Antibiofilm activity and downregulation of icaA and icaD expression by magnesium oxide nanoparticles in clinical Staphylococcus aureus isolates from diabetic foot ulcers

Abstract Background Diabetic foot ulcers (DFUs) are frequently complicated by chronic infections and biofilm formation, with Staphylococcus aureus ( S. aureus ) being one of the predominant pathogens. Because biofilm-associated infections exhibit increased tolerance to antibiotics, alternative or adjunctive therapeutic strategies are needed. This study evaluated the antibacterial, antibiofilm, cytocompatibility, and molecular effects of magnesium oxide nanoparticles (MgO-NPs) against S. aureus isolates recovered from DFUs. Methods Six clinical S. aureus isolates from DFUs and the reference strain S. aureus ATCC 25,923 were included. MgO-NPs were synthesized using a hydrothermal sol–gel method and characterized by XRD, UV–Vis spectroscopy, TEM, FESEM, EDX, BET, and DLS analyses. Antibacterial activity was determined by minimum inhibitory concentration (MIC) testing. Biofilm inhibition was assessed using the microtiter plate crystal violet assay. Cytocompatibility was evaluated by MTT assay using L929 fibroblast cells. Biofilm-associated genes ( icaA , icaB , icaC , and icaD ) were detected by PCR, and the expression of icaA and icaD following exposure to strain-specific sub-MIC concentrations of MgO-NPs was analyzed by quantitative real-time PCR. Results MgO-NPs exhibited MIC values of 512–1024 µg/mL, with corresponding strain-specific sub-MIC concentrations of 256–512 µg/mL. All isolates carried icaA , icaC , and icaD , whereas icaB was detected in 83.3% of strains. Exposure to strain-specific sub-MIC concentrations of MgO-NPs reduced biofilm biomass by an average of approximately 54% across the six clinical isolates. Quantitative real-time PCR demonstrated significant downregulation of icaA and icaD expression in all tested strains, with mean reductions of 40.6% among strain–gene observations exposed to 256 µg/mL and 59.5% among those exposed to 512 µg/mL. These concentrations represented 0.5×MIC for different strains and therefore were not considered directly comparable exposure doses within the same isolate. Cell viability remained above 90% at concentrations up to 512 µg/mL. Conclusion MgO-NPs demonstrated antibacterial and antibiofilm activity and were associated with significant suppression of key biofilm-associated genes while maintaining acceptable cytocompatibility under the conditions evaluated. These findings provide preliminary evidence supporting the potential of MgO-NPs as an adjunctive strategy for managing biofilm-associated S. aureus infections in DFUs. However, the small number of clinical isolates, limited experimental controls, and inability of the crystal violet assay to completely distinguish direct antibiofilm effects from growth-related reductions should be considered. Larger studies incorporating appropriate controls, comprehensive mechanistic analyses, and in vivo models are required to validate these findings and clarify the underlying mechanisms.