{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,11]],"date-time":"2026-09-11T18:07:47Z","timestamp":1789150067587,"version":"build-2803163510"},"reference-count":35,"publisher":"Public Library of Science (PLoS)","issue":"4","license":[{"start":{"date-parts":[[2026,4,20]],"date-time":"2026-04-20T00:00:00Z","timestamp":1776643200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Start-up fund for New Ph.D. Researchers of Suzhou Chien-Shiung Institute of Technology.","award":["60000309"],"award-info":[{"award-number":["60000309"]}]},{"name":"The Natural Science Foundation of Jiangsu Province","award":["BK20220301"],"award-info":[{"award-number":["BK20220301"]}]},{"name":"Jiangsu Province high level innovation and entrepreneurship talent introduction plan","award":["JSSCBS20221009"],"award-info":[{"award-number":["JSSCBS20221009"]}]},{"name":"Natural Science Foundation of the Jiangsu Higher Education Institutions","award":["25KJB320013"],"award-info":[{"award-number":["25KJB320013"]}]},{"name":"Science Projects of Taicang City, China","award":["TC2022JC29"],"award-info":[{"award-number":["TC2022JC29"]}]},{"DOI":"10.13039\/501100013088","name":"Qing Lan Project","doi-asserted-by":"crossref","award":["Su Teacher [2024]14"],"award-info":[{"award-number":["Su Teacher [2024]14"]}],"id":[{"id":"10.13039\/501100013088","id-type":"DOI","asserted-by":"crossref"}]}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS One"],"abstract":"<jats:p>Avian metapneumovirus subgroup A (aMPV\/A) causes widespread infections in commercial poultry worldwide. The development of highly efficient viral vector vaccines is constrained by multiple factors, among which the lack of a mature reverse genetics system and the uncertainty regarding the optimal insertion site for foreign genes represent one of the core technical bottlenecks. Here, we constructed an aMPV\/A reverse genetics system based on the pBR322 vector and inserted EGFP into seven of its intergenic regions to generate recombinant viruses. Viral replication, exogenous gene expression and genetic stability were systematically assessed via multiple assays. The results demonstrated that the expression level at the P\u2011M locus after EGFP insertion was approximately 1.2\u2011fold higher than that of other recombinant viruses. In addition, there was no significant difference in viral titer, and good genetic stability was observed over 20 generations. This study establishes a stable and efficient reverse genetics system for aMPV\/A, which provides a critical technical platform for aMPV\/A molecular mechanism research and novel recombinant vector vaccine development, with important practical value for global aMPV prevention and control.<\/jats:p>","DOI":"10.1371\/journal.pone.0347597","type":"journal-article","created":{"date-parts":[[2026,4,20]],"date-time":"2026-04-20T17:34:07Z","timestamp":1776706447000},"page":"e0347597","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":0,"title":["Identification of an optimal exogenous gene insertion site (P\u2013M) and establishment of a reverse genetics system for aMPV\/A"],"prefix":"10.1371","volume":"21","author":[{"ORCID":"https:\/\/orcid.org\/0009-0003-8411-5437","authenticated-orcid":true,"given":"Yu","family":"Guo","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ting","family":"Xue","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bingqing","family":"Lu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jiqing","family":"Zheng","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yirong","family":"Wang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xueqiao","family":"Xie","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Huiting","family":"Wang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Furui","family":"Cao","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yijun","family":"Zhang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jianhua","family":"Wang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"340","published-online":{"date-parts":[[2026,4,20]]},"reference":[{"issue":"9","key":"pone.0347597.ref001","doi-asserted-by":"crossref","first-page":"1960","DOI":"10.3390\/v15091960","article-title":"Trends and Challenges in the Surveillance and Control of Avian Metapneumovirus","volume":"15","author":"GBC Salles","year":"2023","journal-title":"Viruses"},{"key":"pone.0347597.ref002","unstructured":"Falchieri M. Avian metapneumovirus in poultry \u2013 an overview. CABI Compendium. 2019."},{"issue":"4","key":"pone.0347597.ref003","doi-asserted-by":"crossref","first-page":"42","DOI":"10.52419\/issn2782-6252.2022.4.42","article-title":"Diagnosis of avian metapneumovirus infection using multiplex PCR","author":"SR Abgaryan","year":"2023","journal-title":"Legal regulation in veterinary medicine"},{"issue":"4","key":"pone.0347597.ref004","doi-asserted-by":"crossref","first-page":"100211","DOI":"10.1016\/j.japr.2021.100211","article-title":"Avian metapneumovirus infection in turkeys: a review on turkey rhinotracheitis","volume":"30","author":"K Kaboudi","year":"2021","journal-title":"Journal of Applied Poultry Research"},{"issue":"4","key":"pone.0347597.ref005","doi-asserted-by":"crossref","first-page":"508","DOI":"10.3390\/v16040508","article-title":"Geographical expansion of avian metapneumovirus subtype B: first detection and molecular characterization of avian metapneumovirus subtype B in US poultry","volume":"16","author":"M Luqman","year":"2024","journal-title":"Viruses"},{"issue":"4","key":"pone.0347597.ref006","doi-asserted-by":"crossref","first-page":"311","DOI":"10.1080\/03079457.2019.1584390","article-title":"Host specificity of avian metapneumoviruses","volume":"48","author":"PA Brown","year":"2019","journal-title":"Avian Pathol"},{"issue":"3","key":"pone.0347597.ref007","doi-asserted-by":"crossref","first-page":"971","DOI":"10.47836\/pjtas.46.3.14","article-title":"Avian Metapneumovirus Infection in Poultry Flocks: A Review of Current Knowledge","volume":"46","author":"W A. 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