Assessment of photodynamic therapy efficacy against Escherichia coli-Enterococcus faecalis biofilms using optical coherence tomography.

Significance: In orthopedic trauma surgery, spatially structured biofilm ecosystems of bacteria that colonize orthopedic devices account for up to 65% of all healthcare infections, including tens of millions of people affected in the United States. These biofilm infections typically show increased resistance to antibiotics due to their structure and composition, which contributes significantly to treatment failure. Anti-biofilm approaches are needed together with clinically usable microscopic-resolution imaging techniques for treatment efficacy assessment.

Aim: Antimicrobial photodynamic therapy (aPDT) has been recently proposed to combat clinically relevant biofilms (chronic wound infections, dental biofilms, etc.) using photosensitizers excited with visible light to generate reactive oxygen species that can kill bacteria residing within pathogenic biofilms. We aim to assess the efficacy of this treatment for eradication of biofilms typically present on surfaces of orthopedic devices (e.g., intramedullary nails and osseointegrated prosthetic implants).

Approach: In the first phase reported here, we test aPDT in vitro by growing biofilms of Escherichia coli and Enterococcus faecalis bacteria (two of the seven most common pathogens found in orthopedic trauma patients) inside soft lithography-fabricated microfluidic devices. We treat these biofilms with 5-aminolevulinic acid (5-ALA)-based aPDT, evaluate treatment efficacy with optical coherence tomography, and compare with regular clinical antibiotic treatment outcomes.

Results: The antibacterial efficiency of 5-ALA-based aPDT showed nonlinear dependence on the photosensitizer concentration and the light power density, with low parameters ( $30J/{\mathrm{cm}}^2$ light dose, $100\mathrm{mg}/\mathrm{mL}$ 5-ALA concentration) being significantly more effective than antibiotic-treated groups ( $p<0.01$ ), reaching 99.98% of bacteria killed at $150J/{\mathrm{cm}}^2$ light dose and $200\mathrm{mg}/\mathrm{mL}$ 5-ALA concentration setting.

Conclusions: Performed experiments enable the translation of this portable treatment/imaging platform to the second phase of the study: aPDT treatment response assessment of biofilms grown on orthopedic hardware.