Purpose: To evaluate the performance of a new E. coli O157 isothermal molecular detection method for 1) inclusivity; 2) exclusivity; and 3) comparison to the ISO cultural method.
Methods: Inclusivity evaluation was performed using 60 different E. coli O157 cultures and exclusivity using 60 different non-E. coli O157 cultures. True positive and true negative states were defined by each organism’s genetic or biochemical identity. A method comparison was performed by testing 60 produce samples including various sprouts, fresh leafy greens, cilantro and parsley. Samples were artificially contaminated at a low level or left non-inoculated, then were acclimated before testing. Non-paired matrix samples were analyzed using the ISO 16654 reference method and the new isothermal molecular method after incubation in Buffered Peptone Water-ISO at 41.5°C for 18 hours. All results were confirmed using the ISO method protocol.
Results: Inclusivity and exclusivity of 100% were determined using pure cultures. For the method comparison, there was positive agreement among 28 samples, negative agreement among 31 samples and one negative deviation. According to ISO 16140-2:2015, this result meets the acceptability limit criterion for alternative method performance, demonstrating the capability of the new isothermal molecular method for detection of E. coli O157 in these matrices.
Significance: The new isothermal molecular detection method was evaluated using cultures of known identity, and sample matrices were tested in comparison to a reference cultural method. The new method was determined to be reliable and accurate and to offer substantial advantages to the end user, including a quicker time to result compared to the cultural method.