Detection of Escherichia coli cytotoxins by enzyme-linked immunosorbent assays

1991 ◽  
Vol 37 (8) ◽  
pp. 650-653 ◽  
Author(s):  
Joan I. Speirs ◽  
Mumtaz Akhtar
Keyword(s):  
Escherichia Coli ◽  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Vero Cell ◽  
Key Words ◽  
E Coli ◽  
Treated Cell ◽  
Cell Extracts ◽  
Immunosorbent Assays ◽  
Key Words Escherichia Coli

Sandwich enzyme-linked immunosorbent assays (ELISAs) were developed to detect Escherichia coli cytotoxins. Wells were coated with monoclonal antibodies from hybridomas 13C4 and (or) 11E10, and biotin conjugates of these antibodies were used for detecting verotoxin 1 and Shiga-like toxin II, respectively. Sensitivities were about 100 and 200 cytotoxic doses, respectively. Verotoxin 2 was detected by ELISA with monoclonal antibody 11E10, but at a sensitivity of only about 4000 cytotoxic doses. ELISA results of polymyxin-treated cell extracts from cultures of 67 E. coli strains were in agreement with Vero cell assay as regards the presence and type of toxin. Key words: Escherichia coli, cytotoxin, ELISA.

The effect of fasting and diet on fecal shedding of Escherichia coli O157:H7 by cattle

2000 ◽  
Vol 80 (4) ◽  
pp. 741-744 ◽  
Author(s):  
S. J. Buchko ◽  
R. A. Holley ◽  
W. O. Olson ◽  
V. P. J. Gannon ◽  
D. M. Veira
Keyword(s):  
Escherichia Coli ◽  
Key Words ◽  
Escherichia Coli O157 ◽  
Fecal Shedding ◽  
E Coli ◽  
Feed Withdrawal ◽  
Alfalfa Silage ◽  
Key Words Escherichia Coli

Cattle naturally infected with Escherichia coli O157:H7 were used to assess the effects of diet and feed withdrawal on the fecal shedding of E. coli O157:H7. Animals were fed an 80% concentrate diet (80% barley and 20% alfalfa silage), fasted for 48 h, fed a 100% forage diet (alfalfa silage), fasted for 48 h, and subsequently re-fed 100% forage (alfalfa silage). There were no differences in the numbers of animals positive for the shedding of E. coli O157:H7 when fed an 80% barley diet or an all-forage diet (P > 0.05) or during the fasting periods following each diet (P > 0.05). Upon re-feeding an all-forage diet following a 48-h fast, animals positive for E. coli O157:H7 shedding increased (P < 0.05), with 42.5% of the animals shedding the pathogen after 5 d. Re-feeding 100% forage following fasting appeared to have increased the number of animals shedding E. coli O157:H7 in their feces, which may have been influenced by diet in addition to fasting. Key words: Escherichia coli O157:H7, fasting, diet, cattle, fecal shedding

scholarly journals UJI TOKSISITAS ESCHERICHIA COLI ASAL DAGING TERHADAP SEL VERO

2018 ◽  
Vol 18 (2) ◽  
Author(s):  
Widodo Suwito ◽  
Andriani Andriani
Keyword(s):  
Escherichia Coli ◽  
Vero Cell ◽  
Key Words ◽  
Vero Cells ◽  
E Coli ◽  
Traditional Market ◽  
Cattle Faeces

