{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,22]],"date-time":"2026-08-22T05:23:02Z","timestamp":1787376182118,"version":"build-2736575974"},"reference-count":79,"publisher":"MDPI AG","issue":"8","license":[{"start":{"date-parts":[[2026,8,20]],"date-time":"2026-08-20T00:00:00Z","timestamp":1787184000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100007465","name":"UiT The Arctic University of Norway","doi-asserted-by":"crossref","id":[{"id":"10.13039\/100007465","id-type":"DOI","asserted-by":"crossref"}]},{"id":[{"id":"https:\/\/ror.org\/00wge5k78","id-type":"ROR","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100008119","name":"Norwegian University of Life Sciences","doi-asserted-by":"crossref","id":[{"id":"10.13039\/501100008119","id-type":"DOI","asserted-by":"crossref"}]},{"id":[{"id":"https:\/\/ror.org\/04a1mvv97","id-type":"ROR","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100005416","name":"the Research Council of Norway","doi-asserted-by":"publisher","award":["301401"],"award-info":[{"award-number":["301401"]}],"id":[{"id":"10.13039\/501100005416","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100005416","name":"the Research Council of Norway","doi-asserted-by":"publisher","award":["32519"],"award-info":[{"award-number":["32519"]}],"id":[{"id":"10.13039\/501100005416","id-type":"DOI","asserted-by":"publisher"}]},{"award":["301401"],"award-info":[{"award-number":["301401"]}],"id":[{"id":"https:\/\/ror.org\/00epmv149","id-type":"ROR","asserted-by":"publisher"}]},{"award":["32519"],"award-info":[{"award-number":["32519"]}],"id":[{"id":"https:\/\/ror.org\/00epmv149","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Pharmaceutics"],"abstract":"<jats:p>Background: Pancreas disease, caused by salmonid alphavirus subtype 3 (SAV3), remains a significant threat to Atlantic salmon (Salmo salar L.) aquaculture in Norway despite the widespread use of inactivated vaccines. Molecular adjuvants represent a promising strategy to improve vaccine efficacy by enhancing immune responses. Methods: In this study, we evaluated the immunological and protective effects of two transcription factors, gata3 and eomes encoded by plasmid DNA adjuvants in combination with a low-dose inactivated SAV3 vaccine. Atlantic salmon were immunized with inactivated virus alone or in combination with either gata3- or eomes-encoding plasmids, followed by SAV3 challenge. Vaccine performance was assessed by growth, antibody response (ELISA and virus neutralization assay), and histopathological scoring of heart and pancreas. Results: The effects of gata3 and eomes were context-dependent: eomes reduced pancreas lesions at 4 weeks post-challenge, while both plasmid-adjuvant groups showed improved growth at 10 weeks post-challenge compared to the plasmid control. Although antibody responses were increased in all vaccinated groups at 6 weeks post-immunization, the plasmid control performed comparably to the GATA3 or EOMES groups, and no significant differences were detected in the virus neutralization test. Interestingly, the empty plasmids given to control fish exhibited immunostimulatory activity, warranting further investigation. In vitro and in vivo fluorescence microscopy confirmed expression of the transcription factors in transfected cells and at the injection site. Conclusions: Our results suggest that transcription factor-encoding plasmids can provide selective adjuvant effects, while also highlighting the potential that plasmid DNA can have as adjuvants in fish vaccine development.