scholarly journals espC Pathogenicity Island of Enteropathogenic Escherichia coli Encodes an Enterotoxin

2001 ◽  
Vol 69 (1) ◽  
pp. 315-324 ◽  
Author(s):  
Jay L. Mellies ◽  
Fernando Navarro-Garcia ◽  
Iruka Okeke ◽  
Julie Frederickson ◽  
James P. Nataro ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Developing Countries ◽  
Type Iii Secretion ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Pathogenicity Island ◽  
Enteropathogenic Escherichia Coli ◽  
Ussing Chambers ◽  
E Coli ◽  
Enterocyte Effacement

ABSTRACT At least five proteins are secreted extracellularly by enteropathogenic Escherichia coli (EPEC), a leading cause of infant diarrhea in developing countries. However only one, EspC, is known to be secreted independently of the type III secretion apparatus encoded by genes located within the 35.6-kb locus of enterocyte effacement pathogenicity island. EspC is a member of the autotransporter family of proteins, and the secreted portion of the molecule is 110 kDa. Here we determine that the espC gene is located within a second EPEC pathogenicity island at 60 min on the chromosome of E. coli. We also show that EspC is an enterotoxin, indicated by rises in short-circuit current and potential difference in rat jejunal tissue mounted in Ussing chambers. In addition, preincubation with antiserum against the homologous Pet enterotoxin of enteroaggregative E. coli eliminated EspC enterotoxin activity. Like the EAF plasmid, the espCpathogenicity island was found only in a subset of EPEC, suggesting that EspC may play a role as an accessory virulence factor in some but not all EPEC strains.

scholarly journals Quorum-Sensing Escherichia coli Regulator A: a Regulator of the LysR Family Involved in the Regulation of the Locus of Enterocyte Effacement Pathogenicity Island in Enterohemorrhagic E. coli

2002 ◽  
Vol 70 (6) ◽  
pp. 3085-3093 ◽  
Author(s):  
Vanessa Sperandio ◽  
Caiyi C. Li ◽  
James B. Kaper
Keyword(s):  
Escherichia Coli ◽  
Quorum Sensing ◽  
Type Iii Secretion ◽  
Pathogenicity Island ◽  
The Other ◽  
E Coli ◽  
Type Iii ◽  
Enterocyte Effacement ◽  
K 12 ◽  
Lysr Family

ABSTRACT The locus of enterocyte effacement (LEE) is a chromosomal pathogenicity island that encodes the proteins involved in the formation of the attaching and effacing lesions by enterohemorrhagic Escherichia coli (EHEC) and enteropathogenic E. coli (EPEC). The LEE comprises 41 open reading frames organized in five major operons, LEE1, LEE2, LEE3, tir (LEE5), and LEE4, which encode a type III secretion system, the intimin adhesin, the translocated intimin receptor (Tir), and other effector proteins. The first gene of LEE1 encodes the Ler regulator, which activates all the other genes within the LEE. We previously reported that the LEE genes were activated by quorum sensing through Ler (V. Sperandio, J. L. Mellies, W. Nguyen, S. Shin, and J. B. Kaper, Proc. Natl. Acad. Sci. USA 96:15196-15201, 1999). In this study we report that a putative regulator in the E. coli genome is itself activated by quorum sensing. This regulator is encoded by open reading frame b3243; belongs to the LysR family of regulators; is present in EHEC, EPEC, and E. coli K-12; and shares homology with the AphB and PtxR regulators of Vibrio cholerae and Pseudomonas aeruginosa, respectively. We confirmed the activation of b3243 by quorum sensing by using transcriptional fusions and renamed this regulator quorum-sensing E. coli regulator A (QseA). We observed that QseA activated transcription of ler and therefore of the other LEE genes. An EHEC qseA mutant had a striking reduction of type III secretion activity, which was complemented when qseA was provided in trans. Similar results were also observed with a qseA mutant of EPEC. The QseA regulator is part of the regulatory cascade that regulates EHEC and EPEC virulence genes by quorum sensing.

scholarly journals Cytoskeletal Effects Induced by Pet, the Serine Protease Enterotoxin of Enteroaggregative Escherichia coli

1999 ◽  
Vol 67 (5) ◽  
pp. 2184-2192 ◽  
Author(s):  
Fernando Navarro-García ◽  
Cynthia Sears ◽  
Carlos Eslava ◽  
Alejandro Cravioto ◽  
James P. Nataro
Keyword(s):  
Escherichia Coli ◽  
Serine Protease ◽  
Protease Activity ◽  
Short Circuit ◽  
Ussing Chamber ◽  
Short Circuit Current ◽  
T84 Cells ◽  
Ussing Chambers ◽  
E Coli ◽  
Cytopathic Effects

