Health Canada, Bureau of Microbial Hazards, 251 Sir Frederick Banting Driveway, Ottawa, Ontario, Canada K1A 0K9.
(ORCID: https://orcid.org/0000-0003-2380-2148 [A.G.]).
J Food Prot. 2021 Feb 1;84(2):296-302. doi: 10.4315/JFP-20-233.
Verotoxin-producing Escherichia coli (VTEC; also known as Shiga toxin-producing E. coli) is a significant cause of foodborne illnesses around the world. Due to the serological and genomic diversity of VTEC, methods of detection for VTEC in food samples require detection of verotoxin or its gene vt (also known as stx). The current taxonomy of vt identifies three vt1 (a, c, d) and seven vt2 (a to g) subtypes. PCR detection of vt is convenient and rapid, but protocols may not detect all currently identified variants or subtypes of vt. The Health Canada Compendium of Analytical Methods protocol for the analysis of food for VTEC is MFLP-52. MFLP-52 includes a VT Screening PCR that is used to determine the presumptive presence of VTEC by the detection of vt in food enrichments and to differentiate VTEC from other isolates. The VT Screening PCR was developed prior to the establishment of the current vt taxonomy. An evaluation of VT Screening PCR for detection of the 10 established vt subtypes was performed, and it was discovered that the method could not detect subtypes vt1d and vt2f. Additional primers and a modified protocol were developed, and the modified VT Screening PCR was tested against an inclusivity panel of 50 VTEC strains, including representatives of 10 vt subtypes, and an exclusivity panel of 30 vt-negative E. coli from various sources, to ensure specificity. The reliability of MFLP-52 with the modified VT Screening PCR was assessed by analysis of four priority food matrices (ground beef, lettuce, cheese, and apple cider) inoculated with a VTEC strain at 2 to 5 CFU/25 g. The modified VT Screening PCR was determined to be able to detect all 10 vt subtypes and reliably detect the presence of VTEC in all tested food enrichments.
产志贺毒素大肠杆菌（VTEC；也称为产志贺毒素大肠杆菌）是全球食源性疾病的重要病因。由于 VTEC 的血清学和基因组多样性，食品样本中 VTEC 的检测方法需要检测志贺毒素或其基因 vt（也称为 stx）。目前的 vt 分类学确定了三个 vt1（a、c、d）和七个 vt2（a 到 g）亚型。PCR 检测 vt 方便快捷，但方案可能无法检测到所有目前已识别的变体或 vt 亚型。加拿大卫生部食品中 VTEC 分析方法纲要 MFLP-52 包括 VT 筛选 PCR，用于通过检测食品增菌物中的 vt 来确定 VTEC 的推定存在，并将 VTEC 与其他分离株区分开来。VT 筛选 PCR 是在当前 vt 分类学建立之前开发的。对 VT 筛选 PCR 检测 10 种已建立的 vt 亚型进行了评估，发现该方法无法检测 vt1d 和 vt2f 亚型。开发了额外的引物和修改后的协议，并对修改后的 VT 筛选 PCR 进行了测试，以针对包含 10 种 vt 亚型代表的 50 株 VTEC 菌株的包容性面板和来自各种来源的 30 株 vt 阴性大肠杆菌的排他性面板进行测试，以确保特异性。通过对 4 种优先食品基质（碎牛肉、生菜、奶酪和苹果酒）进行 VTEC 菌株接种（2 至 5 CFU/25 g），评估 MFLP-52 与修改后的 VT 筛选 PCR 的可靠性。修改后的 VT 筛选 PCR 被确定能够检测所有 10 种 vt 亚型，并可靠地检测所有测试食品增菌物中 VTEC 的存在。
