A rapid detection method for Escherichia coli O157:H7 and Salmonella Typhimurium in food based on the combination of loop-mediated isothermal amplification and hybridization chain reaction

Abstract

Foodborne pathogens present a significant threat to public health, highlighting the critical need for rapid, accurate, and highly sensitive detection methods. This study introduces a novel combined method, integrating loop-mediated isothermal amplification (LAMP) and hybridization chain reaction (HCR), for the simultaneous detection of Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium. LAMP amplification was initially performed to amplify the gene Z3276 and invA, followed by a binding reaction with the specific sequence designed for each amplification product. The above binding products were subsequently captured by streptavidin-modified magnetic beads (SA-MB), initiating the HCR reaction on their surface, with the primers H2 labeled with FAM or VIC fluorophores. Finally, fluorescence signals were measured using a microplate reader for a rapid screening of target pathogens. This method showed a high sensitivity of detection and achieved the detection limit of 1-2 log CFU/mL for E. coli O157:H7, and 2-3 log CFU/mL for S. Typhimurium either in model system or in real food such as apple juice. Moreover, this method effectively reduced the whole detection time to less than three hours through the optimization of the LAMP amplification, SA-MB enrichment and HCR signal amplification. The development of the combination usage of LAMP and HCR in this study can realize the direct detection of E. coli O157:H7 and S. Typhimurium in real food without the requirement of a pre-enrichment step, while effectively shortening the reaction time and thus satisfy the demand food safety supervision for rapid detection of foodborne pathogens.