{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,4,1]],"date-time":"2026-04-01T07:26:38Z","timestamp":1775028398814,"version":"3.50.1"},"reference-count":43,"publisher":"MDPI AG","issue":"3","license":[{"start":{"date-parts":[[2023,2,24]],"date-time":"2023-02-24T00:00:00Z","timestamp":1677196800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100008771","name":"Animal and Plant Quarantine Agency","doi-asserted-by":"publisher","award":["N-1543418-2018-22-01"],"award-info":[{"award-number":["N-1543418-2018-22-01"]}],"id":[{"id":"10.13039\/501100008771","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Ministry of Agriculture, Food and Rural Affairs, Republic of Korea","award":["N-1543418-2018-22-01"],"award-info":[{"award-number":["N-1543418-2018-22-01"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Vaccines"],"abstract":"<jats:p>Prior to the identification of low pathogenic avian influenza H9N2 viruses belonging to the Y280 lineage in 2020, Y439 lineage viruses had been circulating in the Republic of Korea since 1996. Here, we developed a whole inactivated vaccine (vac564) by multiple passage of Y439 lineage viruses and then evaluated immunogenicity and protective efficacy in specific-pathogen-free chickens. We found that LBM564 could be produced at high yield in eggs (108.4EID50\/0.1 mL; 1024 hemagglutinin units) and was immunogenic (8.0 \u00b1 1.2 log2) in chickens. The vaccine showed 100% inhibition of virus in the cecal tonsil with no viral shedding detected in either oropharyngeal or cloacal swabs after challenge with homologous virus. However, it did not induce effective protection against challenge with heterologous virus. An imported commercial G1 lineage vaccine inhibited viral replication against Y280 and Y439 lineage viruses in major tissues, although viral shedding in oropharyngeal and cloacal swabs was observed up until 5 dpi after exposure to both challenge viruses. These results suggest that a single vaccination with vac564 could elicit immune responses, showing it to be capable of protecting chickens against the Y439 lineage virus. Thus, our results suggest the need to prepare suitable vaccines for use against newly emerging and re-emerging H9N2 viruses.<\/jats:p>","DOI":"10.3390\/vaccines11030538","type":"journal-article","created":{"date-parts":[[2023,2,24]],"date-time":"2023-02-24T03:57:35Z","timestamp":1677211055000},"page":"538","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Protection of SPF Chickens by H9N2 Y439 and G1 Lineage Vaccine against Homologous and Heterologous Viruses"],"prefix":"10.3390","volume":"11","author":[{"given":"Hyun-Kyu","family":"Cho","sequence":"first","affiliation":[{"name":"Avian Influenza Research & Diagnostic Division, Animal and Plant Quarantine Agency, 177 Hyeoksin 8-ro, Gimcheon-si 39660, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0353-9138","authenticated-orcid":false,"given":"Yong-Myung","family":"Kang","sequence":"additional","affiliation":[{"name":"Avian Influenza Research & Diagnostic Division, Animal and Plant Quarantine Agency, 177 Hyeoksin 8-ro, Gimcheon-si 39660, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5809-364X","authenticated-orcid":false,"given":"Mingeun","family":"Sagong","sequence":"additional","affiliation":[{"name":"Avian Influenza Research & Diagnostic Division, Animal and Plant Quarantine Agency, 177 Hyeoksin 8-ro, Gimcheon-si 39660, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Juhun","family":"Kim","sequence":"additional","affiliation":[{"name":"Bioapp Institute, 394 Jigok-ro, Pohang-si 37668, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Hyunjun","family":"Kim","sequence":"additional","affiliation":[{"name":"Avian Influenza Research & Diagnostic Division, Animal and Plant Quarantine Agency, 177 Hyeoksin 8-ro, Gimcheon-si 39660, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sungjun","family":"An","sequence":"additional","affiliation":[{"name":"Avian Influenza Research & Diagnostic Division, Animal and Plant Quarantine Agency, 177 Hyeoksin 8-ro, Gimcheon-si 39660, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Youn-Jeong","family":"Lee","sequence":"additional","affiliation":[{"name":"Avian Influenza Research & Diagnostic Division, Animal and Plant Quarantine Agency, 177 Hyeoksin 8-ro, Gimcheon-si 39660, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Hyun-Mi","family":"Kang","sequence":"additional","affiliation":[{"name":"Avian Influenza Research & Diagnostic Division, Animal and Plant Quarantine Agency, 177 Hyeoksin 8-ro, Gimcheon-si 39660, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2023,2,24]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"9372","DOI":"10.1128\/JVI.74.20.9372-9380.2000","article-title":"H9N2 influenza viruses possessing H5N1-like internal genomes continue to circulate in poultry in Southeastern China","volume":"74","author":"Guan","year":"2000","journal-title":"J. Virol."