Rapid detection of O157 and major Non-O157 Shiga toxin-producing Escherichia coli serogroups in ground beef, ready-to-eat vegetables, and shredded cheese using VIDAS–WGA real-time PCR after 6-h enrichment

Abstract

Shiga toxin-producing Escherichia coli (STEC) may occur at trace levels in inhibitor-rich foods, which makes culture-independent PCR screening challenging without prolonged enrichment. To overcome this challenge, we evaluated a rapid workflow that combined 6-h enrichment in buffered peptone water with automated VIDAS immunocapture/DNA extraction, whole-genome amplification (WGA), and real-time PCR for same-day STEC detection in ground beef, ready-to-eat vegetables, and shredded cheese. In matrix-based limit-of-detection testing, WGA improved analytical performance compared with VIDAS real-time PCR alone by enabling earlier amplification, with Ct reductions of 2.76–4.66 cycles across matrix–inoculum combinations in which both methods yielded detectable Ct values. For practical assessment at ultra-low contamination, an AOAC-style fractional probability of detection design was applied using 500-g bulk samples divided into 20 test portions (approximately 0–2 CFU per portion). Across matrices, VIDAS–WGA real-time PCR detected 56 of 58 culture-positive fractions (96.6%) with no false positives and showed near-complete agreement with the reference culture method. The analytical workflow took approximately 10 h, excluding sample preparation, instrument setup, and result interpretation. These findings support VIDAS–WGA real-time PCR as a same-day screening approach for VIDAS-targeted STEC serogroups in tested food matrices, although broader non-O157 validation remains warranted. Copyright