Evaluation and comparison of recombinase polymerase amplification coupled with lateral-flow bioassay for Escherichia coli O157:H7 detection using diifeerent genes

Abstract

Shiga toxin-producing Escherichia coli serotype O157:H7 is a food and waterborne zoonotic pathogen causing gastroenteritis in humans. Rapid and simple detection in water and food is imperative to control its spread. However, traditional microbial detection approaches are time-consuming, expensive and complex to operate at the point-of-care without professional training. We present a rapid, simple, sensitive, specific and portable method for detection of E. coli O157:H7 in drinking water, apple juice and milk. We evaluated the effect of gene selection in detecting E. coli O157:H7 using recombinase polymerase amplification coupled with a lateral flow assay using rfbE, fliC and stx gene targets. As low as 100 ag and 1 fg DNA, 4–5 CFU/mL and 101 CFU/mL of E. coli O157:H7 was detected using the stx and rfbE gene targets respectively with 100% specificity, whilst the detection limit was 10 fg DNA and 102 CFU/mL for the fliC gene target, with 72.8% specificity. The RPA-LFA can be completed within 8 min at temperatures between 37 and 42 °C with reduced handling and simple equipment requirements. The test threshold amplification of the target was achieved in 5–30 min of incubation. In conclusion, RPA-LFA represents a potential rapid and effective alternative to conventional methods for the monitoring of E. coli O157:H7 in food and water.