Nano-boosted thymoquinone: moving beyond antibiotics to inhibit Staphylococcus aureus biofilms.

Staphylococcus aureus is a high-priority pathogen requiring novel antimicrobial strategies. Thymoquinone (TQ), a bioactive compound from Nigella sativa, exhibits antimicrobial and antibiofilm properties, but its precise mechanisms remain unclear. This study evaluated free and nano-encapsulated TQ against multidrug-resistant S. aureus isolates. Nano-encapsulation enhanced biofilm penetration, with TQ significantly reducing extracellular DNA (eDNA) at sub-MIC levels without affecting initial adhesion, exopolysaccharides, or enzymatic virulence. Comparative analysis with vancomycin, ciprofloxacin, and azithromycin confirmed TQ's potent antibiofilm activity. Notably, TQ downregulated crtN, essential for staphyloxanthin biosynthesis, and msaB, a key biofilm and stress response regulator. However, at MIC and super-MIC levels, TQ paradoxically increased staphyloxanthin, suggesting a concentration-dependent oxidative stress response. We propose that TQ disrupts biofilm integrity by modulating the msaABCR operon. These findings highlight nano-TQ's therapeutic potential for biofilm-associated infections and underscore the role of natural compounds in combating multidrug-resistant pathogens.