Comparison of Foodproof Real-time PCR Kits to the ISO Method for STEC Screening and O-group Identification

Introduction: Foodborne illness caused by Enterohemorrhagic Escherichia coli (EHEC) claimed ~ 50 lives during one of the largest outbreaks in 2011. EHEC have been encountered in leafy vegetables, sprouted seeds, raw milk and cheeses, as well as fresh, minced, and mixed meat preparations. Most infections have been with E. coli O157; however, other strains of non-O157 E. coli capable of causing sickness and death also should be examined in food.

Following the current ISO method (ISO/TS 13136), not all variants of the stx2 pathogenicity gene are detected, particularly stx2f. Differences in important serotypes exist between Europe and the United States. ISO/TS 13136 requires five serotypes (O26, O103, O111, O145 and O157). Moreover, there will be an extension of regulation (EC) 2073/2005 specifically for sprouts, which requires these five as well as O104.

Purpose: To evaluate the specificity (inclusivity and exclusivity), sensitivity, and robustness of the ISO/TS 13136 method (a PCR-based method) for STEC screening and serotype identification in comparison to the foodproof STEC Screening and O-group Identification Kits.

Methods: ISO primary enrichment for STEC testing was performed. Enrichments were then tested using ISO/TS13136 described PCR-based method or PCR-based STEC screening kit, which involved DNA extraction from the primary enrichment followed by real-time PCR. The serotypes of STEC-positive samples were then determined using the O-group identification kit.  

Results: The sensitivity and robustness of the screening and O-group identification kits fulfill requirements set forth by ISO/TS13136. However, the specificity of the screening kit was superior due to its detection of more Shiga-toxin variants, such as stx2, and the O-group Identification Kit determined 3 more O-groups than the ISO/TS 13136 method currently requires.

Significance: The combination of the foodproof STEC Screening and O-group Identification Kits provided the ability to detect and identify all eight serotypes simultaneously and reduce the time-to-result.