{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,4,17]],"date-time":"2026-04-17T20:58:10Z","timestamp":1776459490673,"version":"3.51.2"},"reference-count":31,"publisher":"Wiley","issue":"1","license":[{"start":{"date-parts":[[2025,11,12]],"date-time":"2025-11-12T00:00:00Z","timestamp":1762905600000},"content-version":"vor","delay-in-days":315,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2025,1,1]],"date-time":"2025-01-01T00:00:00Z","timestamp":1735689600000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/doi.wiley.com\/10.1002\/tdm_license_1.1"}],"funder":[{"DOI":"10.13039\/501100012166","name":"National Key Research and Development Program of China","doi-asserted-by":"publisher","award":["2023YFD1800501"],"award-info":[{"award-number":["2023YFD1800501"]}],"id":[{"id":"10.13039\/501100012166","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["32302892"],"award-info":[{"award-number":["32302892"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100007540","name":"Jiangsu Agricultural Science and Technology Innovation Fund","doi-asserted-by":"publisher","award":["CX(24)2001"],"award-info":[{"award-number":["CX(24)2001"]}],"id":[{"id":"10.13039\/100007540","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100002858","name":"China Postdoctoral Science Foundation","doi-asserted-by":"publisher","award":["2024M751210"],"award-info":[{"award-number":["2024M751210"]}],"id":[{"id":"10.13039\/501100002858","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Transboundary and Emerging Diseases"],"published-print":{"date-parts":[[2025,1]]},"abstract":"<jats:p>\n                    Porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis virus (TGEV), porcine deltacoronavirus (PDCoV), and porcine rotavirus group A (PoRVA) are recognized as major enteric viral pathogens responsible for porcine viral diarrhea. These viruses exhibit similar clinical manifestations, including vomiting, diarrhea, and dehydration, which complicate differential diagnosis. Therefore, there is an urgent need for a highly sensitive and specific diagnostic method to differentiate these pathogens. In this study, we developed a TaqMan\u2010based one\u2010step quadruplex reverse transcription real\u2010time PCR (RT\u2010qPCR) assay for the simultaneous and differential detection of these four porcine diarrhea viruses. The standard curves demonstrated correlation coefficients (\n                    <jats:italic>R<\/jats:italic>\n                    <jats:sup>2<\/jats:sup>\n                    ) exceeded 0.990 across a dynamic range of 10\n                    <jats:sup>7.5<\/jats:sup>\n                    \u2013 10\n                    <jats:sup>2.5<\/jats:sup>\n                    TCID\n                    <jats:sub>50<\/jats:sub>\n                    \/mL, and amplification efficiency ranged from 90% to 110%. The limit of detection (LOD) were 10\n                    <jats:sup>1.5<\/jats:sup>\n                    TCID\n                    <jats:sub>50<\/jats:sub>\n                    \/mL. Specificity analysis showed no cross\u2010reactivity with other related pathogens. The assay exhibited good repeatability, with coefficients of variation (CVs) ranging from 0.15% to 1.41% for intra\u2010assay and 0.09% to 2.09% for inter\u2010assay, respectively. Finally, this method was evaluated for its practicality in the field using 348 clinical fecal samples. The positive rates of PEDV, TGEV, PDCoV, and PoRVA were 43.68%, 0.57%, 26.44%, and 33.91%, respectively. Furthermore, the coinfection rates of PEDV\/PDCoV, PEDV\/PoRVA, PDCoV\/PoRVA, and PEDV\/PDCoV\/PoRVA were14.37%, 13.51%, 2.01%, and 5.75%, respectively. Compared to singleplex RT\u2010qPCR assays, the quadruplex method demonstrated agreement rates ranging from 99.41% to 100% in detecting these four viral pathogens. In conclusion, the developed quadruplex RT\u2010qPCR assay offers a reliable, sensitive, and accurate tool for the identification of four causative agents of porcine viral diarrhea, making it suitable for clinical diagnosis, disease surveillance, and epidemiological studies.\n                  <\/jats:p>","DOI":"10.1155\/tbed\/9454210","type":"journal-article","created":{"date-parts":[[2025,11,12]],"date-time":"2025-11-12T09:42:56Z","timestamp":1762940576000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Development and Application of a TaqMan\u2010Based One\u2010Step Quadruplex Reverse Transcription Real\u2010Time PCR (RT\u2010qPCR) for Differential Detection of Four Porcine Diarrhea Viruses"],"prefix":"10.1155","volume":"2025","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-0721-447X","authenticated-orcid":false,"given":"Wei","family":"Wang","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0009-0006-8483-6877","authenticated-orcid":false,"given":"Zhaokun","family":"Chen","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0009-0002-9158-8163","authenticated-orcid":false,"given":"Dandan","family":"Wang","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5682-1834","authenticated-orcid":false,"given":"Jizong","family":"Li","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9780-5080","authenticated-orcid":false,"given":"Baochao","family":"Fan","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0647-2266","authenticated-orcid":false,"given":"Xuehan","family":"Zhang","