Abstract: Verotoxigenic Escherichia coli (VTEC) is responsible for serious human illnesses. Source of VTEC is cattle faeces which beef contamination. The aims of this study was to determine the ability of E. coli which beef contamination from traditional market to damage the vero cells monolayer. A total of 35 E. coli isolates and vero cells monolayer were used in these study. All isolates E. coli were re-indentified with biochemistry and vero cells monolayer were uesed to determination verotoxigenecity tests. None of E. coli isolates showed damage the vero cells monolayer, so there are not verotoxigenik E. coli. The study showed that all isolate E. coli which beef contamination from tradiotional market none damage the vero cells, so there are not verotoxigenic. Key words: E.coli, beef, vero cell Abstrak: Escherichia coli verotoksigenik (VTEC) menyebabkan penyakit pada manusia. Sumber VTEC adalah feses sapi yang dapat mengkontaminasi daging. Tujuan penelitian ini adalah mengetahui kemampuan E. coli yang diisolasi dari daging sapi di pasar tradisional dalam merusak sel vero monolayer. Sebanyak 35 isolat E. coli dan sel vero monolayer digunakan dalam penelitian ini. Isolat E. coli di identifikasi ulang secara biokimia dan untuk menentukan sifat verotoksigenesitasnya menggunakan sel vero monolayer. Semua isolat E. coli tidak bersifat verotoksigenik karena tidak mampu merusak sel vero. Penelitian ini menunjukkan bahwa E. coli yang mengkontaminasi daging sapi dari pasar tradisional tidak bersifat verotoksigenik.  Kata kunci: E.coli, daging, sel vero. 

Antibiotic resistance indexing of Escherichia coli to identify sources of fecal contamination in water

1990 ◽  
Vol 36 (12) ◽  
pp. 891-894 ◽  
Author(s):  
Charles W. Kaspar ◽  
Janie L. Burgess ◽  
Ivor T. Knight ◽  
R. R. Colwell
Keyword(s):  
Escherichia Coli ◽  
Water Quality ◽  
Antibiotic Resistance ◽  
Key Words ◽  
Fecal Contamination ◽  
Antibiotic Resistant ◽  
E Coli ◽  
Rural Water ◽  
Key Words Escherichia Coli

A total of 202 Escherichia coli isolated from urban and rural water were tested with 11 antibiotics to assess the prevalence of antibiotic resistance from each source. Urban waters harbored higher percentages of resistant E. coli strains than rural waters. Antibiotic-resistant E. coli may offer an index of water quality related to source. Key words: Escherichia coli, antibiotic resistance, indicator.

scholarly journals Murine Monoclonal Antibodies Specific for Lipopolysaccharide of Escherichia coli O26 and O111

2000 ◽  
Vol 66 (9) ◽  
pp. 4124-4127 ◽  
Author(s):  
Mildred Rivera-Betancourt ◽  
James E. Keen
Keyword(s):  
Escherichia Coli ◽  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Bacterial Strains ◽  
Antigen Specificity ◽  
E Coli ◽  
O Antigen ◽  
Murine Monoclonal Antibodies

ABSTRACT Monoclonal antibody (MAb) 12F5 reacted with 35 Escherichia coli O26 isolates and cross-reacted with 1 of 365 non-E. coli O26 isolates. MAb 15C4 reacted with 30 E. coliO111 strains and 8 Salmonella O35 strains (possessing identical O antigen) but not with 362 other bacterial strains. Lipopolysaccharide immunoblots confirmed MAb O-antigen specificity.

scholarly journals Isolation of a Gene Responsible for the Oxidation oftrans-Anethole topara-Anisaldehyde by Pseudomonas putida JYR-1 and Its Expression in Escherichia coli

2012 ◽  
Vol 78 (15) ◽  
pp. 5238-5246 ◽  
Author(s):  
Dongfei Han ◽  
Ji-Young Ryu ◽  
Robert A. Kanaly ◽  
Hor-Gil Hur
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Putida ◽  
Hypothetical Protein ◽  
Aldehyde Group ◽  
Agrobacterium Vitis ◽  
T7 Promoter ◽  
Content Type ◽  
E Coli ◽  
Cell Extracts ◽  
Isoeugenol Monooxygenase

