Assessment of fungal viability after long-wave ultraviolet light irradiation combined with riboflavin administration

Background

Corneal collagen cross-linking (CXL), a technique that combines riboflavin administration with long-wave ultraviolet light irradiation, was primarily developed to increase the biomechanical strength of collagen fibrils of the cornea to avoid the progression of keratoconus. Recently, this method has been proposed to treat selected cases of infectious keratitis.

Methods

To test the protocol used for progressive keratoconus in infectious keratitis, Candida albicans, and Fusarium solani, strains were exposed to irradiation using a wavelength of 365 nm at a power density of 3 mW/cm² for 30 min in the presence of riboflavin photosensitizer. All experiments were performed in triplicate. Qualitative and quantitative measurements of fungal viability used plate cultures and an automated trypan blue dye exclusion method respectively. Fungal cell diameter was also assessed in all groups. Statistical analyses were performed using the triplicate values of each experimental condition.

Results

Experimental findings of photodynamic therapy applied to the cell inactivation of both yeasts and filamentous fungi were compared with control groups. Qualitative results were corroborated with quantitative findings which showed no statistical significance between challenged samples (experimental groups) and the control group (p-value?=?1). In comparison with a control group of live cells, statistical significance was observed when riboflavin solution alone had an effect on the morphologic size of filamentous fungi, while ultraviolet light irradiation alone showed a slight decrease in the cell structure of C. albicans.

Conclusions

The impact of long-wave ultraviolet combined with riboflavin photosensitizer showed no antifungal effect on C. albicans and F. solani. The significant decrease in cell morphology of both filamentous fungi and yeasts submitted to photosensitizing riboflavin and exposure to ultraviolet light, respectively, may be promising in the development and standardization of alternatives for fungal cell inactivation, because of their minimal cytotoxic effects on the corneal surface. The methodological improvement in the preparation and application of individual chemical compounds, such as riboflavin, or physical systems, such as a long-wave light source, as antifungal agents may also assist in establishing promising therapeutic procedures for keratomycosis.