Real-Time Microscopic Observation of Candida Biofilm Development and Effects Due to Micafungin and Fluconazole

To understand the process of Candida biofilm development and the effects of antifungal agents on biofilms, we analyzed real-time data comprising time-lapse images taken at times separated by brief intervals. The growth rate was calculated by measuring the change of biofilm thickness every hour. For the antifungal study, 5-h-old biofilms of Candida albicanswere treated with either micafungin (MCFG) or fluconazole (FLCZ). MCFG began to suppress biofilm growth a fewminutes after the initiation of the treatment, and this effect was maintained over the course of the observation period. In contrast, the suppressive effects of FLCZ on biofilm growth took longer to manifest: biofilms grew in the first 5 h after treatment, and then their growth was sup-pressed over the next 10 h, finally producing results similar to those observed withMCFG. MCFGwas also involved in the dis-ruption of cells in the biofilms, releasing string-like structures (undefined extracellular component) from the burst hyphae. Thus, MCFG inhibited the detachment of yeast cell clusters from the tips of hyphae. In contrast, FLCZ did not disrupt biofilm cells. MCFG also showed fast antifungal activity against Candida parapsilosis biofilms. In conclusion, our results show that inhi-bition of glucan synthesis due toMCFG contributed not only to fungicidal activity but also to the immediate suppression of bio-film growth, while FLCZ suppressed growth by inhibiting ergosterol synthesis. Therefore, those characteristic differences should be considered when treating clinical biofilm infections. Pathogenic fungi in the genus Candida can cause both superfi-cial and serious systemic diseases and are now widely recog-nized as important agents of hospital-acquired infection (1).Can