Purpose: The purpose of this study was to conduct an NF Validation by AFNOR Certification extension study to validate use of the SureTect Esherichia coli O157:H7 PCR Assay on the Applied Biosystems™ 7500 Fast PCR Instrument with Applied Biosystems™ RapidFinder™ Express version 2.0 Software (the alternative method) with seasoned and unseasoned raw beef meats.
Methods: A method comparison study and relative limit of detection (RLOD) study was conducted using the SureTect™ standard protocol as the alternative method. Twenty five g samples were enriched in 225 ml of pre-warmed (41.5±1 °C) BPW and incubated at 41.5±1 °C. Direct lysis was conducted following 8 and 24 hours enrichment. Following direct lysis, PCR was run and results were automatically interpreted by the software. The reference method was conducted according to ISO 16654:2001.
Results: Sixty raw beef samples were tested using the alternative and ISO 16654:2001 reference methods. For the RLOD study, an isolate of E. coli O157:H7 was spiked into a raw beef trim matrix and tested according to the alternative method protocol and the ISO 16654:2001 reference method. The alternative method demonstrated equivalent performance for the raw beef meat samples analysed to the ISO 16654:2001 reference method. The alternative method showed the same RLOD (0.4-0.7 CFU/25 g) to the ISO 16654:2001 reference method (0.4-0.7 CFU/25 g) at both 8 and 24 hours incubation.
Significance: The alternative method proved to be a suitable substitute to the ISO 16654:2001 reference method for E. coli O157:H7 detection in seasoned and unseasoned raw beef meats.