Oropouche virus (OROV) is reemerging in the Americas, along with a growing threat to global public health. Recent outbreaks have witnessed the first reported fatalities, vertical transmissions, and intercontinental importations of OROV, underscoring its expanding risk. Despite this, no vaccines or specific therapeutics are available, and fundamental research on OROV vaccinology and antigenicity remains limited. Here, we show that co-expression of the M polyprotein and nucleocapsid protein (NP) drives the assembly of OROV virus-like particles (VLPs) with high immunogenicity. Using the prototype strain OROV/sloth/Brazil/PA-UG-BeAn19991/1960, we developed an mRNA vaccine, M/N-vac, encoding these VLPs. Immunization with M/N-vac in mice elicited robust OROV-specific IgG and pseudovirus-neutralizing antibodies that cross-reacted with a contemporary circulating strain, hOROV/Brazil/AM-UKY-AM0088/2024. The vaccine also induced a durable, antigen-specific Th1-biased cellular immune response characterized by high-level interferon-gamma secretion. Additionally, we identified a highly conserved potential immunodominant epitope in BALB/c, N2-3, within the nucleocapsid protein. Furthermore, the VLP-encoding mRNA vaccine induced stronger OROV-specific humoral and cellular immune responses than the VLP protein vaccine.results based on immunization with M/N-vac demonstrate VLP-based vaccines to be a promising broad-spectrum strategy against OROV while providing novel insights into their antigenicity and design.IMPORTANCEOropouche virus (OROV) is a reemerging pathogen with no approved countermeasures, and it poses a growing public health threat. In response, we have developed a virus-like particle-based mRNA vaccine that elicits potent and durable neutralizing antibodies against both historical and circulating OROV strains, alongside a robust Th1-biased cellular immune response. This study reports the design and development of a critically needed vaccine candidate and provides fundamental insights into OROV antigenicity, thus demonstrating the utility of the mRNA platform for rapid response to emerging viral threats. In vivo
