{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,8]],"date-time":"2025-10-08T11:11:39Z","timestamp":1759921899480,"version":"build-2065373602"},"reference-count":42,"publisher":"MDPI AG","issue":"1","license":[{"start":{"date-parts":[[2025,1,15]],"date-time":"2025-01-15T00:00:00Z","timestamp":1736899200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001691","name":"JSPS KAKENHI","doi-asserted-by":"publisher","award":["26860271","17K08801"],"award-info":[{"award-number":["26860271","17K08801"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Tokyo Metropolitan Government, Japan","award":["26860271","17K08801"],"award-info":[{"award-number":["26860271","17K08801"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Vaccines"],"abstract":"<jats:p>Background\/Objectives: In preparation for a potential pandemic caused by the H5N1 highly pathogenic avian influenza (HPAI) virus, pre-pandemic vaccines against several viral clades have been developed and stocked worldwide. Although these vaccines are well tolerated, their immunogenicity and cross-reactivity with viruses of different clades can be improved. Methods: To address this aspect, we generated recombinant influenza vaccines against H5-subtype viruses using two different strains of highly attenuated vaccinia virus (VACV) vectors. Results: rLC16m8-mcl2.2 hemagglutinin (HA) and rLC16m8-mcl2.3.4 HA consisted of a recombinant LC16m8 vector encoding the HA protein from clade 2.2 or clade 2.3.4 viruses (respectively); rDIs-mcl2.2 HA consisted of a recombinant DIs vector encoding the HA protein from clade 2.2. A single dose of rLC16m8-mcl2.2 HA showed rapid (1 week after vaccination) and long-term protection (20 months post-vaccination) in mice against the HPAI H5N1 virus. Moreover, cynomolgus macaques immunized with rLC16m8-mcl2.2 HA exhibited long-term protection when challenged with a heterologous clade of the HPAI H5N1 virus. Although the DIs strain is unable to grow in most mammalian cells, rDIs-mcl2.2 HA also showed rapid and long-lasting effects against HPAI H5N1 virus infection. Notably, the protective efficacy of rDIs-mcl2.2 HA was comparable to that of rLC16m8-mcl2.2 HA. Furthermore, these vaccines protected animals previously immunized with VACVs from a lethal challenge with the HPAI H5N1 virus. Conclusions: These results suggest that both rLC16m8-mcl2.2 HA and rDIs-mcl2.2 HA are effective in preventing HPAI H5N1 virus infection, and rDIs-mcl2.2 HA is a promising vaccine candidate against H5 HA-subtype viruses.<\/jats:p>","DOI":"10.3390\/vaccines13010074","type":"journal-article","created":{"date-parts":[[2025,1,15]],"date-time":"2025-01-15T05:59:42Z","timestamp":1736920782000},"page":"74","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Single Dose of Attenuated Vaccinia Viruses Expressing H5 Hemagglutinin Affords Rapid and Long-Term Protection Against Lethal Infection with Highly Pathogenic Avian Influenza A H5N1 Virus in Mice and Monkeys"],"prefix":"10.3390","volume":"13","author":[{"given":"Fumihiko","family":"Yasui","sequence":"first","affiliation":[{"name":"Department of Microbiology and Cell Biology, Tokyo Metropolitan Institute of Medical Science, 2-1-6, Kamikitazawa, Setagaya-ku, Tokyo 156-8506, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Keisuke","family":"Munekata","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Cell Biology, Tokyo Metropolitan Institute of Medical Science, 2-1-6, Kamikitazawa, Setagaya-ku, Tokyo 156-8506, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5533-4129","authenticated-orcid":false,"given":"Tomoko","family":"Fujiyuki","sequence":"additional","affiliation":[{"name":"Infectious Disease Control Science, Institute of Industrial Science, The University of Tokyo, 4-6-1, Komaba, Meguro-ku, Tokyo 153-8505, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Takeshi","family":"Kuraishi","sequence":"additional","affiliation":[{"name":"Animal Laboratory of Injurious Animals, The Institute of Medical Science, The University of Tokyo, 802, Tean