Fiber-2, not Fiber-1, is the principal protective immunogen of fowl adenovirus serotype 4: insights from a comparative efficacy study of chimeric virus-based inactivated vaccines.

Fowl adenovirus serotype 4 (FAdV-4) uniquely encodes two fiber proteins (fiber-1 and fiber-2), while FAdV-8b possesses only one. Although our previous study showed that an inactivated vaccine based on a chimeric virus with FAdV-4's fiber-1 replaced by FAdV-8b's fiber (rFAdV-4-Δfiber1-fiber/8b) conferred broad protection against both serotypes, the relative contribution of each FAdV-4 fiber to protective immunity remained unclear, impeding the rational design of optimized bivalent vaccines. To address this, we constructed a novel chimeric virus, rFAdV-4-Δfiber2-fiber/8b, by replacing the fiber-2 gene of FAdV-4 with the fiber gene of FAdV-8b. Inactivated oil-emulsion vaccines prepared from both chimeric viruses were compared in SPF chickens. The vaccine retaining native fiber-2 (rFAdV-4-Δfiber1-fiber/8b) induced significantly higher neutralizing antibody titers against FAdV-4, provided superior clinical protection, and more effectively reduced viral loads in key organs and shedding post-challenge, compared to the fiber-2-replaced vaccine. Both vaccines protected against FAdV-8b challenge. Our findings establish that fiber-2 is the principal protective immunogen of FAdV-4, underscoring the necessity of preserving it in the viral backbone when engineering chimeric FAdV-4-based bivalent vaccines. This study provides a fundamental rationale and identifies a superior vaccine candidate (rFAdV-4-Δfiber1-fiber/8b) for the concurrent control of hepatitis-hydropericardium syndrome and inclusion body hepatitis.