scholarly journals Functional Heterogeneity of RpoS in Stress Tolerance of Enterohemorrhagic Escherichia coli Strains

2006 ◽  
Vol 72 (7) ◽  
pp. 4978-4986 ◽  
Author(s):  
Arvind A. Bhagwat ◽  
Jasmine Tan ◽  
Manan Sharma ◽  
Mahendra Kothary ◽  
Sharon Low ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Dna Sequences ◽  
Stress Responses ◽  
Glycogen Synthesis ◽  
Acid Tolerance ◽  
Acid Resistance ◽  
Growth Conditions ◽  
Enterohemorrhagic Escherichia Coli ◽  
Alkaline Stress ◽  
E Coli

ABSTRACT The stationary-phase sigma factor (RpoS) regulates many cellular responses to environmental stress conditions such as heat, acid, and alkali shocks. On the other hand, mutations at the rpoS locus have frequently been detected among pathogenic as well as commensal strains of Escherichia coli. The objective of this study was to perform a functional analysis of the RpoS-mediated stress responses of enterohemorrhagic E. coli strains from food-borne outbreaks. E. coli strains belonging to serotypes O157:H7, O111:H11, and O26:H11 exhibited polymorphisms for two phenotypes widely used to monitor rpoS mutations, heat tolerance and glycogen synthesis, as well as for two others, alkali tolerance and adherence to Caco-2 cells. However, these strains synthesized the oxidative acid resistance system through an rpoS-dependent pathway. During the transition from mildly acidic growth conditions (pH 5.5) to alkaline stress (pH 10.2), cell survival was dependent on rpoS functionality. Some strains were able to overcome negative regulation by RpoS and induced higher β-galactosidase activity without compromising their acid resistance. There were no major differences in the DNA sequences in the rpoS coding regions among the tested strains. The heterogeneity of rpoS-dependent phenotypes observed for stress-related phenotypes was also evident in the Caco-2 cell adherence assay. Wild-type O157:H7 strains with native rpoS were less adherent than rpoS-complemented counterpart strains, suggesting that rpoS functionality is needed. These results show that some pathogenic E. coli strains can maintain their acid tolerance capability while compromising other RpoS-dependent stress responses. Such adaptation processes may have significant impact on a pathogen's survival in food processing environments, as well in the host's stomach and intestine.

pH-Dependent Stationary-Phase Acid Resistance Response of Enterohemorrhagic Escherichia coli in the Presence of Various Acidulants†

1999 ◽  
Vol 62 (3) ◽  
pp. 211-218 ◽  
Author(s):  
ROBERT L. BUCHANAN ◽  
SHARON G. EDELSON
Keyword(s):  
Escherichia Coli ◽  
Stationary Phase ◽  
Stress Responses ◽  
Physiological Stress ◽  
Accurate Determination ◽  
Acid Resistance ◽  
Enterohemorrhagic Escherichia Coli ◽  
Resistance Response ◽  
E Coli ◽  
Ph Dependent

The effect of acidulant identity on the pH-dependent stationary-phase acid resistance response of enterohemorrhagic Escherichia coli was studied. Nine strains of E. coli (seven O157:H7, one O111:H-, and one biotype 1 reference strain) were cultured individually for 18 h at 37°C in tryptic soy broth (TSB) plus 1% dextrose and in TSB without dextrose to yield acid resistance induced and noninduced stationary-phase cells, respectively. These cultures were then inoculated into brain heart infusion broth (BHI) supplemented with 0.5% citric, malic, lactic, or acetic acid and adjusted to pH 3.0 with HCl. The BHI tubes were incubated at 37°C for up to 7 h and samples were removed after 0, 2, 5, and 7 h and plated for counting CFU on BHI agar and MacConkey agar (MA). The results were compared to data previously obtained with HCl only. Acid resistance varied substantially among the isolates, being dependent on the strain, the acidulant, and the induction of pH-dependent acid resistance. Hydrochloric acid was consistently the least damaging to cells; lactic acid was the most detrimental. The relative activity of the other acids was strain dependent. Inducing pH-dependent acid resistance increased the already substantial acid tolerance of stationary-phase E. coli. The extent of injury also varied with acid and strain, with as much as a 5-log-cycle differential between BHI agar and MA CFU counts. The accurate determination of the survival of enterohemorrhagic E. coli in acidic foods must take into account the biological variability of the microorganism with respect to its acid resistance and its ability to enhance survival through the induction of physiological stress responses.