Sude, Setouchi-cho, Oshima-gun, Kagoshima 894-1531, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kenzaburo","family":"Yamaji","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Cell Biology, Tokyo Metropolitan Institute of Medical Science, 2-1-6, Kamikitazawa, Setagaya-ku, Tokyo 156-8506, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Tomoko","family":"Honda","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Cell Biology, Tokyo Metropolitan Institute of Medical Science, 2-1-6, Kamikitazawa, Setagaya-ku, Tokyo 156-8506, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sumiko","family":"Gomi","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Cell Biology, Tokyo Metropolitan Institute of Medical Science, 2-1-6, Kamikitazawa, Setagaya-ku, Tokyo 156-8506, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Misako","family":"Yoneda","sequence":"additional","affiliation":[{"name":"Infectious Disease Control Science, Institute of Industrial Science, The University of Tokyo, 4-6-1, Komaba, Meguro-ku, Tokyo 153-8505, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Takahiro","family":"Sanada","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Cell Biology, Tokyo Metropolitan Institute of Medical Science, 2-1-6, Kamikitazawa, Setagaya-ku, Tokyo 156-8506, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3812-0094","authenticated-orcid":false,"given":"Koji","family":"Ishii","sequence":"additional","affiliation":[{"name":"Center for Quality Management Systems, National Institute of Infectious Diseases, 4-7-1, Gakuen, Musashi-murayama, Tokyo 208-0011, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7021-1688","authenticated-orcid":false,"given":"Yoshihiro","family":"Sakoda","sequence":"additional","affiliation":[{"name":"Laboratory of Microbiology, Faculty of Veterinary Medicine, Hokkaido University, Kita 18 Nishi 9, Kita-ku, Sapporo 060-0818, Japan"},{"name":"Institute for Vaccine Research and Development (HU-IVReD), Hokkaido University, Sapporo 001-0021, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Hiroshi","family":"Kida","sequence":"additional","affiliation":[{"name":"Institute for Vaccine Research and Development (HU-IVReD), Hokkaido University, Sapporo 001-0021, Japan"},{"name":"International Institute for Zoonosis Control, Hokkaido University, Sapporo 001-0020, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Shosaku","family":"Hattori","sequence":"additional","affiliation":[{"name":"Animal Laboratory of Injurious Animals, The Institute of Medical Science, The University of Tokyo, 802, Tean Sude, Setouchi-cho, Oshima-gun, Kagoshima 894-1531, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Chieko","family":"Kai","sequence":"additional","affiliation":[{"name":"Infectious Disease Control Science, Institute of Industrial Science, The University of Tokyo, 4-6-1, Komaba, Meguro-ku, Tokyo 153-8505, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2130-5837","authenticated-orcid":false,"given":"Michinori","family":"Kohara","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Cell Biology, Tokyo Metropolitan Institute of Medical Science, 2-1-6, Kamikitazawa, Setagaya-ku, Tokyo 156-8506, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2025,1,15]]},"reference":[{"key":"ref_1","unstructured":"(2024, October 30). Cumulative Number of Confirmed Human Cases for Avian Influenza A(H5N1) Reported to WHO, 2003\u20132024, 27 September 2024. Available online: https:\/\/www.who.int\/publications\/m\/item\/cumulative-number-of-confirmed-human-cases-for-avian-influenza-a(h5n1)-reported-to-who--2003-2024--27-september-2024."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"786","DOI":"10.3201\/eid2904.221538","article-title":"Highly Pathogenic Avian Influenza A(H5N1) Virus Outbreak in New England Seals, United States","volume":"29","author":"Puryear","year":"2023","journal-title":"Emerg. Infect. Dis."