Whole proteome-integrated and vaccinomics-based next generation mRNA vaccine design against Pseudomonas aeruginosa-A hierarchical subtractive proteomics approach.

Pseudomonas aeruginosa (P. aeruginosa) is a multidrug-resistant opportunistic pathogen responsible for chronic obstructive pulmonary disease (COPD), cystic fibrosis, and ventilator-associated pneumonia (VAP), leading to cancer. Developing an efficacious vaccine remains the most promising strategy for combating P. aeruginosa infections. In this study, we employed an advanced in silico strategy to design a highly efficient and stable mRNA vaccine using immunoinformatics tools. Whole proteome data were utilized to identify highly immunogenic vaccine candidates using subtractive proteomics. Three extracellular proteins were prioritized for T- and linear B-cell epitope prediction. Beta-definsin protein sequence was incorporated as an adjuvant at the N-terminus of the construct. A total of 3 CTL, 3 HTL, and 3 linear B cell highly immunogenic epitopes were combined using specific linkers to design this multi-peptide construct. The 5' and 3' UTR sequences, Kozak sequence with a stop codon, and signal peptides followed by a poly-A tail were incorporated into the above vaccine construct to create our final mRNA vaccine. The vaccines exhibited antigenicity scores >0.88, ensuring high antigenicity with no allergenic or toxic. Physiochemical properties analysis revealed high solubility and thermostability. Three-dimensional structural analysis determined high-quality structures. Vaccine-receptor docking and molecular dynamic simulations demonstrated strong molecular interactions, stable binding affinities, dynamic nature, and structural stability of this vaccine, with significant immunogenic responses of the immune system against the vaccine. The immunological simulation indicates successful cellular and humoral immune responses to defend against P. aeruginosa infection. Validation of the study outcomes necessitates both experimental and clinical testing.