Antifungal susceptibility testing in three-dimensional keratinocyte culture model

Background: Standardization of antifungal susceptibility testing for dermatophytes is important and several variables, such as inoculum size, length and temperature of incubation, media, and end point determination has recently been established. However, a more improved test model which can evaluate bioavailability and has clinical relevance is still needed. Objective: We performed antifungal susceptibility testing by using three-dimensional keratinoctyte culture model, living skin equivalent (LSE), as an in vitro model. Methods: LSE was prepared and various concentrations of antifungals were added to media. Microconidia of Trichophyton mentagrophytes was inoculated onto LSE and incubated for 6 days at 35°C. Results: Inhibition of fungal proliferation and invasion were observed at 0.1 μg/ml of terbinafine, 0.01 μg/ml of itraconazole solution, 0.1 μg/ml of itraconazole powder, and 10 μg/ml of fluconazole, respectively. Conclusion: Our culture model is similar to in vivo situation and the results were relatively well accordant to those of other previous studies. Our LSE model is considered as a promising in vitro system for evaluating the activity and safety of antifungal agents. However, further study using more various species of dermatophytes and more strains is still needed.