ABSTRACT We have previously described enteroaggregative Escherichia coli (EAEC) strains that induce cytotoxic effects on T84 cells, ligated rat ileal loops, and human intestine in culture. Such strains secrete a 104-kDa protein termed Pet (for plasmid-encoded toxin). We have also shown previously that the Pet toxin induces rises in short-circuit current and decreases the electrical resistance in rat jejunum mounted in an Ussing chamber. The nucleotide sequence of thepet gene revealed that Pet is a member of the autotransporter class of secreted proteins. Here we show that a concentrated supernatant of E. coli HB101 harboring the minimal pet clone pCEFN1 induces temperature-, time- and dose-dependent cytopathic effects on HEp-2 cells and HT29 C1 cells in culture. The effects were characterized by release of the cellular focal contacts from the glass substratum, followed by complete rounding of the cells and detachment from the glass. Staining of the Pet-treated cells with Live/Dead viability stain revealed that >90% of rounded cells were viable. Pet-intoxicated HEp-2 and HT29 cells stained with fluorescein-labeled phalloidin revealed contraction of the cytoskeleton and loss of actin stress fibers. However, the effects of Pet were not inhibited by cytoskeleton-altering drugs, including colchicine, taxol, cytochalasin D, and phallicidin. The Pet protein induced proteolysis in zymogram gels, and preincubation with the serine protease inhibitor phenylmethylsulfonyl fluoride resulted in complete abrogation of Pet cytopathic effects. We introduced a mutation in a predicted catalytic serine residue and found that the mutant (Pet S260I) was deficient in protease activity and did not produce cytopathic effects, cytoskeletal damage, or enterotoxic effects in Ussing chambers. These data suggest that Pet is a cytoskeleton-altering toxin and that its protease activity is involved in each of the observed phenotypes.

Distribution of espI among clinical enterohaemorrhagic and enteropathogenic Escherichia coli isolates

2004 ◽  
Vol 53 (11) ◽  
pp. 1145-1149 ◽  
Author(s):  
Rosanna Mundy ◽  
Claire Jenkins ◽  
Jun Yu ◽  
Henry Smith ◽  
Gad Frankel
Keyword(s):  
Escherichia Coli ◽  
Type Iii Secretion ◽  
Severe Disease ◽  
Pathogenicity Island ◽  
Effector Proteins ◽  
Effector Protein ◽  
Enteropathogenic Escherichia Coli ◽  
Type Iii Effector ◽  
Type Iii

Enterohaemorrhagic (EHEC) and enteropathogenic (EPEC) Escherichia coli are important diarrhoeagenic pathogens; infection is dependent on translocation of a number of type III effector proteins. Until recently all the known effectors were encoded on the LEE pathogenicity island, which also encodes the adhesin intimin and the type III secretion apparatus. Recently, a novel non-LEE effector protein, EspI/NleA, which is required for full virulence in vivo and is encoded on a prophage, was identified. The aim of this study was to determine the distribution of espI among clinical EHEC and EPEC isolates. espI was detected in 86 % and 53 % of LEE+ EHEC and EPEC strains, respectively. Moreover, the espI gene was more commonly found in patients suffering from a more severe disease.

scholarly journals Virulence of Enteropathogenic Escherichia coli, a Global Pathogen

2003 ◽  
Vol 16 (3) ◽  
pp. 365-378 ◽  
Author(s):  
S. C. Clarke ◽  
R. D. Haigh ◽  
P. P. E. Freestone ◽  
P. H. Williams
Keyword(s):  
Escherichia Coli ◽  
Host Cell ◽  
Type Iii Secretion ◽  
Pathogenic Bacteria ◽  
Diarrheal Disease ◽  
Lesion Site ◽  
Intimate Contact ◽  
E Coli ◽  
Enterocyte Effacement ◽  
Broad Overview

SUMMARY Enteropathogenic Escherichia coli (EPEC) remains an important cause of diarrheal disease worldwide. Research into EPEC is intense and provides a good virulence model of other E. coli infections as well as other pathogenic bacteria. Although the virulence mechanisms are now better understood, they are extremely complex and much remains to be learnt. The pathogenesis of EPEC depends on the formation of an ultrastructural lesion in which the bacteria make intimate contact with the host apical enterocyte membrane. The formation of this lesion is a consequence of the ability of EPEC to adhere in a localized manner to the host cell, aided by bundle-forming pili. Tyrosine phosphorylation and signal transduction events occur within the host cell at the lesion site, leading to a disruption of the host cell mechanisms and, consequently, to diarrhea. These result from the action of highly regulated EPEC secreted proteins which are released via a type III secretion system, many genes of which are located within a pathogenicity island known as the locus of enterocyte effacement. Over the last few years, dramatic increases in our knowledge of EPEC virulence have taken place. This review therefore aims to provide a broad overview of and update to the virulence aspects of EPEC.

scholarly journals 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

2017 ◽  
Vol 85 (12) ◽  
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.

