From Bacterial Extract to Breakthrough Therapy: Pseudomonas fluorescens-Enabled Green Synthesis of pH-Responsive Chitosan–Silver Hybrid Nanoparticles for Next-Generation Pulmonary Drug Delivery Anti-MDR Treatment

<b>Background:</b> Multidrug-resistant (MDR) pulmonary infections represent a critical global health challenge, necessitating innovative therapeutic approaches. Green synthesis methodologies offer sustainable alternatives for nanoparticle fabrication while addressing antimicrobial resistance. <b>Methods:</b> Stimuli-responsive chitosan-silver hybrid nanoparticles (CS-Ag HNPs) were biosynthesized using <i>Pseudomonas fluorescens</i> bacterial extracts and loaded with ciprofloxacin for targeted pulmonary delivery. Comprehensive characterization included dynamic light scattering, transmission electron microscopy, UV-visible spectroscopy, and aerodynamic assessment via next-generation impactor. Antimicrobial efficacy was evaluated against MDR <i>Pseudomonas aeruginosa</i> and <i>Klebsiella pneumoniae</i>, including biofilm disruption studies, and biocompatibility was assessed. Molecular docking analysis elucidated binding mechanisms. Cytotoxicity and epithelial barrier integrity were evaluated using Calu-3 cell models. <b>Results:</b> The biosynthesized NPs exhibited optimal physicochemical properties (180 ± 20 nm, PDI 0.21 ± 0.04, ζ-potential + 32.4 ± 3.1 mV) with high encapsulation efficiency (68.2 ± 4.0%). Aerodynamic analysis revealed excellent inhalation characteristics (MMAD 2.6 μm, FPF 65 ± 5%). The hybrid system demonstrated 4-fold enhanced antimicrobial activity against MDR pathogens and significant biofilm disruption (70% for <i>P. aeruginosa</i>, 65% for <i>K. pneumoniae</i>) compared to free ciprofloxacin. Cell viability remained ≥85% at therapeutic concentrations. Molecular docking revealed enhanced drug-target binding affinity (-11.2 vs. -9.3 kcal/mol) and multi-residue interactions. <b>Conclusions:</b> Green-synthesized CS-Ag HNPs represent a promising sustainable platform for combating pulmonary MDR infections through enhanced antimicrobial efficacy and optimal aerodynamic properties.