Mechanisms in E. coli heat-stable toxin (STA)-mediated duodenal mucosal bicarbonate secretion in health and cystic fibrosis (CF)

DNA probes that identify genes coding for heat-labile type I (LT-I) and heat-stable type 1 (ST-I) enterotoxins, enteropathogenic Escherichia coli adherence factor (EAF), and Shigella-like, invasiveness (INV) are used to evaluate the sensitivity and specificity of stool blots in comparison with the sensitivity and specificity of colony blots in detecting enteropathoghens. The sensitivities of the probes in stool blots are 91.7% for the LT-I probe, 76.9% for the ST-I probes, 78.9% for the EAF probe, and 45.5% for the INV probe. The specificity of all probes is higher than 95%. In general, the stool blot method identifies as many if not more LT-I-, ST-I-, and EAF-producing E. coli infections than the colony blots. Key words: DNA probes, stool blots, enteropathogens, diagnosis.

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Erratum

Epidemiology and Infection ◽

10.1017/s0950268803009592 ◽

2003 ◽

Vol 130 (3) ◽

pp. 573-573

Author(s):

Z. ZHOU ◽

J. OGASAWARA ◽

Y. NISHIKAWA

Keyword(s):

Escherichia Coli ◽

E Coli ◽

Heat Stable ◽

Epidemiol Infect

Epidemiol. Infect. 128 (2002), 363–371An outbreak of gastroenteritis in Osaka, Japan due toEscherichia coliserogroup O166[ratio ]H15 that had a coding gene for enteroaggregativeE. coliheat-stable enterotoxin 1 (EAST1)Tables 1 and 2 were omitted

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Evolution of diarrheagenic Escherichia coli pathotypes in India

Journal of Laboratory Physicians ◽

10.4103/jlp.jlp_58_19 ◽

2019 ◽

Vol 11 (04) ◽

pp. 346-351

Author(s):

Pankaj Singh ◽

Sharda C. Metgud ◽

Subarna Roy ◽

Shashank Purwar

Keyword(s):

Escherichia Coli ◽

Cross Sectional Study ◽

Clinical Settings ◽

Accurate Identification ◽

Cross Sectional ◽

E Coli ◽

Medical College ◽

Diarrheagenic Escherichia Coli ◽

Proper Management ◽

Heat Stable

Abstract CONTEXT: Diarrheagenic Escherichia coli (DEC) is the leading cause of infectious diarrhea in developing countries. On the basis of virulence and phenotypic characteristics, the DEC is categorized into multiple pathotypes. Each pathotype has different pathogenesis and geographical distribution. Thus, the proper management of disease relies on rapid and accurate identification of DEC pathotypes. AIMS: The aim of the study was to determine the prevalence of DEC pathotypes in India. MATERIALS AND METHODS: A cross-sectional study was carried out between January 2008 and December 2012 at Jawaharlal Nehru Medical College and KLES Dr. Prabhakar Kore Hospital and Medical Research Center, Belgaum (Karnataka), India. A total of 300 stool samples were collected from diarrhea patients with age >3 months. The DEC was identified by both conventional and molecular methods. RESULTS: Of 300 samples, E. coli were detected in 198 (66%) and 170 (56.6%) samples by culture and polymerase chain reaction, respectively. Among DEC (n = 198) isolates, eae gene (59.5%) was the most prevalent followed by stx (27.7%), east (27.2%), elt (12.6%), est (10.6%), ipaH (5.5%), and eagg (1.5%) genes. On the basis of virulence genes, enteropathogenic E. coli (33.8%) was the most common pathotype followed by Shiga toxin-producing E. coli (STEC, 23.2%), enterotoxigenic E. coli (ETEC, 13.6%), enteroinvasive E. coli (5.5%), enteroaggregative heat-stable enterotoxin 1-harboring E. coli (EAST1EC, 4.5%), STEC/ETEC (3.5%), STEC/enteroaggregative E. coli (STEC/EAEC, 1.0%), and EAEC (0.05%). CONCLUSIONS: The hybrid DEC is potentially more virulent than basic pathotypes. The pathotyping should be included in clinical settings for the proper management of DEC-associated diarrhea.