<\/jats:p>","DOI":"10.3390\/pharmaceutics18081033","type":"journal-article","created":{"date-parts":[[2026,8,20]],"date-time":"2026-08-20T12:57:53Z","timestamp":1787230673000},"page":"1033","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Plasmid-DNA Encoded gata3 and eomes Increase Antibody Response and Performance of an Inactivated Salmonid Alphavirus Subtype 3 Vaccine"],"prefix":"10.3390","volume":"18","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-4674-7279","authenticated-orcid":false,"given":"Filipe","family":"Figueiredo","sequence":"first","affiliation":[{"name":"Norwegian College of Fishery Science, Faculty of Biosciences, Fisheries and Economics, UiT-The Arctic University of Norway, N-9019 Troms\u00f8, Norway"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kristian Gillebo","family":"S\u00f8rmo","sequence":"additional","affiliation":[{"name":"Norwegian College of Fishery Science, Faculty of Biosciences, Fisheries and Economics, UiT-The Arctic University of Norway, N-9019 Troms\u00f8, Norway"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Aleksander","family":"Pettersen","sequence":"additional","affiliation":[{"name":"Norwegian College of Fishery Science, Faculty of Biosciences, Fisheries and Economics, UiT-The Arctic University of Norway, N-9019 Troms\u00f8, Norway"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8106-836X","authenticated-orcid":false,"given":"Heng","family":"Chi","sequence":"additional","affiliation":[{"name":"Norwegian College of Fishery Science, Faculty of Biosciences, Fisheries and Economics, UiT-The Arctic University of Norway, N-9019 Troms\u00f8, Norway"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1787-5498","authenticated-orcid":false,"given":"Jaya Kumari","family":"Swain","sequence":"additional","affiliation":[{"name":"Norwegian College of Fishery Science, Faculty of Biosciences, Fisheries and Economics, UiT-The Arctic University of Norway, N-9019 Troms\u00f8, Norway"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0001-3764-6011","authenticated-orcid":false,"given":"Toni","family":"Erkinharju","sequence":"additional","affiliation":[{"name":"Norwegian Veterinary Institute, N-1431 \u00c5s, Norway"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Trilochan","family":"Swain","sequence":"additional","affiliation":[{"name":"Norwegian College of Fishery Science, Faculty of Biosciences, Fisheries and Economics, UiT-The Arctic University of Norway, N-9019 Troms\u00f8, Norway"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8389-2216","authenticated-orcid":false,"given":"Shripathi","family":"Bhat","sequence":"additional","affiliation":[{"name":"Norwegian College of Fishery Science, Faculty of Biosciences, Fisheries and Economics, UiT-The Arctic University of Norway, N-9019 Troms\u00f8, Norway"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0681-1854","authenticated-orcid":false,"given":"Kim","family":"Pr\u00e6bel","sequence":"additional","affiliation":[{"name":"Norwegian College of Fishery Science, Faculty of Biosciences, Fisheries and Economics, UiT-The Arctic University of Norway, N-9019 Troms\u00f8, Norway"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9043-7438","authenticated-orcid":false,"given":"Stefano","family":"Peruzzi","sequence":"additional","affiliation":[{"name":"Department of Arctic and Marine Biology, Faculty of Biosciences, Fisheries and Economics, UiT\u2013The Arctic University of Norway, N-9019 Troms\u00f8, Norway"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0782-7858","authenticated-orcid":false,"given":"Jacques","family":"Godfroid","sequence":"additional","affiliation":[{"name":"Department of Arctic and Marine Biology, Faculty of Biosciences, Fisheries and Economics, UiT\u2013The Arctic University of Norway, N-9019 Troms\u00f8, Norway"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3538-3657","authenticated-orcid":false,"given":"\u00d8ystein","family":"Evensen","sequence":"additional","affiliation":[{"name":"Department of Paraclinical Sciences, Faculty of Veterinary Medicine, Norwegian University of Life Sciences, N-1431 \u00c5s, Norway"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1021-6396","authenticated-orcid":false,"given":"Hetron Mweemba","family":"Munang\u2019andu","sequence":"additional","affiliation":[{"name":"Faculty of Biosciences and Aquaculture, Nord University, N-8026 Bod\u00f8, Norway"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6181-9859","authenticated-orcid":false,"given":"Roy Ambli","family":"Dalmo","sequence":"additional","affiliation":[{"name":"Norwegian College of Fishery Science, Faculty of Biosciences, Fisheries and Economics, UiT-The Arctic University of Norway, N-9019 Troms\u00f8, Norway"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,8,20]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"Buchmann, K., and Secombes, C.J. (2022). Current Use and Need for New Fish Vaccines. Principles of Fish Immunology: From Cells and Molecules to Host Protection, Springer International Publishing.","DOI":"10.1007\/978-3-030-85420-1"},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Miccoli, A., Manni, M., Picchietti, S., and Scapigliati, G. (2021). State-of-the-Art Vaccine Research for Aquaculture Use: The Case of Three Economically Relevant Fish Species. Vaccines, 9.","DOI":"10.3390\/vaccines9020140"},{"key":"ref_3","doi-asserted-by":"crossref","unstructured":"Du, Y., Hu, X., Miao, L., and Chen, J. (2022). Current status and development prospects of aquatic vaccines. Front. Immunol., 13.","DOI":"10.3389\/fimmu.2022.1040336"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1971","DOI":"10.1007\/s10499-022-00884-w","article-title":"A review on the recent advances and application of vaccines against fish pathogens in aquaculture","volume":"30","author":"Mondal","year":"2022","journal-title":"Aquac. Int."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"116","DOI":"10.1016\/j.fsi.2020.12.002","article-title":"Effect of a novel DNA vaccine against pancreas disease caused by salmonid alphavirus subtype 3 in Atlantic salmon (Salmo salar)","volume":"108","author":"Thorarinsson","year":"2021","journal-title":"Fish Shellfish Immunol."},{"key":"ref_6","first-page":"1","article-title":"Salmonid Alphavirus (SAV)","volume":"62","author":"Schulz","year":"2018","journal-title":"J. Vet. Res."},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Macqueen, D.J., Eve, O., Gundappa, M.K., Daniels, R.R., Gallagher, M.D., Alexandersen, S., and Karlsen, M. (2021). Genomic Epidemiology of Salmonid Alphavirus in Norwegian Aquaculture Reveals Recent Subtype-2 Transmission Dynamics and Novel Subtype-3 Lineages. Viruses, 13.","DOI":"10.3390\/v13122549"},{"key":"ref_8","unstructured":"Sommerset, I., Wiik-Nielsen, J., Moldal, T., Oliveira, V., Svendsen, J., Haukaas, A., and Brun, E. (2024). Norwegian Fish Health Report 2023, Norwegian Veterinary Institute."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"3167","DOI":"10.1016\/j.vaccine.2019.04.055","article-title":"Vaccine adjuvants: Understanding the structure and mechanism of adjuvanticity","volume":"37","author":"Shi","year":"2019","journal-title":"Vaccine"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"1740","DOI":"10.1016\/j.fsi.2013.02.029","article-title":"Adjuvants and immunostimulants in fish vaccines: Current knowledge and future perspectives","volume":"35","author":"Tafalla","year":"2013","journal-title":"Fish Shellfish Immunol."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"225","DOI":"10.1016\/0959-8030(93)90036-B","article-title":"Viral vaccines for aquaculture","volume":"3","author":"Leong","year":"1993","journal-title":"Annu. Rev. Fish Dis."