title: Quantification of viable Escherichia coli O157:H7 in meat products by duplex real-time PCR assays.
abstract: Rapid and specific detection of viable Escherichia coli O157:H7 cells in ready-to-eat (RTE) meat products, by duplex quantitative PCR (qPCR) procedures with mRNA and SYBR Green and TaqMan methodologies were developed. Specific primers and probes were designed based on the serotype of E. coli O157:H7, fliCh7 and rfbE genes. No cross-reactivity with other microorganisms was observed. The detection limit of the assays was 10(1) or 10(2)CFU/g for artificially contaminated meat products, and after a 4h enrichment period at 37 °C, the detection limit decreased to about 1 CFU/g. Time-to completion of the assay was approximately 8h. Thus, these qPCR methods offer a useful, rapid and efficient tool for screening viable E. coli O157:H7 in RTE meat products. This tool could also be proposed for monitoring these foodborne pathogens in HACCP programs.
authors.affiliation: Food Hygiene and Safety, Faculty of Veterinary Science, University of Extremadura, Avda. de la Universidad, s/n., 10003 Cáceres, Spain.
authors.forename: Rubén
authors.initials: R
authors.lastname: Gordillo
authors.forename: Alicia
authors.initials: A
authors.lastname: Rodríguez
authors.forename: María L
authors.initials: ML
authors.lastname: Werning
authors.forename: Elena
authors.initials: E
authors.lastname: Bermúdez
authors.forename: Mar
authors.initials: M
authors.lastname: Rodríguez
mesh_terms: D015169:Colony Count, Microbial; D004269:DNA, Bacterial / Q000302:isolation & purification; D019453:Escherichia coli O157 / Q000302:isolation & purification*; D005506:Food Contamination / Q000032:analysis; D005516:Food Microbiology; D008461:Meat Products / Q000032:analysis / Q000382:microbiology*; D050296:Microbial Viability; D060888:Real-Time Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA
chemical_list: D004269:DNA, Bacterial
keywords: DNA; E. coli O157:H7; RTE meat products; mRNA; qPCR