Methods for Screening and Isolating Extremely Heat-Resistant Escherichia coli from Meat Sources

Meat animals harbor diverse E. coli populations in their digestive tracts and can serve as sources of pathogenic E. coli. The consumption of meat and produce contaminated with virulent E. coli from animal sources is associated with human illnesses and outbreaks. Heat treatment is an antimicrobial intervention that is commonly used during meat processing to ensure effective reductions in microbial load. Extreme heat resistance (XHR) has been reported among meat-borne E. coli and is mainly attributed to an ~15-19 kb genetic element known as the transmissible locus of stress tolerance (tLST). XHR E. coli can resist treatments used during meat processing and cooking. Therefore, the detection of heat-resistant E. coli is important for devising effective control measures to prevent meat spoilage and ensure meat safety. Here, we present methods used to (1) screen for tLST genes by multiplex PCR and (2) screen and isolate XHR E. coli from meat sources. The mode of heat exposure affects the outcome of XHR testing. Hence, the protocols were optimized to achieve maximum agreement between the tLST genotype and the XHR phenotype.