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"veae031","DOI":"10.1093\/ve\/veae031","article-title":"Highly pathogenic avian influenza H5N1 virus infections in pinnipeds and seabirds in Uruguay: Implications for bird\u2013mammal transmission in South America","volume":"10","author":"Marandino","year":"2024","journal-title":"Virus Evol."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"2028","DOI":"10.1056\/NEJMc2405371","article-title":"Highly Pathogenic Avian Influenza A(H5N1) Virus Infection in a Dairy Farm Worker","volume":"390","author":"Uyeki","year":"2024","journal-title":"N. Engl. J. Med."},{"key":"ref_5","unstructured":"(2024, October 30). USDA Animal and Plant Health Inspection Service, Available online: https:\/\/www.aphis.usda.gov\/livestock-poultry-disease\/avian\/avian-influenza\/hpai-detections\/livestock."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"1959","DOI":"10.1128\/JVI.80.4.1959-1964.2006","article-title":"Protection of Mice and Poultry from Lethal H5N1 Avian Influenza Virus through Adenovirus-Based Immunization","volume":"80","author":"Gao","year":"2006","journal-title":"J. Virol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"150","DOI":"10.1128\/JVI.01514-06","article-title":"Newcastle Disease Virus-Based Live Attenuated Vaccine Completely Protects Chickens and Mice from Lethal Challenge of Homologous and Heterologous H5N1 Avian Influenza Viruses","volume":"81","author":"Ge","year":"2007","journal-title":"J. Virol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"1598","DOI":"10.1086\/517614","article-title":"Recombinant Modified Vaccinia Virus Ankara\u2013Based Vaccine Induces Protective Immunity in Mice against Infection with Influenza Virus H5N1","volume":"195","author":"Kreijtz","year":"2007","journal-title":"J. Infect. Dis."},{"key":"ref_9","doi-asserted-by":"crossref","unstructured":"Bull, J.J., Nuismer, S.L., and Antia, R. (2019). Recombinant vector vaccine evolution. PLoS Comput. Biol., 15.","DOI":"10.1371\/journal.pcbi.1006857"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1111\/imm.12829","article-title":"Novel viral vectors in infectious diseases","volume":"153","author":"Humphreys","year":"2018","journal-title":"Immunology"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"5179","DOI":"10.1128\/JVI.02642-05","article-title":"LC16m8, a Highly Attenuated Vaccinia Virus Vaccine Lacking Expression of the Membrane Protein B5R, Protects Monkeys from Monkeypox","volume":"80","author":"Saijo","year":"2006","journal-title":"J. Virol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"11873","DOI":"10.1128\/JVI.79.18.11873-11891.2005","article-title":"An Attenuated LC16m8 Smallpox Vaccine: Analysis of Full-Genome Sequence and Induction of Immune Protection","volume":"79","author":"Morikawa","year":"2005","journal-title":"J. Virol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1025","DOI":"10.1001\/jama.2009.289","article-title":"Clinical and Immunological Response to Attenuated Tissue-Cultured Smallpox Vaccine LC16m8","volume":"301","author":"Saito","year":"2009","journal-title":"JAMA"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"7009","DOI":"10.1016\/j.vaccine.2006.03.087","article-title":"LC16m8: An attenuated smallpox vaccine","volume":"24","author":"Kenner","year":"2006","journal-title":"Vaccine"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"1395","DOI":"10.1093\/infdis\/jir527","article-title":"Safety and Immunogenicity of LC16m8, an Attenuated Smallpox Vaccine in Vaccinia-Naive Adults","volume":"204","author":"Kennedy","year":"2011","journal-title":"J. Infect. Dis."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"6337","DOI":"10.4049\/jimmunol.181.9.6337","article-title":"Prior Immunization with Severe Acute Respiratory Syndrome (SARS)-Associated Coronavirus (SARS-CoV) Nucleocapsid Protein Causes Severe Pneumonia in Mice Infected with SARS-CoV","volume":"181","author":"Yasui","year":"2008","journal-title":"J. Immunol."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"37915","DOI":"10.1038\/srep37915","article-title":"Sensitization with vaccinia virus encoding H5N1 hemagglutinin restores immune potential against H5N1 influenza virus","volume":"6","author":"Yasui","year":"2016","journal-title":"Sci. Rep."