},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Pusch, E.A., and Suarez, D.L. (2018). The multifaceted zoonotic risk of H9N2 avian influenza. Vet. Sci., 5.","DOI":"10.3390\/vetsci5040082"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"67","DOI":"10.1016\/j.vaccine.2008.07.064","article-title":"Animal influenza epidemiology","volume":"26","author":"Ducatez","year":"2008","journal-title":"Vaccine"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"26","DOI":"10.7774\/cevr.2013.2.1.26","article-title":"H9N2 avian influenza virus in Korea: Evolution and vaccination","volume":"2","author":"Lee","year":"2013","journal-title":"Clin. Exp. Vaccine Res."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"67","DOI":"10.4142\/jvs.2008.9.1.67","article-title":"An inactivated vaccine to control the current H9N2 low pathogenic avian influenza in Korea","volume":"9","author":"Choi","year":"2008","journal-title":"J. Vet. Sci."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"508","DOI":"10.1637\/11123-050615-Reg","article-title":"Vaccine efficacy against a new avian influenza (H9N2) field isolate from the Middle East (serology and challenge studies)","volume":"59","author":"Gharaibeh","year":"2015","journal-title":"Avian Dis."},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Jin, F., Dong, X., Wan, Z., Ren, D., Liu, M., Geng, T., Zhang, J., Gao, W., Shao, H., and Qin, A. (2019). A single mutation N166D in hemagglutinin affects antigenicity and pathogenesis of H9N2 avian influenza virus. Viruses, 11.","DOI":"10.3390\/v11080709"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"9363","DOI":"10.1073\/pnas.96.16.9363","article-title":"Molecular characterization of H9N2 influenza viruses: Were they the donors of the \u201cinternal\u201d genes of H5N1 viruses in Hong Kong?","volume":"96","author":"Guan","year":"1999","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"83","DOI":"10.1186\/s13567-018-0575-1","article-title":"A G1-lineage H9N2 virus with oviduct tropism causes chronic pathological changes in the infundibulum and a long-lasting drop in egg production","volume":"49","author":"Bonfante","year":"2018","journal-title":"Vet. Res."},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Peacock, T.P., James, J., Sealy, J.E., and Iqbal, M. (2019). A global perspective on H9N2 avian influenza virus. Viruses, 11.","DOI":"10.20944\/preprints201906.0085.v1"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"215","DOI":"10.1016\/j.vetmic.2010.05.010","article-title":"Genotypic evolution and antigenic drift of H9N2 influenza viruses in China from 1994 to 2008","volume":"146","author":"Sun","year":"2010","journal-title":"Vet. Microbiol."},{"key":"ref_12","first-page":"1","article-title":"Genetic evolution of influenza H9N2 viruses isolated from various hosts in China from 1994 to 2013","volume":"6","author":"Li","year":"2017","journal-title":"Emerg. Microbes Infect."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"8609","DOI":"10.1128\/JVI.78.16.8609-8614.2004","article-title":"Continuing evolution of H9N2 influenza viruses in Southeastern China","volume":"78","author":"Choi","year":"2004","journal-title":"J. Virol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"548","DOI":"10.1099\/jgv.0.000657","article-title":"Novel reassortant H9N2 viruses in pigeons and evidence for antigenic diversity of H9N2 viruses isolated from quails in Egypt","volume":"98","author":"Kandeil","year":"2017","journal-title":"J. Gen. Virol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"e000171","DOI":"10.1136\/vetreco-2016-000171","article-title":"H9N2 low pathogenic avian influenza in Pakistan (2012\u20132015)","volume":"3","author":"Lee","year":"2016","journal-title":"Vet. Rec."