Detection of enterotoxigenic Escherichia coli in stool specimens by polymerase chain reaction

Yavzori, Miri; Porath, N; Ochana, O; Dagan, Ron; Orni-Wasserlauf, Ruth; Cohen, Dani

doi:10.1016/s0732-8893(98)00040-6

Cited by 19 publications

(15 citation statements)

References 28 publications

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“…Schultsz et al [24] used this technique for detecting ETEC in smears of lactosepositive cultures on solid selective media. We employed a newer technique for detecting LT and ST DNA sequences directly in stool samples [19]. This method allowed us to rapidly identify the causative agent of this epidemic.…”

Section: Discussionmentioning

confidence: 99%

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A Waterborne Outbreak of Gastroenteritis in the Golan Heights due to Enterotoxigenic Escherichia coli

et al. 2000

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This report suggests that the involvement of ETEC in the etiology of waterborne diarrheal outbreaks may be underestimated, probably due to the difficulties involved in the laboratory identification of this enteropathogen. Adoption of our rapid method for the identification of ETEC, which is applicable to routine diagnostic laboratories, facilitates pathogen detection within hours, and allows early intervention in cases of widespread diarrheal epidemics.

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Section: Discussionmentioning

confidence: 99%

“…The presence in stools of genes encoding for ETEC ST and LT was determined by a PCR protocol recently developed in our laboratory [19]. Briefly, after preincubation of the swabs in brain heart infusion broth for 4 h, the fecal suspension was boiled to lyse the bacterial cells and release their DNA.…”

Section: Laboratory Analysismentioning

confidence: 99%

A Waterborne Outbreak of Gastroenteritis in the Golan Heights due to Enterotoxigenic Escherichia coli

et al. 2000

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“…Later, enzyme-linked immunosorbent assays and membrane-based DNA hybridization assays, which improved speed and ease of use, became popular. Further enhancements in methods for the detection of ETEC gave rise to PCR assays (18,24,26,28). These assays were attractive alternatives to the earlier methods because of their sensitivity and specificity, as well as their speed and use of readily available reagents.…”

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confidence: 99%

Real-Time Fluorescence PCR Assays for Detection and Characterization of Heat-Labile I and Heat-Stable I Enterotoxin Genes from Enterotoxigenic Escherichia coli

et al. 2004

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To facilitate the diagnosis of enterotoxigenic Escherichia coli (ETEC) infections in humansEnterotoxigenic Escherichia coli (ETEC) is a major cause of sporadic diarrheal disease in humans, affecting mainly children in developing countries (1,3,21) and travelers from industrialized countries visiting tropical or subtropical areas (9). ETEC strains have also caused waterborne outbreaks on cruise ships (6), food-borne outbreaks at schools (16) and restaurants (5, 11), and outbreaks among persons serving in the military (12,20,27). ETEC strains from humans cause mild or severe watery diarrhea by producing a heat-labile enterotoxin (LT I) (similar in structure to cholera toxin), heat-stable enterotoxins (ST Ia and/or ST Ib), or both (17).Initially, detection of ETEC strains was accomplished with animal assays and cell culture techniques that required specific antibodies to reliably demonstrate the presence of the target enterotoxins. Later, enzyme-linked immunosorbent assays and membrane-based DNA hybridization assays, which improved speed and ease of use, became popular. Further enhancements in methods for the detection of ETEC gave rise to PCR assays (18,24,26,28). These assays were attractive alternatives to the earlier methods because of their sensitivity and specificity, as well as their speed and use of readily available reagents. Recent advances in PCR amplification and fluorescence-based detection technologies have brought significant improvements to conventional block cycler PCR assays, thereby enabling the simultaneous, sequence-specific detection of multiple products in real time. The development of thermal cyclers with the ability to rapidly amplify genes and the capacity to continuously monitor the accumulation of specific PCR products labeled with fluorescent probes obviates the need for labor-intensive agarose gel procedures and provides a real-time, reliable identification of target genes (2,7,14,15,22,23).In the present study, we describe the development and evaluation of a set of real-time PCR assays for the LightCycler (LC) (Roche Diagnostics, Mannheim, Germany), using primers and hybridization probes that target the LT I, ST Ia (also referred to as STp), and ST Ib (also referred to as STh) genes. We used the hybridization probe format for these assays because it permitted a melting curve analysis to be performed after the amplification phase to achieve additional characterization of the target genes. The availability of rapid and specific assays for the detection and characterization of ETEC should

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“…PCR analysis for screening drinking water and environmental samples has been reported (46)(47)(48) and has been utilized to identify E. coli in primary water specimens (15,27), stool specimens (33,56), and outbreaks (23,24). In contrast, isolation of E. coli O157:H7 from water and other environmental samples is laborious.…”

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Detection, Isolation, and Molecular Subtyping of Escherichia coli O157:H7 and Campylobacter jejuni Associated with a Large Waterborne Outbreak

et al. 2003

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The largest reported outbreak of waterborne Escherichia coli O157:H7 in the United States occurred in upstate New York following a county fair in August 1999. Culture methods were used to isolate E. coli O157:H7 from specimens from 128 of 775 patients with suspected infections. Campylobacter jejuni was also isolated from stools of 44 persons who developed diarrheal illness after attending this fair. There was one case of a confirmed coinfection with E. coli O157:H7 and C. jejuni. Molecular detection of stx 1 and stx 2 Shiga toxin genes, immunomagnetic separation (IMS), and selective culture enrichment were utilized to detect and isolate E. coli O157:H7 from an unchlorinated well and its distribution points, a dry well, and a nearby septic tank. PCR for stx 1 and stx 2 was shown to provide a useful screen for toxin-producing E. coli O157:H7, and IMS subculture improved recovery. Pulsed-field gel electrophoresis (PFGE) was used to compare patient and environmental E. coli O157:H7 isolates. Among patient isolates, 117 of 128 (91.5%) were type 1 or 1a (three or fewer bands different). Among the water distribution system isolates, 13 of 19 (68%) were type 1 or 1a. Additionally, PFGE of C. jejuni isolates revealed that 29 of 35 (83%) had indistinguishable PFGE patterns. The PFGE results implicated the water distribution system as the main source of the E. coli O157:H7 outbreak. This investigation demonstrates the potential for outbreaks involving more than one pathogen and the importance of analyzing isolates from multiple patients and environmental samples to develop a better understanding of bacterial transmission during an outbreak.

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Detection of enterotoxigenic Escherichia coli in stool specimens by polymerase chain reaction

Cited by 19 publications

References 28 publications

A Waterborne Outbreak of Gastroenteritis in the Golan Heights due to Enterotoxigenic Escherichia coli

A Waterborne Outbreak of Gastroenteritis in the Golan Heights due to Enterotoxigenic Escherichia coli

Real-Time Fluorescence PCR Assays for Detection and Characterization of Heat-Labile I and Heat-Stable I Enterotoxin Genes from Enterotoxigenic Escherichia coli

Detection, Isolation, and Molecular Subtyping of Escherichia coli O157:H7 and Campylobacter jejuni Associated with a Large Waterborne Outbreak

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