scholarly journals Global Analysis of Posttranscriptional Regulation by GlmY and GlmZ in Enterohemorrhagic Escherichia coli O157:H7

2015 ◽  
Vol 83 (4) ◽  
pp. 1286-1295 ◽  
Author(s):  
Charley C. Gruber ◽  
Vanessa Sperandio
Keyword(s):  
Escherichia Coli ◽  
Rna Sequencing ◽  
Posttranscriptional Regulation ◽  
Global Analysis ◽  
Acid Resistance ◽  
Enterohemorrhagic Escherichia Coli ◽  
Global Changes ◽  
Content Type ◽  
Enterocyte Effacement ◽  
K 12

EnterohemorrhagicEscherichia coli(EHEC) is a significant human pathogen and is the cause of bloody diarrhea and hemolytic-uremic syndrome. The virulence repertoire of EHEC includes the genes within the locus of enterocyte effacement (LEE) that are largely organized in five operons,LEE1toLEE5, which encode a type III secretion system, several effectors, chaperones, and regulatory proteins. In addition, EHEC also encodes several non-LEE-encoded effectors and fimbrial operons. The virulence genes of this pathogen are under a large amount of posttranscriptional regulation. The small RNAs (sRNAs) GlmY and GlmZ activate the translation of glucosamine synthase (GlmS) inE. coliK-12, and in EHEC they destabilize the 3′ fragments of theLEE4andLEE5operons and promote translation of the non-LEE-encoded effector EspFu. We investigated the global changes of EHEC gene expression governed by GlmY and GlmZ using RNA sequencing and gene arrays. This study extends the known effects of GlmY and GlmZ regulation to show that they promote expression of the curli adhesin, repress the expression of tryptophan metabolism genes, and promote the expression of acid resistance genes and the non-LEE-encoded effector NleA. In addition, seven novel EHEC-specific sRNAs were identified using RNA sequencing, and three of them—sRNA56, sRNA103, and sRNA350—were shown to regulate urease, fimbria, and the LEE, respectively. These findings expand the knowledge of posttranscriptional regulation in EHEC.

scholarly journals Evolutionary Silence of the Acid Chaperone Protein HdeB in Enterohemorrhagic Escherichia coli O157:H7

2011 ◽  
Vol 78 (4) ◽  
pp. 1004-1014 ◽  
Author(s):  
Michelle Q. Carter ◽  
Jacqueline W. Louie ◽  
Clifton K. Fagerquist ◽  
Omar Sultan ◽  
William G. Miller ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Acid Resistance ◽  
Significant Loss ◽  
Enterohemorrhagic Escherichia Coli ◽  
Survival Strategies ◽  
Start Codon ◽  
Divergent Evolution ◽  
Content Type ◽  
E Coli ◽  
K 12

ABSTRACTThe periplasmic chaperones HdeA and HdeB are known to be important for cell survival at low pH (pH < 3) inEscherichia coliandShigellaspp. Here we investigated the roles of HdeA and HdeB in the survival of various enterohemorrhagicE. coli(EHEC) following exposure to pH 2.0. Similar to K-12 strains, the acid protections conferred by HdeA and HdeB in EHEC O145 were significant: loss of HdeA and HdeB led to over 100- to 1,000-fold reductions in acid survival, depending on the growth condition of prechallenge cells. However, this protection was much less inE. coliO157:H7 strains. Deletion ofhdeBdid not affect the acid survival of cells, and deletion ofhdeAled to less than a 5-fold decrease in survival. Sequence analysis of thehdeABoperon revealed a point mutation at the putative start codon of thehdeBgene in all 26E. coliO157:H7 strains analyzed, which shifted the ATG start codon to ATA. This mutation correlated with the lack of HdeB inE. coliO157:H7; however, the plasmid-borne O157-hdeBwas able to restore partially the acid resistance in anE. coliO145ΔhdeABmutant, suggesting the potential function of O157-HdeB as an acid chaperone. We conclude thatE. coliO157:H7 strains have evolved acid survival strategies independent of the HdeA/B chaperones and are more acid resistant than nonpathogenic K-12 for cells grown under nonfavorable culturing conditions such as in Luria-Bertani no-salt broth at 28°C. These results suggest a divergent evolution of acid resistance mechanisms withinE. coli.

scholarly journals Coordinate Control of the Locus of Enterocyte Effacement and Enterohemolysin Genes by Multiple Common Virulence Regulators in Enterohemorrhagic Escherichia coli

