Immunoinformatic design of a multi-epitope vaccine candidate targeting multidrug-resistant Pseudomonas aeruginosa

Pseudomonas aeruginosa, a World Health Organization (WHO) priority pathogen, causes severe multidrug-resistant (MDR) infections in immunocompromised and hospitalized patients, highlighting the critical need for effective vaccines. To this end, we designed a novel multi-epitope vaccine by integrating immunodominant domains from three key virulence proteins (PcrV, OprE, and CupC2) with the cholera toxin B subunit (CTB) as a mucosal adjuvant. Immunoinformatics was used to predict epitopes and construct a vaccine with EAAAKEAAAK linkers. In silico assessments included antigenicity, allergenicity, solubility, molecular docking with TLR2/TLR4, 150 ns MD simulations, PDBsum interaction analysis, and C-ImmSim immune profiling. Both the adjuvanted and non-adjuvanted candidates were predicted to be non-allergenic, with antigenicity scores of 0.6557 and 0.7338, respectively, indicating strong immunogenic potential. Both constructs were also predicted to exhibit high solubility, with scores exceeding the threshold (0.927 and 0.729). Each vaccine candidate showed robust interactions with TLR2 and TLR4; however, the adjuvanted construct demonstrated superior binding affinity to both receptors. Molecular docking of the adjuvanted vaccine candidate followed by 150 ns molecular dynamics (MD) simulations confirmed stable and strong interactions with TLR2 and TLR4, as evidenced by RMSD values of 0.64 nm and 0.73 nm, and binding energies of ΔG = − 15.9 kcal/mol and − 12.7 kcal/mol, respectively further supporting their high immunogenic potential. Moreover, in silico immune simulation data predicted rapid induction of protective antibodies and cytokines and revealed that adjuvant-enhanced responses substantially improved the stability of the TLR2 complex. These computational results suggest that our multi-antigen vaccine candidate may have potential against Pseudomonas aeruginosa infections. Further validation through gene synthesis, murine immunogenicity studies (e.g., BALB/c models), and challenge experiments is required.