{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,22]],"date-time":"2026-10-22T23:38:51Z","timestamp":1792712331193,"version":"4.5.0"},"reference-count":63,"publisher":"MDPI AG","issue":"9","license":[{"start":{"date-parts":[[2025,9,22]],"date-time":"2025-09-22T00:00:00Z","timestamp":1758499200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100014440","name":"Ministerio de Ciencia, Innovaci\u00f3n y Universidades (Spain)","doi-asserted-by":"publisher","award":["PID2021-124872OB-100"],"award-info":[{"award-number":["PID2021-124872OB-100"]}],"id":[{"id":"10.13039\/100014440","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100014440","name":"Ministerio de Ciencia, Innovaci\u00f3n y Universidades (Spain)","doi-asserted-by":"publisher","award":["101136346 EUPAHW"],"award-info":[{"award-number":["101136346 EUPAHW"]}],"id":[{"id":"10.13039\/100014440","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000780","name":"European Union\u2019s Horizon Europe Project","doi-asserted-by":"publisher","award":["PID2021-124872OB-100"],"award-info":[{"award-number":["PID2021-124872OB-100"]}],"id":[{"id":"10.13039\/501100000780","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000780","name":"European Union\u2019s Horizon Europe Project","doi-asserted-by":"publisher","award":["101136346 EUPAHW"],"award-info":[{"award-number":["101136346 EUPAHW"]}],"id":[{"id":"10.13039\/501100000780","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Vaccines"],"abstract":"<jats:p>Background\/Objectives: Bluetongue virus (BTV) is an emerging arbovirus causing significant economic losses in the ruminant industry. Current vaccines offer limited cross-protection against heterologous serotypes and do not enable differentiation between infected and vaccinated animals (DIVA). Subunit-based vaccines provide a potential DIVA-compatible solution. This study aimed to develop a vaccination protocol expressing BTV structural proteins VP7 or VP2 using antibiotic-resistance-free DNA plasmids and replication-defective adenovirus vectors. Methods: We evaluated homologous DNA prime\u2013boost and heterologous DNA prime\u2013adenovirus boost strategies in a murine model, assessing adaptive immune responses and protection against virulent BTV challenge. Results: The heterologous DNA\u2013adenovirus prime\u2013boost strategy expressing both antigens conferred full protection, preventing viremia, while homologous DNA-DNA prime\u2013boost provided only partial protection. Both VP7 and VP2 elicited cellular and humoral immune responses, but the heterologous strategy significantly enhanced anti-BTV IgG, neutralizing antibody titers, and T cell activation. CD8+ T cell responses showed the strongest correlation with viral load reduction, suggesting that cellular immunity to conserved VP7 could serve as a platform for cross-protection against multiple BTV serotypes. Conclusions: These findings highlight the potential of heterologous DNA\u2013adenovirus vaccination as an effective DIVA-compatible strategy for BTV control. By inducing strong and protective immune responses, this approach could improve disease surveillance and management, ultimately reducing the impact of BTV on livestock industries.<\/jats:p>","DOI":"10.3390\/vaccines13090991","type":"journal-article","created":{"date-parts":[[2025,9,22]],"date-time":"2025-09-22T17:08:17Z","timestamp":1758560897000},"page":"991","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Heterologous DNA\u2013Adenovirus Prime\u2013Boost Strategy Expressing Bluetongue Virus VP2 and VP7 Proteins Protects Against Virulent Challenge"],"prefix":"10.3390","volume":"13","author":[{"ORCID":"https:\/\/orcid.org\/0009-0009-8655-6205","authenticated-orcid":false,"given":"Pablo","family":"Nogales-Altozano","sequence":"first","affiliation":[{"name":"Centro de Investigaci\u00f3n en Sanidad Animal, Instituto de Investigaci\u00f3n y Tecnolog\u00eda Agraria y Alimentaria, Consejo Superior de Investigaciones Cient\u00edficas (CISA-INIA-CSIC), Valdeolmos, 28130 Madrid, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Laro","family":"G\u00f3mez-Marcos","sequence":"additional","affiliation":[{"name":"Centro de Investigaci\u00f3n en Sanidad Animal, Instituto de Investigaci\u00f3n y Tecnolog\u00eda Agraria y Alimentaria, Consejo Superior de Investigaciones Cient\u00edficas (CISA-INIA-CSIC), Valdeolmos, 28130 Madrid, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ana Bel\u00e9n","family":"Carl\u00f3n","sequence":"additional","affiliation":[{"name":"Centro de Investigaci\u00f3n en Sanidad Animal, Instituto de Investigaci\u00f3n y Tecnolog\u00eda Agraria y Alimentaria, Consejo Superior de Investigaciones Cient\u00edficas (CISA-INIA-CSIC), Valdeolmos, 28130 Madrid, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andr\u00e9s","family":"Louloudes-L\u00e1zaro","sequence":"additional","affiliation":[{"name":"Centro de Investigaci\u00f3n en Sanidad Animal, Instituto de Investigaci\u00f3n y Tecnolog\u00eda Agraria y Alimentaria, Consejo Superior de Investigaciones Cient\u00edficas (CISA-INIA-CSIC), Valdeolmos, 28130 Madrid, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alicia","family":"Rivera-Rodr\u00edguez","sequence":"additional","affiliation":[{"name":"Centro de Investigaci\u00f3n en Sanidad Animal, Instituto de Investigaci\u00f3n y Tecnolog\u00eda Agraria y Alimentaria, Consejo Superior de Investigaciones Cient\u00edficas (CISA-INIA-CSIC), Valdeolmos, 28130 Madrid, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7293-7239","authenticated-orcid":false,"given":"Jaime","family":"Larraga","sequence":"additional","affiliation":[{"name":"Centro de Investigaciones Biol\u00f3gicas Margarita Salas (CIBMS-CSIC), 28040 Madrid, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0729-8941","authenticated-orcid":false,"given":"Pedro J.","family":"Alcolea","sequence":"additional","affiliation":[{"name":"Centro de Investigaciones Biol\u00f3gicas Margarita Salas (CIBMS-CSIC), 28040 Madrid, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ana","family":"Alonso","sequence":"additional","affiliation":[{"name":"Centro de Investigaciones Biol\u00f3gicas Margarita Salas (CIBMS-CSIC), 28040 Madrid, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1260-7400","authenticated-orcid":false,"given":"Vicente","family":"Larraga","sequence":"additional","affiliation":[{"name":"Centro de Investigaciones Biol\u00f3gicas Margarita Salas (CIBMS-CSIC), 28040 Madrid, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1486-1656","authenticated-orcid":false,"given":"Ver\u00f3nica","family":"Mart\u00edn","sequence":"additional","affiliation":[{"name":"Centro de Investigaci\u00f3n en Sanidad Animal, Instituto de Investigaci\u00f3n y Tecnolog\u00eda Agraria y Alimentaria, Consejo Superior de Investigaciones Cient\u00edficas (CISA-INIA-CSIC), Valdeolmos, 28130 Madrid, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4055-3967","authenticated-orcid":false,"given":"Jos\u00e9 M.","family":"Rojas","sequence":"additional","affiliation":[{"name":"Centro de Investigaci\u00f3n en Sanidad Animal, Instituto de Investigaci\u00f3n y Tecnolog\u00eda Agraria y Alimentaria, Consejo Superior de Investigaciones Cient\u00edficas (CISA-INIA-CSIC), Valdeolmos, 28130 Madrid, 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