Correlation between antifungal drug susceptibility and morphological changes of Candida albicans and Candida parapsilosis biofilms
Abstract number: R2260
Katragkou A., Kruhlak M., Simitsopoulou M., Chatzimoschou A., Georgiadou E., Cotten C., Walsh T., Roilides E.
Objective: While the effects of older antifungals on Candida biofilms (BF) are well-documented, less is known about the effects of newer triazoles and echinocandins against Candida BF. Our objective was to determine the activities of voriconazole (VRC), posaconazole (PSC), caspofungin (CAS) and anidulafungin (ANID) against Candida albicans (CA) and Candida parapsilosis (CP) BF, to compare these activities with their planktonic (PL) counterparts and to correlate the susceptibility with the corresponding morphological changes.
Methods: Two CA and 1 CP clinical BF-producing strains were used for susceptibility testing. A Green Fluorescent Protein-tagged CA strain was used for microscopy. BF were grown on silicone elastomer disks in 12-well or 96-well plates at 37°C under constant shaking for 4872 h. Mature BF were incubated in RPMI-1640 containing no drug (controls), VRC, PSC, CAS or ANID at two-fold dilutions for 24 h. BF formation and antifungal activities were assessed by XTT assay as changes in BF metabolic activity. BF MICs were determined as minimum antifungal concentrations causing 50% reduction in the BF metabolic activity compared to controls. PL MICs were determined by CLSI M27-A2 method. Confocal laser scanning microscopy (CLSM) was used to evaluate the effects of VRC and ANID on BF morphology. ANOVA with post-test analysis was employed.
Results: PL MICs for CA-M61, CA-GDH2346 and CP-PA/71 were of VRC 0.01, 4, 0.03, of PSC 0.001, 4;0.001, 0.01, of CAS 0.06, 0.03, 0.06 and of ANID 0.003, 0.003, 0.125 mg/l, respectively. By comparison, the BF MICs of VRC were >256, >256, 256, of PSC >64, >64, >256, of CAS 0.06, 0.03, 1 and of ANID 0.5, 0.12, 2. The maximum inhibitory effect of CAS against CA BF was noted for both strains at 2 mg/l (37±9% vs. control p < 0.01) and of ANID at 0.5 or 8 mg/l (48.5±3.4% or 40.3±4.1% vs. control, p < 0.001) depending on the strain. The corresponding values of both CAS and ANID for CP BF were 16 mg/l (41.2±9%, p < 0.001; 47.6±7%, p = 0.01). CLSM showed that VRC-treated CA BF were morphologically similar to untreated controls; whereas, ANID-treated CA BF were greatly distorted with shorter hyphae, looser network and vacuoles on the cell wall. However, complete sterility of the silicone substrate did not occur.
Conclusions: Echinocandins exhibit relatively low MICs for Candida BF; whereas, MICs of triazoles are very high. These findings correlate with the morphological changes observed in drug-treated BF and provide insight into mechanisms underlying BF resistance.
|Session name:||18th European Congress of Clinical Microbiology and Infectious Diseases|
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