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"381","DOI":"10.1038\/192381a0","article-title":"A New Mutant of Dermovaccinia Virus","volume":"192","author":"Tagaya","year":"1961","journal-title":"Nature"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"433","DOI":"10.1006\/viro.2002.1622","article-title":"Structural Analysis of Vaccinia Virus DIs Strain: Application as a New Replication-Deficient Viral Vector","volume":"302","author":"Ishiia","year":"2002","journal-title":"Virology"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"368","DOI":"10.1016\/j.virol.2006.03.020","article-title":"Induction of protective immunity against severe acute respiratory syndrome coronavirus (SARS-CoV) infection using highly attenuated recombinant vaccinia virus DIs","volume":"351","author":"Ishii","year":"2006","journal-title":"Virology"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"2485","DOI":"10.1099\/vir.0.021659-0","article-title":"Pathogenicity of highly pathogenic avian H5N1 influenza A viruses isolated from humans between 2003 and 2008 in northern Vietnam","volume":"91","author":"Le","year":"2010","journal-title":"J. Gen. Virol."},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Fujiyuki, T., Yoneda, M., Yasui, F., Kuraishi, T., Hattori, S., Kwon, H.-J., Munekata, K., Kiso, Y., Kida, H., and Kohara, M. (2013). Experimental Infection of Macaques with a Wild Water Bird-Derived Highly Pathogenic Avian Influenza Virus (H5N1). PLoS ONE, 8.","DOI":"10.1371\/journal.pone.0083551"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"3166","DOI":"10.1073\/pnas.0308391100","article-title":"Pathogenicity and immunogenicity of influenza viruses with genes from the 1918 pandemic virus","volume":"101","author":"Tumpey","year":"2004","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"75","DOI":"10.1016\/j.jviromet.2011.08.015","article-title":"Rapid typing of influenza viruses using super high-speed quantitative real-time PCR","volume":"178","author":"Sakurai","year":"2011","journal-title":"J. Virol. Methods"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"1131","DOI":"10.1038\/nm917","article-title":"Duration of antiviral immunity after smallpox vaccination","volume":"9","author":"Hammarlund","year":"2003","journal-title":"Nat. Med."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"1058","DOI":"10.1016\/j.amjmed.2008.08.019","article-title":"Immunity from Smallpox Vaccine Persists for Decades: A Longitudinal Study","volume":"121","author":"Taub","year":"2008","journal-title":"Am. J. Med."},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Antigua, K.J.C., Choi, W.-S., Baek, Y.H., and Song, M.-S. (2019). The Emergence and Decennary Distribution of Clade 2.3.4.4 HPAI H5Nx. Microorganisms, 7.","DOI":"10.3390\/microorganisms7060156"},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"WHO\/OIE\/FAO\/H5N1 Evolution Working Group (2008). Toward a Unified Nomenclature System for Highly Pathogenic Avian Influenza Virus (H5N1). Emerg. Infect. Dis., 14, e1.","DOI":"10.3201\/eid1407.071681"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"263","DOI":"10.1111\/tbed.13347","article-title":"Amino acid substitutions in antigenic region B of hemagglutinin play a critical role in the antigenic drift of subclade 2.3.4.4 highly pathogenic H5NX influenza viruses","volume":"67","author":"Li","year":"2019","journal-title":"Transbound. Emerg. Dis."},{"key":"ref_30","unstructured":"(2024, October 30). Evolution of a Pandemic A(H1N1) 2009. Available online: https:\/\/iris.who.int\/handle\/10665\/78414."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"1729","DOI":"10.1093\/infdis\/jir172","article-title":"Dose-Sparing H5N1 A\/Indonesia\/05\/2005 Pre-pandemic Influenza Vaccine in Adults and Elderly Adults: A Phase III, Placebo-Controlled, Randomized Study","volume":"203","author":"Langley","year":"2011","journal-title":"J. Infect. Dis."