Optimization of Bacterial Concentration by Filtration for Rapid Detection of Foodborne Escherichia coli O157:H7 Using Real-Time PCR Without Microbial Culture Enrichment.

Abstract

Escherichia coli O157:H7 is an important foodborne pathogen and has been implicated in numerous food poisoning outbreaks worldwide. Although several microbiological and molecular methods have been developed to detect E. coli O157:H7, the difficulty to rapidly detect low levels of the foodborne bacteria persists. Here, the optimization of a filtration technique to concentrate and rapidly detect E. coli O157:H7 was conducted. Using homogenates prepared from freshly cut lettuce and cabbage samples, the E. coli O157:H7 concentration efficiencies of seven membrane filters were compared. Mixed cellulose ester (MCE) and polyvinylidene difluoride (PVDF) filters demonstrated the highest bacterial recoveries. In addition, the optimal E. coli O157:H7 detachment method from MCE filters after filtration was investigated. Tapping for 80 s was demonstrated to be the most effective method for detaching bacteria from the filters. Further, the possibility of the rapid detection of low levels of E. coli O157:H7 in lettuce and cabbage was evaluated using real-time polymerase chain reaction after bacterial concentration using MCE and PVDF filters. The use of MCE filters enabled the detection of 10° CFU/g (5 CFU/g) of E. coli O157:H7 within 2 hr without microbial enrichment culture. Therefore, concentration by filtration can be used for the rapid detection of low levels of foodborne pathogens. PRACTICAL APPLICATION: The modified method, which has been verified in this study, has been optimized to reduce the analysis time and to detect very low concentrations of E. coli O157:H7 within 2 hr. All these detection systems have a direct economic impact on the food analysis of producers, health authorities, or third-party laboratories.