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"108784","DOI":"10.1016\/j.fsi.2023.108784","article-title":"New vaccination strategies are required for effective control of winter ulcer disease caused by emerging variant strains of Moritella viscosa in Atlantic salmon","volume":"137","author":"Furevik","year":"2023","journal-title":"Fish Shellfish Immunol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"S34","DOI":"10.1016\/S0264-410X(02)00169-X","article-title":"Mechanisms of stimulation of the immune response by aluminum adjuvants","volume":"20","author":"HogenEsch","year":"2002","journal-title":"Vaccine"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"735259","DOI":"10.1016\/j.aquaculture.2020.735259","article-title":"The efficacy and side-effects of oil-based adjuvants emulsified Vibrio anguillarum bivalent inactivated vaccine in turbot (Scophthalmus maximus) under production mode","volume":"524","author":"Li","year":"2020","journal-title":"Aquaculture"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"5930","DOI":"10.1038\/s41598-017-06324-7","article-title":"Alternatives to mineral oil adjuvants in vaccines against Aeromonas salmonicida subsp. salmonicida in rainbow trout offer reductions in adverse effects","volume":"7","author":"Villumsen","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"148","DOI":"10.3390\/vaccines3010148","article-title":"Choice and Design of Adjuvants for Parenteral and Mucosal Vaccines","volume":"3","author":"Savelkoul","year":"2015","journal-title":"Vaccines"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"456","DOI":"10.1016\/S0952-7915(00)00120-5","article-title":"Immunological concepts of vaccine adjuvant activity: Commentary","volume":"12","author":"Schijns","year":"2000","journal-title":"Curr. Opin. Immunol."},{"key":"ref_18","doi-asserted-by":"crossref","unstructured":"Tammas, I., Bitchava, K., and Gelasakis, A.I. (2024). Advances in Vaccine Adjuvants for Teleost Fish: Implications for Aquatic Welfare and the Potential of Nanoparticle-Based Formulations. Vaccines, 12.","DOI":"10.20944\/preprints202411.0110.v1"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"283","DOI":"10.1038\/s41392-023-01557-7","article-title":"Vaccine adjuvants: Mechanisms and platforms","volume":"8","author":"Zhao","year":"2023","journal-title":"Signal Transduct. Target. Ther."},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Awate, S., Babiuk, L.A., and Mutwiri, G. (2013). Mechanisms of Action of Adjuvants. Front. Immunol., 4.","DOI":"10.3389\/fimmu.2013.00114"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"17","DOI":"10.21775\/cimb.022.017","article-title":"Molecular Adjuvants for DNA Vaccines","volume":"22","author":"Li","year":"2017","journal-title":"Curr. Issues Mol. Biol."},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Hojecki, C.E., Tursi, N.J., Livingston, C., Weiner, D.B., and Gary, E.N. (2025). Advances in molecular adjuvants for nucleic acid vaccines. Front. Immunol., 16.","DOI":"10.3389\/fimmu.2025.1646800"},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"B\u00f8gwald, J., and Dalmo, R.A. (2019). Review on Immersion Vaccines for Fish: An Update 2019. Microorganisms, 7.","DOI":"10.3390\/microorganisms7120627"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1111\/jfd.12727","article-title":"DNA vaccines for fish: Review and perspectives on correlates of protection","volume":"41","author":"Dalmo","year":"2018","journal-title":"J. Fish Dis."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"996","DOI":"10.1111\/raq.12509","article-title":"An overview of cytokine used as adjuvants in fish: Current state and future trends","volume":"13","author":"Guo","year":"2021","journal-title":"Rev. Aquac."