A viability assay for Candida albicans based on the electron transfer mediator 2,6-dichlorophenolindophenol

Area of Science:

• Mycology
• Biochemistry
• Microbiology

Background:

• Candida albicans is an opportunistic fungal pathogen known for high respiratory activity.
• Existing viability assays may not effectively differentiate between fungal species with varying respiratory pathways.
• 2,6-dichlorophenolindophenol (DCIP) is a membrane-permeable electron transfer agent used in enzymatic assays.

Purpose of the Study:

• To establish a DCIP-based viability test for Candida albicans.
• To investigate the role of complex I in DCIP reduction by C. albicans.
• To differentiate complex I-negative yeasts from complex I-positive yeasts using DCIP assays.

Main Methods:

• Established a viability test utilizing 2,6-dichlorophenolindophenol (DCIP) and NADH dehydrogenases.
• Employed specific respiratory chain inhibitors, including rotenone (complex I inhibitor).
• Conducted comparative studies with complex I-negative yeasts Saccharomyces cerevisiae and Candida glabrata.

Main Results:

• DCIP reduction in C. albicans was significantly inhibited by rotenone, indicating strong dependence on complex I activity (approx. 30% residual activity).
• Complex I-negative yeasts (S. cerevisiae, C. glabrata) exhibited lower basal DCIP turnover rates compared to C. albicans.
• Glucose supplementation increased DCIP turnover in complex I-negative yeasts to levels comparable to C. albicans, while galactose did not.

Conclusions:

• The DCIP viability assay is largely dependent on complex I activity in Candida albicans.
• Optimized assay conditions allow for the distinction between complex I-positive and complex I-negative yeasts.
• Metabolic responses to carbon sources like glucose can be used to differentiate yeast species in DCIP assays.