{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,31]],"date-time":"2026-07-31T15:59:10Z","timestamp":1785513550072,"version":"3.56.0"},"reference-count":32,"publisher":"MDPI AG","issue":"15","license":[{"start":{"date-parts":[[2026,7,31]],"date-time":"2026-07-31T00:00:00Z","timestamp":1785456000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Science Technology Program of State Administration for Market Regulation","award":["2025MK053"],"award-info":[{"award-number":["2025MK053"]}]},{"award":["2025MK053"],"award-info":[{"award-number":["2025MK053"]}],"id":[{"id":"https:\/\/ror.org\/00aaqtw69","id-type":"ROR","asserted-by":"publisher"}]},{"name":"Jiangsu Excellent Postdoctoral Program","award":["2025ZB879"],"award-info":[{"award-number":["2025ZB879"]}]},{"name":"Jiangsu Provincial Administration for Market Regulation Science and Technology Plan Project","award":["KJ2026010"],"award-info":[{"award-number":["KJ2026010"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Foods"],"abstract":"<jats:p>Probiotics play an important role in maintaining human gastrointestinal health and immune function, driving a rapidly expanding global market for probiotic-fortified foods. Lacticaseibacillus paracasei is one of the most widely applied probiotic strains in functional foods and dairy formulations, with well-documented health-promoting properties. For probiotic foods, authenticity requires rigorous verification for efficacy and compliance, which depends on rapid strain-specific identification. However, conventional culture-based methods are time-consuming and cannot distinguish closely related Lacticaseibacillus species, while PCR-based assays require expensive thermal cyclers and trained personnel, barring deployment in resource-limited settings. Here, we report the development and validation of a MIRA-CRISPR\/Cas12a fluorescence assay for rapid, species-specific, and highly sensitive on-site detection of Lacticaseibacillus paracasei. This integrated workflow combines 8 min rapid crude DNA extraction, 30 min isothermal MIRA pre-amplification, and 20 min CRISPR\/Cas12a signal amplification with naked-eye fluorescence readout under blue light excitation, and requires only miniaturized portable equipment. The assay achieved a reliable limit of detection of 100 CFU\/mL in milk-matrix spiked samples, showing no cross-reactivity across 17 tested lactic acid bacteria strains. In a validation set of 48 food samples, our method demonstrated 100% concordance with the gold-standard species-specific qPCR assay. This field-deployable method is appropriate for on-site quality control in probiotic manufacturing, label compliance verification, and frontline market regulatory inspections.<\/jats:p>","DOI":"10.3390\/foods15152709","type":"journal-article","created":{"date-parts":[[2026,7,31]],"date-time":"2026-07-31T15:02:01Z","timestamp":1785510121000},"page":"2709","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Rapid On-Site Detection of Lacticaseibacillus paracasei Using a Portable MIRA-CRISPR\/Cas12a Naked-Eye Fluorescence Assay"],"prefix":"10.3390","volume":"15","author":[{"given":"Kai","family":"Liao","sequence":"first","affiliation":[{"name":"Key Laboratory of Food Contact Materials Safety, State Administration for Market Regulation, Jiangsu Product Quality Testing and Inspection Institute, Nanjing 210007, China"},{"name":"Key Laboratory of Microbiological Metrology, Measurement & Bio-Product Quality Security, State Administration for Market Regulation, College of Life Sciences, China Jiliang University, Hangzhou 310018, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yang","family":"Zhang","sequence":"additional","affiliation":[{"name":"Key Laboratory of Food Contact Materials Safety, State Administration for Market Regulation, Jiangsu Product Quality Testing and Inspection Institute, Nanjing 210007, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaosu","family":"Xiong","sequence":"additional","affiliation":[{"name":"Key Laboratory of Food Contact Materials Safety, State Administration for Market Regulation, Jiangsu Product Quality Testing and Inspection Institute, Nanjing 210007, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zhonglu","family":"She","sequence":"additional","affiliation":[{"name":"Key Laboratory of Food Contact Materials Safety, State Administration for Market Regulation, Jiangsu Product Quality Testing and Inspection Institute, Nanjing 210007, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yufeng","family":"Wang","sequence":"additional","affiliation":[{"name":"Key Laboratory of Microbiological Metrology, Measurement & Bio-Product Quality Security, State Administration for Market Regulation, College of Life Sciences, China Jiliang University, Hangzhou 310018, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5827-3699","authenticated-orcid":false,"given":"Zihong","family":"Ye","sequence":"additional","affiliation":[{"name":"Key Laboratory of Microbiological Metrology, Measurement & Bio-Product Quality Security, State Administration for Market Regulation, College of Life Sciences, China Jiliang University, Hangzhou 310018, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zefeng","family":"Shan","sequence":"additional","affiliation":[{"name":"Taizhou Institute of Agricultural Science, Jiangsu Academy of Agricultural Sciences, Taizhou 225300, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yuxin","family":"Li","sequence":"additional","affiliation":[{"name":"MOE Joint International Research Laboratory of Animal Health and Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaoran","family":"Jia","sequence":"additional","affiliation":[{"name":"MOE Joint International Research Laboratory of Animal Health and Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaojun","family":"Zhu","sequence":"additional","affiliation":[{"name":"Key Laboratory of Food Contact Materials Safety, State Administration for Market Regulation, Jiangsu Product Quality Testing and Inspection Institute, Nanjing 210007, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0002-1797-9027","authenticated-orcid":false,"given":"Feng","family":"Xue","sequence":"additional","affiliation":[{"name":"MOE Joint International Research Laboratory of Animal Health and Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jie","family":"Zou","sequence":"additional","affiliation":[{"name":"Key Laboratory of Food Contact Materials Safety, State Administration for Market Regulation, Jiangsu Product Quality Testing and Inspection Institute, Nanjing 210007, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaoqiang","family":"Zhang","sequence":"additional","affiliation":[{"name":"Key Laboratory of Food Contact Materials Safety, State Administration for Market Regulation, Jiangsu Product Quality Testing and Inspection Institute, Nanjing 210007, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3763-1183","authenticated-orcid":false,"given":"Wei","family":"Chen","sequence":"additional","affiliation":[{"name":"School of Food and Biological Engineering, Hefei University of Technology, Hefei 230009, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,7,31]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"649","DOI":"10.1038\/s41575-021-00440-6","article-title":"The International Scientific Association of Probiotics and Prebiotics (ISAPP) consensus statement on the definition and scope of postbiotics","volume":"18","author":"Salminen","year":"2021","journal-title":"Nat. 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