2011 ◽  
Vol 79 (11) ◽  
pp. 4628-4637 ◽  
Author(s):  
Sunao Iyoda ◽  
Naoko Honda ◽  
Takehito Saitoh ◽  
Ken Shimuta ◽  
Jun Terajima ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Regulatory Region ◽  
Regulatory System ◽  
Enterohemorrhagic Escherichia Coli ◽  
Mobility Shift ◽  
Content Type ◽  
Positive Regulators ◽  
Enterocyte Effacement ◽  
Gel Mobility Shift ◽  
K 12

ABSTRACTThe locus of enterocyte effacement (LEE) pathogenicity island is required for the intimate adhesion of enterohemorrhagicEscherichia coli(EHEC) to the intestinal epithelial cells. GrlR and GrlA are LEE-encoded negative and positive regulators, respectively. The interaction of these two regulators is important for controlling the transcription of LEE genes through Ler, a LEE-encoded central activator for the LEE. The GrlR-GrlA regulatory system controls not only LEE but also the expression of the flagellar and enterohemolysin (Ehx) genes in EHEC. Since Ehx levels were markedly induced in agrlRmutant but not in agrlR grlAdouble mutant and significantly increased by overexpression of GrlA in alermutant, GrlA is responsible for this regulation (T. Saitoh et al., J. Bacteriol.190:4822-4830, 2008). In this study, additional investigations of the regulation ofehxgene expression determined that Ler also acts as an activator for Ehx expression without requiring GrlA function. We recently reported that the LysR-type regulator LrhA positively controls LEE expression (N. Honda et al., Mol. Microbiol.74:1393-1411, 2009). The hemolytic activity of thelrhAmutant strain of EHEC was lower than that of the wild-type strain, and LrhA markedly inducedehxtranscription in anE. coliK-12 strain, suggesting that LrhA also activates the transcription ofehxwithout GrlA and Ler. Gel mobility shift assays demonstrated that Ler and LrhA directly bind to the regulatory region ofehxC. Together, these results indicate that transcription ofehxis positively regulated by Ler, GrlA, and LrhA, which all act as positive regulators for LEE expression.

scholarly journals Global Regulator of Virulence A (GrvA) Coordinates Expression of Discrete Pathogenic Mechanisms in Enterohemorrhagic Escherichia coli through Interactions with GadW-GadE

2015 ◽  
Vol 198 (3) ◽  
pp. 394-409 ◽  
Author(s):  
Jason K. Morgan ◽  
Ronan K. Carroll ◽  
Carly M. Harro ◽  
Khoury W. Vendura ◽  
Lindsey N. Shaw ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Acid Resistance ◽  
Enterohemorrhagic Escherichia Coli ◽  
Sequencing Analysis ◽  
Global Regulator ◽  
Altered Expression ◽  
Content Type ◽  
Pathogenic Mechanisms ◽  
Gut Colonization ◽  
Intestinal Pathogens

ABSTRACTGlobal regulator of virulence A (GrvA) is a ToxR-family transcriptional regulator that activates locus of enterocyte effacement (LEE)-dependent adherence in enterohemorrhagicEscherichia coli(EHEC). LEE activation by GrvA requires the Rcs phosphorelay response regulator RcsB and is sensitive to physiologically relevant concentrations of bicarbonate, a known stimulant of virulence systems in intestinal pathogens. This study determines the genomic scale of GrvA-dependent regulation and uncovers details of the molecular mechanism underlying GrvA-dependent regulation of pathogenic mechanisms in EHEC. In agrvA-null background of EHEC strain TW14359, RNA sequencing analysis revealed the altered expression of over 700 genes, including the downregulation of LEE- and non-LEE-encoded effectors and the upregulation of genes for glutamate-dependent acid resistance (GDAR). Upregulation of GDAR genes corresponded with a marked increase in acid resistance. GrvA-dependent regulation of GDAR and the LEE requiredgadE, the central activator of GDAR genes and a direct repressor of the LEE. Control ofgadEby GrvA was further determined to occur through downregulation of thegadEactivator GadW. This interaction of GrvA with GadW-GadE represses the acid resistance phenotype, while it concomitantly activates the LEE-dependent adherence and secretion of immune subversion effectors. The results of this study significantly broaden the scope of GrvA-dependent regulation and its role in EHEC pathogenesis.IMPORTANCEEnterohemorrhagicEscherichia coli(EHEC) is an intestinal human pathogen causing acute hemorrhagic colitis and life-threatening hemolytic-uremic syndrome. For successful transmission and gut colonization, EHEC relies on the glutamate-dependent acid resistance (GDAR) system and a type III secretion apparatus, encoded on the LEE pathogenicity island. This study investigates the mechanism whereby the DNA-binding regulator GrvA coordinates activation of the LEE with repression of GDAR. Investigating how these systems are regulated leads to an understanding of pathogenic behavior and novel strategies aimed at disease prevention and control.

