To develop an effective Pseudomonas aeruginosa outer-membrane-vesicle (OMV) vaccine, we eliminated multiple virulence factors from a wild-type (WT) P. aeruginosa strain, PA103, to generate a recombinant strain, PA-m14. Strain PA-m14 was tailored with a pSMV83 plasmid carrying the pcrV-hitAT fusion gene to produce OMVs. The recombinant OMVs (termed OMV-PH) enclosed increased amounts of the PcrV-HitAT bivalent antigen (PH) and exhibited lower toxicity than did the OMVs from PA103. Intramuscular vaccination with OMV-PH from PA-m14(pSMV83) afforded 70% protection against intranasal challenge with 6.5 × 106 CFU (∼30 50% lethal doses [LD50]) of PA103, while immunization using OMVs without the PH antigen (termed OMV-NA) or the PH antigen alone failed to offer effective protection against the same challenge. Further immune analysis showed that OMV-PH immunization significantly stimulated potent antigen-specific humoral and T-cell (Th1/Th17) responses over those with PH or OMV-NA immunization in mice and that these more-potent responses can effectively hinder P. aeruginosa infection. Undiluted antisera from OMV-PH-immunized mice displayed significantly more opsonophagocytic killing of WT PA103 than antisera from PH antigen- or OMV-NA-immunized mice. Moreover, OMV-PH immunization afforded significant antibody-independent cross-protection to mice against PAO1 and the AMC-PA10 clinical isolate. Taking our findings together, the recombinant P. aeruginosa OMV delivering the bivalent PH antigen exhibits high immunogenicity and may be a promising next-generation vaccine candidate against P. aeruginosa infection.
Keywords: P. aeruginosa; nanoparticle; outer membrane vesicles; protective immunity; vaccine.