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"2378","DOI":"10.1016\/j.vaccine.2008.02.068","article-title":"Safety and reactogenicity profile of an adjuvanted H5N1 pandemic candidate vaccine in adults within a phase III safety trial","volume":"26","author":"Bayas","year":"2008","journal-title":"Vaccine"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"78","DOI":"10.1016\/j.virusres.2013.05.006","article-title":"H5N1 vaccines in humans","volume":"178","author":"Baz","year":"2013","journal-title":"Virus Res."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"6642","DOI":"10.1016\/j.vaccine.2009.03.015","article-title":"Safety and immunogenicity of inactivated, Vero cell culture-derived whole virus influenza A\/H5N1 vaccine given alone or with aluminum hydroxide adjuvant in healthy adults","volume":"27","author":"Keitel","year":"2009","journal-title":"Vaccine"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"467","DOI":"10.1016\/S0140-6736(98)01182-9","article-title":"Clinical features and rapid viral diagnosis of human disease associated with avian influenza A H5N1 virus","volume":"351","author":"Yuen","year":"1998","journal-title":"Lancet"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"244","DOI":"10.1007\/s13337-020-00593-z","article-title":"Human infection with Avian influenza A virus in Nepal: Requisite for timely management and preparedness","volume":"31","author":"Acharya","year":"2020","journal-title":"VirusDisease"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"1585","DOI":"10.1084\/jem.20032083","article-title":"Distinct Time Effects of Vaccination on Long-Term Proliferative and IFN-\u03b3\u2013producing T Cell Memory to Smallpox in Humans","volume":"199","author":"Combadiere","year":"2004","journal-title":"J. Exp. Med."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"3249","DOI":"10.1099\/vir.0.83156-0","article-title":"Genomic sequence of chorioallantois vaccinia virus Ankara, the ancestor of modified vaccinia virus Ankara","volume":"88","author":"Schmidt","year":"2007","journal-title":"J. Gen. Virol."},{"key":"ref_39","doi-asserted-by":"crossref","unstructured":"Kreijtz, J.H.C.M., Suezer, Y., de Mutsert, G., van Amerongen, G., Schwantes, A., Brand, J.M.A.v.D., Fouchier, R.A.M., L\u00f6wer, J., Osterhaus, A.D.M.E., and Sutter, G. (2009). MVA-Based H5N1 Vaccine Affords Cross-Clade Protection in Mice against Influenza A\/H5N1 Viruses at Low Doses and after Single Immunization. PLoS ONE, 4.","DOI":"10.1371\/journal.pone.0007790"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"1196","DOI":"10.1016\/S1473-3099(14)70963-6","article-title":"Safety and immunogenicity of a modified-vaccinia-virus-Ankara-based influenza A H5N1 vaccine: A randomised, double-blind phase 1\/2a clinical trial","volume":"14","author":"Kreijtz","year":"2014","journal-title":"Lancet Infect. Dis."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"614","DOI":"10.1093\/infdis\/jiy214","article-title":"Induction of Cross-Clade Antibody and T-Cell Responses by a Modified Vaccinia Virus Ankara\u2013Based Influenza A(H5N1) Vaccine in a Randomized Phase 1\/2a Clinical Trial","volume":"218","author":"Altenburg","year":"2018","journal-title":"J. Infect. Dis."},{"key":"ref_42","unstructured":"Fenner, F., Henderso, D.A., Atira, I., Jezek, Z., and Ladnyi, I.D. (1988). Smallpox and It Eradication, World Health Organization."}],"container-title":["Vaccines"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-393X\/13\/1\/74\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,8]],"date-time":"2025-10-08T10:29:01Z","timestamp":1759919341000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-393X\/13\/1\/74"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,1,15]]},"references-count":42,"journal-issue":{"issue":"1","published-online":{"date-parts":[[2025,1]]}},"alternative-id":["vaccines13010074"],"URL":"https:\/\/doi.org\/10.3390\/vaccines13010074","relation":{},"ISSN":["2076-393X"],"issn-type":[{"type":"electronic","value":"2076-393X"}],"subject":[],"published":{"date-parts":[[2025,1,15]]}}}