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"328","DOI":"10.1016\/j.fsi.2016.04.027","article-title":"IFN-adjuvanted DNA vaccine against infectious salmon anemia virus: Antibody kinetics and longevity of IFN expression","volume":"54","author":"Robertsen","year":"2016","journal-title":"Fish Shellfish Immunol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"934","DOI":"10.1016\/S1359-6446(03)02864-2","article-title":"Vaccine adjuvants: Role and mechanisms of action in vaccine immunogenicity","volume":"8","author":"Marciani","year":"2003","journal-title":"Drug Discov. Today"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"337","DOI":"10.1016\/j.immuni.2011.08.012","article-title":"An essential role of the transcription factor GATA-3 for the function of regulatory T cells","volume":"35","author":"Wang","year":"2011","journal-title":"Immunity"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"25","DOI":"10.1385\/IR:28:1:25","article-title":"The function role of GATA-3 in Th1 and Th2 differentiation","volume":"28","author":"Zhou","year":"2003","journal-title":"Immunol. Res."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"1041","DOI":"10.1126\/science.1090148","article-title":"Control of effector CD8+ T cell function by the transcription factor Eomesodermin","volume":"302","author":"Pearce","year":"2003","journal-title":"Science"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"1305","DOI":"10.1016\/S1357-2725(97)00085-X","article-title":"Transcription factors: An overview","volume":"29","author":"Latchman","year":"1997","journal-title":"Int. J. Biochem. Cell Biol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"2449","DOI":"10.1016\/j.molcel.2023.06.012","article-title":"Transcription factors interact with RNA to regulate genes","volume":"83","author":"Oksuz","year":"2023","journal-title":"Mol. Cell"},{"key":"ref_33","doi-asserted-by":"crossref","unstructured":"Slettjord, T.H., Sekkenes, H.J., Chi, H., B\u00f8gwald, J., Swain, T., Dalmo, R.A., and Kumari Swain, J. (2020). Overexpression of T-bet, GATA-3 and TGF-\u00df Induces IFN-\u03b3, IL-4\/13A, and IL-17A Expression in Atlantic Salmon. Biology, 9.","DOI":"10.3390\/biology9040082"},{"key":"ref_34","doi-asserted-by":"crossref","unstructured":"Kumari, J., B\u00f8gwald, J., and Dalmo, R.A. (2013). Eomesodermin of Atlantic Salmon: An Important Regulator of Cytolytic Gene and Interferon Gamma Expression in Spleen Lymphocytes. PLoS ONE, 8.","DOI":"10.1371\/journal.pone.0055893"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"3099","DOI":"10.1016\/j.molimm.2009.06.008","article-title":"Transcription factor GATA-3 in Atlantic salmon (Salmo salar): Molecular characterization, promoter activity and expression analysis","volume":"46","author":"Kumari","year":"2009","journal-title":"Mol. Immunol."},{"key":"ref_36","doi-asserted-by":"crossref","unstructured":"Kumari, J., Zhang, Z., Swain, T., Chi, H., Niu, C., B\u00f8gwald, J., and Dalmo, R.A. (2015). Transcription Factor T-Bet in Atlantic Salmon: Characterization and Gene Expression in Mucosal Tissues during Aeromonas Salmonicida Infection. Front. Immunol., 6.","DOI":"10.3389\/fimmu.2015.00345"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"671","DOI":"10.1007\/BF02618871","article-title":"Fish cell lines: Establishment and characterization of nine cell lines from salmonids","volume":"20","author":"Lannan","year":"1984","journal-title":"In Vitro"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"266","DOI":"10.1016\/j.fsi.2017.07.025","article-title":"CHSE-214: A model for studying extracellular dsRNA sensing in vitro","volume":"68","author":"Monjo","year":"2017","journal-title":"Fish Shellfish Immunol."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"66","DOI":"10.1186\/s12985-016-0520-8","article-title":"Atlantic salmon (Salmo salar L.) post-smolts challenged two or nine weeks after seawater-transfer show differences in their susceptibility to salmonid alphavirus subtype 3 (SAV3)","volume":"13","author":"Jarungsriapisit","year":"2016","journal-title":"Virol. J."