related RFLP pattern clusters were observed: one with 9 strains (8 isolates of bovine origin and one strain obtained from pigs) and the other one comprises the remaining 5 E. coli isolates (4 of porcine origin and one strain recovered from cattle). The results suggest that pigs, besides cattle, may be a reservoir of E. coli O157 strains potentially pathogenic to humans. Moreover, epidemiologically unrelated isolates of the O157 serogroup, recovered from different animal species, showed a clonal relationship as demonstrated by the RFLP analysis.

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