An Independent Laboratory Evaluation of the Mericon E. coli Detection Workflows for AOAC-RI PTM Status

Introduction: Regulation of pathogenic E. coli in foods is receiving heightened attention under new food safety requirements. The mericon E. coli O157 Screen Plus and E. coli STEC O-type detection methods combine straightforward sample prep with real-time PCR detection. A choice of two different DNA prep procedures can be utilized; the manual mericon DNA Bacteria Kit or the automated QIAsymphony mericon Bacteria Kit. The resultant purified DNA is combined with mericon PCR Master Mix and real-time PCR is conducted on the Rotor-Gene Q platform.

Purpose: To conduct a method comparison, inclusivity/exclusivity and ruggedness studies on the mericon E. coli kits as part of the AOAC Research Institute PTM process.

Methods: For the method comparison, 3 foods were analyzed (raw ground beef (70% lean), beef trim, and fresh spinach). Each matrix was inoculated with a different serotype of pathogenic E. coli at 2 levels (0.2-2 CFU/test portion and 2-5 CFU/test portion) and cultured for 10 +/- 1 h in mTSB at 41°C. For each sample, DNA was extracted by both methods, analyzed by both mericon real-time PCR assays and compared to the MLG or BAM reference methods. Test kits were also evaluated for ruggedness, inclusivity and exclusivity.

Results: For inclusivity, all 50 strains were positively detected.  For exclusivity, all 30 strains were correctly excluded. The ruggedness study indicated that minor changes to the DNA extraction and PCR analysis did not affect the outcome.  The method comparison demonstrated no significant differences between the number of positive samples detected by the mericon and reference methods for all 3 matrices. The mericon E. coli methods were granted AOAC RI PTM status.

Significance: This new method is an efficient, rapid and reliable alternative to the traditional methods of detecting E. coli virulence factors and Big 7 STEC O-type in several foods.