scholarly journals Long Polar Fimbriae of Enterohemorrhagic Escherichia coli O157:H7 Bind to Extracellular Matrix Proteins

2011 ◽  
Vol 79 (9) ◽  
pp. 3744-3750 ◽  
Author(s):  
Mauricio J. Farfan ◽  
Lidia Cantero ◽  
Roberto Vidal ◽  
Douglas J. Botkin ◽  
Alfredo G. Torres
Keyword(s):  
Escherichia Coli ◽  
Extracellular Matrix ◽  
Enterohemorrhagic Escherichia Coli ◽  
Matrix Proteins ◽  
Intestinal Cells ◽  
T84 Cells ◽  
Content Type ◽  
Fimbrial Subunit ◽  
Ecm Proteins

ABSTRACTAdherence to intestinal cells is a key process in infection caused by enterohemorrhagicEscherichia coli(EHEC). Several adhesion factors that mediate the binding of EHEC to intestinal cells have been described, but the receptors involved in their recognition are not fully characterized. Extracellular matrix (ECM) proteins might act as receptors involved in the recognition of enteric pathogens, including EHEC. In this study, we sought to characterize the binding of EHEC O157:H7 to ECM proteins commonly present in the intestine. We found that EHEC prototype strains as well as other clinical isolates adhered more abundantly to surfaces coated with fibronectin, laminin, and collagen IV. Further characterization of this phenotype, by using antiserum raised against the LpfA1 putative major fimbrial subunit and by addition of mannose, showed that a reduced binding of EHEC to ECM proteins was observed in a long polar fimbria (lpf) mutant. We also found that the two regulators, H-NS and Ler, had an effect in EHEC Lpf-mediated binding to ECM, supporting the roles of these tightly regulated fimbriae as adherence factors. Purified Lpf major subunit bound to all of the ECM proteins tested. Finally, increased bacterial adherence was observed when T84 cells, preincubated with ECM proteins, were infected with EHEC. Taken together, these findings suggest that the interaction of Lpf and ECM proteins contributes to the EHEC colonization of the gastrointestinal tract.

scholarly journals Identification and Characterization of a Phase-Variable Element That Regulates the Autotransporter UpaE in UropathogenicEscherichia coli

mBio ◽  
2018 ◽  
Vol 9 (4) ◽  
Author(s):  
E. J. Battaglioli ◽  
K. G. K. Goh ◽  
T. S. Atruktsang ◽  
K. Schwartz ◽  
M. A. Schembri ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Extracellular Matrix ◽  
Urinary Tract Infection ◽  
Reference Strain ◽  
Invertible Element ◽  
Matrix Proteins ◽  
Phase Variable ◽  
Content Type ◽  
Third Phase ◽  
Strain Cft073

ABSTRACTUropathogenicEscherichia coli(UPEC) is the most common etiologic agent of uncomplicated urinary tract infection (UTI). An important mechanism of gene regulation in UPEC is phase variation that involves inversion of a promoter-containing DNA element via enzymatic activity of tyrosine recombinases, resulting in biphasic, ON or OFF expression of target genes. The UPEC reference strain CFT073 has five tyrosine site-specific recombinases that function at two previously characterized promoter inversion systems,fimSandhyxS. Three of the five recombinases are located proximally to their cognate target elements, which is typical of promoter inversion systems. The genes for the other two recombinases, IpuA and IpuB, are located distal from these sites. Here, we identified and characterized a third phase-variable invertible element in CFT073,ipuS, located proximal toipuAandipuB. The inversion ofipuSis catalyzed by four of the five CFT073 recombinases. Orientation of the element drives transcription of a two-gene operon containingipuR, a predicted LuxR-type regulator, andupaE, a predicted autotransporter. We show that the predicted autotransporter UpaE is surface located and facilitates biofilm formation as well as adhesion to extracellular matrix proteins in a K-12 recombinant background. Consistent with this phenotype, theipuSON condition in CFT073 results in defective swimming motility, increased adherence to human kidney epithelial cells, and a positive competitive kidney colonization advantage in experimental mouse UTIs. Overall, the identification of a third phase switch in UPEC that is regulated by a shared set of recombinases describes a complex phase-variable virulence network in UPEC.IMPORTANCEUropathogenicEscherichia coli(UPEC) is the most common cause of urinary tract infection (UTI). ON versus OFF phase switching by inversion of small DNA elements at two chromosome sites in UPEC regulates the expression of important virulence factors, including the type 1 fimbria adhesion organelle. In this report, we describe a third invertible element,ipuS, in the UPEC reference strain CFT073. The inversion ofipuScontrols the phase-variable expression ofupaE, an autotransporter gene that encodes a surface protein involved in adherence to extracellular matrix proteins and colonization of the kidneys in a murine model of UTI.