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.

scholarly journals Characterization of Enterohemorrhagic Escherichia coli Strains Based on Acid Resistance Phenotypes

2005 ◽  
Vol 73 (8) ◽  
pp. 4993-5003 ◽  
Author(s):  
Arvind A. Bhagwat ◽  
Lynn Chan ◽  
Rachel Han ◽  
Jasmine Tan ◽  
Mahendra Kothary ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Natural Populations ◽  
Acid Resistance ◽  
The United States ◽  
Growth Conditions ◽  
Enterohemorrhagic Escherichia Coli ◽  
Aerobic Growth ◽  
E Coli ◽  
Low Copy Number ◽  
Fermentative Growth

ABSTRACT Acid resistance is perceived to be an important property of enterohemorrhagic Escherichia coli strains, enabling the organisms to survive passage through the acidic environment of the stomach so that they may colonize the mammalian gastrointestinal tract and cause disease. Accordingly, the organism has developed at least three genetically and physiologically distinct acid resistance systems which provide different levels of protection. The glutamate-dependent acid resistance (GDAR) system utilizes extracellular glutamate to protect cells during extreme acid challenges and is believed to provide the highest protection from stomach acidity. In this study, the GDAR system of 82 pathogenic E. coli isolates from 34 countries and 23 states within the United States was examined. Twenty-nine isolates were found to be defective in inducing GDAR under aerobic growth conditions, while five other isolates were defective in GDAR under aerobic, as well as fermentative, growth conditions. We introduced rpoS on a low-copy-number plasmid into 26 isolates and were able to restore GDAR in 20 acid-sensitive isolates under aerobic growth conditions. Four isolates were found to be defective in the newly discovered LuxR-like regulator GadE (formerly YhiE). Defects in other isolates could be due to a mutation(s) in a gene(s) with an as yet undefined role in acid resistance since GadE and/or RpoS could not restore acid resistance. These results show that in addition to mutant alleles of rpoS, mutations in gadE exist in natural populations of pathogenic E. coli. Such mutations most likely alter the infectivity of individual isolates and may play a significant role in determining the infective dose of enterohemorrhagic E. coli.

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.

Effects of pH and Acid Resistance on the Radiation Resistance of Enterohemorrhagic Escherichia coli

1999 ◽  
Vol 62 (3) ◽  
pp. 219-228 ◽  
Author(s):  
ROBERT L. BUCHANAN ◽  
SHARON G. EDELSON ◽  
GLENN BOYD
Keyword(s):  
Escherichia Coli ◽  
Stationary Phase ◽  
Radiation Resistance ◽  
Brain Heart Infusion ◽  
Acid Resistance ◽  
Enterohemorrhagic Escherichia Coli ◽  
E Coli ◽  
Effects Of Ph ◽  
Ph Dependent ◽  
Subsequent Storage

The effects of pH and the induction of pH-dependent stationary-phase acid resistance on the radiation resistance of Escherichia coli were determined for seven enterohemorrhagic strains and one nonenterohemorrhagic strain. The isolates were grown in acidogenic or nonacidogenic media to pH levels of approximately 4.7 and 7.2, respectively. The cells were then transferred to brain heart infusion (BHI) broth adjusted to pH 4.0, 4.5, 5.0, and 5.5 (with HCl) that was preequilibrated to 2°C, and cultures were then irradiated using a 137Cs source. Surviving cells and the extent of injury were determined by plating on BHI and MacConkey agars both immediately after irradiation and after subsequent storage at 2°C for 7 days. Decreasing the pH of the BHI in which E. coli was irradiated had relatively little effect on the microorganism's radiation resistance. Substantial differences in radiation resistance were noted among strains, and induction of acid resistance consistently increased radiation resistance. Comparison of E. coli levels immediately after irradiation and after 7 days of refrigerated storage suggested that irradiation enhanced pH-mediated inactivation of the pathogen. These results demonstrate that prior growth under conditions that induce a pH-dependent stationary phase cross-protects E. coli against radiation inactivation and must be taken into account when determining the microorganism's irradiation D value.

scholarly journals Functional Overlap but Lack of Complete Cross-Complementation of Streptococcus mutans and Escherichia coli YidC Orthologs

2008 ◽  
Vol 190 (7) ◽  
pp. 2458-2469 ◽  
Author(s):  
Yuxia Dong ◽  
Sara R. Palmer ◽  
Adnan Hasona ◽  
Shushi Nagamori ◽  
H. Ronald Kaback ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Streptococcus Mutans ◽  
Protein A ◽  
Membrane Integrity ◽  
Acid Tolerance ◽  
Growth Conditions ◽  
Stress Sensitivity ◽  
Protein Insertion ◽  
E Coli ◽  
Membrane Protein Insertion