ABSTRACTA plasmid, pTA163, inEscherichia colicontained an approximately 34-kb gene fragment fromPseudomonas putidaJYR-1 that included the genes responsible for the metabolism oftrans-anethole to protocatechuic acid. Three Tn5-disrupted open reading frame 10 (ORF 10) mutants of plasmid pTA163 lost their abilities to catalyzetrans-anethole. Heterologously expressed ORF 10 (1,047 nucleotides [nt]) under a T7 promoter inE. colicatalyzed oxidative cleavage of a propenyl group oftrans-anethole to an aldehyde group, resulting in the production ofpara-anisaldehyde, and this gene was designatedtao(trans-anetholeoxygenase). The deduced amino acid sequence of TAO had the highest identity (34%) to a hypothetical protein ofAgrobacterium vitisS4 and likely contained a flavin-binding site. Preferred incorporation of an oxygen molecule from water intop-anisaldehyde using18O-labeling experiments indicated stereo preference of TAO for hydrolysis of the epoxide group. Interestingly, unlike the narrow substrate range of isoeugenol monooxygenase fromPseudomonas putidaIE27 andPseudomonas nitroreducensJin1, TAO fromP. putidaJYR-1 catalyzed isoeugenol,O-methyl isoeugenol, and isosafrole, all of which contain the 2-propenyl functional group on the aromatic ring structure. Addition of NAD(P)H to the ultrafiltered cell extracts ofE. coli(pTA163) increased the activity of TAO. Due to the relaxed substrate range of TAO, it may be utilized for the production of various fragrance compounds from plant phenylpropanoids in the future.

Comparison of colony and stool blots for detection of enteropathogens by DNA probes

1991 ◽  
Vol 37 (5) ◽  
pp. 407-410
Author(s):  
Mônica A. M. Vieira ◽  
Beatriz E. C. Guth ◽  
Tânia A. T. Gomes
Keyword(s):  
Escherichia Coli ◽  
Key Words ◽  
Sensitivity And Specificity ◽  
Dna Probes ◽  
Type I ◽  
Enteropathogenic Escherichia Coli ◽  
Key Words Dna ◽  
E Coli ◽  
Heat Stable

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.

scholarly journals Protective Roles of γδ T Cells and Interleukin-15 in Escherichia coli Infection in Mice

1998 ◽  
Vol 66 (7) ◽  
pp. 3270-3278 ◽  
Author(s):  
M. Takano ◽  
H. Nishimura ◽  
Y. Kimura ◽  
Y. Mokuno ◽  
J. Washizu ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Monoclonal Antibody ◽  
T Cells ◽  
T Cell Receptor ◽  
Host Defense ◽  
Γδ T Cells ◽  
Cell Receptor ◽  
E Coli ◽  
Escherichia Coli Infection ◽  
Interleukin 15

The number of γδ T cells in the peritoneal cavity was increased after an intraperitoneal (i.p.) infection with Escherichia coli in lipopolysaccharide (LPS)-responsive C3H/HeN mice but not in LPS-hyporesponsive C3H/HeJ mice. The γδ T cells preferentially expressed invariant Vγ6 and Vδ1 chains and proliferated to produce a large amount of gamma interferon in the presence of LPS. Mice depleted of γδ T cells by T-cell receptor δ gene mutation showed impaired resistance against E. coli as assessed by bacterial growth. Macrophages from C3H/HeN mice infected with E. coli expressed higher levels of interleukin-15 (IL-15) mRNA than those from the infected C3H/HeJ mice. Administration of anti-IL-15 monoclonal antibody inhibited, albeit partially, the appearance of γδ T cells in C3H/HeN mice after E. coli infection and diminished the host defense against the infection. These results suggest that LPS-stimulated γδ T cells play an important role in the host defense against E. coli infection and that IL-15 may be partly involved in the protection via an increase in the γδ T cells.

