Summary
Propolis, a resinous product of plant origin collected by honeybees, has long been used in traditional medicine for its antimicrobial properties. However, its antifungal mechanism of action against Candida albicans remains incompletely understood. This study aimed to elucidate the antifungal mechanism of ethanolic extracts of Polish propolis (EEP) and a defined mixture of its key flavonoid constituents against C. albicans. Antifungal activity was assessed using Time-kill assay and broth microdilution method under different medium supplementation conditions. Cellular responses were analysed by fluorescence microscopy (FM) and flow cytometry (FCM) in regard to: membrane integrity, reactive oxygen species (ROS) accumulation, mitochondrial membrane potential changes, cytosolic Ca{superscript 2} levels, and morphological transition. Transcriptomic changes were evaluated by RNA sequencing (RNA-seq) and validated by quantitative reverse transcription PCR (RT-qPCR). EEP exhibited concentration-dependent, extract-specific antifungal activity, with selected samples showing rapid fungicidal effects. Mechanistic studies demonstrated membrane permeabilization, reactive oxygen species (ROS) accumulation, mitochondrial dysfunction, disruption of Ca{superscript 2} homeostasis, and inhibition of hyphal formation. Ergosterol supplementation reduced antifungal efficacy, indicating membrane sterols as primary targets. Transcriptomic analysis revealed downregulation of genes associated with DNA replication, transcription, and biosynthesis, alongside upregulation of stress-response pathways, including oxidative stress, protein folding, and mitochondrial processes. Polish propolis exerts antifungal activity through a multi-target mechanism involving membrane disruption and induction of cellular stress. Transcriptomic data indicate coordinated suppression of essential cellular functions and activation of stress-response pathways, supporting a system-level disruption of fungal homeostasis.
Outcomes reported
Propolis, a resinous product of plant origin collected by honeybees, has long been used in traditional medicine for its antimicrobial properties. However, its antifungal mechanism of action against Candida albicans remains incompletely understood. This study aimed to elucidate the antifungal mechanism of ethanolic extracts of Polish propolis (EEP) and a defined mixture of its key flavonoid constituents against C. albicans. Antifungal activity was assessed using Time-kill assay and broth microdilution method under different medium supplementation conditions. Cellular responses were analysed by fluorescence microscopy (FM) and flow cytometry (FCM) in regard to: membrane integrity, reactive oxygen species (ROS) accumulation, mitochondrial membrane potential changes, cytosolic Ca{superscript 2} levels, and morphological transition. Transcriptomic changes were evaluated by RNA sequencing (RNA-seq) and validated by quantitative reverse transcription PCR (RT-qPCR). EEP exhibited concentration-dependent, extract-specific antifungal activity, with selected samples showing rapid fungicidal effects. Mechanistic studies demonstrated membrane permeabilization, reactive oxygen species (ROS) accumulation, mitochondrial dysfunction, disruption of Ca{superscript 2} homeostasis, and inhibition of hyphal formation. Ergosterol supplementation reduced antifungal efficacy, indicating membrane sterols as primary targets. Transcriptomic analysis revealed downregulation of genes associated with DNA replication, transcription, and biosynthesis, alongside upregulation of stress-response pathways, including oxidative stress, protein folding, and mitochondrial processes. Polish propolis exerts antifungal activity through a multi-target mechanism involving membrane disruption and induction of cellular stress. Transcriptomic data indicate coordinated suppression of essential cellular functions and activation of stress-response pathways, supporting a system-level disruption of fungal homeostasis.
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