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"125","DOI":"10.3354\/dao072125","article-title":"Virological, serological and histopathological evaluation of fish strain susceptibility to experimental infection with salmonid alphavirus","volume":"72","author":"McLoughlin","year":"2006","journal-title":"Dis. Aquat. Org."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"173","DOI":"10.1128\/JVI.74.1.173-183.2000","article-title":"Rainbow Trout Sleeping Disease Virus Is an Atypical Alphavirus","volume":"74","author":"Villoing","year":"2000","journal-title":"J. Virol."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"184","DOI":"10.1016\/j.jviromet.2005.08.012","article-title":"Sensitive and specific detection of Salmonid alphavirus using real-time PCR (TaqMan\u00ae)","volume":"131","author":"Hodneland","year":"2006","journal-title":"J. Virol. Methods"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"402","DOI":"10.1006\/meth.2001.1262","article-title":"Analysis of Relative Gene Expression Data Using Real-Time Quantitative PCR and the 2\u2212\u0394\u0394CT Method","volume":"25","author":"Livak","year":"2001","journal-title":"Methods"},{"key":"ref_44","unstructured":"R Development Core Team (R: A Language and Environment for Statistical Computing, 2024). R: A Language and Environment for Statistical Computing, Version 4.4.0."},{"key":"ref_45","unstructured":"Kassambara, A. (2026, June 01). ggpubr: \u2018ggplot2\u2019 Based Publication Ready Plots, Available online: https:\/\/CRAN.R-project.org\/package=ggpubr."},{"key":"ref_46","unstructured":"Wickham, H., and Bryan, J. (2026, June 01). readxl: Read Excel Files, Available online: https:\/\/CRAN.R-project.org\/package=readxl."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"1686","DOI":"10.21105\/joss.01686","article-title":"Welcome to the Tidyverse","volume":"4","author":"Wickham","year":"2019","journal-title":"J. Open Source Softw."},{"key":"ref_48","unstructured":"Kassambara, A. (2026, June 01). rstatix: Pipe-Friendly Framework for Basic Statistical Tests, Available online: https:\/\/CRAN.R-project.org\/package=rstatix."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"2815","DOI":"10.21105\/joss.02815","article-title":"effectsize: Estimation of Effect Size Indices and Standardized Parameters","volume":"5","author":"Makowski","year":"2020","journal-title":"J. Open Source Softw."},{"key":"ref_50","unstructured":"Wickam, H., Pedersen, T.L., and Seidel, D. (2026, June 01). scales: Scale Functions for Visualization, Available online: https:\/\/CRAN.R-project.org\/package=scales."},{"key":"ref_51","unstructured":"Mills, B.R. (2026, June 01). MoMAColors: Color Palettes Inspired by Artwork at the Museum of Modern Art in New York City, Available online: https:\/\/github.com\/BlakeRMills\/MoMAColors."},{"key":"ref_52","unstructured":"Pedersen, T.L. (2026, June 01). patchwork: The Composer of Plots, Available online: https:\/\/CRAN.R-project.org\/package=patchwork."},{"key":"ref_53","doi-asserted-by":"crossref","unstructured":"Cohen, J. (1977). CHAPTER 2\u2014The t Test for Means. Statistical Power Analysis for the Behavioral Sciences, Academic Press.","DOI":"10.1016\/B978-0-12-179060-8.50007-4"},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"28","DOI":"10.1016\/j.fsi.2019.07.021","article-title":"T-box transcription factor eomesodermin\/Tbr2 in Atlantic cod (Gadus morhua L.): Molecular characterization, promoter structure and function analysis","volume":"93","author":"Chi","year":"2019","journal-title":"Fish Shellfish Immunol."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"215","DOI":"10.1016\/j.fsi.2013.11.004","article-title":"Transcription analysis of two Eomesodermin genes in lymphocyte subsets of two teleost species","volume":"36","author":"Takizawa","year":"2014","journal-title":"Fish Shellfish Immunol."