Sensitive and Rapid Detection of Escherichia coli O157: H7 From Beef Samples Based on Recombinase Aided Amplification Assisted CRISPR/Cas12a System.

This article focuses on the development and evaluation of a recombinase aided amplification (RAA) assisted CRISPR/Cas12a fluorescence platform (RAA-CRISPR/Cas12a) for the rapid, specific, and sensitive detection of Escherichia coli O157:H7 in beef samples. Targeting the rfbE gene, the method demonstrated high specificity by distinguishing E. coli O157:H7 from 23 non-target bacterial strains and achieved a limit of detection of 5.4 × 10² CFU/mL in pure cultures, outperforming RAA-gel electrophoresis and real-time PCR (RT-PCR) methods. In spiked ground beef samples, the assay detected as low as 9 CFU/25 g after 4 hours of enrichment, with results consistent with RT-PCR and traditional culture-based methods when applied to 93 real beef samples. The assay requires only simple isothermal equipment, enabling completion within 30 minutes post-enrichment, and shows promise as a portable tool for foodborne pathogen diagnostics in both laboratory and field settings.