Evaluation of Larger Test Portion Sizes for Escherichia coli Shiga Toxin Producer (STEC) on the Detection by Immunomagnetic Separation and Real-Time PCR in Meat and Vegetables

Abstract

The aims of this study were (i) to evaluate the performance of the Assurance® GDS method combining immunomagnetic separation and real-time PCR for STEC detection in pooled samples (up to 375g) of vegetables and meat, and (ii) to compare its performances to that of the reference method ISO/TS-13136:2012 (25 g sample size) in artificially contaminated samples. The alternative Assurance® GDS method includes enrichment in proprietary broth at 41.5°C for 10 h followed by primary screening of TOP 7 STEC containing eae, stx genes and O157:H7 markers using MPX-Top 7 (IMS+PCR), secondary screening for serogroup identification using MPX-ID/EHEC-ID (IMS+PCR). And finally, cultural confirmation from same IMS beads on two selective agars is set up. For reference method, the enrichment was performed in BPW at 41.5°C for 18 h. A total of 120 samples of meat and vegetables, including 20 uninoculated and 100 samples spiked with stressed cells (<12 CFU/25 or 375g) of STEC, were analyzed using both methods. Our results showed that the Assurance® GDS method LOD50 ranged from 0.12 CFU/375 to 0.522 CFU/375 g and the LOD95 from 0.518 CFU/375 to 2.257 CFU/375 g. These data are similar to the LOD50 data of the reference method.