{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,4]],"date-time":"2026-07-04T15:52:35Z","timestamp":1783180355877,"version":"3.54.6"},"reference-count":34,"publisher":"MDPI AG","issue":"4","license":[{"start":{"date-parts":[[2026,4,16]],"date-time":"2026-04-16T00:00:00Z","timestamp":1776297600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100012166","name":"National Key Research and Development Program of China","doi-asserted-by":"publisher","award":["2023YFE0106100"],"award-info":[{"award-number":["2023YFE0106100"]}],"id":[{"id":"10.13039\/501100012166","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100006407","name":"Natural Science Foundation of Henan Province","doi-asserted-by":"publisher","award":["232300421009"],"award-info":[{"award-number":["232300421009"]}],"id":[{"id":"10.13039\/501100006407","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Elite Talents Development Program of Henan Academy of Agricultural Sciences","award":["2026RC03"],"award-info":[{"award-number":["2026RC03"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Viruses"],"abstract":"<jats:p>Marek\u2019s disease (MD), caused by pathogenic Marek\u2019s disease virus serotype 1 (MDV-1), is one of the most important avian immunosuppressive and neoplastic diseases and has led to huge economic losses to the poultry industry worldwide. Rapid and accurate clinical diagnosis is of great significance for efficient control of the disease. Herein, we have established a multiplex PCR (mPCR) method to simply differentiate all of the three types of MDV, using five specific primers targeting to MDV-1 oncogene meq or MDV-2 and MDV-3\/HVT gB genes. Simultaneously, it can detect any type of virulent or vaccine MDV strains in one PCR reaction, with amplicons of the short (S) and long (L)-meq of MDV-1 strains, and the gB of MDV-2 and HVT vaccine strains. Non-specific amplifications of avian leukosis virus (ALV), reticuloendotheliosis virus (REV), or fowl adenovirus virus 4 (FAdV-4) were not observed, indicating a good specificity of this method. A total of 522 clinical samples of tumor-bearing or suspected diseased birds collected from 30 poultry farms were detected. The results demonstrated that the newly developed mPCR method accurately detected and differentiated epidemic MDV-1 infections and vaccine strains, and provided nearly 100% consistency for detecting clinical wild-type infections compared with conventional PCR amplification of the meq gene. Collectively, our data has provided a highly efficient method for early differential diagnosis of MD clinical cases, virus identification and future evaluation of vaccination efficacy in healthy chicken flocks, which would be meaningful for efficient control of the disease.<\/jats:p>","DOI":"10.3390\/v18040471","type":"journal-article","created":{"date-parts":[[2026,4,16]],"date-time":"2026-04-16T11:35:42Z","timestamp":1776339342000},"page":"471","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Development of a Multiplex PCR Method for Efficient Differential Diagnosis of Clinical Cases and Vaccine Immunization of Marek\u2019s Disease"],"prefix":"10.3390","volume":"18","author":[{"given":"Wen-Kai","family":"Zhang","sequence":"first","affiliation":[{"name":"Institute for Animal Health & UK-China Centre of Excellence for Research on Avian Diseases, Henan Academy of Agricultural Sciences, Zhengzhou 450002, China"},{"name":"Laboratory of Functional Microbiology and Animal Health, College of Animal Science and Technology, Henan University of Science and Technology, Luoyang 471003, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2784-4169","authenticated-orcid":false,"given":"Man","family":"Teng","sequence":"additional","affiliation":[{"name":"Institute for Animal Health & UK-China Centre of Excellence for Research on Avian Diseases, Henan Academy of Agricultural Sciences, Zhengzhou 450002, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lu-Ping","family":"Zheng","sequence":"additional","affiliation":[{"name":"Institute for Animal Health & UK-China Centre of Excellence for Research on Avian Diseases, Henan Academy of Agricultural Sciences, Zhengzhou 450002, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bin","family":"Shi","sequence":"additional","affiliation":[{"name":"Institute for Animal Health & UK-China Centre of Excellence for Research on Avian Diseases, Henan Academy of Agricultural Sciences, Zhengzhou 450002, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wei-Dong","family":"Wang","sequence":"additional","affiliation":[{"name":"Institute for Animal Health & UK-China Centre of Excellence for Research on Avian Diseases, Henan Academy of Agricultural Sciences, Zhengzhou 450002, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Gui-Xi","family":"Li","sequence":"additional","affiliation":[{"name":"Shangqiu Center for Animal Quarantine and Disease Control and Prevention, Shangqiu 476100, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yong-Xu","family":"Zhao","sequence":"additional","affiliation":[{"name":"Scientific Support and Investigation Unit, Ceva Animal Health, Hangzhou 310018, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zhen","family":"Yang","sequence":"additional","affiliation":[{"name":"Jinyu Youbang Biotechnology (Jiangsu) Co., Ltd., Yangzhou 225007, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zu-Hua","family":"Yu","sequence":"additional","affiliation":[{"name":"Laboratory of Functional Microbiology and Animal Health, College of Animal Science and Technology, Henan University of Science and Technology, Luoyang 471003, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7351-8180","authenticated-orcid":false,"given":"Jun","family":"Luo","sequence":"additional","affiliation":[{"name":"Institute for Animal Health & UK-China Centre of Excellence for Research on Avian Diseases, Henan Academy of Agricultural Sciences, Zhengzhou 450002, China"},{"name":"Laboratory of Functional Microbiology and Animal Health, College of Animal Science and Technology, Henan University of Science and Technology, Luoyang 471003, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,4,16]]},"reference":[{"key":"ref_1","unstructured":"Nair, V., Gimeno, I., and Dunn, J. 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