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Equine veterinary journal2025; 58(2); 591-600; doi: 10.1111/evj.70090

Adjuvant combination enhances immune response of equine influenza virus vaccine in horses.

Abstract: Despite routine vaccination, equine influenza outbreaks occur due to disparities of virus strains between vaccine and field, and immunity gaps between vaccinations. Objective: To evaluate whether a combination of adjuvants can enhance immune responses to equine influenza virus (EIV) vaccines. Methods: In vivo experimental study. Methods: Eight mixed-breed horses were immunised intramuscularly with a recombinant canarypox-based EIV vaccine, either alone or in combination with monophosphoryl lipid A (MPL) and polyinosinic-polycytidylic acid (poly I:C). A single vaccine dose was administered, and serum samples were collected before immunisation and at 2, 8, and 24 weeks post-immunisation to assess the humoral immune response. Peripheral blood mononuclear cells (PBMCs) were isolated to evaluate immunomodulatory cytokine expression and characterise recall cellular immune responses. Additionally, bone marrow and lymph node cells were analysed to assess memory cell responses. Results: The combination of MPL and poly I:C significantly enhanced serum IgG antibody concentrations and haemagglutination inhibition (HI) titres against H3N8 A/equine/Kyonggi/SA01/2011 (Florida sub-lineage clade 1) and maintained these for up to 24 weeks post-immunisation. This combination led to increased cytokine mRNA expression and interferon-gamma (IFN-γ) responses in PBMCs. Enhanced memory B cell responses were also observed in the bone marrow and lymph nodes at 8 weeks post-immunisation. MPL and poly I:C induced higher frequencies of CD4 and CD8 T cells following in vitro restimulation with H3N8 EIV. Conclusions: A viral challenge experiment was not included. Conclusions: The use of MPL and poly I:C as adjuvants in EIV vaccines significantly improved immune responses by enhancing humoral immunity, memory cell responses, and antigen-specific cellular immunity. 背景:马流感(Equine influenza，EI)是一种由甲型流感病毒H3N8亚型引起的高度传染性呼吸系统疾病。尽管进行常规疫苗接种，但由于疫苗株与野外毒株之间的差异，以及疫苗接种间的免疫空窗期，马流感(EI)仍有爆发的风险。 目的:评估佐剂组合能否增强马流感病毒(EIV)疫苗的免疫反应。 研究设计:体内实验研究。 方法:8匹混种马通过肌肉注射免疫重组金丝雀痘病毒载体的EIV疫苗，接种方式为单独接种疫苗或与单磷酰脂A(MPL)和聚肌胞酸(poly I:C)联合使用。疫苗接种仅进行一次，血清样本在接种前、以及接种后第2、8和24周采集，用于评估体液免疫反应。从外周血分离的单核细胞(Peripheral blood mononuclear cells， PBMCs)中检测免疫调节细胞因子表达并评估细胞免疫记忆反应。此外，本研究还采样骨髓和淋巴结细胞以评估记忆细胞反应。 结果:MPL与poly I:C的组合显著增强了针对H3N8 A/equine/Kyonggi/SA01/2011(佛罗里达支系1型分支)的血清IgG抗体水平和血凝抑制(HI)滴度，并在接种后维持该水平长达24周。此外，该佐剂组合还提高了PBMC细胞因子mRNA的表达和γ干扰素(IFN‐γ)的反应。在免疫后第8周时，在骨髓和淋巴结中也观察到了增强的记忆B细胞反应。而且，在体外使用H3N8 EIV再刺激后，该佐剂组合诱导了更高频率的CD4⁺和CD8⁺ T细胞反应。 主要局限:本研究未进行攻毒实验。 结论:使用MPL和poly I:C作为EIV疫苗的佐剂可通过提升体液免疫、记忆细胞反应及抗原特异性细胞免疫，从而显著提高整体免疫反应。.
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Publication Date: 2025-09-02 PubMed ID: 40898646DOI: 10.1111/evj.70090Google Scholar: Lookup
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Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

Overview

  • This study investigated whether combining two adjuvants—monophosphoryl lipid A (MPL) and polyinosinic-polycytidylic acid (poly I:C)—with an equine influenza virus (EIV) vaccine enhances the immune response in horses.
  • The research demonstrated that this adjuvant combination significantly improved both humoral and cellular immunity, including long-lasting antibody levels and memory cell responses.

Background

  • Equine influenza (EI) is a highly contagious respiratory disease caused by the H3N8 subtype of the influenza A virus affecting horses.
  • Despite routine vaccination programs, outbreaks still occur. This is mainly due to two factors:
    • Mismatch between vaccine virus strains and the circulating field strains
    • Immunity gaps between vaccination intervals
  • Improving vaccine efficacy is essential to prevent EI outbreaks and protect horse health.
  • Adjuvants are substances added to vaccines to enhance the immune response; in this context, MPL and poly I:C are known immunostimulatory molecules.

Objective

  • To evaluate whether a combination of the two adjuvants, MPL and poly I:C, can enhance immune responses to a recombinant canarypox-based equine influenza virus vaccine in horses.

