Purpose: The purpose of this study was to develop a collection of positive-control strains for the detection of E. coli O157:H7 and the six USDA-regulated serogroups of non-O157 STEC.
Methods: A unique DNA target sequence and a gene for spectinomycin resistance were integrated into the chromosomes of a strain of O157:H7 and strains of the six non-O157 STEC by allelic exchange to generate positive-control strains for STEC detection. End-point and real-time PCR methods were developed for the specific detection of the control strains. In addition, the strains were tested for their potential use in modeling the growth of STEC in the presence of foodborne background flora by incorporating spectinomycin into selective plating media.
Results: The developed end-point and real-time PCR methods demonstrated excellent inclusivity and exclusivity when tested against 42 strains of STEC O157, at least six strains of each of the USDA-regulated non-O157 STEC, and more than 50 strains of other bacteria representing 29 species and 21 genera. Plating the STEC control strains on modified Rainbow agar containing spectinomycin eliminated the growth of ground beef background flora that otherwise grew on modified Rainbow agar.
Significance: The genetically modified STEC strains are useful as positive control strains for the detection of STEC and will be useful for the modeling the growth of STEC in foods.