Detection of O25b-ST131 clone, CTX-M-1 and CTX-M-15 genes via real-time PCR in Escherichia coli strains in patients with UTIs obtained from a university hospital in Istanbul

Shiga toxin-producing Escherichia coli (STEC) are associated with acute gastroenteritis worldwide, which induces a high economic burden on both healthcare and individuals. Culture-independent diagnostic tests (CIDT) in frontline microbiology laboratories have been implemented in Alberta since 2019. The objectives of this study were to determine the association between gene detection and culture positivity over time using STEC microbiological clearance samples and also to establish the frequency of specimen submission. Both stx genes’ amplification by real-time PCR was performed with DNA extracted from stool samples using the easyMAG system. Stools were inoculated onto chromogenic agar for culture. An association between gene detection and culture positivity was found to be independent of which stx gene was present. CIDT can provide rapid reporting with less hands-on time and technical expertise. However, culture is still important for surveillance and early cluster detection. In addition, stool submissions could be reduced from daily to every 3–5 days until a sample is negative by culture.

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Real-Time PCR Determination of Escherichia coli Genomic DNA Contamination in Plasmid Preparations

Analytical Biochemistry ◽

10.1006/abio.2001.5480 ◽

2002 ◽

Vol 301 (1) ◽

pp. 151-153 ◽

Cited By ~ 10

Author(s):

Adrián Vilalta ◽

Vanessa Whitlow ◽

Terrie Martin

Keyword(s):

Escherichia Coli ◽

Real Time ◽

Real Time Pcr ◽

Genomic Dna ◽

Dna Contamination

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Optimization of a 3′-minor groove binder-DNA probe targeting the uidA gene for rapid identification of Escherichia coli O157:H7 using real-time PCR

Molecular and Cellular Probes ◽

10.1016/j.mcp.2003.07.001 ◽

2003 ◽

Vol 17 (6) ◽

pp. 275-280 ◽

Cited By ~ 27

Author(s):

Ken J Yoshitomi ◽

Karen C Jinneman ◽

Stephen D Weagant

Keyword(s):

Escherichia Coli ◽

Real Time ◽

Real Time Pcr ◽

Minor Groove ◽

Rapid Identification ◽

Dna Probe ◽

Escherichia Coli O157 ◽

Uida Gene ◽

Minor Groove Binder

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Real-time PCR for differentiation of F18 variants among enterotoxigenic and Shiga toxin-producing Escherichia coli from piglets with diarrhoea and oedema disease

The Veterinary Journal ◽

10.1016/j.tvjl.2013.07.021 ◽

2013 ◽

Vol 198 (2) ◽

pp. 538-540 ◽

Cited By ~ 8

Author(s):

Jae-Won Byun ◽

Byeong Yeal Jung ◽

Ha-Young Kim ◽

John M. Fairbrother ◽

Myoung-Heon Lee ◽

...

Keyword(s):

Escherichia Coli ◽

Real Time ◽

Shiga Toxin ◽

Real Time Pcr ◽

Oedema Disease

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Application of quantitative real-time PCR compared to filtration methods for the enumeration of Escherichia coli in surface waters within Vietnam

Journal of Water and Health ◽

10.2166/wh.2016.173 ◽

2016 ◽

Vol 15 (1) ◽

pp. 155-162 ◽

Cited By ~ 5

Author(s):

Pierangeli G. Vital ◽

Nguyen Thi Van Ha ◽

Le Thi Hong Tuyet ◽

Kenneth W. Widmer

Keyword(s):

Escherichia Coli ◽

Real Time ◽

Surface Waters ◽

Real Time Pcr ◽

Membrane Filtration ◽

Quantitative Real Time Pcr ◽

Wet Season ◽

Culture Methods ◽

Fecal Indicator ◽

Filtration Method

Surface water samples in Vietnam were collected from the Saigon River, rural and suburban canals, and urban runoff canals in Ho Chi Minh City, Vietnam, and were processed to enumerate Escherichia coli. Quantification was done through membrane filtration and quantitative real-time polymerase chain reaction (PCR). Mean log colony-forming unit (CFU)/100 ml E. coli counts in the dry season for river/suburban canals and urban canals were log 2.8 and 3.7, respectively, using a membrane filtration method, while using Taqman quantitative real-time PCR they were log 2.4 and 2.8 for river/suburban canals and urban canals, respectively. For the wet season, data determined by the membrane filtration method in river/suburban canals and urban canals samples had mean counts of log 3.7 and 4.1, respectively. While mean log CFU/100 ml counts in the wet season using quantitative PCR were log 3 and 2, respectively. Additionally, the urban canal samples were significantly lower than those determined by conventional culture methods for the wet season. These results show that while quantitative real-time PCR can be used to determine levels of fecal indicator bacteria in surface waters, there are some limitations to its application and it may be impacted by sources of runoff based on surveyed samples.