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"828","DOI":"10.1637\/0005-2086-47.s3.828","article-title":"Avian influenza (H9N2) outbreak in Iran","volume":"47","author":"Nili","year":"2003","journal-title":"Avian Dis."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"135","DOI":"10.1016\/j.virol.2019.01.002","article-title":"Molecular characterization and receptor binding specificity of H9N2 avian influenza viruses based on poultry-related environmental surveillance in China between 2013 and 2016","volume":"529","author":"Zou","year":"2019","journal-title":"Virology"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"125","DOI":"10.1017\/S0043933915002603","article-title":"Low pathogenic avian influenza H9N2: World-wide distribution","volume":"72","author":"Agha","year":"2016","journal-title":"Worlds Poult. Sci."},{"key":"ref_19","first-page":"379","article-title":"An occurrence of non-highly pathogenic avian influenza in Korea","volume":"47","author":"Mo","year":"2003","journal-title":"Avian Dis."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"527","DOI":"10.2307\/1593091","article-title":"Sequence analysis of the hemagglutinin gene of H9N2 Korean avian influenza viruses and assessment of the pathogenic potential of isolate MS96","volume":"44","author":"Lee","year":"2000","journal-title":"Avian Dis."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"313","DOI":"10.1016\/j.virol.2006.09.025","article-title":"Continuing evolution of H9 influenza viruses in Korean poultry","volume":"359","author":"Lee","year":"2007","journal-title":"Virology"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"225","DOI":"10.1016\/j.virol.2015.11.023","article-title":"Poultry vaccination directed evolution of H9N2 low pathogenic avian influenza viruses in Korea","volume":"488","author":"Lee","year":"2016","journal-title":"Virology"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"e21","DOI":"10.4142\/jvs.2021.22.e21","article-title":"Genetic characterization of H9N2 avian influenza virus previously unrecognized in Korea","volume":"22","author":"Heo","year":"2021","journal-title":"J. Vet. Sci."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"327","DOI":"10.1080\/03079457.2021.1929833","article-title":"First report of field cases of Y280-like LPAI H9N2 strains in South Korean poultry farms: Pathological findings and genetic characterization","volume":"50","author":"Lai","year":"2021","journal-title":"Avian Pathol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"101318","DOI":"10.1016\/j.psj.2021.101318","article-title":"Pathogenicity of H9N2 low pathogenic avian influenza viruses of different lineages isolated from live bird markets tested in three animal models: SPF chickens, Korean native chickens, and ducks","volume":"100","author":"Kye","year":"2021","journal-title":"Poult. Sci."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"6201","DOI":"10.1016\/j.vaccine.2021.08.089","article-title":"Development of a recombinant H9N2 influenza vaccine candidate against the Y280 lineage field virus and its protective efficacy","volume":"39","author":"Kim","year":"2021","journal-title":"Vaccine"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"6213","DOI":"10.1016\/j.vaccine.2021.09.028","article-title":"Cross-protective efficacy of inactivated whole influenza vaccines against Korean Y280 and Y439 lineage H9N2 viruses in mice","volume":"39","author":"Park","year":"2021","journal-title":"Vaccine"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"268","DOI":"10.1016\/j.vetmic.2008.02.005","article-title":"Generation and evaluation of reassortant influenza vaccines made by reverse genetics for H9N2 avian influenza in Korea","volume":"130","author":"Song","year":"2008","journal-title":"Vet. Microbiol."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"2537","DOI":"10.1016\/j.vaccine.2016.04.022","article-title":"Recombinant Newcastle disease virus expressing H9 HA protects chickens against heterologous avian influenza H9N2 virus challenge","volume":"34","author":"Nagy","year":"2016","journal-title":"Vaccine"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"50","DOI":"10.1016\/j.antiviral.2011.04.007","article-title":"Immune responses of chickens inoculated with a recombinant fowlpox vaccine coexpressing HA of H9N2 avian influenza virus and chicken IL-18","volume":"91","author":"Chen","year":"2011","journal-title":"Antivir. Res."