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

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Abstract

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.

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Acknowledgments

The authors acknowledge a financial support given by an unrestricted grant from Research to Prevent Blindness Inc, NY, NY to the Wilmer Ophthalmological Institute to conduct this study (laboratory supplies).

Financial support

Research to Prevent Blindness Inc, New York, NY provided financial support to the Wilmer Ophthalmological Institute to conduct this study (laboratory supplies). Fabio RS Carvalho was supported by a postdoctoral fellowship from Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (PNPD/CAPES).

Conflict of interest

No conflicting relationship exists for any author.

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Authors and Affiliations

  1. Department of Ophthalmology, Paulista School of Medicine – Federal University of Sao Paulo – UNIFESP, Sao Paulo, Brazil

    Renata T. Kashiwabuchi, Fabio R. S. Carvalho, Flavio Hirai & Mauro S. Campos

  2. The Wilmer Ophthalmological Institute, The Johns Hopkins University, School of Medicine, Baltimore, MD, USA

    Yasin A. Khan & Peter J. McDonnell

  3. Rua Botucatu, 820; Vila Clementino, Sao Paulo, SP, Brazil, 04023-062

    Renata T. Kashiwabuchi

Authors
  1. Renata T. Kashiwabuchi
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  2. Fabio R. S. Carvalho
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  3. Yasin A. Khan
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  4. Flavio Hirai
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  5. Mauro S. Campos
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  6. Peter J. McDonnell
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Corresponding author

Correspondence to Renata T. Kashiwabuchi.

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All authors have full control of all primary data, and we agree to allow Graefe’s Archive for Clinical and Experimental Ophthalmology to review our data upon request.

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Kashiwabuchi, R.T., Carvalho, F.R.S., Khan, Y.A. et al. Assessment of fungal viability after long-wave ultraviolet light irradiation combined with riboflavin administration. Graefes Arch Clin Exp Ophthalmol 251, 521–527 (2013). https://doi.org/10.1007/s00417-012-2209-z

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  • DOI: https://doi.org/10.1007/s00417-012-2209-z

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