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 3 h 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.