Characterization of the synergistic interaction of Escherichia coli heat-stable toxin and carbachol

1991 ◽  
Vol 261 (4) ◽  
pp. G592-G601 ◽  
Author(s):  
S. A. Levine ◽  
M. Donowitz ◽  
A. J. Watson ◽  
G. W. Sharp ◽  
J. K. Crane ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Short Circuit ◽  
Second Messengers ◽  
Short Circuit Current ◽  
Ussing Chambers ◽  
Simultaneous Addition ◽  
Heat Stable ◽  
Cholinergic Agent ◽  
Synergistic Response ◽  
Intracellular Mediators

STa, the heat-stable enterotoxin of Escherichia coli, is a specific activator of membrane-bound guanylyl cyclase and stimulates secretion of Cl- in a human colonic carcinoma cell line (T84). We investigated the effect of the cholinergic agent carbachol on the secretory response to STa. T84 cell monolayers were studied under voltage-clamped conditions in modified Ussing chambers. Simultaneous addition of STa and carbachol resulted in a biphasic synergistic response characterized by a brief peak in short-circuit current (Isc) followed by a prolonged plateau phase lasting up to 90 min. A synergistic response was also seen with sequential addition of the agonists, and was altered by the order and timing of agonist addition. Pretreatment with STa enhanced the synergistic response to carbachol, while the reverse order of additions produced synergy only when STa was added during or immediately after the Isc response to carbachol. Synergy occurred only with a concentration of STa sufficient to produce an Isc response alone. However, a concentration of carbachol that caused neither an increase in Isc nor intracellular Ca2+ mobilization was sufficient to evoke a synergistic response. Addition of 8-bromoguanosine 3',5'-cyclic monophosphate also produced a synergistic Isc response with carbachol, although maximal synergism was seen with simultaneous addition. Augmentation of the intracellular Ca2+ response to carbachol by STa is not the mechanism of synergy. Although the mechanism of synergy is not understood, these studies suggest that STa-induced cGMP interacts with other second messengers to produce the synergistic response, and that multiple intracellular mediators may influence the ability of STa to cause disease.

scholarly journals 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

2006 ◽  
Vol 52 (2) ◽  
pp. 192-201 ◽  
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.

scholarly journals Regulators Encoded in the Escherichia coli Type III Secretion System 2 Gene Cluster Influence Expression of Genes within the Locus for Enterocyte Effacement in Enterohemorrhagic E. coli O157:H7

2004 ◽  
Vol 72 (12) ◽  
pp. 7282-7293 ◽  
Author(s):  
Lihong Zhang ◽  
Roy R. Chaudhuri ◽  
Chrystala Constantinidou ◽  
Jon L. Hobman ◽  
Mala D. Patel ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Gene Cluster ◽  
Type Iii Secretion ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Regulatory Genes ◽  
E Coli ◽  
Type Iii ◽  
System 2 ◽  
Enterocyte Effacement

ABSTRACT Enterohemorrhagic Escherichia coli (EHEC) O157:H7 subverts host cells through a type III secretion system encoded by the locus for enterocyte effacement (LEE). Genome sequencing of this pathotype revealed the existence of a gene cluster encoding components of a second cryptic type III secretion system, E. coli type III secretion system 2 (ETT2). Recently, we showed that the ETT2 gene cluster is present in whole or in part in the majority of E. coli strains but is unable to encode a functional secretion system in most strains, including EHEC O157:H7. However, here we show that mutational inhibition of two regulatory genes (ECs3720 or etrA and ECs3734 or eivF) from the ETT2 cluster in EHEC O157:H7 leads to greatly increased secretion of proteins encoded by the LEE and to increased adhesion to human intestinal cells. Studies in which transcriptional fusions and microarrays were used indicated that EtrA and EivF exert profound negative effects on gene transcription within the LEE. Consistent with these observations, expression of these regulators in an EHEC O26:H- strain led to suppression of protein secretion under LEE-inducing conditions. These findings provide fresh examples of the influence of mobile genetic elements on regulation of the LEE and of cross talk between type III secretion system gene clusters. In addition, they provide a cautionary tale because they show that the effects of regulatory genes can outlive widespread decay of other genes in a functionally coherent gene cluster, a phenomenon that we have named the “Cheshire cat effect.” It also seems likely that variations in the ETT2 regulator repertoire might account for strain-to-strain variation in secretion of LEE-encoded proteins.

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