},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"13","DOI":"10.2478\/acb-2023-0002","article-title":"Components of the fascia\u2013cells and extracellular matrix","volume":"11","author":"Kosiorowski","year":"2023","journal-title":"Med. J. Cell Biol."},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"890","DOI":"10.1016\/j.fsi.2013.06.030","article-title":"Transgene and immune gene expression following intramuscular injection of Atlantic salmon (Salmo salar L.) with DNA-releasing PLGA nano- and microparticles","volume":"35","author":"Fredriksen","year":"2013","journal-title":"Fish Shellfish Immunol."},{"key":"ref_58","doi-asserted-by":"crossref","first-page":"90","DOI":"10.1016\/j.fsi.2007.09.006","article-title":"Specific uptake of plasmid DNA without reporter gene expression in Atlantic salmon (Salmo salar L.) kidney after intramuscular administration","volume":"24","author":"Tonheim","year":"2008","journal-title":"Fish Shellfish Immunol."},{"key":"ref_59","doi-asserted-by":"crossref","unstructured":"Kumar, A., Middha, S.K., Menon, S.V., Paital, B., Gokarn, S., Nelli, M., Rajanikanth, R.B., Chandra, H.M., Mugunthan, S.P., and Kantwa, S.M. (2024). Current Challenges of Vaccination in Fish Health Management. Animals, 14.","DOI":"10.3390\/ani14182692"},{"key":"ref_60","doi-asserted-by":"crossref","first-page":"313","DOI":"10.1586\/14760584.2016.1124762","article-title":"Molecular mechanisms for enhanced DNA vaccine immunogenicity","volume":"15","author":"Li","year":"2016","journal-title":"Expert Rev. Vaccines"},{"key":"ref_61","doi-asserted-by":"crossref","first-page":"255","DOI":"10.1038\/s41590-018-0052-z","article-title":"Toll-like receptor 9 antagonizes antibody affinity maturation","volume":"19","author":"Akkaya","year":"2018","journal-title":"Nat. Immunol."},{"key":"ref_62","doi-asserted-by":"crossref","first-page":"3565","DOI":"10.1038\/s41598-018-21895-9","article-title":"Profiling Atlantic salmon B cell populations: CpG-mediated TLR-ligation enhances IgM secretion and modulates immune gene expression","volume":"8","author":"Jenberie","year":"2018","journal-title":"Sci. Rep."},{"key":"ref_63","doi-asserted-by":"crossref","unstructured":"Zhu, J. (2018). Seventeen-Year Journey Working with a Master. Front. Immunol., 9.","DOI":"10.3389\/fimmu.2018.00960"},{"key":"ref_64","doi-asserted-by":"crossref","first-page":"318","DOI":"10.1016\/j.cyto.2015.02.007","article-title":"B cells responses and cytokine production are regulated by their immune microenvironment","volume":"74","author":"Vazquez","year":"2015","journal-title":"Cytokine"},{"key":"ref_65","doi-asserted-by":"crossref","first-page":"125","DOI":"10.1038\/nri2476","article-title":"GATA3 and the T-cell lineage: Essential functions before and after T-helper-2-cell differentiation","volume":"9","author":"Ho","year":"2009","journal-title":"Nat. Rev. Immunol."},{"key":"ref_66","doi-asserted-by":"crossref","first-page":"27","DOI":"10.1016\/S1074-7613(00)80156-9","article-title":"Stat6-Independent GATA-3 Autoactivation Directs IL-4-Independent Th2 Development and Commitment","volume":"12","author":"Ouyang","year":"2000","journal-title":"Immunity"},{"key":"ref_67","doi-asserted-by":"crossref","first-page":"8700","DOI":"10.4049\/jimmunol.181.12.8700","article-title":"T-bet and eomesodermin play critical roles in directing T cell differentiation to Th1 versus Th17","volume":"181","author":"Yang","year":"2008","journal-title":"J. Immunol."},{"key":"ref_68","doi-asserted-by":"crossref","first-page":"3","DOI":"10.1038\/sj.cr.7310002","article-title":"GATA-3 promotes Th2 responses through three different mechanisms: Induction of Th2 cytokine production, selective growth of Th2 cells and inhibition of Th1 cell-specific factors","volume":"16","author":"Zhu","year":"2006","journal-title":"Cell Res."