A viral-vectored multipathogen vaccine platform based on replication-competent vaccinia virus and adeno-associated virus

Multi pathogen vaccines have emerged as a promising strategy to improve vaccine coverage, simplify immunization logistics, and address overlapping global health threats. However, few vaccine platforms have demonstrated robust and durable immune responses against pathogens spanning distinct biological classes. Here, we developed and evaluated a viral-vectored multipathogen vaccine platform based on a heterologous prime–boost regimen combining the highly attenuated vaccinia virus LC16m8Δ (m8Δ) and adeno-associated virus serotype 1 (AAV1). As a preclinical proof-of-concept, antigen-specific immune responses against Plasmodium falciparum and SARS-CoV-2 (Omicron variant) were evaluated, while neutralizing activity against mpox virus was assessed to examine the cross-reactive immunity conferred by the vaccinia virus vector. Immunogenicity, protective efficacy, and transmission-blocking activity were evaluated in murine models and mosquito feeding assays. The heterologous m8Δ prime/AAV1 boost regimen induced robust and long-lasting antigen-specific antibody responses that were maintained for up to 32 weeks. Complete sterile protection against transgenic Plasmodium berghei sporozoite challenge was achieved, together with greater than 90% transmission-blocking efficacy in mosquito feeding assays. In parallel, potent neutralizing antibody responses against SARS-CoV-2 Omicron and cross-neutralizing activity against mpox virus were observed. These findings demonstrate the feasibility of the m8Δ/AAV1 platform as a versatile viral-vectored multipathogen vaccine platform capable of integrating protozoan and viral antigens, providing a rationale for further preclinical optimization and clinical evaluation of next generation multipathogen vaccines targeting both endemic and emerging infectious diseases.