{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,16]],"date-time":"2026-09-16T17:04:18Z","timestamp":1789578258539,"version":"build-2803163510"},"reference-count":29,"publisher":"MDPI AG","issue":"9","license":[{"start":{"date-parts":[[2026,9,16]],"date-time":"2026-09-16T00:00:00Z","timestamp":1789516800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["32473207"],"award-info":[{"award-number":["32473207"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]},{"award":["32473207"],"award-info":[{"award-number":["32473207"]}],"id":[{"id":"https:\/\/ror.org\/01h0zpd94","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Vet. Sci.","Veterinary Sciences"],"abstract":"<jats:p>Cyprinid herpesvirus 2 (CyHV-2) is a highly lethal pathogen responsible for substantial economic losses in farmed cyprinids worldwide. However, rapid and field-deployable diagnostic tools remain limited. Here, we developed a one-tube recombinase polymerase amplification (RPA)-CRISPR\/Cas12a-based fluorescence assay for the visual detection of CyHV-2. The system integrates RPA pre-amplification targeting the highly conserved helicase gene hphS, Cas12a-mediated specific recognition and collateral cleavage of a fluorophore-quencher-labeled ssDNA reporter, and visual readout under a portable blue light transilluminator. After optimizing the concentrations of primers, crRNA, Cas12a and the reaction time, the detection limit of this method reached 1 copy\/\u03bcL, and it showed no cross-reaction with other common aquatic pathogens. Clinical evaluation using 100 tissue samples demonstrated 100% concordance with quantitative real-time PCR (qPCR). This method does not require expensive equipment and can be completed within approximately one hour. This RPA-CRISPR\/Cas12a platform offers a rapid and specific solution for on-site CyHV-2 surveillance, especially suitable for aquaculture environments with limited resources, and is expected to have broader applications in the detection of aquatic pathogens.<\/jats:p>","DOI":"10.3390\/vetsci13090970","type":"journal-article","created":{"date-parts":[[2026,9,16]],"date-time":"2026-09-16T16:07:29Z","timestamp":1789574849000},"page":"970","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Rapid Visual Detection of Cyprinid Herpesvirus 2 Based on One-Tube RPA-CRISPR\/Cas12a System"],"prefix":"10.3390","volume":"13","author":[{"given":"Jiaqi","family":"Li","sequence":"first","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"},{"role":"writing-original-draft","vocabulary":"credit"}]},{"given":"Wen","family":"Zou","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"},{"role":"data-curation","vocabulary":"credit"}]},{"given":"Yuping","family":"Tang","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"},{"role":"data-curation","vocabulary":"credit"}]},{"given":"Mingyang","family":"Liu","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"},{"role":"resources","vocabulary":"credit"}]},{"given":"Yongjie","family":"Liu","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"},{"role":"supervision","vocabulary":"credit"}]},{"given":"Yuhao","family":"Dong","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"},{"role":"funding-acquisition","vocabulary":"credit"}]}],"member":"1968","published-online":{"date-parts":[[2026,9,16]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"73","DOI":"10.2478\/jvetres-2024-0008","article-title":"Occurrence of herpesvirus in fish","volume":"68","author":"Bergmann","year":"2024","journal-title":"J. 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