Abstract
Purpose: :
To evaluate the disinfectant effect of methylene blue (MB) mediated photodynamic therapy (PDT) on a pathogenic strain of Acanthamoeba known to cause infectious keratitis.
Methods: :
Acanthamoeba castellanii (ATCC50370) trophozoites used in this study were treated under one of four experimental conditions: PDT light and MB (L+M+), PDT light and physiologic solution (L+M-), MB only (L-M+), and physiologic solution (L-M-). M+ trophozoites were incubated in either 0.25 or 0.5 mM MB for 10 minutes. L+ organisms were exposed to PDT light for 30 minutes following incubation in solution. A halogen lamp (660 nm) with a maximum output of 6 mW/cm2 was used as the PDT light source. After treatment, anti-acanthamoeba activity was evaluated by checking the respiratory activity of the trophozoites. The trophozoites were stained with 5-cyano-2,3-ditolyl-tetrazolium chloride, and the magnitude of respiratory activity was determined by measuring the fluorescence intensity with a fluorescence microplate reader. We also evaluated the effect of crosslinking by treating Acanthamoeba for 30 minutes in vitro with a combination of 0.1% riboflavin and ultraviolet illumination (UVA, 365 nm) at an irradiance of 3 mM/cm2.
Results: :
MB-PDT suppressed the respiratory activity of the trophozoites in a MB-concentration dependent manner at total light doses of 10.8 J/cm2. The respiratory activity of each group as a percentage of that of L-M- is as follows: L+M+ 11.6% (0.5 mM), 60.9% (0.25 mM); L-M+ 116.5% (0.5 mM), 105.5% (0.25mM); L+M- 107.6%; and L-M- 100%. (L+M+ vs. L-M- P<0.05). There was no significant difference between the respiratory activity of trophozoites treated with crosslinking reagents and untreated control trophozoites.
Conclusions: :
MB-PDT was effective against Acanthamoeba in vitro. In contrast, crosslinking produced by riboflavin and UVA exhibited almost no disinfectant effect on the Acanthamoeba.
Keywords: Acanthamoeba • keratitis • photodynamic therapy