Experimental Approaches to Prophylaxis of MRSA Biofilm Formation on 3D Scaffolds Modified with Rough Calcium Phosphate Coating

Introduction: Microbial biofilm formation on surgical implants significantly ex-acerbates implant-associated infections (IAIs) and often leads to implant failure. This study aimed to investigate the ex vivo effects of elevated temperature and antibiotics, or zinc in-corporation, on methicillin-resistant Staphylococcus aureus (MRSA) biofilm growth on ti-tanium (Ti)-based 3D plates fabricated by selective laser melting (SLM) and coated with a rough calcium phosphate (CaP) layer using the micro-arc oxidation technique. Materials and Methods: MRSA strain 43300 broth cultures were exposed to heating at 45°C for 60 min or 56°C for 30 min, with or without vancomycin (VMN). The control culture (250 microbial cells per 1 ml of isotonic sodium chloride) was maintained at 37°C. Ti-based discs (10 mm in diameter and 1 mm in thickness) with structured surfaces were coated with a CaP layer via micro-arc oxidation. The Ti-based samples were added to MRSA broth cul-tures for 2 hours, then transferred to fresh serum-free medium and incubated for 48 hours at 37 °C. Bacterial biofilm optical density on the CaP coating was quantified by computer mor-phometry after fixation and Gram staining. Results: A single ex vivo administration of VMN at its minimum inhibitory concentration (MIC, 2 μg/ml) was ineffective against MRSA biofilm growth. The therapeutic VMN con-centration (20 μg/ml) reduced the biofilm optical density by 17%. Heating of MRSA broth cultures showed no significant bacteriostatic effect, and combining hyperthermia with VMN did not enhance antimicrobial efficacy. In contrast, Zn-doped (~0.31 at.%) CaP coatings inhibited MRSA biofilm development at 37°C. The bacteriostatic effect (5%) exceeded that of VMN at MIC (0%) but was lower than that observed for the therapeutic VMN dose (17%). Discussion: The negatively charged CaP coating on titanium implants may promote MRSA biofilm formation through electrostatic interactions with bacterial cell surfaces. Hyperther-mia at 45°C or 56°C, either alone or in combination with vancomycin, showed limited effi-cacy, possibly due to bacterial thermotolerance and antibiotic resistance. Incorporation of zinc into CaP coatings demonstrated a modest bacteriostatic effect, suggesting potential as an adjunct strategy alongside conventional antibiotics such as vancomycin. Conclusion: Zn-doped rough CaP coatings on Ti-based implants show promise for use in bone tissue engineering, where there is a risk of implant-associated infection, owing to the bacteriostatic activity of zinc at low concentrations.