Single-tube detection of a foodborne bacterial pathogen using user-friendly portable device

Timely and reliable detection of foodborne bacterial pathogen is crucial for reducing disease burden in low- and middle-income countries. However, laboratory-based methods are often inaccessibility in resource-limited settings. Here, we developed a single-tube assay and a low-cost palm-sized device for on-site detection of the representative foodborne bacterial pathogen, Salmonella Enteritidis. Our assay incorporates the advantages of protein-nucleic acid signal transduction, EXPonential Amplification Reaction (EXPAR), and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 12a (Cas12a). After systematically investigating the compatibility of these components, we developed a "three-in-one" integration reaction, termed ST-EXPAR-CRISPR assay. This assay requires only one tube, one controlled temperature (39 °C) and simple operation, eliminating the need for bacterial isolation, nucleic acid extraction, or washing steps. ST-EXPAR-CRISPR assay is capable of detecting as few as 37 CFU/mL of target bacterium. Using our kit and portable device, untrained volunteers successfully detected contamination in food samples outdoors. The simplicity of the detection process and minimal hardware requirements make our assay highly promising for application in point-of-care and on-site scenarios. Moreover, the ST-EXPAR-CRISPR assay can be easily modified to detect other targets by changing the nucleic acid sequence with low research and development cost, potentially reducing the global disease burden.