{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,24]],"date-time":"2026-09-24T21:11:00Z","timestamp":1790284260640,"version":"4.1.0"},"reference-count":39,"publisher":"MDPI AG","issue":"8","license":[{"start":{"date-parts":[[2022,7,30]],"date-time":"2022-07-30T00:00:00Z","timestamp":1659139200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Jiaxing Municipal Science and Technology Project","award":["2020AY30024"],"award-info":[{"award-number":["2020AY30024"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Pathogens"],"abstract":"<jats:p>Molecular diagnostic assays for cholera detection have superior sensitivity to conventional assays and are now being accepted as the new standard method, especially the real-time PCR\/RT-PCR. However, limited throughput capacity and long detection duration prevent them from detecting more specimens and more targets in one turnaround time simultaneously. In this study, we utilized nucleic acid extraction-free, direct RT-PCR and high-speed amplification to develop a novel multiplex assay, a quadplex direct one-tube real-time RT-PCR assay, for rapid detection of the serogroup and cholera toxin toxigenicity of Vibrio cholerae targeting the epsM, ctxA, rfb-O1, and rfb-O139 genes. Performance of the multiplex assay was evaluated by comparison with the monoplex real-time PCR assay according to the China Cholera Prevention Manual. Detection data from clinical specimens showed that the new assay had good diagnostic sensitivities for epsM (100%, n = 301), ctxA (100%, n = 125), rfb-O1 (100%, n = 85), and rfb-O139 (97.87%, n = 49). Analysis of the analytical sensitivities with serial dilutions of positive standards showed that the detection limits of the new assay for Vibrio cholerae\u00a0epsM,ctxA,rfb-O1, and rfb-O139 were up to 200, 590, 115, and 1052 copies per mL lower than the monoplex real-time PCR (910, 345, and 1616 copies\/mL respectively, for ctxA,rfb-O1, and rfb-O139). The results indicate that the multiplex assay is a rapid, sensitive, specific, and easy-to-use detection tool for Vibrio cholerae, especially suitable for rapid identification and screening detection of mass specimens.<\/jats:p>","DOI":"10.3390\/pathogens11080865","type":"journal-article","created":{"date-parts":[[2022,8,1]],"date-time":"2022-08-01T04:04:00Z","timestamp":1659326640000},"page":"865","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":5,"title":["Direct and Rapid Identification of Vibrio Cholerae Serogroup and Toxigenicity by a Novel Multiplex Real-Time Assay"],"prefix":"10.3390","volume":"11","author":[{"given":"Yong","family":"Yan","sequence":"first","affiliation":[{"name":"Jiaxing Key Laboratory of Pathogenic Microbiology, Jiaxing Municipal Center for Disease Control and Prevention, Wenqiao Road 486, Jiaxing 314050, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Li","family":"Zhan","sequence":"additional","affiliation":[{"name":"Institute of Microbiology, Zhejiang Provincial Center for Disease Control and Prevention, Binsheng Road 3399, Hangzhou 310051, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Guoying","family":"Zhu","sequence":"additional","affiliation":[{"name":"Jiaxing Key Laboratory of Pathogenic Microbiology, Jiaxing Municipal Center for Disease Control and Prevention, Wenqiao Road 486, Jiaxing 314050, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Junyan","family":"Zhang","sequence":"additional","affiliation":[{"name":"Institute of Microbiology, Zhejiang Provincial Center for Disease Control and Prevention, Binsheng Road 3399, Hangzhou 310051, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ping","family":"Li","sequence":"additional","affiliation":[{"name":"Jiaxing Key Laboratory of Pathogenic Microbiology, Jiaxing Municipal Center for Disease Control and Prevention, Wenqiao Road 486, Jiaxing 314050, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lixia","family":"Chen","sequence":"additional","affiliation":[{"name":"Jiaxing Key Laboratory of Pathogenic Microbiology, Jiaxing Municipal Center for Disease Control and Prevention, Wenqiao Road 486, Jiaxing 314050, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Peiyan","family":"He","sequence":"additional","affiliation":[{"name":"Jiaxing Key Laboratory of Pathogenic Microbiology, Jiaxing Municipal Center for Disease Control and Prevention, Wenqiao Road 486, Jiaxing 314050, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jianyong","family":"Luo","sequence":"additional","affiliation":[{"name":"Jiaxing Key Laboratory of Pathogenic Microbiology, Jiaxing Municipal Center for Disease Control and Prevention, Wenqiao Road 486, Jiaxing 314050, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zhongwen","family":"Chen","sequence":"additional","affiliation":[{"name":"Jiaxing Key Laboratory of Pathogenic Microbiology, Jiaxing Municipal Center for Disease Control and Prevention, Wenqiao Road 486, Jiaxing 314050, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2022,7,30]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"2212","DOI":"10.1128\/JCM.00189-12","article-title":"Vibrio cholerae classical biotype strains reveal distinct signatures in Mexico","volume":"50","author":"Alam","year":"2012","journal-title":"J. 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