Development of filtration-based RPA–CRISPR/Cas12a system for rapid, sensitive and visualized detection of Salmonella in ready-to-eat salads

Abstract

Salmonella contamination of ready-to-eat (RTE) salads, a pathogen that causes foodborne diseases worldwide, continues to increase. Since RTE salads are consumed without any processing processes that remove pathogens, such as heat treatment, rapid and accurate testing is essential to effectively prevent such foodborne contamination. Therefore, in this study, I developed the recombinase polymerase amplification (RPA)–clustered regularly interspaced short palindromic repeats (CRISPR)/Cas12a system to amplify Salmonella gene (invA) in 20 min through RPA and visually detect it within 5 min using CRISPR/Cas12a. I successfully visually detected 101 CFU/mL of Salmonella pure culture using this system. Furthermore, for food sample preparation, I introduced the filtration based approach as a pretreatment method, which successfully reduced detection time and improved detection sensitivity. In conclusion, the developed filtration-based RPA CRISPR/Cas12a system enabled the visual detection of Salmonella in salads as low as 101 CFU/g. This system, which integrates filtration-based RPA-CRISPR/Cas12a, is a rapid, sensitive, and visually intuitive method for detecting Salmonella in RTE salads. It is also a useful technology with high applicability in POCT.