},{"key":"ref_69","doi-asserted-by":"crossref","first-page":"778","DOI":"10.1038\/ni.2362","article-title":"The transcription factor Sox4 is a downstream target of signaling by the cytokine TGF-\u03b2 and suppresses TH2 differentiation","volume":"13","author":"Kuwahara","year":"2012","journal-title":"Nat. Immunol."},{"key":"ref_70","doi-asserted-by":"crossref","first-page":"323","DOI":"10.1016\/S1074-7613(00)80185-5","article-title":"ROG, Repressor of GATA, Regulates the Expression of Cytokine Genes","volume":"12","author":"Miaw","year":"2000","journal-title":"Immunity"},{"key":"ref_71","doi-asserted-by":"crossref","first-page":"1461","DOI":"10.1084\/jem.194.10.1461","article-title":"Friend of GATA-1 Represses GATA-3\u2013dependent Activity in CD4+ T Cells","volume":"194","author":"Zhou","year":"2001","journal-title":"J. Exp. Med."},{"key":"ref_72","doi-asserted-by":"crossref","first-page":"157895","DOI":"10.1155\/2014\/157895","article-title":"Neutralization of Virus Infectivity by Antibodies: Old Problems in New Perspectives","volume":"2014","author":"Klasse","year":"2014","journal-title":"Adv. Biol."},{"key":"ref_73","unstructured":"Mahy, B.W.J., and Van Regenmortel, M.H.V. (2008). Immunopathology. Encyclopedia of Virology, Academic Press. [3rd ed.]."},{"key":"ref_74","doi-asserted-by":"crossref","first-page":"1871","DOI":"10.1007\/s00705-007-1006-1","article-title":"Tissue tropism of salmonid alphaviruses (subtypes SAV1 and SAV3) in experimentally challenged Atlantic salmon (Salmo salar L.)","volume":"152","author":"Andersen","year":"2007","journal-title":"Arch. Virol."},{"key":"ref_75","doi-asserted-by":"crossref","first-page":"132","DOI":"10.1016\/j.fsi.2018.03.060","article-title":"Correlates of protective immunity for fish vaccines","volume":"85","author":"Evensen","year":"2019","journal-title":"Fish Shellfish Immunol."},{"key":"ref_76","doi-asserted-by":"crossref","unstructured":"Thorarinsson, R., Ramstad, A., Wolf, J.C., Sindre, H., Skjerve, E., Rimstad, E., Evensen, \u00d8., and Rodriguez, J.F. (2024). Effect of pancreas disease vaccines on infection levels and virus transmission in Atlantic salmon (Salmo salar) challenged with salmonid alphavirus, genotype 2. Front. Immunol., 15.","DOI":"10.3389\/fimmu.2024.1342816"},{"key":"ref_77","doi-asserted-by":"crossref","first-page":"e70003","DOI":"10.1111\/jfd.70003","article-title":"Controlled Field Trial of Pancreas Disease Vaccines in Farmed Atlantic Salmon: Effects on Growth and Mortality During a SAV2 Outbreak","volume":"48","author":"Pettersen","year":"2025","journal-title":"J. Fish Dis."},{"key":"ref_78","doi-asserted-by":"crossref","first-page":"297","DOI":"10.1016\/j.fsi.2014.11.003","article-title":"Antiviral and metabolic gene expression responses to viral infection in Atlantic salmon (Salmo salar)","volume":"42","author":"Heidari","year":"2015","journal-title":"Fish Shellfish Immunol."},{"key":"ref_79","doi-asserted-by":"crossref","first-page":"5688","DOI":"10.1016\/j.vaccine.2012.05.069","article-title":"Efficacy and safety of an inactivated vaccine against Salmonid alphavirus (family Togaviridae)","volume":"30","author":"Karlsen","year":"2012","journal-title":"Vaccine"}],"container-title":["Pharmaceutics"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1999-4923\/18\/8\/1033\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,8,22]],"date-time":"2026-08-22T04:25:25Z","timestamp":1787372725000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1999-4923\/18\/8\/1033"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,8,20]]},"references-count":79,"journal-issue":{"issue":"8","published-online":{"date-parts":[[2026,8]]}},"alternative-id":["pharmaceutics18081033"],"URL":"https:\/\/doi.org\/10.3390\/pharmaceutics18081033","relation":{},"ISSN":["1999-4923"],"issn-type":[{"value":"1999-4923","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,8,20]]}}}