scholarly journals Involvement of PatE, a Prophage-Encoded AraC-Like Regulator, in the Transcriptional Activation of Acid Resistance Pathways of Enterohemorrhagic Escherichia coli Strain EDL933

2012 ◽  
Vol 78 (15) ◽  
pp. 5083-5092 ◽  
Author(s):  
Jennifer K. Bender ◽  
Judyta Praszkier ◽  
Matthew J. Wakefield ◽  
Kathryn Holt ◽  
Marija Tauschek ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Transcriptional Activation ◽  
Regulatory Protein ◽  
Acid Resistance ◽  
Enterohemorrhagic Escherichia Coli ◽  
Transcriptional Analysis ◽  
Mobility Shift ◽  
Content Type ◽  
Infectious Dose ◽  
Genes Encoding

ABSTRACTEnterohemorrhagicEscherichia coli(EHEC) O157:H7 is a lethal human intestinal pathogen that causes hemorrhagic colitis and the hemolytic-uremic syndrome. EHEC is transmitted by the fecal-oral route and has a lower infectious dose than most other enteric bacterial pathogens in that fewer than 100 CFU are able to cause disease. This low infectious dose has been attributed to the ability of EHEC to survive in the acidic environment of the human stomach.In silicoanalysis of the genome of EHEC O157:H7 strain EDL933 revealed a gene,patE, for a putative AraC-like regulatory protein within the prophage island, CP-933H. Transcriptional analysis inE. colishowed that the expression ofpatEis induced during stationary phase. Data from microarray assays demonstrated that PatE activates the transcription of genes encoding proteins of acid resistance pathways. In addition, PatE downregulated the expression of a number of genes encoding heat shock proteins and the type III secretion pathway of EDL933. Transcriptional analysis and electrophoretic mobility shift assays suggested that PatE also activates the transcription of the gene for the acid stress chaperonehdeAby binding to its promoter region. Finally, assays of acid tolerance showed that increasing the expression of PatE in EHEC greatly enhanced the ability of the bacteria to survive in different acidic environments. Together, these findings indicate that EHEC strain EDL933 carries a prophage-encoded regulatory system that contributes to acid resistance.

scholarly journals Cross-Reactive Protection against Enterohemorrhagic Escherichia coli Infection by Enteropathogenic E. coli in a Mouse Model

2011 ◽  
Vol 79 (6) ◽  
pp. 2224-2233 ◽  
Author(s):  
Carla Calderon Toledo ◽  
Ida Arvidsson ◽  
Diana Karpman
Keyword(s):  
Escherichia Coli ◽  
Weight Loss ◽  
Mouse Model ◽  
Enterohemorrhagic Escherichia Coli ◽  
Epec Strain ◽  
Content Type ◽  
E Coli ◽  
Renal Histology ◽  
Escherichia Coli Infection ◽  
Enterocyte Effacement

ABSTRACTEnteropathogenicEscherichia coli(EPEC) and enterohemorrhagicE. coli(EHEC) are related attaching and effacing (A/E) pathogens. The genes responsible for the A/E pathology are carried on a chromosomal pathogenicity island termed the locus of enterocyte effacement (LEE). Both pathogens share a high degree of homology in the LEE and additional O islands. EHEC prevalence is much lower in areas where EPEC is endemic. This may be due to the development of antibodies against common EPEC and EHEC antigens. This study investigated the hypothesis that EPEC infections may protect against EHEC infections. We used a mouse model to inoculate BALB/c mice intragastrically, first with EPEC and then with EHEC (E. coliO157:H7). Four control groups received either a nonpathogenicE. coli(NPEC) strain followed by EHEC (NPEC/EHEC), phosphate-buffered saline (PBS) followed by EHEC (PBS/EHEC), EPEC/PBS, or PBS/PBS. Mice were monitored for weight loss and symptoms. EPEC colonized the intestine after challenge, and mice developed serum antibodies to intimin andE. colisecreted protein B (encoded in the LEE). Prechallenge with an EPEC strain had a protective effect after EHEC infection, as only a few mice developed mild symptoms, from which they recovered. These mice had an increase in body weight similar to that in control animals, and tissue morphology exhibited mild intestinal changes and normal renal histology. All mice that were not prechallenged with the EPEC strain developed mild to severe symptoms after EHEC infection, with weight loss as well as intestinal and renal histopathological changes. These data suggest that EPEC may protect against EHEC infection in this mouse model.