scholarly journals Enterohemorrhagic Escherichia coli Colonization of Human Colonic EpitheliumIn VitroandEx Vivo

2014 ◽  
Vol 83 (3) ◽  
pp. 942-949 ◽  
Author(s):  
Steven B. Lewis ◽  
Vivienne Cook ◽  
Richard Tighe ◽  
Stephanie Schüller
Keyword(s):  
Escherichia Coli ◽  
Human Colon ◽  
Enterohemorrhagic Escherichia Coli ◽  
Care Needs ◽  
T84 Cells ◽  
Host Cell Membrane ◽  
Content Type ◽  
Colonic Biopsy

EnterohemorrhagicEscherichia coli(EHEC) is an important foodborne pathogen causing gastroenteritis and more severe complications, such as hemorrhagic colitis and hemolytic uremic syndrome. Pathology is most pronounced in the colon, but to date there is no direct clinical evidence showing EHEC binding to the colonic epithelium in patients. In this study, we investigated EHEC adherence to the human colon by usingin vitroorgan culture (IVOC) of colonic biopsy samples and polarized T84 colon carcinoma cells. We show for the first time that EHEC colonizes human colonic biopsy samples by forming typical attaching and effacing (A/E) lesions which are dependent on EHEC type III secretion (T3S) and binding of the outer membrane protein intimin to the translocated intimin receptor (Tir). A/E lesion formation was dependent on oxygen levels and suppressed under oxygen-rich culture conditions routinely used for IVOC. In contrast, EHEC adherence to polarized T84 cells occurred independently of T3S and intimin and did not involve Tir translocation into the host cell membrane. Colonization of neither biopsy samples nor T84 cells was significantly affected by expression of Shiga toxins. Our study suggests that EHEC colonizes and forms stable A/E lesions on the human colon, which are likely to contribute to intestinal pathology during infection. Furthermore, care needs to be taken when using cell culture models, as they might not reflect thein vivosituation.

scholarly journals Molecular Characterization of the EhaG and UpaG Trimeric Autotransporter Proteins from Pathogenic Escherichia coli

2012 ◽  
Vol 78 (7) ◽  
pp. 2179-2189 ◽  
Author(s):  
Makrina Totsika ◽  
Timothy J. Wells ◽  
Christophe Beloin ◽  
Jaione Valle ◽  
Luke P. Allsopp ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Epithelial Cells ◽  
Cell Surface ◽  
Specific Binding ◽  
Negative Regulator ◽  
Passenger Domain ◽  
Content Type ◽  
E Coli ◽  
Ecm Proteins

ABSTRACTTrimeric autotransporter proteins (TAAs) are important virulence factors of many Gram-negative bacterial pathogens. A common feature of most TAAs is the ability to mediate adherence to eukaryotic cells or extracellular matrix (ECM) proteins via a cell surface-exposed passenger domain. Here we describe the characterization of EhaG, a TAA identified from enterohemorrhagicEscherichia coli(EHEC) O157:H7. EhaG is a positional orthologue of the recently characterized UpaG TAA from uropathogenicE. coli(UPEC). Similarly to UpaG, EhaG localized at the bacterial cell surface and promoted cell aggregation, biofilm formation, and adherence to a range of ECM proteins. However, the two orthologues display differential cellular binding: EhaG mediates specific adhesion to colorectal epithelial cells while UpaG promotes specific binding to bladder epithelial cells. The EhaG and UpaG TAAs contain extensive sequence divergence in their respective passenger domains that could account for these differences. Indeed, sequence analyses of UpaG and EhaG homologues from severalE. coligenomes revealed grouping of the proteins in clades almost exclusively represented by distinctE. colipathotypes. The expression of EhaG (in EHEC) and UpaG (in UPEC) was also investigated and shown to be significantly enhanced in anhnsisogenic mutant, suggesting that H-NS acts as a negative regulator of both TAAs. Thus, while the EhaG and UpaG TAAs contain some conserved binding and regulatory features, they also possess important differences that correlate with the distinct pathogenic lifestyles of EHEC and UPEC.

scholarly journals Characterization of a Novel Microcin That Kills Enterohemorrhagic Escherichia coli O157:H7 and O26