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9323-0758","authenticated-orcid":false,"given":"Min","family":"Sun","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7707-6981","authenticated-orcid":false,"given":"Yongxiang","family":"Zhao","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3786-5687","authenticated-orcid":false,"given":"Jinzhu","family":"Zhou","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2743-539X","authenticated-orcid":false,"given":"Hongqi","family":"Shang","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1318-7081","authenticated-orcid":false,"given":"Bin","family":"Li","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0009-0006-1897-3718","authenticated-orcid":false,"given":"Junming","family":"Zhou","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2025,11,12]]},"reference":[{"key":"e_1_2_14_1_2","doi-asserted-by":"publisher","DOI":"10.3390\/v15061238"},{"key":"e_1_2_14_2_2","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2024.1380849"},{"key":"e_1_2_14_3_2","doi-asserted-by":"publisher","DOI":"10.3390\/v15010154"},{"key":"e_1_2_14_4_2","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2024.1390328"},{"key":"e_1_2_14_5_2","doi-asserted-by":"publisher","DOI":"10.3390\/vetsci9110634"},{"key":"e_1_2_14_6_2","doi-asserted-by":"publisher","DOI":"10.3390\/v14102217"},{"key":"e_1_2_14_7_2","doi-asserted-by":"publisher","DOI":"10.1007\/BF01317606"},{"key":"e_1_2_14_8_2","doi-asserted-by":"publisher","DOI":"10.1186\/s12985-015-0421-2"},{"key":"e_1_2_14_9_2","doi-asserted-by":"publisher","DOI":"10.3390\/v14112434"},{"key":"e_1_2_14_10_2","doi-asserted-by":"publisher","DOI":"10.1007\/s12250-021-00364-0"},{"key":"e_1_2_14_11_2","first-page":"257","article-title":"A Transmissible Gastroenteritis in Pigs","volume":"108","author":"Doyle L. P.","year":"1946","journal-title":"Journal of the American Veterinary Medical Association"},{"key":"e_1_2_14_12_2","doi-asserted-by":"publisher","DOI":"10.1007\/s11259-021-09808-0"},{"key":"e_1_2_14_13_2","doi-asserted-by":"publisher","DOI":"10.1186\/s12917-022-03554-4"},{"key":"e_1_2_14_14_2","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.06540-11"},{"key":"e_1_2_14_15_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.micpath.2023.106118"},{"key":"e_1_2_14_16_2","doi-asserted-by":"publisher","DOI":"10.3390\/v9030048"},{"key":"e_1_2_14_17_2","doi-asserted-by":"publisher","DOI":"10.1128\/jvi.00212-24"},{"key":"e_1_2_14_18_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.micpath.2024.106646"},{"key":"e_1_2_14_19_2","doi-asserted-by":"publisher","DOI":"10.1186\/s44149-024-00137-8"},{"key":"e_1_2_14_20_2","doi-asserted-by":"publisher","DOI":"10.1128\/mbio.02958-23"},{"key":"e_1_2_14_21_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.virol.2020.03.001"},{"key":"e_1_2_14_22_2","doi-asserted-by":"publisher","DOI":"10.3390\/v13122403"},{"key":"e_1_2_14_23_2","doi-asserted-by":"publisher","DOI":"10.1373\/clinchem.2008.112797"},{"key":"e_1_2_14_24_2","doi-asserted-by":"publisher","DOI":"10.3389\/fvets.2023.1158585"},{"key":"e_1_2_14_25_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.virusres.2022.198954"},{"key":"e_1_2_14_26_2","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2022.1074513"},{"key":"e_1_2_14_27_2","doi-asserted-by":"publisher","DOI":"10.3390\/vetsci10060402"},{"key":"e_1_2_14_28_2","doi-asserted-by":"publisher","DOI":"10.3390\/v17040493"},{"key":"e_1_2_14_29_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.virol.2025.110582"},{"key":"e_1_2_14_30_2","doi-asserted-by":"publisher","DOI":"10.1080\/22221751.2024.2332653"},{"key":"e_1_2_14_31_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.talanta.2024.127428"}],"container-title":["Transboundary and Emerging Diseases"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/pdf\/10.1155\/tbed\/9454210","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/full-xml\/10.1155\/tbed\/9454210","content-type":"application\/xml","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/pdf\/10.1155\/tbed\/9454210","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,3,8]],"date-time":"2026-03-08T20:58:42Z","timestamp":1773003522000},"score":1,"resource":{"primary":{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/10.1155\/tbed\/9454210"}},"subtitle":[],"editor":[{"given":"Nan-Hua","family":"Chen","sequence":"additional","affiliation":[],"role":[{"role":"editor","vocabulary":"crossref"}]}],"short-title":[],"issued":{"date-parts":[[2025,1]]},"references-count":31,"journal-issue":{"issue":"1","published-print":{"date-parts":[[2025,1]]}},"alternative-id":["10.1155\/tbed\/9454210"],"URL":"https:\/\/doi.org\/10.1155\/tbed\/9454210","archive":["Portico"],"relation":{},"ISSN":["1865-1674","1865-1682"],"issn-type":[{"value":"1865-1674","type":"print"},{"value":"1865-1682","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,1]]},"assertion":[{"value":"2025-08-26","order":0,"name":"received","label":"Received","group":{"name":"publication_history","label":"Publication History"}},{"value":"2025-09-29","order":2,"name":"accepted","label":"Accepted","group":{"name":"publication_history","label":"Publication History"}},{"value":"2025-11-12","order":3,"name":"published","label":"Published","group":{"name":"publication_history","label":"Publication History"}}],"article-number":"9454210"}}