Detection of viable Listeria monocytogenes with a 5' nuclease PCR assay

Article Abstract:

A 5' nuclease assay for the rapid and specific detection of viable Listeria monocytogenes has been developed, targeting the hemolysin A transcript found only in the organism. The viability of L monocytogenes was reduced by heating at a variety of temperatures and times up to a maximum of a 9-D inactivation. The primer's location had a marked effect on the assay's utility, with primers located in the most distal regions of the hlyA transcript apparently correlating with the number of CFU. The assay with primers that included the 3' end of the transcript accurately indicated viability as measured by CFU determination or staining with 5-sulfofluorescein diacetate.

Design and construction, Microbiological assay

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Detection of Listeria monocytogenes in cheese with the magnetic immuno-polymerase chain reaction assay

Article Abstract:

The magnetic immuno-polymerase chain reaction (PCR) assay (MIPA) was used to detect Listeria monocytogenes in food. This method combines magnetic separation of Listeria cells by magnetic particles coated with specific monoclonal antibodies (MAb) and amplification of bacterial DNA by PCR. A MIPA employing MAb 55 and the listeriolysin O gene primer set detected L. monocytogenes after 24 h culture of samples contaminated with 40 colony forming units per 25 g of cheese. It was shown that 1 CFU of L. monocytogenes per g of cheese after two enrichments can be detected within 55 h.

Cheese

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A PCR-based assay for the detection of Escherichia coli Shiga-like toxin genes in ground beef

Article Abstract:

A PCR-based quantitative assay involving the 5' to 3' nuclease activity of Thermus aquaticus DNA polymerase is useful for the identification of Shiga-like toxin I-producing Escherichia coli (SLTIEC) in ground beef. The polymerase breaks down an internal oligonucleotide probe labeled with a fluorescent reporter dye and a quencher dye. The assay has two PCR primers that produce a 497-bp amplicon specific for the sltI gene. The assay can detect 10 +/- 5 CFU of SLTIEC per PCR.

Food, Escherichia coli, Food microbiology, Identification and classification, Beef

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