scholarly journals Enterohemorrhagic Escherichia coli Virulence Regulation by Two Bacterial Adrenergic Kinases, QseC and QseE

2011 ◽  
Vol 80 (2) ◽  
pp. 688-703 ◽  
Author(s):  
Jacqueline Njoroge ◽  
Vanessa Sperandio
Keyword(s):  
Escherichia Coli ◽  
Response Regulator ◽  
Enterohemorrhagic Escherichia Coli ◽  
Dependent Manner ◽  
Content Type ◽  
E Coli ◽  
Virulence Regulation ◽  
Sensor Kinases ◽  
Virulence Factor Gene ◽  
Enterocyte Effacement

ABSTRACTThe human pathogen enterohemorrhagicEscherichia coli(EHEC) O157:H7 has two histidine sensor kinases, QseC and QseE, which respond to the mammalian adrenergic hormones epinephrine and norepinephrine by increasing their autophosphorylation. Although QseC and QseE are present in nonpathogenic strains ofE. coli, EHEC exploits these kinases for virulence regulation. To further investigate the full extent of epinephrine and its sensors' impact on EHEC virulence, we performed transcriptomic and phenotypic analyses of single and double deletions ofqseCandqseEgenes in the absence or presence of epinephrine. We showed that in EHEC, epinephrine sensing seems to occur primarily through QseC and QseE. We also observed that QseC and QseE regulate expression of the locus of enterocyte effacement (LEE) genes positively and negatively, respectively. LEE activation, which is required for the formation of the characteristic attaching and effacing (A/E) lesions by EHEC on epithelial cells, is epinephrine dependent. Regulation of the LEE and the non-LEE-contained virulence factor genenleAby QseE is indirect, through transcription inhibition of the RcsB response regulator. Finally, we show that coincubation of HeLa cells with epinephrine increases EHEC infectivity in a QseC- and QseE-dependent manner. These results genetically and phenotypically map the contributions of the two adrenergic sensors QseC and QseE to EHEC pathogenesis.

scholarly journals The Acyl-Homoserine Lactone Synthase YenI from Yersinia enterocolitica Modulates Virulence Gene Expression in Enterohemorrhagic Escherichia coli O157:H7

2013 ◽  
Vol 81 (11) ◽  
pp. 4192-4199 ◽  
Author(s):  
Y N. Nguyen ◽  
Haiqing Sheng ◽  
Rambabu Dakarapu ◽  
John R. Falck ◽  
Carolyn J. Hovde ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Yersinia Enterocolitica ◽  
Virulence Gene ◽  
Acid Resistance ◽  
Homoserine Lactone ◽  
Enterohemorrhagic Escherichia Coli ◽  
Content Type ◽  
Fecal Shedding ◽  
Virulence Gene Expression

ABSTRACTThe human pathogen enterohemorrhagicEscherichia coli(EHEC) O157:H7 colonizes the rectoanal junction (RAJ) in cattle, its natural reservoir. Colonization at the RAJ poses a serious risk for fecal shedding and contamination of the environment. We previously demonstrated that EHEC senses acyl-homoserine lactones (AHLs) produced by the microbiota in the rumen to activate thegadacid resistance genes necessary for survival through the acidic stomachs in cattle and to repress the locus of enterocyte effacement (LEE) genes important for colonization of the RAJ, but unnecessary in the rumen. Devoid of AHLs, the RAJ is the prominent site of colonization of EHEC in cattle. To determine if the presence of AHLs in the RAJ could repress colonization at this site, we engineered EHEC to express theYersinia enterocoliticaAHL synthase geneyenI, which constitutively produces AHLs, to mimic a constant exposure of AHLs in the environment. TheyenI+EHEC produces oxo-C6-homoserine lactone (oxo-C6-HSL) and had a significant reduction in LEE expression, effector protein secretion, and attaching and effacing (A/E) lesion formationin vitrocompared to the wild type (WT). TheyenI+EHEC also activated expression of thegadgenes. To assess whether AHL production, which decreases LEE expression, would decrease RAJ colonization by EHEC, cattle were challenged at the RAJ with WT oryenI+EHEC. Although theyenI+EHEC colonized the RAJ with efficiency equal to that of the WT, there was a trend for the cattle to shed the WT strain longer than theyenI+EHEC.