2012 ◽  
Vol 78 (18) ◽  
pp. 6592-6599 ◽  
Author(s):  
Lauren J. Eberhart ◽  
James R. Deringer ◽  
Kelly A. Brayton ◽  
Ashish A. Sawant ◽  
Thomas E. Besser ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Enterohemorrhagic Escherichia Coli ◽  
Logarithmic Phase ◽  
Type I ◽  
Escherichia Coli O157 ◽  
Content Type ◽  
E Coli ◽  
Secretion Pathway ◽  
Dependent Inhibition

ABSTRACTA novel phenotype was recently identified in which specific strains ofEscherichia coliinhibit competingE. colistrains via a mechanism that was designated “proximity-dependent inhibition” (PDI). PDI-expressing (PDI+)E. coliis known to inhibit susceptible (PDI−)E. colistrains, including several enterohemorrhagic (EHEC) and enterotoxigenic (ETEC)E. colistrains. In this study, every strain from a genetically diverse panel ofE. coliO157:H7 (n= 25) and additional strains ofE. coliserovar O26 were susceptible to the PDI phenotype. LIVE/DEAD staining was consistent with inhibition by killing of susceptible cells. Comparative genome analysis identified the genetic component of PDI, which is composed of a plasmid-borne (Incl1) operon encoding a putative microcin and associated genes for transport, immunity, and microcin activation. Transfer of the plasmid to a PDI−strain resulted in transfer of the phenotype, and deletion of the genes within the operon resulted in loss of the inhibition phenotype. Deletion of chromosomally encodedtolCalso resulted in loss of the inhibitory phenotype, and this confirmed that the putative microcin is most likely secreted via a type I secretion pathway. Deletion of an unrelated plasmid gene did not affect the PDI phenotype. Quantitative reverse transcription (RT)-PCR demonstrated that microcin expression is correlated with logarithmic-phase growth. The ability to inhibit a diversity ofE. colistrains indicates that this microcin may influence gut community composition and could be useful for control of important enteric pathogens.

scholarly journals The Major Pilin Subunit of the AAF/II Fimbriae from Enteroaggregative Escherichia coli Mediates Binding to Extracellular Matrix Proteins

2008 ◽  
Vol 76 (10) ◽  
pp. 4378-4384 ◽  
Author(s):  
Mauricio J. Farfan ◽  
Keith G. Inman ◽  
James P. Nataro
Keyword(s):  
Escherichia Coli ◽  
Extracellular Matrix ◽  
Extracellular Matrix Proteins ◽  
Prototype Strain ◽  
Matrix Proteins ◽  
Dependent Manner ◽  
T84 Cells ◽  
Cell Monolayers ◽  
Type Iv ◽  
Pilin Gene

ABSTRACT Enteroaggregative Escherichia coli (EAEC) adherence to human intestinal tissue is mediated by aggregative adherence fimbriae (AAF); however, the receptors involved in EAEC adherence remain uncharacterized. Adhesion to extracellular matrix proteins is commonly observed among enteric pathogens, so we addressed the hypothesis that EAEC may bind to extracellular matrix proteins commonly found in the intestine. We found that EAEC prototype strain 042 adhered more abundantly to surfaces that were precoated with the extracellular matrix proteins fibronectin, laminin, and type IV collagen. Differences in fibronectin binding of almost 2 orders of magnitude were observed between EAEC 042 and a mutant in the AAF/II major pilin gene, aafA. Purified AafA, refolded as a donor strand complementation construct, bound fibronectin in a dose-dependent manner. Addition of fibronectin to the apical surfaces of polarized T84 cell monolayers augmented EAEC 042 adherence, and this effect required expression of aafA. Finally, increased bacterial adherence was observed when apical secretion of fibronectin was induced by adenosine in polarized T84 cells. Binding to fibronectin may contribute to colonization of the gastrointestinal tract by EAEC.

scholarly journals Identification of Cell Surface-Exposed Proteins Involved in the Fimbria-Mediated Adherence of Enteroaggregative Escherichia coli to Intestinal Cells

2014 ◽  
Vol 82 (4) ◽  
pp. 1719-1724 ◽  
Author(s):  
Mariana Izquierdo ◽  
Fernando Navarro-Garcia ◽  
Raul Nava-Acosta ◽  
James P. Nataro ◽  
Fernando Ruiz-Perez ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Prototype Strain ◽  
Intestinal Cells ◽  
Intestinal Epithelial ◽  
T84 Cells ◽  
Content Type ◽  
Intestinal Epithelia ◽  
Liquid Chromatography Tandem Mass ◽  
Interfering Rna

