High Fluence Increases the Antibacterial Efficacy of PACK Cross-Linking

Photoactivated chromophore for keratitis cross-linking (PACK-CXL) is used as an adjunct therapy to antibiotic medication in infectious keratitis. This experimental study aimed at quantifying the PACK-CXL efficacy as a function of UV fluence using several bacterial strains and irradiated volumes.

Six distinct bacterial strains, including standardized strains and clinically isolated strains from patients with keratitis, were analyzed. Bacterial concentrations between 10 and 10 cells/mL were used (simulating small corneal ulcers). Volumes of either 11 μL (≈285 μm stromal thickness) or 40 μL (≈1000 μm stromal thickness) were irradiated within a microtiter plate at different fluences (5.4-27 J/cm) and irradiances (3, 9 and 18 mW/cm). The ratio of bacterial killing (B†) was determined to evaluate the antimicrobial efficacy of PACK-CXL.

B† was similar (51 ± 11%) in bacterial concentrations between 10 and 10 per ml. In 11 μL volume, Staphylococcus aureus (SA) 8325-4 ATCC 29213, Bacillus subtilis (BS) 212901, and Pseudomonas aeruginosa (PA) 2016-866624 were most sensitive to PACK-CXL at 5.4 J/cm (on average B† = 49 ± 8%), whereas Klebsiella oxytoca (KO) 2016-86624 (B† = 25%) was least sensitive. When irradiating a larger volume, B† was on average lower in 40 μL (19 ± 18%), compared with 11 μL (45 ± 17%, P < 0.001). By contrast, applying a higher UV fluence increased B† of SA ATCC 29213, from 50% at 5.4 J/cm to 92% at 10.8 J/cm, to 100% at 16.2 J/cm and above.

Applying higher UV fluences substantially increases the bacterial killing rates. Safety limits for clinical application require further investigation.