scholarly journals Comparative Genomics and Immunoinformatics Approach for the Identification of Vaccine Candidates for Enterohemorrhagic Escherichia coli O157:H7

2014 ◽  
Vol 82 (5) ◽  
pp. 2016-2026 ◽  
Author(s):  
Víctor A. García-Angulo ◽  
Anjana Kalita ◽  
Mridul Kalita ◽  
Luis Lozano ◽  
Alfredo G. Torres
Keyword(s):  
Escherichia Coli ◽  
Comparative Genomics ◽  
Enterohemorrhagic Escherichia Coli ◽  
Th2 Cytokines ◽  
Structural Protein ◽  
Gene Encoding ◽  
Content Type ◽  
Vaccine Candidates ◽  
Genes Encoding ◽  
K 12

ABSTRACTEnterohemorrhagicEscherichia coli(EHEC) O157:H7 strains are major human food-borne pathogens, responsible for bloody diarrhea and hemolytic-uremic syndrome worldwide. Thus far, there is no vaccine for humans against EHEC infections. In this study, a comparative genomics analysis was performed to identify EHEC-specific antigens useful as potential vaccines. The genes present in both EHEC EDL933 and Sakai strains but absent in nonpathogenicE. coliK-12 and HS strains were subjected to anin silicoanalysis to identify secreted or surface-expressed proteins. We obtained a total of 65 gene-encoding protein candidates, which were subjected to immunoinformatics analysis. Our criteria of selection aided in categorizing the candidates as high, medium, and low priority. Three members of each group were randomly selected and cloned into pVAX-1.Candidates were pooled accordingly to their priority group and tested for immunogenicity against EHEC O157:H7 using a murine model of gastrointestinal infection. The high-priority (HP) pool, containing genes encoding a Lom-like protein (pVAX-31), a putative pilin subunit (pVAX-12), and a fragment of the type III secretion structural protein EscC (pVAX-56.2), was able to induce the production of EHEC IgG and sIgA in sera and feces. HP candidate-immunized mice displayed elevated levels of Th2 cytokines and diminished cecum colonization after wild-type challenge. Individually tested HP vaccine candidates showed that pVAX-12 and pVAX-56.2 significantly induced Th2 cytokines and production of fecal EHEC sIgA, with pVAX-56.2 reducing EHEC cecum colonization. We describe here a bioinformatics approach able to identify novel vaccine candidates potentially useful for preventing EHEC O157:H7 infections.

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.

scholarly journals Distinct Acid Resistance and Survival Fitness Displayed by Curli Variants of Enterohemorrhagic Escherichia coli O157:H7

2011 ◽  
Vol 77 (11) ◽  
pp. 3685-3695 ◽  
Author(s):  
Michelle Q. Carter ◽  
Maria T. Brandl ◽  
Jacqueline W. Louie ◽  
Jennifer L. Kyle ◽  
Diana K. Carychao ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Acid Resistance ◽  
Cell Aggregation ◽  
Oxygen Depletion ◽  
Growth Conditions ◽  
Enterohemorrhagic Escherichia Coli ◽  
Surface Attachment ◽  
Content Type ◽  
E Coli ◽  
Physiological Properties

ABSTRACTCurli are adhesive fimbriae ofEnterobacteriaceaeand are involved in surface attachment, cell aggregation, and biofilm formation. Here, we report that both inter- and intrastrain variations in curli production are widespread in enterohemorrhagicEscherichia coliO157:H7. The relative proportions of curli-producing variants (C+) and curli-deficient variants (C−) in anE. coliO157:H7 cell population varied depending on the growth conditions. In variants derived from the 2006 U.S. spinach outbreak strains, the shift between the C+and C−subpopulations occurred mostly in response to starvation and was unidirectional from C−to C+; in variants derived from the 1993 hamburger outbreak strains, the shift occurred primarily in response to oxygen depletion and was bidirectional. Furthermore, curli variants derived from the same strain displayed marked differences in survival fitness: C+variants grew to higher concentrations in nutrient-limited conditions than C−variants, whereas C−variants were significantly more acid resistant than C+variants. This difference in acid resistance does not appear to be linked to the curli fimbriaeper se, since acsgAdeletion mutant in either a C+or a C−variant exhibited an acid resistance similar to that of its parental strain. Our data suggest that natural curli variants ofE. coliO157:H7 carry several distinct physiological properties that are important for their environmental survival. Maintenance of curli variants in anE. coliO157:H7 population may provide a survival strategy in which C+variants are selected in a nutrient-limited environment, whereas C−variants are selected in an acidic environment, such as the stomach of an animal host, including that of a human.

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