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Detection and quantification of Escherichia coli O157:H7 in environmental samples by real-time PCR

Journal of Applied Microbiology ◽

10.1046/j.1365-2672.2003.01848.x ◽

2003 ◽

Vol 94 (3) ◽

pp. 421-431 ◽

Cited By ~ 115

Author(s):

A.M. Ibekwe ◽

C.M. Grieve

Keyword(s):

Escherichia Coli ◽

Real Time ◽

Real Time Pcr ◽

Environmental Samples ◽

Escherichia Coli O157 ◽

Detection And Quantification

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Novel multiplex real-time PCR assays reveal a high prevalence of diarrhoeagenic Escherichia coli pathotypes in healthy and diarrhoeal children in the South of Vietnam

10.21203/rs.3.rs-18430/v1 ◽

2020 ◽

Author(s):

Vu Thuy Duong ◽

Le Thi Phuong Tu ◽

Ha Thanh Tuyen ◽

Le Thi Quynh Nhi ◽

James I Campbell ◽

...

Keyword(s):

Escherichia Coli ◽

Real Time ◽

Real Time Pcr ◽

Clinical Samples ◽

Middle Income ◽

Middle Income Countries ◽

Virulence Markers ◽

Low Middle Income Countries ◽

High Prevalence ◽

Pcr Assays

Abstract BackgroundDiarrhoeagenic Escherichia coli (DEC) infections are common in children in low-middle income countries (LMICs). However, detecting the various DEC pathotypes is complex as they cannot be differentiated by classical microbiology. We developed four multiplex real-time PCR assays were to detect virulence markers of six DEC pathotypes; specificity was tested using DEC controls and other enteric pathogens. PCR amplicons from the six E. coli pathotypes were purified and amplified to be used to optimize PCR reactions and to calculate reproducibility. After validation, these assays were applied to clinical samples from healthy and diarrhoeal Vietnamese children and associated with clinical data. ResultsThe multiplex real-time PCRs were found to be reproducible, and specific. At least one DEC variant was detected in 34.7% (978/2,815) of the faecal samples from diarrhoeal children; EAEC, EIEC and atypical EPEC were most frequent Notably, 41.2% (205/498) of samples from non-diarrhoeal children was positive with a DEC pathotype. In this population, only EIEC, which was detected in 34.3% (99/289) of diarrhoeal samples vs. 0.8% (4/498) non-diarrhoeal samples (p<0.001), was significantly associated with diarrhoea. Multiplex real-time PCR when applied to clinical samples is an efficient and high-throughput approach to DEC pathotypes. ConclusionsThis approach revealed high carriage rates of DEC pathotypes among Vietnamese children. We describe a novel diagnostic approach for DEC, which provides baseline data for future surveillance studies assessing DEC burden in LMICs.

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Quantitative detection of Escherichia coli from urine of patients with bacteriuria by real-time PCR

Molecular Diagnosis ◽

10.1007/bf03260062 ◽

2004 ◽

Vol 8 (3) ◽

pp. 179-184 ◽

Cited By ~ 10

Author(s):

Nobuyuki Hinata ◽

Toshiro Shirakawa ◽

Hiroshi Okada ◽

Katsumi Shigemura ◽

Sadao Kamidono ◽

...

Keyword(s):

Escherichia Coli ◽

Real Time ◽

Real Time Pcr ◽

Quantitative Detection

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Comparison of a New Enrichment Procedure for Shiga Toxin–Producing Escherichia coli with Five Standard Methods

Journal of Food Protection ◽

10.4315/0362-028x-68.8.1593 ◽

2005 ◽

Vol 68 (8) ◽

pp. 1593-1599 ◽

Cited By ~ 13

Author(s):

MICHAEL A. GRANT

Keyword(s):

Escherichia Coli ◽

Real Time ◽

Shiga Toxin ◽

Real Time Pcr ◽

Target Cells ◽

Macconkey Agar ◽

Standard Methods ◽

E Coli ◽

Canadian Health ◽

Health Products

A new procedure for enrichment of Escherichia coli O157:H7 and other Shiga toxin–producing E. coli was compared to five standard methods: the British Public Health Laboratory Service, International Standard Method, U.S. Department of Agriculture, Canadian Health Products and Food Branch, and U.S. Food and Drug Administration. The new procedure was comparable to the standard methods in its ability to detect target cells inoculated into foods at approximately 1 CFU g−1. Comparisons were also made of the ability of the six enrichment procedures to detect E. coli O157:H7 against a large background of competitor microorganisms. In these experiments the new procedure yielded more target cells than the other five enrichments by two to three orders of magnitude as determined by enumeration on sorbitol MacConkey agar with tellurite and cefixime and Rainbow agar with tellurite and novobiocin and by verification of presumptive colonies by real-time PCR. For example, the population of enterohemorrhagic E. coli strain 6341 recovered on sorbitol MacConkey agar with tellurite and cefixime after enrichment with the experimental method was 2.42 × 108 CFU ml−1 and 1.80 × 106 CFU ml−1 after enrichment with the Canadian Health Products and Food Branch method, the second most effective in this experiment. In addition, broth cultures resulting from each of the six enrichment procedures were used to prepare templates for real-time PCR detection of stx1/stx2. Resulting threshold cycle (Ct) values after the experimental enrichment were similar to positive control values, whereas the five standard methods produced delayed Ct values or were not detected.

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