ABSTRACT Oxa/YidC/Alb family proteins are chaperones involved in membrane protein insertion and assembly. Streptococcus mutans has two YidC paralogs. Elimination of yidC2, but not yidC1, results in stress sensitivity with decreased membrane-associated F1Fo ATPase activity and an inability to initiate growth at low pH or high salt concentrations (A. Hasona, P. J. Crowley, C. M. Levesque, R. W. Mair, D. G. Cvitkovitch, A. S. Bleiweis, and L. J. Brady, Proc. Natl. Acad. Sci. USA 102:17466-17471, 2005). We now show that Escherichia coli YidC complements for acid tolerance, and partially for salt tolerance, in S. mutans lacking yidC2 and that S. mutans YidC1 or YidC2 complements growth in liquid medium, restores the proton motive force, and functions to assemble the F1Fo ATPase in a previously engineered E. coli YidC depletion strain (J. C. Samuelson, M. Chen, F. Jiang, I. Moller, M. Wiedmann, A. Kuhn, G. J. Phillips, and R. E. Dalbey, Nature 406:637-641, 2000). Both YidC1 and YidC2 also promote membrane insertion of known YidC substrates in E. coli; however, complete membrane integrity is not fully replicated, as evidenced by induction of phage shock protein A. While both function to rescue E. coli growth in broth, a different result is observed on agar plates: growth of the YidC depletion strain is largely restored by 247YidC2, a hybrid S. mutans YidC2 fused to the YidC targeting region, but not by a similar chimera, 247YidC1, nor by YidC1 or YidC2. Simultaneous expression of YidC1 and YidC2 improves complementation on plates. This study demonstrates functional redundancy between YidC orthologs in gram-negative and gram-positive organisms but also highlights differences in their activity depending on growth conditions and species background, suggesting that the complete functional spectrum of each is optimized for the specific bacteria and environment in which they reside.

A Multiplex PCR for Rapid Identification of Shiga-Like Toxin-Producing Escherichia coli O157:H7 Isolated from Foods‡

1996 ◽  
Vol 59 (6) ◽  
pp. 570-576 ◽  
Author(s):  
MING Y. DENG ◽  
PINA M. FRATAMICO
Keyword(s):  
Escherichia Coli ◽  
Multiplex Pcr ◽  
Dna Sequences ◽  
Bacterial Species ◽  
Colorimetric Detection ◽  
Food Samples ◽  
Enterohemorrhagic Escherichia Coli ◽  
Experimental Conditions ◽  
Serological Tests ◽  
E Coli

For rapid and specific identification of enterohemorrhagic Escherichia coli (EHEC) serotype O157:H7 isolated from food samples, experimental conditions for a multiplex polymerase chain reaction (PCR) were optimized and a multiple digoxigenin (DIG)-labeled oligonucleotide probe hybridization (DLOPH) assay was developed. A suspect colony from MacConkey sorbitol agar containing 5-bromo-4-chloro-3-indoxyl-β-d-glucuronide (MSA-BCIG) was used for the multiplex PCR. Three different DNA sequences of E. coli O157:H7 were amplified simultaneously in the PCR: a specific fragment of an attaching and effacing gene (eae gene), conserved sequences of Shiga-like toxins (SLT) I and II, and a fragment of the 60-MDa plasmid. The identities of PCR products were confirmed by hybridization using DIG-labeled internal oligonucleotide probes and colorimetric detection with anti-DIG Fab fragments conjugated to alkaline phosphatase. This method yielded positive results with all reference strains of EHEC serogroup O157, including serotypes O157:H7, O157:NM, and O157:H−, and negative results were obtained with all strains of nontoxigenic E. coli serogroup O157, other serotypes of E. coli, and other bacterial species. The detection limit of the method was 65 colony-forming units (CFU) of E. coli O157:H7. All 29 cultures of EHEC O157:H7 isolated from meat samples and identified by biochemical and serological tests were positive in the multiplex PCR. EHEC O157:H7 was identified from all of 70 experimentally inoculated ground beef and milk samples which had initial inocula of 4 to 9 CFU/g (ml) and were subjected to a 6-h enrichment culturing. The multiplex PCR procedure could be very useful for routine examinations of food samples for the presence of EHEC O157.

scholarly journals Urease of Enterohemorrhagic Escherichia coli: Evidence for Regulation by Fur and a trans-Acting Factor

2002 ◽  
Vol 70 (2) ◽  
pp. 1027-1031 ◽  
Author(s):  
Susan R. Heimer ◽  
Rod A. Welch ◽  
Nicole T. Perna ◽  
György Pósfai ◽  
Peter S. Evans ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Acid Tolerance ◽  
Enterohemorrhagic Escherichia Coli ◽  
Gene Fragment ◽  
Enzymatic Assays ◽  
E Coli ◽  
Urease Gene ◽  
Genomic Analyses ◽  
Serovar Typhimurium ◽  
K 12

