Biofilm-forming capacity and biofilm antifungal drug resistance of Candida bloodstream isolates

Abstract number: P1367

Chatzimoschou A., Katragkou A., Simitsopoulou M., Iosifides E., Georgiadou E., Antachopoulos C., Roilides E.

Objective: Candidaemia is one of the most important nosocomial bloodstream infections. Biofilm (BF) formation is considered a virulence factor responsible for catheter-related candidaemia. Our aim was to investigate the capacity of bloodstream Candida albicans (CA) and Candida parapsilosis (CP) isolates to form BF and to correlate the degree of BF formation with the resistance of BF to voriconazole (VRC) and posaconazole (PSC).

Methods: 5×10⁵ planktonic (PL) cells/mL were grown in YNB medium with 2% glucose at 37^°C for 24 h. For biofilm formation, 10⁶ cells/mL were grown on silicone disks placed at the bottom of 96-well plates in RPMI-1640 under constant shaking at 37^°C for 48–72 h. BF production was then evaluated by XTT metabolic assay, safranin staining and light microscopy (LM). Documented BF producers (CA-M61 and CP/PA71) were used as positive controls (metabolic activity by XTT assay: 100%). Isolates that a) showed XTT conversion 80% of positive controls, b) stained with safranin and c) produced microscopically a dense network of fungal elements were considered high BF producers. XTT conversion of <80% defined non BF producers, while conversion 80% with inconsistent safranin and LM findings defined low BF producers. PL MICs of VRC and PSC were assessed by CLSI M27-A2 method. BF MICs were determined as the minimum antifungal concentration causing 50% reduction in the metabolic activity of BF as compared to drug-free controls. All isolates were tested in triplicate at 3 different experiments.

Results: A total of 43 bloodstream Candida isolates (29 CA and 14 CP) were examined. BF production was detected in 89% of CA isolates vs. 43% of CP (p < 0.01). Among CA isolates, 72% and 17% were high and low BF producers, respectively; among CP isolates, the high and low BF producers were 36% and 7%. PL MIC90 (mg/L) of VRC was 0.037 and of PSC 0.063 for CA isolates, while they were 0.031 and 0.063 for CP isolates, respectively. BF MIC90 of both VRC and PSC were 1024 mg/L for CA and CP. No correlation was observed between BF-forming capacity and BF azole MIC pattern.

Conclusion: BF-forming capacity is a frequent characteristic especially among CA clinical isolates. Both high and low BF producers exhibited high MIC to azoles, which may account for treatment failures with these agents in BF-related bloodstream infections.