Production, characterization, and antimicrobial activity of polyhydroxyalkanoates synthesized by <i>Bacillus</i> species against skin pathogens

Polyhydroxyalkanoates (PHAs) are biodegradable polyesters with promising biomedical applications, particularly for combating antibiotic-resistant skin infections, in the development of wound dressings and other healthcare materials. The growing challenge of antibiotic-resistant skin infections has been managed by exploiting PHAs as an alternative to synthetic materials due to their sustainability, eco-friendly nature and antimicrobial functionality. This research used an orthogonal experimental design for PHB optimization, and the results demonstrated that ammonium sulphate (2 g L^-1), glucose, 10% NaCl, and 67 h incubation maximized PHB yield (46.7%). Citric acid supplementation (2-4 g L^-1) enhanced acetyl-CoA flux, increasing PHB synthesis. The extracted PHB was characterized using FTIR, ¹H-NMR, and ¹³C-NMR spectroscopies, confirming the presence of poly-3-hydroxybutyrate (PHB). Thermal and structural characterization of the synthesized PHB confirmed its semi-crystalline nature and enhanced thermal stability. TGA, DSC, and XRD analyses revealed high degradation temperatures, distinct melting and crystallization transitions, and well-defined diffraction peaks, indicating a stable and structurally ordered polymer. PHB exhibited potent antimicrobial activity against skin pathogens, with zones of inhibition of 16 mm for <i>Staphylococcus aureus</i>, 14 mm for <i>S. epidermidis</i>, and 10 mm for <i>Candida albicans</i> at 1000 μg per disk. The MIC values were 400 μg mL^-1 for <i>S. aureus</i>, 600 μg mL^-1 for <i>S. epidermidis</i>, and 1000 μg mL^-1 for <i>C. albicans</i>. Time-kill assays showed complete eradication of <i>S. aureus</i> within 8 h. PHB also disrupted biofilms, achieving 60% inhibition (co-incubation) and 70% eradication (preformed biofilms) at MIC concentrations. The XTT assay revealed PHA's dose-dependent <i>anti</i>-biofilm activity. Co-incubation with 600 μg mL^-1 PHB strongly inhibited biofilms (24-55% viability), while 1000 μg mL^-1 nearly eradicated bacterial biofilms (12-16% viability). PHB also disrupted the preformed biofilms, showing stronger activity against Gram-positive bacteria than against <i>C. albicans</i>. This study underscores the potential of <i>B. megaterium</i>-derived PHB as an eco-friendly antimicrobial agent against resistant skin pathogens, aligning with global efforts to replace synthetic plastics and antibiotics.