ABSTRACT Recent genomic analyses of Escherichia coli O157:H7 strain EDL933 revealed two loci encoding urease gene homologues (ureDABCEFG), which are absent in nonpathogenic E. coli strain K-12. This report demonstrates that the cloned EDL933 ure gene cluster is capable of synthesizing urease in an E. coli DH5α background. However, when the gene fragment is transformed back into the native EDL933 background, the enzymatic activity of the cloned determinants is undetectable. We speculate that an unidentified trans-acting factor in enterohemorrhagic E. coli (EHEC) is responsible for this regulation of ure expression. In addition, Fur-like recognition sites are present in three independent O157:H7 isolates upstream of ureD and ureA. Enzymatic assays confirmed a difference in urease expression of cloned EHEC ure clusters in E. coli MC3100Δfur. Likewise, interruption of fur in O157:H7 isolate IN1 significantly diminished urease activity. We propose that, similar to the function of Fur in regulating the acid response of Salmonella enterica serovar Typhimurium, it modulates urease expression in EHEC, perhaps contributing to the acid tolerance of the organism.

scholarly journals Effect of Temperature, pH and Plasmids on In Vitro Biofilm Formation in Escherichia coli

Acta Naturae ◽  
2018 ◽  
Vol 10 (4) ◽  
pp. 129-132 ◽  
Author(s):  
A. Mathlouthi ◽  
E. Pennacchietti ◽  
D. De Biase
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Acid Resistance ◽  
Transcriptional Regulators ◽  
Growth Conditions ◽  
Luria Bertani ◽  
Effect Of Temperature ◽  
E Coli ◽  
Static Conditions

Acid resistance (AR) in Escherichia coli is an important trait that protects this microorganism from the deleterious effect of low-pH environments. Reports on biofilm formation in E. coli K12 showed that the genes participating in AR were differentially expressed. Herein, we investigated the relationship between AR genes, in particular those coding for specific transcriptional regulators, and their biofilm-forming ability at the phenotypic level. The latter was measured in 96-well plates by staining the bacteria attached to the well, following 24-hour growth under static conditions, with crystal violet. The growth conditions were as follows: Luria Bertani (LB) medium at neutral and acidic pH, at 37C or 25C. We observed that the three major transcriptional regulators of the AR genes (gadX, gadE, gadW) only marginally affected biofilm formation in E. coli. However, a striking and novel finding was the different abilities of all the tested E. coli strains to form a biofilm depending on the temperature and pH of the medium: LB, pH 7.4, strongly supported biofilm formation at 25C, with biofilm being hardly detectable at 37C. On the contrary, LB, pH 5.5, best supported biofilm formation at 37C. Moreover, we observed that when E. coli carried a plasmid, the presence of the plasmid itself affected the ability to develop a biofilm, typically by increasing its formation. This phenomenon varies from plasmid to plasmid, depends on growth conditions, and, to the best of our knowledge, remains largely uninvestigated.

Effect of Stress on Non-O157 Shiga Toxin–Producing Escherichia coli†

2012 ◽  
Vol 75 (12) ◽  
pp. 2241-2250 ◽  
Author(s):  
JAMES L. SMITH ◽  
PINA M. FRATAMICO
Keyword(s):  
Escherichia Coli ◽  
Foodborne Pathogens ◽  
Stress Responses ◽  
Shiga Toxin ◽  
Acid Stress ◽  
Acid Tolerance ◽  
Arginine Decarboxylase ◽  
E Coli ◽  
Tolerance Response ◽  
Inspection Service

Non-O157 Shiga toxin–producing Escherichia coli (non-O157 STEC) strains have emerged as important foodborne pathogens worldwide. Non-O157 STEC serogroups O26, O45, O103, O111, O121, and O145 have been declared as adulterants in beef by the U.S. Department of Agriculture Food Safety and Inspection Service. While documentation is limited, treatments including heat and acid that have been shown to inactivate E. coli O157:H7 will likely also destroy non-O157 STEC; however, non-O157 STEC strains show variability in their responses to stress. It has been shown that non-O157 STEC may survive in fermented sausages and cheeses, and treatments such as high pressure may be necessary to eliminate non-O157 STEC from these products. The mechanisms used by non-O157 STEC to resist acid environments are similar to those used by O157:H7 strains and include the acid tolerance response, the oxidative system, and the glutamate and arginine decarboxylase systems. However, one study demonstrated that some non-O157 STEC strains utilize a chaperone-based acid stress response (HdeA and HdeB) to combat acidic conditions, which is lacking in E. coli O157:H7. Genomic studies suggest that while non-O157 STEC can cause diseases similar to those caused by E. coli O157:H7, O157 and non-O157 STECs have different evolutionary histories. Non-O157 STECs are a heterogeneous group of organisms, and there is currently a limited amount of information on their virulence, fitness, and stress responses, rendering it difficult to draw firm conclusions on their behavior when exposed to stress in the environment, in food, and during processing.

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