Application Guide - QIAxcel Advanced

108 QIAxcel Advanced Application Guide 10/2016 Introduction Shiga toxin-producing E. coli (STEC) causes serious gastrointestinal disease and death in humans. The seven major E. coli serogroups that give rise to infections in humans are O26, O45, O103, O111, O121, O145 and O157 (1, 2). Cattle are considered one of the major asymptomatic STEC carriers, and the bacteria propagate in their gut and are shed in the feces, which is a major source of food contamination and human infection (3). Early detection of STEC is crucial for prevention and intervention strategies. Nowadays most tests for STEC in veterinary laboratories are performed using real-time PCR, but this method is limited by the number of targets it can detect. Most real-time cyclers are limited to five channels, and in addition, developing multiplex qPCR using more than three channels is technically challenging. Detecting more targets by real-time PCR would require multiple reactions and result in significant cost increases. We have developed an 11-gene multiplex PCR assay that detects four major E. coli virulence genes (stx1, stx2, eae, ehxA) along with the seven major STEC serogroups mentioned above. The level of throughput and the interpretation accuracy of this assay have been dramatically improved by using the QIAxcel Advanced, a fast capillary electrophoresis system with high resolution and reproducibility. High-throughput detection of Shiga toxin-producing Escherichia coli using multiplex PCR and the QIAxcel® Advanced system Jianfa Bai, Ph.D., Director of Molecular Research and Development Kansas State Veterinary Diagnostic Laboratory, Department of Diagnostic Medicine/Pathobiology, College of Veterinary Medicine, Kansas State University, 1800 Denison Avenue, Manhattan, KS 66506-5606, USA Using the QIAxcel Advanced system after a multiplex PCR allowed us to perform efficient and reliable high-throughput screening of cattle feces for the presence of four major E. coli virulence genes and the seven major Shiga toxin-producing serogroups that give rise to infection in humans.

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