CRISPR/Cas12a-Triggered\nVisible-Light-Driven Photoelectrochemical\nAssay with Single-Nucleotide Resolution for Drug-Resistant Foodborne Salmonella Detection

The prevalence of foodborne pathogenic bacteria, especially\ndrug-resistant\nstrains, such as Salmonella enterica, poses serious threats to public health, highlighting the requirement\nfor the development of rapid and precise detection methods. Herein,\na CRISPR/Cas12a-triggered visible-light-driven photoelectrochemical\n(PEC) assay (CasPEC) was developed using a SiO<sub>2</sub>-quenched\nBiVO<sub>4</sub>/MoS<sub>2</sub> p/n-type heterojunction as the photoactive\nmaterial. The CRISPR/Cas12a recognition endowed the CasPEC assay with\nhigh specificity capable of resolving single-nucleotide polymorphisms\n(SNPs) and identifying SNP-involved drug-resistant bacteria. SiO<sub>2</sub> was linked to the surface of the BiVO<sub>4</sub>/MoS<sub>2</sub> heterojunction by single-stranded DNA (ssDNA), which would\nbe cleaved by target-activated CRISPR/Cas12a. This cleavage of ssDNA\nresulted in the detachment of SiO<sub>2</sub>, thereby achieving a\n“signal-on” PEC output. Leveraging the multiple-turnover\nCRISPR cleavage and the outstanding photoactive performance of PEC\nsignaling, the CasPEC assay for S. enterica showed a detection limit of 103 colony-forming units (CFU)/mL and\nthe ability to detect as few as 0.01% drug-resistant strains. The\nCasPEC assay can accurately sense the S. enterica contamination in complex food matrices, including beef and milk.\nThese findings demonstrated the great potential of the CasPEC assay\nfor detecting pathogenic bacterial contamination in food, particularly\nconcerning food safety related to SNP-involved drug-resistant bacteria.