Nitrogen concentration affects amphotericin B and fluconazole tolerance of pathogenic cryptococci

Area of Science:

• Mycology
• Antimicrobial Resistance
• Environmental Microbiology

Background:

• Pathogenic cryptococci exhibit phenotypic plasticity in response to environmental stressors.
• Environmental factors can alter cryptococcal antifungal susceptibility, capsule, melanin, and ergosterol levels.

Purpose of the Study:

• To investigate the impact of nitrogen limitation on cryptococcal virulence factors and antifungal drug susceptibility.
• To determine how varying nitrogen concentrations affect capsule, melanin, and ergosterol biosynthesis in Cryptococcus neoformans and Cryptococcus gattii.

Main Methods:

• Cryptococcal strains were cultured in media with varying ammonium chloride (NH4Cl) concentrations (high, control, low).
• Capsule thickness was measured using India ink staining.
• Melanin and ergosterol content were quantified spectrophotometrically.
• Antifungal susceptibility was assessed using time-kill assays and fluconazole (FLU) heteroresistance testing against amphotericin B (AmB) and FLU.

Main Results:

• Low nitrogen concentrations significantly enhanced ergosterol and capsule biosynthesis.
• Melanin formation showed a variable response to nitrogen levels.
• Cultures grown in low nitrogen conditions exhibited increased survival rates against both AmB and FLU.
• A higher frequency of FLU heteroresistance was observed in low nitrogen cultures.

Conclusions:

• Nitrogen availability is a critical environmental factor influencing cryptococcal ergosterol and capsule production.
• Reduced nitrogen levels can lead to increased tolerance to key antifungal medications, including amphotericin B and fluconazole.
• Environmental nitrogen concentration may play a significant role in the development of antifungal drug tolerance in pathogenic cryptococci.