Strategies for the detection of Escherichia coli O157:H7 in foods

Deisingh, Anil K.; Thompson, Michael

doi:10.1111/j.1365-2672.2003.02170.x

Cited by 175 publications

(109 citation statements)

References 46 publications

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“…scherichia coli O157:H7 is a major food-borne pathogen responsible for a number of outbreaks in animals, poultry, and humans worldwide (1,7,14). It is estimated that E. coli O157:H7 outbreaks infect over 73,500 people annually in the United States (24), and this pathogen has not only become an important food safety concern but also a serious medical and public health problem.…”

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confidence: 99%

“…It also has the potential for high-throughput selective detection of viable E. coli O157:H7 cells in other food matrices and, thus, will have an impact on the accurate microbiological and epidemiological monitoring of food safety and environmental sources. E scherichia coli O157:H7 is a major food-borne pathogen responsible for a number of outbreaks in animals, poultry, and humans worldwide (1,7,14). It is estimated that E. coli O157:H7 outbreaks infect over 73,500 people annually in the United States (24), and this pathogen has not only become an important food safety concern but also a serious medical and public health problem.…”

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confidence: 99%

“…More recently, real-time PCR is gaining popularity for its enhanced sensitivity and specificity and its speedy turnaround time. Numerous real-time PCR-based methods have been reported for rapid and sensitive detection of E. coli O157:H7 (4,7,9,10). One of the major obstacles for the PCR-based methodologies for sensitive and specific detection of E. coli O157:H7 is the selection of unique genetic makers for E. coli O157:H7.…”

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Real-Time PCR Methodology for Selective Detection of Viable Escherichia coli O157:H7 Cells by Targeting Z3276 as a Genetic Marker

Chen

2012

Appl Environ Microbiol

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The goal of this study was to develop a sensitive, specific, and accurate method for the selective detection of viable Escherichia coli O157:H7 cells in foods. A unique open reading frame (ORF), Z3276, was identified as a specific genetic marker for the detection of E. coli O157:H7. We developed a real-time PCR assay with primers and probe targeting ORF Z3276 and confirmed that this assay was sensitive and specific for E. coli O157:H7 strains (n ‫؍‬ 298). Using this assay, we can detect amounts of genomic DNA of E. coli O157:H7 as low as a few CFU equivalents. Moreover, we have developed a new propidium monoazide (PMA)-real-time PCR protocol that allows for the clear differentiation of viable from dead cells. In addition, the protocol was adapted to a 96-well plate format for easy and consistent handling of a large number of samples. Amplification of DNA from PMA-treated dead cells was almost completely inhibited, in contrast to the virtually unaffected amplification of DNA from PMA-treated viable cells. With beef spiked simultaneously with 8 ؋ 10 7 dead cells/g and 80 CFU viable cells/g, we were able to selectively detect viable E. coli O157:H7 cells with an 8-h enrichment. In conclusion, this PMA-real-time PCR assay offers a sensitive and specific means to selectively detect viable E. coli O157:H7 cells in spiked beef. It also has the potential for high-throughput selective detection of viable E. coli O157:H7 cells in other food matrices and, thus, will have an impact on the accurate microbiological and epidemiological monitoring of food safety and environmental sources. E scherichia coli O157:H7 is a major food-borne pathogen responsible for a number of outbreaks in animals, poultry, and humans worldwide (1,7,14). It is estimated that E. coli O157:H7 outbreaks infect over 73,500 people annually in the United States (24), and this pathogen has not only become an important food safety concern but also a serious medical and public health problem. In order to effectively handle future outbreaks in a timely manner, it is necessary to have ample availability of sensitive, specific, and reliable methodologies that can be used for the rapid and selective detection of E. coli O157:H7.To date, great efforts have been made to develop appropriate methodologies for the detection of E. coli O157:H7. The traditional culture methods use selective media or chromogenic agar to detect E. coli O157:H7 (23). However, major limitations of this method are that it takes days to obtain the culture while providing little accurate information about the nature of the strain or isolate itself (39). PCR has been widely used for the detection of E. coli O157:H7 from foods and environmental samples. More recently, real-time PCR is gaining popularity for its enhanced sensitivity and specificity and its speedy turnaround time. Numerous real-time PCR-based methods have been reported for rapid and sensitive detection of E. coli O157:H7 (4, 7, 9, 10). One of the major obstacles for the PCR-based methodologies for sensitive and specific detection...

