Effect of Photodynamic Inactivation (PDI) using Riboflavin-Conjugated Antibody against Staphylococcus aureus

Purpose: Crosslinking/riboflavin-UVA photodynamic therapy is a potential treatment alternative in antibiotic resistant infectious keratitis. For photodynamic therapy a specific (against bacteria) conjugated antibody may be used in order to increase the effect of the treatment. In our present study we analysed the impact of photodynamic inactivation using riboflavin-conjugated antibody or riboflavin alone on Staphylococcus aureus, in vitro. Methods: Staphylococcus aureus (S. aureus) was incubated in 1:100 diluted riboflavin-conjugated antibody (R-AB) for 30 minutes in darkness. Following UVA-light illumination (375nm) with an energy dose of 2, 3, 4 and 8 J/cm(2), bacteria were brought to blood agar Plates for 24 hours before colony-forming unit (CFU) counting. In an additional group, we incubated bacteria to 0, 0.05 or 0.1% riboflavin 5-phosphate as described above followed by illumination using UVA light (375nm) with an energy dose of 2 J/cm(2), before CFU counting. Results: The number of CFU decreased significantly (inactivation of 36%, p=0.022) using 1:100 diluted riboflavin-conjugated antibody and 2 J/cm(2)UVA-light illumination, compared to untreated controls. The use of 3, 4 und 8 J/cm(2) energy dose and R-AB in 1:100 dilution did not further change the decrease of CFU (inactivation of 39, 39 and 40%; p=0.016; p=0.016; p=0.015). The use of 0.05% or 0.1% riboflavin 5-phosphate alone and UVA-light illumination reduced the CFU count significantly (inactivation of 73 and 55%; p=0.002; p=0.005), compared to untreated controls. Conclusions: The use of riboflavin-conjugated antibody or 0.05% or 0.1% riboflavin 5-phosphate and UVA-light illumination reduces the number of CFU of S. aureus. However, none of these photodynamic therapies reached the necessary 99% killing rate of these bacteria. Further work is needed to increase the efficacy of riboflavin-conjugated antibodies against antibiotic resistant bacteria.