Otitis media (OM) is a leading cause of childhood morbidity and hearing impairment, particularly affecting Aboriginal and Torres Strait Islander children in Australia. Despite its global prevalence, no licensed vaccine is currently available to prevent OM. Pneumococcal vaccines offer limited protection due to incomplete serotype coverage and their inability to target key non-pneumococcal otopathogens, including non-typeable Haemophilus influenzae and Moraxella catarrhalis. The reliance on antibiotic therapy has further exacerbated antimicrobial resistance, reinforcing the urgent need for innovative and broadly protective vaccine approaches. Among OM-associated pathogens, M. catarrhalis remains particularly underrepresented in vaccine development efforts, with no licensed antigens currently available.
Outer membrane vesicles (OMVs) represent a promising vaccine platform due to their ability to present multiple native antigens in their natural conformations, along with inherent immunostimulatory properties. In this study, we developed genetically modified M. catarrhalis strains lacking immunodominant, strain-variable surface proteins to generate engineered OMVs enriched for conserved antigens. These knockout mutants were constructed using targeted genetic approaches to minimize antigenic variability and enhance cross-strain coverage.
OMVs were isolated from the engineered strains using ultracentrifugation and extensively characterized for yield and composition. Total protein content was quantified using bicinchoninic acid (BCA) assays, while proteomic profiling by mass spectrometry confirmed the enrichment of conserved antigenic components within the vesicle preparations. The immunogenicity of these engineered OMVs was evaluated in a murine immunization model.
Immunization with engineered OMVs elicited strong antigen-specific antibody responses, as measured by ELISA against multiple M. catarrhalis strains. IgG subclass analysis demonstrated a Th2- and a Th1-type immune response, indicative of effective immune activation. Functional assessment using serum bactericidal assays revealed that antibodies generated in response to OMV immunization were capable of killing diverse M. catarrhalis strains, supporting the potential of this platform to overcome pathogen heterogeneity.
In summary, bioengineered M. catarrhalis OMVs represent a versatile and multivalent vaccine strategy capable of inducing robust and functional immune responses. This approach offers a promising pathway toward the development of a broadly protective vaccine against otitis media.