An intranasal bivalent vaccine induces antigen-specific humoral and Th1/Th17 cellular immunity and protects young and aged mice against MRSA and Pseudomonas aeruginosa skin and soft tissue infections

ABSTRACT Staphylococcus aureus (SA) and Pseudomonas aeruginosa (PA) are ESKAPE pathogens that are responsible for severe skin and soft tissue infections (SSTIs), particularly in the elderly and those with immunocompromising conditions. Rising rates of methicillin-resistant S. aureus (MRSA) and carbapenem-resistant PA have rendered standard antibiotics less effective. Unfortunately, there are currently no licensed vaccines for either pathogen. To address this need, we developed a bivalent intranasal (IN) vaccine formulation combining L-PaF, a fusion of PA T3SS proteins PcrV and PopB, with the mucosal adjuvant LTA1, NEAT2 (N2), the protective heme-transporting domain of SA IsdB protein, and a novel TLR-4 agonist, which is a detoxified lipid A analog designated Bacterial Enzymatic Combinatorial Chemistry 438 (BECC438b), formulated in an oil-in-water medImmune emulsion (ME). Mice immunized with L-PaF/ME/N2/BECC438b generated robust antigen-specific IgG and IgA responses, which correlated with strong opsonophagocytic killing of both MRSA and PA strains. Vaccination also induced interleukin IL-17A, IL-2, and IL-22 responses in spleen cells from both young and elderly mice, supporting Th1/Th17-mediated immunity. In stringent murine SSTI models, immunized animals exhibited significantly reduced dermonecrosis, abscess volume, and bacterial burden, with approximately a 10-fold reduction in abscess CFUs compared to non-immunized controls. Notably, significant protection was observed in both young and elderly mice despite age-related immunosenescence, with elderly vaccinated animals showing substantially reduced abscess formation and bacterial burden relative to non-immunized controls. These results suggest that the bivalent L-PaF/ME/N2/BECC438b formulation has potential as a broad-spectrum vaccine against both MRSA- and PA-mediated SSTIs across different age groups.