Effect of antifungal agents, lysozyme and human antimicrobial peptide LL-37 on clinical Candida isolates with high biofilm production

Abstract

Introduction. Candida species can form biofilms on tissues and medical devices, making them less susceptible to antifungal agents. Hypothesis/Gap Statement. Antifungal combination may be an effective strategy to fight against Candida biofilm. Aim. In this study, we investigated the in vitro activity of fluconazole, caspofungin and amphotericin B, alone and in combination, against 17 clinical Candida tropicalis and 6 Candida parapsilosis isolates with high biofilm formation. We also tested LL-37 and lysozyme for anti-biofilm activity against a selected C. tropicalis isolate. Methodology. Candida biofilms were prepared using the 96-well plate-based method. The minimum biofilm eradication concentrations were determined for single and combined antifungal drugs. The activity of LL-37 and lysozyme was determined by visual reading for planktonic cells and using the XTT assay for biofilms. Results. Under biofilm conditions, fluconazole plus caspofungin showed synergistic effects against 60.9% (14 of 23) of the tested isolates, including 70.6% of C. tropicalis [fractional inhibitory concentration index (FICI), 0.26–1.03] and 33.3% of C. parapsilosis (FICI, 0.04–2.03) isolates. Using this combination, no antagonism was observed. Amphotericin B plus caspofungin showed no effects against 78.3% (18 of 23) of the tested isolates. Amphotericin B plus fluconazole showed no effects against 65.2% (15 of 23) of the tested isolates and may have led to antagonism against 2 C. tropicalis and 2 C. parapsilosis isolates. LL-37 and lysozyme had no effect on biofilms of the selected C. tropicalis isolate. Conclusions. We found that fluconazole plus caspofungin led to a synergistic effect against C. tropicalis and C. parapsilosis biofilms. The efficacy of the antifungal combination therapies of the proposed schemes against biofilm-associated Candida infections requires careful and constant evaluation.