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Epidemiology of E. coli in Cystic Fibrosis Airways Demonstrates the Capacity for Persistent Infection but Not Patient-Patient Transmission

Frontiers in Microbiology ◽

10.3389/fmicb.2020.00475 ◽

2020 ◽

Vol 11 ◽

Author(s):

Conrad Izydorczyk ◽

Barbara Waddell ◽

Brett D. Edwards ◽

Jasper Greysson-Wong ◽

Michael G. Surette ◽

...

Keyword(s):

Cystic Fibrosis ◽

Persistent Infection ◽

E Coli

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Full Capacity of Recombinant Escherichia coliHeat-Stable Enterotoxin Fusion Proteins for Extracellular Secretion, Antigenicity, Disulfide Bond Formation, and Activity

Infection and Immunity ◽

10.1128/iai.68.7.4064-4074.2000 ◽

2000 ◽

Vol 68 (7) ◽

pp. 4064-4074 ◽

Cited By ~ 16

Author(s):

Isabelle Batisson ◽

Maurice Der Vartanian ◽

Brigitte Gaillard-Martinie ◽

Michel Contrepois

Keyword(s):

Disulfide Bonds ◽

Secretory Pathway ◽

Biological Properties ◽

Leader Peptide ◽

Dependent Manner ◽

Reporter Protein ◽

E Coli ◽

Heat Stable

ABSTRACT We have successfully used the major subunit ClpG ofEscherichia coli CS31A fimbriae as an antigenic and immunogenic exposure-delivery vector for various heterologous peptides varying in nature and length. However, the ability of ClpG as a carrier to maintain in vitro and in vivo the native biological properties of passenger peptide has not yet been reported. To address this possibility, we genetically fused peptides containing all or part of the E. coli human heat-stable enterotoxin (STh) sequence to the amino or carboxyl ends of ClpG. Using antibodies to the ClpG and STh portions for detecting the hybrids; AMS (4-acetamido-4′-maleimidylstilbene-2,2′-disulfonate), a potent free thiol-trapping reagent, for determining the redox state of STh in the fusion; and the suckling mouse assay for enterotoxicity, we demonstrated that all ClpG-STh proteins were secreted in vitro and in vivo outside the E. coli cells in a heat-stable active oxidized (disulfide-bonded) form. Indeed, in contrast to many earlier studies, blocking the natural NH2 or COOH extremities of STh had, in all cases, no drastic effect on cell release and toxin activity. Only antigenicity of STh C-terminally extended with ClpG was strongly affected in a conformation-dependent manner. These results suggest that the STh activity was not altered by the chimeric structure, and therefore that, like the natural toxin, STh in the fusion had a spatial structure flexible enough to be compatible with secretion and enterotoxicity (folding and STh receptor recognition). Our study also indicates that disulfide bonds were essential for enterotoxicity but not for release, that spontaneous oxidation by molecular oxygen occurred in vitro in the medium, and that the E. coli cell-bound toxin activity in vivo resulted from an effective export processing of hybrids and not cell lysis. None of the ClpG-STh subunits formed hybrid CS31A-STh fimbriae at the cell surface ofE. coli, and a strong decrease in the toxin activity was observed in the absence of CS31A helper proteins. In fact, chimeras translocated across the outer membrane as a free folded monomer once they were guided into the periplasm by the ClpG leader peptide through the CS31A-dependent secretory pathway. In summary, ClpG appears highly attractive as a carrier reporter protein for basic and applied research through the engineering of E. coli for culture supernatant delivery of an active cysteine-containing protein, such as the heat-stable enterotoxin.

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Properties of human testis-specific lactate dehydrogenase expressed from Escherichia coli

Biochemical Journal ◽

10.1042/bj2730587 ◽

1991 ◽

Vol 273 (3) ◽

pp. 587-592 ◽

Cited By ~ 22

Author(s):

K M LeVan ◽

E Goldberg

Keyword(s):

Escherichia Coli ◽

Lactate Dehydrogenase ◽

Prokaryotic Expression ◽

Specific Activity ◽

Human Testis ◽

Reading Frame ◽

E Coli ◽

Heat Stable ◽

Prokaryotic Expression Vector ◽

Tac Promoter

The cDNA encoding the C4 isoenzyme of lactate dehydrogenase (LDH-C4) was engineered for expression in Escherichia coli. The Ldh-c open reading frame was constructed as a cassette for production of the native protein. The modified Ldh-c cDNA was subcloned into the prokaryotic expression vector pKK223-3. Transformed E. coli cells were grown to mid-exponential phase, and induced with isopropyl beta-D-thiogalactopyranoside for positive regulation of the tac promoter. Induced cells expressed the 35 kDa subunit, which spontaneously formed the enzymically active 140 kDa tetramer. Human LDH-C4 was purified over 200-fold from litre cultures of cells by AMP and oxamate affinity chromatography to a specific activity of 106 units/mg. The enzyme was inhibited by pyruvate concentrations above 0.3 mM, had a Km for pyruvate of 0.03 mM, a turnover number (nmol of NADH oxidized/mol of LDH-C4 per min at 25 degrees C) of 14,000 and was heat-stable.

