Tracking spatio-temporal dynamics of early immune responses to an intranasal OMV-based pneumococcal vaccine candidate...

Outer membrane vesicle (OMV)-based vaccines elicit strong immune responses and have emerged as a versatile platform for targeting multiple pathogens, yet the mechanisms underlying their efficacy remain incompletely understood. Here, we investigated the early immune response following intranasal administration of an OMV-based pneumococcal vaccine candidate in mice. Using in vivo bioluminescence imaging, spectral flow cytometry, and high-resolution microscopy, we tracked OMV biodistribution and immune activation throughout the respiratory tract from 1 to 72 h post-immunization. OMVs persisted in the nasal cavity for up to 48 h and rapidly recruited Ly6G^hi neutrophils and myeloid-derived suppressor cells, followed by activation of local T cells. MHCII expression was significantly upregulated on Ly6G^hi neutrophils in nasal tissue and coincided with a marked expansion at 24 h. In the lungs, alveolar macrophages and plasmacytoid dendritic cells emerged as early responders. OMV exposure also induced costimulatory molecule expression across multiple myeloid cell subsets. Together, these findings reveal distinct spatio-temporal patterns of innate and adaptive immune activation at mucosal sites, providing mechanistic insight into OMV-induced mucosal immunity and underscoring their potential as a versatile vaccine platform.