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Objective To develop a rapid and sensitive detection technology for pathogenic microorganisms in food packaging based on the clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated protein 13a (Cas13a) system. Methods The method leveraged the specific recognition and signal amplification characteristics of the CRISPR-Cas13a technology. Multiple guide RNAs were designed to target different sites on pathogenic RNA, enabling a single RNA molecule to activate multiple Cas13a enzymes simultaneously. This cascade triggered trans-cleavage of RNA reporter molecules in the reaction mixture, generating a detectable fluorescent signal. The tandem design synergistically enhanced signal amplification efficiency, thereby improving detection sensitivity. Results The limits of detection of this method for pathogenic and RNA were 800 copies/mL and 18.7 pmol/L, respectively, and the detection could be completed in a short time, accurately distinguishing different pathogens. Testing food packaging samples contaminated with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pseudvirus showed recovery rates ranging from 89.71% to 102.56%. Conclusion This study presents a rapid, cost-effective and easy-to-operate method for detecting pathogenic microbial contamination on food packaging surfaces, indicating high accuracy, offering potential application value for monitoring microbial risks in foodborne transmission pathways.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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