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Objectives and questions: Despite the use of conventional local and systemic antibiotic therapies, biomaterial-associated infections (BAI) remain one of the most common complications in trauma patients. Alternative antimicrobial strategies are urgently needed. In trauma surgery, the use bioactive surfaces is a promising approach for decontamination and reducing bacterial attachment. This study aims to demonstrate the efficacy of phenothiazine-based photosensitizers (PS) against Staphylococcus aureus implant-related infections using two strategies: i) Evaluating photodynamic treatment (PDT) based on selected PS against bacteria attached to the surface of clinical metallic implants. ii) Synthesizing PS-loaded polymethylmethacrylate (PMMA)-based bone cements with varying PS concentrations to prevent bacterial attachment.

Material and methods: In vitro experiments were conducted to assess PDT efficiency by determining the minimal photo-bactericidal concentrations against clinical S. aureus strains in planktonic and biofilm condition. The DCFH-DA fluorescent probe was used to quantify reactive oxygen species (ROS) production. Based on these findings, PS-loaded PMMA formulations were prepared by using Palacos R bone cement. Eluates were collected at various time points to assess PS release (concentration and ROS production) and investigate their antibacterial and antibiofilm activity. In vivo experiments using Galleria mellonella infection models were conducted to examine PS and ROS biodistribution, as well as to evaluate PDT efficacy in preventing implant-related infections in larvae.

Results: The combination of PS and light irradiation successfully photoinactivated clinical <em>S. aureus</em> strains. A complete bacterial eradication (8 logCFU/mL) were achieved from 16 to 78 µM, depending on the selected phenothiazine. Based on these results, a direct correlation was observed between the PS efficiency and its ability to produce ROS. Interestingly, PDT treatment significantly reduced bacterial counts after early to mature biofilm formation (4 to 48 h) on K-wire surfaces. When PS was incorporated into bone cement, an increase concentration release from the bone cement was observed over 5 days, regardless the PS-loaded concentration, i-e from 0.25 to 1% (w/w). However, a minimum concentration of 0.5% PS-PMMA was required to release sufficient PS to achieve a significant reduction in bacterial load. in vivo biocompatibility studies demonstrated that PDT was well-tolerated in G. mellonella.

Discussion and conclusions: Based on our strategies, PDT demonstrates encouraging results to prevent bacterial attachment on the surface of implant and could serve as a valuable addition to conventional antibiotic-based BAI prevention strategies. Further in vivo studies using this animal model are ongoing to evaluate the antibiofilm efficacy of this approach.