ABSTRACTFimbria-mediated adherence to the intestinal epithelia is a key step in enteroaggregativeEscherichia coli(EAEC) pathogenesis. To date, four fimbriae have been described for EAEC; aggregative adherence fimbria II (AAF/II) is the most important adherence factor for EAEC prototype strain 042. Previously, we described results showing that extracellular matrix (ECM) components might be involved in the recognition of AAF/II fimbriae by intestinal cells. In this study, we sought to identify novel potential receptors on intestinal epithelial cells recognized by the AAF/II fimbriae. Purified AafA-dscprotein, the major subunit of AAF/II fimbriae, was incubated with a monolayer of T84 cells, cross-linked to the surface-exposed T84 cell proteins, and immunoprecipitated by using anti-AafA antibodies. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis of cellular proteins bound to AafA-dscprotein identified laminin (previously recognized as a potential receptor for AAF/II) and cytokeratin 8 (CK8). Involvement of the major subunit of AAF/II fimbriae (AafA protein) in the binding to recombinant CK8 was confirmed by adherence assays with purified AAF/II fimbriae, AafA-dscprotein, and strain 042. Moreover, HEp-2 cells transfected with CK8 small interfering RNA (siRNA) showed reduced 042 adherence compared with cells transfected with scrambled siRNA as a control. Adherence of 042 to HEp-2 cells preincubated with antibodies against ECM proteins or CK8 was substantially reduced. Altogether, our results supported the idea of a role of CK8 as a potential receptor for EAEC.

scholarly journals Presence of Enterohemorrhagic Escherichia coli ST678/O104:H4 in France Prior to 2011

2011 ◽  
Vol 77 (24) ◽  
pp. 8784-8786 ◽  
Author(s):  
Stefan Monecke ◽  
Patricia Mariani-Kurkdjian ◽  
Edouard Bingen ◽  
François-Xavier Weill ◽  
Charlotte Balière ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Enterohemorrhagic Escherichia Coli ◽  
Content Type

ABSTRACTTwo isolates of enterohemorrhagicEscherichia coli(EHEC) O104:H4 were isolated in France in 2004 and 2009. Both were characterized and compared to the strain which caused the German outbreak in 2011 and to other O104:H4 strains. This suggests that different O104:H4 EHEC strains were present several years prior to the 2011 outbreak.

scholarly journals Characterization of Epidemic IncI1-Iγ Plasmids Harboring Ambler Class A and C Genes in Escherichia coli and Salmonella enterica from Animals and Humans

2015 ◽  
Vol 59 (9) ◽  
pp. 5357-5365 ◽  
Author(s):  
Hilde Smith ◽  
Alex Bossers ◽  
Frank Harders ◽  
Guanghui Wu ◽  
Neil Woodford ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Salmonella Enterica ◽  
Phylogenetic Trees ◽  
Functional Study ◽  
Content Type ◽  
Gene Associations ◽  
Genes Encoding ◽  
Marked Resistance

ABSTRACTThe aim of the study was to identify the plasmid-encoded factors contributing to the emergence and spread of epidemic IncI1-Iγ plasmids obtained fromEscherichia coliandSalmonella entericaisolates from animal and human reservoirs. For this, 251 IncI1-Iγ plasmids carrying various extended-spectrum β-lactamase (ESBL) or AmpC β-lactamase genes were compared using plasmid multilocus sequence typing (pMLST). Thirty-two of these plasmids belonging to different pMLST types were sequenced using Roche 454 and Illumina platforms. Epidemic IncI1-Iγ plasmids could be assigned to various dominant clades, whereas rarely detected plasmids clustered together as a distinct clade. Similar phylogenetic trees were obtained using only the plasmid backbone sequences, showing that the differences observed between the plasmids belonging to distinct clades resulted mainly from differences between their backbone sequences. Plasmids belonging to the various clades differed particularly in the presence/absence of genes encoding partitioning and addiction systems, which contribute to stable inheritance during cell division and plasmid maintenance. Despite this, plasmids belonging to the various phylogenetic clades also showed marked resistance gene associations, indicating the circulation of successful plasmid-gene combinations. The variation intraYandexcAgenes found in IncI1-Iγ plasmids is conserved within pMLST sequence types and plays a role in incompatibility, although functional study is needed to elucidate the role of these genes in plasmid epidemiology.

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