Expression of bluetongue virus core proteins VP2 and VP7 by bovine herpesvirus-4 confers protection against virulent challenge in a mouse model

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Abstract Bluetongue virus (BTV) remains a major threat to livestock health and agricultural economies worldwide due to its multiple serotypes, vector transmission, and wide geographic distribution. Vaccination is essential for controlling BTV outbreaks; however, developing vaccines that provide broad protection while complying with the requirements of differentiating infected from vaccinated animals (DIVA) remains challenging. In this study, we evaluated the antigenicity and protective efficacy of two BTV proteins, VP2 and VP7, delivered using a bovine herpesvirus-4 (BoHV-4)-based viral vector. Groups of mice lacking interferon-α/β receptor (IFNAR −/− ) (C57BL/6 background) were immunized using a prime/boost regimen with BoHV-4 vectors expressing VP2, VP7, or both antigens. All vaccinated groups developed antigen-specific humoral and cellular immune responses. Following challenge with virulent BTV-8, complete protection was observed in the VP2 and VP7 single-antigen groups, while 80% protection was achieved with the combined VP2 + VP7 vaccination. Total anti-BTV IgG levels increased after booster immunization, with the highest pre-challenge titers detected in the combined vaccination group. After challenge, IgG levels rose further in VP2-immunized mice, whereas VP7-immunized animals showed comparatively lower titers. All vaccinated mice mounted BTV-specific cellular responses, as demonstrated by interferon (IFN)-γ enzyme-linked immunoadsorption assay (ELISPOT) and intracellular cytokine staining. T cells expressing IFN-γ, IL-2, TNF-α, or CD107a were detected, with evidence of polyfunctional responses across groups. Notably, VP7 immunization induced stronger cellular responses, including increased cytotoxic potential in CD4 + and CD8 + T cells. These findings demonstrate that BoHV-4 vectors expressing VP2 or VP7 elicit robust humoral and cellular immunity and confer protection against BTV, highlighting their potential as a versatile vaccine platform.

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