Methods

  • Study Design:
    • In vivo experimental study using eight mixed-breed horses.
    • Each horse received a single intramuscular immunization of the recombinant canarypox-based EIV vaccine either alone or combined with MPL and poly I:C adjuvants.
  • Sampling:
    • Serum samples collected pre-immunization and at 2, 8, and 24 weeks post-immunization to measure humoral immune responses.
    • Peripheral blood mononuclear cells (PBMCs) isolated to assess cytokine gene expression and cellular immunity.
    • Bone marrow and lymph node cells sampled to evaluate memory B cell responses.
  • Immune Response Assessment:
    • Serum IgG antibody levels against the H3N8 EIV strain assessed.
    • Hemagglutination inhibition (HI) assays measured antibody functional activity.
    • Quantitative PCR used to evaluate expression of immunomodulatory cytokines, including interferon-gamma (IFN-γ), in PBMCs.
    • Cellular immunity measured by the frequency of CD4+ and CD8+ T cells after in vitro antigen restimulation.

Results

  • The combination of MPL and poly I:C significantly:
    • Increased serum IgG antibody levels against the H3N8 A/equine/Kyonggi/SA01/2011 strain (Florida sub-lineage clade 1).
    • Raised hemagglutination inhibition (HI) antibody titres, indicating better functional antibody response.
    • Maintained elevated antibody levels for up to 24 weeks post-immunization, suggesting durable immunity.
  • Enhanced cellular immune responses:
    • Increased mRNA expression levels of immunomodulatory cytokines including IFN-γ in PBMCs.
    • Higher frequencies of antigen-specific CD4+ and CD8+ T cells were observed after in vitro restimulation with EIV antigen.
  • Memory B cell responses:
    • Stronger memory B cell activity was found in bone marrow and lymph nodes at 8 weeks post-immunization.

Limitations

  • The study did not include a viral challenge experiment to directly assess protection against EIV infection.
  • The sample size was limited to eight horses, which may affect the generalizability of findings.

Conclusions

  • The addition of MPL and poly I:C adjuvants to the recombinant canarypox-based EIV vaccine significantly improved the immune response in horses.
  • This enhancement was seen in:
    • Humoral immunity, via increased and lasting antibody titers.
    • Memory B cell responses in key immune tissues.
    • Antigen-specific cellular immunity, especially CD4+ and CD8+ T cell responses and cytokine production.
  • These findings suggest that using MPL and poly I:C in combination may improve vaccine effectiveness and offer better protection against equine influenza outbreaks.
  • Further studies, including viral challenge experiments, are needed to confirm protective efficacy.

Cite This Article

APA
Dong-Ha L, Eun-Bee L, Jong-Pil S, Eun-Ju K. (2025). Adjuvant combination enhances immune response of equine influenza virus vaccine in horses. Equine Vet J, 58(2), 591-600. https://doi.org/10.1111/evj.70090

Publication

Equine veterinary journal
ISSN: 2042-3306
NlmUniqueID: 0173320
Country: United States
Language: English
Volume: 58
Issue: 2
Pages: 591-600

Researcher Affiliations

Dong-Ha, Lee
  • College of Veterinary Medicine and Veterinary Medical Research Institute, Jeju National University, Jeju, Republic of Korea.
Eun-Bee, Lee
  • College of Veterinary Medicine and Veterinary Medical Research Institute, Jeju National University, Jeju, Republic of Korea.
Jong-Pil, Seo
  • College of Veterinary Medicine and Veterinary Medical Research Institute, Jeju National University, Jeju, Republic of Korea.
Eun-Ju, Ko
  • College of Veterinary Medicine and Veterinary Medical Research Institute, Jeju National University, Jeju, Republic of Korea.
  • Interdisciplinary Graduate Program in Advanced Convergence Technology & Science, Jeju National University, Jeju, Republic of Korea.

MeSH Terms

  • Animals
  • Horses
  • Influenza Vaccines / immunology
  • Influenza Vaccines / administration & dosage
  • Horse Diseases / prevention & control
  • Horse Diseases / virology
  • Horse Diseases / immunology
  • Orthomyxoviridae Infections / veterinary
  • Orthomyxoviridae Infections / prevention & control
  • Orthomyxoviridae Infections / immunology
  • Poly I-C / administration & dosage
  • Poly I-C / pharmacology
  • Influenza A Virus, H3N8 Subtype / immunology
  • Adjuvants, Immunologic / pharmacology
  • Adjuvants, Immunologic / administration & dosage
  • Lipid A / analogs & derivatives
  • Lipid A / administration & dosage
  • Lipid A / pharmacology
  • Antibodies, Viral / blood
  • Female
  • Male

Grant Funding

  • RS-2019-NR040080 / Basic Science Research Program to the Research Institute for Basic Sciences (RIBS) of Jeju National University through the NRF, funded by the Ministry of Education
  • RS-2023-00211504 / National Research Foundation of Korea (NRF) grant funded by the Korean government (MSIT)

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