Culture-independent detection and confirmation of Shiga-toxin-producing E. coli with next-day reporting
Shiga toxin-producing E. coli (STEC) is a major foodborne pathogen. Its detection requires the preliminary testing, culture isolation and confirmation testing on single colonies. The whole process of obtaining a result normally takes a week or longer. Digital PCR aliquots a tube of reaction into thousands of tiny chambers and the majority of occupied chambers contain a single copy of the target. Experiments indicate that we can mobilize intact E. coli cells into the chamber, lyse the cells and PCR amplify the O-antigens and virulence genes from the same chamber. Thus, we can confirm if the virulence genes are carried by the given O-group E. coli without culture isolation of the bacterial strain. We have generated data using pure culture, culture-spiked bovine feces and ground beef and successfully differentiated STEC and non-STEC E. coli strains. Instead of a week-long process, this procedure is able to provide next-day results.