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Escherichia coli heat-stable toxin: its effect on motility of the small intestine

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1982.242.4.g360 ◽

1982 ◽

Vol 242 (4) ◽

pp. G360-G363 ◽

Cited By ~ 1

Author(s):

J. R. Mathias ◽

J. Nogueira ◽

J. L. Martin ◽

G. M. Carlson ◽

R. A. Giannella

Keyword(s):

Escherichia Coli ◽

Small Intestine ◽

Action Potential ◽

Myoelectric Activity ◽

Complex Activity ◽

E Coli ◽

Heat Stable ◽

Rabbit Small Intestine ◽

Weight Substance

Escherichia coli heat-stable enterotoxin is a low-molecular-weight substance that has been shown to induce the active secretion of fluid and electrolytes in the small intestine. In this study, we have characterized the effects of purified E. coli heat-stable toxin (ST, strain 18D, serotype 042:K86:H37) on the motility of rabbit small intestine by using myoelectric recording techniques. Substances, such as cholera toxin, that activate the adenylate cyclase-cAMP system induced predominantly migrating action-potential complex activity. E. coli ST, a toxin that activates the guanylate cyclase-cGMP system, was infused into isolated in vivo ileal loops of New Zealand White rabbits. Inactivated toxin was also studied by exposing the ST to 1 mM dithiothreitol for 90 min. Active E. coli ST induced only repetitive bursts of action potentials. When the toxin was inactivated with dithiothreitol, no alteration in myoelectric activity was observed. We speculate that repetitive bursts of action-potential activity may represent a virulent factor of the bacterium, altering motor activity to slow transit and allowing for bacterial proliferation and invasion.

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LeTetR Positively Regulates 3-Hydroxylation of the Antifungal HSAF and Its Analogs in Lysobacter enzymogenes OH11

Molecules ◽

10.3390/molecules25102286 ◽

2020 ◽

Vol 25 (10) ◽

pp. 2286

Author(s):

Lingjun Yu ◽

Vimmy Khetrapal ◽

Fengquan Liu ◽

Liangcheng Du

Keyword(s):

Fatty Acid ◽

Antifungal Activity ◽

Biocontrol Agent ◽

Biosynthetic Gene Cluster ◽

E Coli ◽

Lysobacter Enzymogenes ◽

Heat Stable ◽

Fatty Acid Hydroxylase ◽

Hydroxylase Gene ◽

Antibiotic Compounds

The biocontrol agent Lysobacter enzymogenes OH11 produces several structurally distinct antibiotic compounds, including the antifungal HSAF (Heat Stable Antifungal Factor) and alteramides, along with their 3-dehydroxyl precursors (3-deOH). We previously showed that the 3-hydroxylation is the final step of the biosynthesis and is also a key structural moiety for the antifungal activity. However, the procedure through which OH11 regulates the 3-hydroxylation is still not clear. In OH11, the gene orf3232 was predicted to encode a TetR regulator (LeTetR) with unknown function. Here, we deleted orf3232 and found that the LeTetR mutant produced very little HSAF and alteramides, while the 3-deOH compounds were not significantly affected. The production of HSAF and alteramides was restored in orf3232-complemented mutant. qRT-PCR showed that the deletion of orf3232 impaired the transcription of a putative fatty acid hydroxylase gene, orf2195, but did not directly affect the expression of the HSAF biosynthetic gene cluster (hsaf). When an enzyme extract from E. coli expressing the fatty acid hydroxylase gene, hsaf-orf7, was added to the LeTetR mutant, the production of HSAF and alteramides increased by 13–14 fold. This study revealed a rare function of the TetR family regulator, which positively controls the final step of the antifungal biosynthesis and thus controls the antifungal activity of the biocontrol agent.

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