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Novel Real-Time PCR Method To Detect Escherichia coli O157:H7 in Raw Milk Cheese and Raw Ground Meat

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Abstract:

Raw milk, raw milk cheeses, and raw ground meat have been implicated in Escherichia coli O157:H7 outbreaks. Developing methods to detect these bacteria in raw milk and meat products is a major challenge for food safety. The aim of our study was to develop a real-time PCR assay to detect E. coli O157:H7 in raw milk cheeses and raw ground meat. Well-known primers targeting a mutation at position +93 of the uidA gene in E. coli O157:H7 were chosen, and a specific TaqMan–minor groove binder probe was designed. This probe targets another mutation, at position +191 of the uidA gene in E. coli O157:H7. The first step in the study was to evaluate the specificity of this probe with 156 different O157:H7/NM strains and 48 non-O157:H7/NM strains of E. coli. The sensitivity of the method was evaluated by pre- and postinoculation of cheeses and meat enrichments with different E. coli O157:H7 strains. All the E. coli O157:H7 isolates tested were positive, and none of the other bacteria were detected. Our results indicate that this method is sensitive enough to detect 102 E. coli O157:H7 isolates per ml of cheese or meat enrichment broth (24 h at 41.5° C) and is more sensitive than the International Organization for Standardization reference method. We can conclude that this new real-time PCR protocol is a useful tool for rapid, specific, and sensitive detection of E. coli O157:H7 in raw milk and raw ground meat products.

Document Type: Research Article

DOI: http://dx.doi.org/10.4315/0362-028X.JFP-11-498

Affiliations: 1: Unité de Recherche CALITYSS/Equipe EMSA, VetAgro Sup, Université de Lyon, 1 avenue Claude Bourgelat, 69290 Marcy l'Etoile, France., Email: s.miszczycha@vetagro-sup.fr 2: Unité de Recherche CALITYSS/Equipe EMSA, Laboratoire LMAP/Laboratoire National de Référence STEC, VetAgro Sup, Université de Lyon, 1 avenue Claude Bourgelat, 69290 Marcy l'Etoile, France 3: Unité de Recherche CALITYSS/Equipe EMSA, VetAgro Sup, Université de Lyon, 1 avenue Claude Bourgelat, 69290 Marcy l'Etoile, France

Publication date: August 1, 2012

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    First published in 1937, the Journal of Food Protection®, is a refereed monthly publication. Each issue contains scientific research and authoritative review articles reporting on a variety of topics in food science pertaining to food safety and quality. The Journal is internationally recognized as the leading publication in the field of food microbiology with a readership exceeding 11,000 scientists from 70 countries. The Journal of Food Protection® is indexed in Index Medicus, Current Contents, BIOSIS, PubMed, Medline, and many others.

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