Use of a Shiga Toxin (Stx)-Enzyme-Linked Immunosorbent Assay and Immunoblot for Detection and Isolation of Stx-Producing Escherichia coli from Naturally Contaminated Beef

2000 ◽  
Vol 63 (9) ◽  
pp. 1167-1172 ◽  
Author(s):  
HEBA NASHED ATALLA ◽  
ROGER JOHNSON ◽  
SCOTT MCEWEN ◽  
R. W. USBORNE ◽  
C. L. GYLES
Keyword(s):  
Escherichia Coli ◽  
Vero Cell ◽  
Immunosorbent Assay ◽  
Shiga Toxin ◽  
Total Coliform ◽  
Complete Agreement ◽  
Enzyme Linked Immunosorbent Assay ◽  
Spearman Correlation ◽  
Southern Ontario ◽  
E Coli

The purpose of this study was to evaluate an enzyme-linked immunosorbent assay (ELISA) and an immunoblot procedure for detection and isolation of Shiga toxin-producing Escherichia coli (STEC) from beef, and to correlate the presence of STEC in beef with E. coli and total coliform counts. A total of 120 samples of boneless beef supplied to a meat processor in southern Ontario were tested for the presence of STEC, E. coli, and total coliforms. Following enrichment in modified tryptic soy broth, samples were screened for Shiga toxin (Stx) by a Stx-ELISA and a Vero cell assay (VCA). Samples that were positive in the Stx-ELISA were subjected to the Stx-immunoblot for STEC isolation. Overall, 33.3% of samples were positive in the VCA, and 34.2% were positive in the Stx-ELISA. There was almost complete agreement between the Stx-ELISA and the VCA results (kappa = 0.98). The sensitivity and specificity of the Stx-ELISA with respect to the VCA were 100% and 98.75%, respectively. STEC were isolated by the Stx-immunoblot from 87.8% of the samples that were positive in the Stx-ELISA. The STEC isolates belonged to 19 serotypes, with serotype O113:H21 accounting for 10 of 41 isolates. No STEC of serotype O157:H7 were isolated. There was a significant correlation between E. coli counts and total coliform counts (Spearman correlation coefficient = 0.68, P &lt; 0.01). The E. coli count was positively correlated with detection of STEC by both the Stx-ELISA and the VCA (P &lt; 0.01).

scholarly journals Identification of a Reactivating Factor for Adenosylcobalamin-Dependent Ethanolamine Ammonia Lyase

2004 ◽  
Vol 186 (20) ◽  
pp. 6845-6854 ◽  
Author(s):  
Koichi Mori ◽  
Reiko Bando ◽  
Naoki Hieda ◽  
Tetsuo Toraya
Keyword(s):  
Escherichia Coli ◽  
Molecular Weight ◽  
Protein S ◽  
Amino Acid Residues ◽  
Permeabilized Cells ◽  
E Coli ◽  
Cell Extracts ◽  
Auxiliary Protein

ABSTRACT The holoenzyme of adenosylcobalamin-dependent ethanolamine ammonia lyase undergoes suicidal inactivation during catalysis as well as inactivation in the absence of substrate. The inactivation involves the irreversible cleavage of the Co-C bond of the coenzyme. We found that the inactivated holoenzyme undergoes rapid and continuous reactivation in the presence of ATP, Mg2+, and free adenosylcobalamin in permeabilized cells (in situ), homogenate, and cell extracts of Escherichia coli. The reactivation was observed in the permeabilized E. coli cells carrying a plasmid containing the E. coli eut operon as well. From coexpression experiments, it was demonstrated that the eutA gene, adjacent to the 5′ end of ethanolamine ammonia lyase genes (eutBC), is essential for reactivation. It encodes a polypeptide consisting of 467 amino acid residues with predicted molecular weight of 49,599. No evidence was obtained that shows the presence of the auxiliary protein(s) potentiating the reactivation or associating with EutA. It was demonstrated with purified recombinant EutA that both the suicidally inactivated and O2-inactivated holoethanolamine ammonia lyase underwent rapid reactivation in vitro by EutA in the presence of adenosylcobalamin, ATP, and Mg2+. The inactive enzyme-cyanocobalamin complex was also activated in situ and in vitro by EutA under the same conditions. Thus, it was concluded that EutA is the only component of the reactivating factor for ethanolamine ammonia lyase and that reactivation and activation occur through the exchange of modified coenzyme for free intact adenosylcobalamin.

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