Key Laboratory of Biomass Chemical Engineering of Ministry of Education, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, China.
College of Pharmaceutical Science, Zhejiang University of Technology, Hangzhou, China.
J Food Sci. 2022 Sep;87(9):4137-4147. doi: 10.1111/1750-3841.16287. Epub 2022 Aug 20.
Salmonella is a pathogen that comes from different animal-originated foods and poses a significant threat to human health. The present detection methods for Salmonella are time-consuming and labor-intensive and requires skilled workers and specialized instruments. In this study, the conservative invA was selected as the target gene, and a quantitative detection method for Salmonella with wide availability and user-friendliness was established based on CRISPR Cas12a and a personal glucose meter (PGM). The indirect signal transformation from the original target DNA to the final glucose signal was achieved through RAA, CRISPR Cas12a reaction, enzymic reaction, and glucose signal reading by a PGM (accu-chek type from Roche). This PGMs-CRISPR assay showed a detection sensitivity of Salmonella as low as 5 colony-forming units (CFU)/reaction in either pure culture or artificially contaminated food samples and exhibited specificity between Salmonella isolates and non-Salmonella isolates. Furthermore, quantitative detection of Salmonella in spiked milk samples was also achieved within the range from 1 to 1 × 10 CFU/reaction. Subsequently, the correlation and consistency between the PGMs-CRISPR assay and quantitative polymerase chain reaction (qPCR) in detection of Salmonella in spiked milk samples were achieved. Therefore, a highly sensitive, portable, quantitative, and user-friendly detection method based on CRISPR Cas12a and PGMs was developed in this study for Salmonella detection and identification. PRACTICAL APPLICATION: A sensitive, rapid, user-friendly, and quantitative detection method based on CRISPR Cas12a for Salmonella in food has been developed in this study, which is of great significance to food safety supervision and management.
沙门氏菌是一种来自不同动物源性食品的病原体，对人类健康构成重大威胁。目前用于检测沙门氏菌的方法既耗时又费力，需要熟练的工人和专门的仪器。在本研究中，选择保守的 invA 作为靶基因，基于 CRISPR Cas12a 和个人血糖仪（PGM）建立了一种具有广泛可用性和用户友好性的沙门氏菌定量检测方法。通过 RAA、CRISPR Cas12a 反应、酶反应和 PGM（罗氏 accu-chek 型）读取葡萄糖信号，实现了原始靶 DNA 到最终葡萄糖信号的间接信号转换。这种 PGMs-CRISPR 测定法在纯培养或人工污染的食品样品中对沙门氏菌的检测灵敏度低至 5 个菌落形成单位（CFU）/反应，并且在沙门氏菌分离株和非沙门氏菌分离株之间表现出特异性。此外，还实现了对添加牛奶样品中沙门氏菌的定量检测，范围为 1 至 1×10 CFU/反应。随后，在添加牛奶样品中沙门氏菌的检测中，PGMs-CRISPR 测定法和定量聚合酶链反应（qPCR）之间的相关性和一致性得以实现。因此，本研究基于 CRISPR Cas12a 和 PGMs 开发了一种用于沙门氏菌检测和鉴定的高灵敏度、便携式、定量和用户友好的检测方法。
本研究开发了一种基于 CRISPR Cas12a 的用于食品中沙门氏菌的敏感、快速、用户友好和定量检测方法，对食品安全监管具有重要意义。