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"7","DOI":"10.1016\/j.virol.2019.01.004","article-title":"Recombinant turkey herpesvirus expressing H9 hemagglutinin providing protection against H9N2 avian influenza","volume":"529","author":"Liu","year":"2019","journal-title":"Virology"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"4003","DOI":"10.1016\/j.vaccine.2011.03.067","article-title":"H9N2 avian influenza virus-like particle vaccine provides protective immunity and a strategy for the differentiation of infected from vaccinated animals","volume":"29","author":"Lee","year":"2011","journal-title":"Vaccine"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"1681","DOI":"10.1586\/erv.09.128","article-title":"Continuous cell lines as a production system for influenza vaccines","volume":"8","author":"Genzel","year":"2009","journal-title":"Expert Rev. Vaccines"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"63","DOI":"10.1186\/s40643-020-00352-4","article-title":"Highly efficient production of an influenza H9N2 vaccine using MDCK suspension cells","volume":"7","author":"Wu","year":"2020","journal-title":"Bioresour. Bioprocess"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"476","DOI":"10.1128\/CMR.00097-12","article-title":"Traditional and new influenza vaccines","volume":"26","author":"Wong","year":"2013","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"85","DOI":"10.5501\/wjv.v5.i2.85","article-title":"Determination of 50% endpoint titer using a simple formula","volume":"5","author":"Ramakrishnan","year":"2016","journal-title":"World J. Virol."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"493","DOI":"10.1093\/oxfordjournals.aje.a118408","article-title":"A simple method of estimating fifty per cent endpoints","volume":"27","author":"Reed","year":"1938","journal-title":"Am. J. Epidemiol."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"8148","DOI":"10.1038\/ncomms9148","article-title":"Development of high-yield influenza A virus vaccine viruses","volume":"6","author":"Ping","year":"2015","journal-title":"Nat. Commun."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"1099","DOI":"10.1016\/S0140-6736(04)15892-3","article-title":"Responsiveness to a pandemic alert: Use of reverse genetics for rapid development of influenza vaccines","volume":"363","author":"Webby","year":"2004","journal-title":"Lancet"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"174","DOI":"10.1196\/annals.1373.021","article-title":"Principles for vaccine protection in chickens and domestic waterfowl against avian influenza","volume":"1081","author":"Swayne","year":"2006","journal-title":"Ann. N. Y. Acad. Sci."},{"key":"ref_41","doi-asserted-by":"crossref","unstructured":"Zhang, A., Lai, H., Xu, J., Huang, W., Liu, Y., Zhao, D., and Chen, R. (2017). Evaluation of the protective efficacy of Poly I:C as an adjuvant for H9N2 subtype avian influenza inactivated vaccine and its mechanism of action in ducks. PLoS ONE, 12.","DOI":"10.1371\/journal.pone.0170681"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"1433","DOI":"10.1007\/s00705-008-0131-9","article-title":"Genetic analysis of H9N2 avian influenza viruses isolated from India","volume":"153","author":"Tosh","year":"2008","journal-title":"Arch. Virol."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"165","DOI":"10.1016\/S0378-1135(00)00176-0","article-title":"Vaccine protect chickens against H5 highly pathogenic avian influenza in the face of genetic changes in field viruses over multiple years","volume":"74","author":"Swayne","year":"2000","journal-title":"Vet. Microbiol."}],"container-title":["Vaccines"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-393X\/11\/3\/538\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T18:41:22Z","timestamp":1760121682000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-393X\/11\/3\/538"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,2,24]]},"references-count":43,"journal-issue":{"issue":"3","published-online":{"date-parts":[[2023,3]]}},"alternative-id":["vaccines11030538"],"URL":"https:\/\/doi.org\/10.3390\/vaccines11030538","relation":{},"ISSN":["2076-393X"],"issn-type":[{"value":"2076-393X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2023,2,24]]}}}