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Updated: Jun 18, 2025
A Soluble Tetrazolium-Based Reduction Assay to Evaluate the Effect of Antibodies on Candida tropicalis Biofilms
Published on: September 16, 2022
Screening of the global health priority BoxⓇ reveals potential new disinfectants against the emerging
Yasmim Isabel Retore1, Fabíola Lucini1, Larissa Rodrigues Pimentel1
1Health Sciences Research Laboratory, Federal University of Grande Dourados, Dourados, Mato Grosso do Sul, Brazil.
Background:
Candida auris has been identified by the World Health Organization as a critical pathogen due to its invasive nature, resistance to multiple drugs, and high mortality rates in hospital outbreaks. This fungus can persist on surfaces and human skin for extended periods, complicating infection control efforts. The need for effective disinfection strategies is urgent, as current disinfectants are often ineffective against C. auris biofilms.
Objective:
The study aimed to identify potential disinfectants from a collection of 240 compounds in the Global Health Priority Box® that are effective against C. auris, particularly strains resistant to existing options.
Methods:
The research employed a screening protocol using a fluconazole-resistant strain of C. auris (149/23). Antifungal activity was assessed using the microdilution method to determine Minimum Inhibitory Concentrations (MICs) and Minimum Fungicidal Concentrations (MFCs). Additional assays were conducted to evaluate biofilm inhibition, biofilm eradication, cell membrane integrity, nucleotide leakage, sorbitol protection assay, efflux pump inhibition, and hemolysis assay.
Results:
Two compounds, Hydramethylnon (MMV1577471) and Flufenerim (MMV1794206), demonstrated significant inhibitory effects against C. auris. Hydramethylnon exhibited potent antifungal activity, inhibiting up to 93 % of fungal growth with an MFC of 16 μg/mL. Flufenerim inhibited up to 58 % of fungal growth, showing fungistatic action with an MFC greater than 4 μg/mL. Biofilm inhibition tests showed that both compounds significantly inhibited biofilm formation, with increased efficacy at higher concentrations. Both compounds showed eradication rates in both stages. Furthermore, Hydramethylnon and Flufenerim did not affect cell membrane integrity or nucleotide leakage, suggesting a mode of action not reliant on disrupting these cellular components. The sorbitol protection assay revealed that neither compound caused cell wall damage. In the efflux pump inhibition assay, Hydramethylnon did not activate efflux pumps, while Flufenerim activated efflux pumps, reducing its effectiveness. Hemocompatibility assay showed safety.
Conclusion:
The study highlights Hydramethylnon and Flufenerim as promising candidates for further development as disinfectants, offering potential solutions to the urgent need for effective disinfection agents against C. auris. The findings underscore the value of screening compound collections to identify novel antifungal agents and understand their mechanisms of action, thereby contributing to the advancement of new disinfection strategies in healthcare settings.
Insights
Two novel compounds, Hydramethylnon and Flufenerim, show promise in combating the critical pathogen Candida auris. These disinfectants effectively inhibit fungal growth and biofilms, offering new hope for infection control in healthcare settings.
Area of Science:
- Medical Mycology
- Infectious Diseases
- Antimicrobial Discovery
Background:
- Candida auris is a critical global pathogen identified by the WHO, known for its drug resistance and high mortality.
- Its persistence on surfaces and skin complicates infection control, necessitating novel disinfection strategies.
- Existing disinfectants are often ineffective against Candida auris biofilms, highlighting an urgent need for new solutions.
Purpose of the Study:
- To screen a library of 240 compounds for efficacy against Candida auris.
- To identify potential disinfectants effective against drug-resistant strains of C. auris.
- To evaluate the mechanism of action of promising antifungal compounds.
Main Methods:
- A fluconazole-resistant strain of C. auris was used for screening.
- Minimum Inhibitory Concentrations (MICs) and Minimum Fungicidal Concentrations (MFCs) were determined.
- Assays included biofilm inhibition/eradication, cell membrane integrity, nucleotide leakage, cell wall damage, efflux pump activity, and hemocompatibility.
Main Results:
- Hydramethylnon and Flufenerim demonstrated significant antifungal activity against C. auris.
- Hydramethylnon inhibited up to 93% of fungal growth (MFC: 16 μg/mL), while Flufenerim inhibited up to 58% (MFC > 4 μg/mL).
- Both compounds inhibited and eradicated biofilms; Hydramethylnon showed no adverse effects on cell integrity or wall, while Flufenerim activated efflux pumps.
Conclusions:
- Hydramethylnon and Flufenerim are promising candidates for developing new disinfectants against C. auris.
- These findings support the screening of compound libraries for novel antifungal agents.
- The study contributes to advancing disinfection strategies for healthcare settings to combat C. auris.
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