Background and Aims:
Otitis media (OM) is the most common childhood disease diagnosed among Australian children and the leading cause of hearing loss in Aboriginal and Torres Strait Islander children. Outer membrane vesicles (OMVs) represent a promising new adjuvant platform because they naturally mimic bacterial membranes and provide strong innate immune stimulation while also displaying high immunogenicity. In this project, we aimed to harness OMVs as an adjuvant system, while genetically modifying them to remove immunodominant, strain-variable proteins, thereby enhancing their potential to promote cross-protective responses when combined with conserved recombinant antigens.

Methods:
Novel recombinant protein antigens from NTHi and M. catarrhalis were cloned, expressed in E. coli, and purified by immobilized metal affinity chromatography. Genetically modified M. catarrhalis OMVs were produced by engineering knockout mutants, purified by ultracentrifugation, and characterized for yield and composition. Mice were immunized with recombinant antigens formulated with these OMVs, and sera were analyzed by ELISA, and bactericidal assays with complement deposition assays in progress.

Results:
Recombinant NTHi and M. catarrhalis antigens were successfully expressed and purified. Genetically modified OMVs with reduced strain variability were generated and characterized. ELISA data indicate that OMV-adjuvanted formulations elicit strong antigen-specific antibody responses in mice and provide broader coverage against both homologous and heterologous strains.

Conclusions:
This study highlights the use of engineered OMVs as a novel adjuvant platform to enhance immunogenicity and broaden protection against diverse OM pathogens. The integration of recombinant protein antigens with adjuvant-active OMVs offers a promising new strategy to overcome the limitations of current vaccines for OM.