Rapid, Visual, and Sequence-Specific Detection of Salmonella in Egg Liquid with vis-NEAA, a CRISPR/Cas12 Empowered New Strategy

Salmonella is one of the main pathogenic factors that cause foodborne diseases. Rapid and accurate detection of Salmonella in food is of great importance to ensure food safety. Nicking enzyme-assisted amplification (NEAA) is one of the promising isothermal amplification methods finishing the in vitro amplification in similar to 10 min; however, it suffers from nonspecific amplification a lot (similar to 70% products are noises). In this paper, we introduced CRISPR/Cas12a to specifically recognize the NEAA amplicons and transduce the signals into turned-on fluorescent visual readouts (vis-NEAA). Impressively, with this method, the high efficiency of NEAA has been taken great advantage and the nonspecific products were successfully bypassed at the same time. In comparison to NEAA-gel electrophoresis, vis-NEAA showed complete fidelity toward the presence of specific products, while for real-time PCR, it possesses equivalent sensitivity and specificity but saves similar to 80% of the time. A level of 80 CFU/mL Salmonella in spiked eggs can be detected on-site in similar to 20 min.

Automated summary

In this study, we introduced CRISPR/Cas12a to specifically recognize the amplicons of NEAA and convert the signals into fluorescent readouts. The vis-NEAA method takes advantage of the high efficiency of NEAA and successfully bypasses nonspecific amplification. Compared to traditional methods, vis-NEAA is efficient, accurate, and fast, allowing for the detection of low concentrations of Salmonella in egg samples within 20 minutes.