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Real-Time PCR Methodology for Selective Detection of Viable Escherichia coli O157:H7 Cells by Targeting Z3276 as a Genetic Marker

Chen

2012

Appl Environ Microbiol

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“…Therefore, a simple, rapid, and sensitive detection of trace amounts of E. coli O157:H7 and other bacterial pathogens is critical for minimizing or eliminating potential infections. Traditional methods for the detection of trace amounts of bacteria require amplification or enrichment of the target bacteria in the sample (1,7,8). These methods tend to be laborious and time-consuming because of the complicated assay procedures (9)(10)(11)(12)(13)(14)(15)(16)(17)(18).…”

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A rapid bioassay for single bacterial cell quantitation using bioconjugated nanoparticles

Zhao

Hilliard

Mechery

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

529

356

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The rapid and sensitive determination of pathogenic bacteria is extremely important in biotechnology, medical diagnosis, and the current fight against bioterrorism. Current methods either lack ultrasensitivity or take a long time for analysis. Here, we report a bioconjugated nanoparticle-based bioassay for in situ pathogen quantification down to single bacterium within 20 min. The bioconjugated nanoparticle provides an extremely high fluorescent signal for bioanalysis and can be easily incorporated with biorecognition molecules, such as antibody. The antibody-conjugated nanoparticles can readily and specifically identify a variety of bacterium, such as Escherichia coli O157:H7, through antibody-antigen interaction and recognition. The single-bacterium-detection capability within 20 min has been confirmed by the plate-counting method and realized by using two independent optical techniques. The two detection methods correlated extremely well. Furthermore, we were able to detect multiple bacterial samples with high throughput by using a 384-well microplate format. To show the usefulness of this assay, we have accurately detected 1-400 E. coli O157 bacterial cells in spiked ground beef samples. Our results demonstrate the potential for a broad application of bioconjugated nanoparticles in practical biotechnological and medical applications in various biodetection systems. The ultimate power of integrating bionanotechnology into complex biological systems will emerge as a revolutionary tool for ultrasensitive detection of disease markers and infectious agents.

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“…In recent years, immunomagnetic separation (IMS) methods have become widespread which will result in less false negatives and false positives (Aminul et al, 2006; Deisingh et al, 2004). The method of immunomagnetic separation (IMS) of E. coli O157:H7 from enrichment culture using paramagnetic particles coated with E. coli O157 antibodies has been developed for the selective isolation of this organism from foods contaminated with background flora (Vernozy-Rozand et al, 1998).…”

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confidence: 99%

Comparison of Methods for Detection of Escherichia coli O157:H7 in Ground Beef and Radish Sprouts

Lee¹,

Hyeon²,

Heo³

et al. 2010

Korean Journal for Food Science of Animal Resources

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Strategies for the detection of Escherichia coli O157:H7 in foods

Cited by 175 publications

References 46 publications

Real-Time PCR Methodology for Selective Detection of Viable Escherichia coli O157:H7 Cells by Targeting Z3276 as a Genetic Marker

Real-Time PCR Methodology for Selective Detection of Viable Escherichia coli O157:H7 Cells by Targeting Z3276 as a Genetic Marker

A rapid bioassay for single bacterial cell quantitation using bioconjugated nanoparticles

Comparison of Methods for Detection of Escherichia coli O157:H7 in Ground Beef and Radish Sprouts

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