Antifungal and antibiofilm activities of flavonoids against Candida albicans: Focus on 3,2'-dihydroxyflavone as a potential therapeutic agent

Abstract

Effective management of microbial biofilms holds significance within food and medical environments. Candida albicans, an opportunistic fungus, forms mucosal biofilms closely linked to candidiasis and drug-resistant infections due to their drug tolerance. Morphologic change from yeast to filamentous cells is a key virulence factor and a prerequisite for biofilm development. This study investigated the anti-fungal and antibiofilm activities of 20 flavonoids against C. albicans. With their known antioxidant capabilities, flavonoids hold promise in combating infections associated with biofilms. Among them, flavone and its derivatives exhibited moderate antifungal activity, 3,2'-dihydroxyflavone (3,2'-DHF) at 1 μg/mL exhibited strong antibiofilm activity (MIC 50 μg/mL). In addition, 3,2'-DHF dramatically inhibited cell aggregation and germ tube/hyphae formation. Transcriptomic analyses revealed that flavone and 3,2'-DHF behaved differently, as 3,2'-DHF downregulated the expressions of germ tube/hyphae-forming and biofilm-related genes (ECE1, HWP1, TEC1, and UME6) but upregulated the biofilm/hyphal regulators (CHK1, IFD6, UCF1, and YWP1). Tests evaluating toxicity with plant and nematode models revealed that flavone and 3,2'-DHF exhibited mild toxicity. Current results indicate that hydroxylated flavone derivatives can enhance anti-fungal and antibiofilm activities and provide a source of potential anti-fungal agents against drug-resistant C. albicans.

Keywords: Anti-fungal; Biofilm; Candida albicans; Flavone; Flavonoids; Hyphae.

Fig. 1

Antibiofilm activity of flavonoids. Structures of the flavonoids studied (A). Biofilm formation by C. albicans DAY185 in the presence of flavonoids at 10 or 50 μg/mL (B). Error bars indicate standard deviations. *, P < 0.05 vs. nontreated controls (None).

Fig. 2

Antibiofilm activities of 3,2′-dihydroxyflavone (3,2′-DHF) and flavone. Antibiofilm activities of 3,2′-DHF (A) and flavone (B) and effects of 3,2′-DHF (C) and flavone (D) on the metabolic activity of C. albicans DAY185 via XTT reduction assay. 3-Dimensional biofilm images of C. albicans DAY185 in the presence of 3,2′-DHF or flavone (E). Black scale bar represents 100 μm *, P < 0.05 vs. nontreated controls (None).

Fig. 3

Inhibitory effects of 3,2′-dihydroxyflavone (3,2′-DHF) and flavone on germ tube/hyphal development and cell aggregation by C. albicans DAY185. Germ tube/hyphae formation (A) and cell aggregation (B) using live cell imaging microscopy. Images were captured at 10 to 40× magnifications. Yellow and white bars represent 50 and 200 μm, respectively. Blue triangles indicate germ tube/hyphae formation. Nontreated controls (None).

Fig. 4

Inhibitory effects of 3,2′-dihydroxyflavone (3,2′-DHF) and flavone on hyphal growth and biofilm formation by C. albicans DAY185 as determined by SEM. The yellow scale bar represents 50 μm. Nontreated controls (None).

Fig. 5

Differential gene expressions and mechanisms responsible for the effects of 3,2′-dihydroxyflavone (3,2′-DHF) and flavone on C. albicans DAY185. Fold changes represent the transcriptional variances identified in cells subjected to treatment compared to those untreated (None), as determined through qRT-PCR. C. albicans cells were treated with 3,2′-DHF or flavone at 10 μg/mL for 6 h without shaking (A). RDN18 was used as the housekeeping gene. *, p < 0.05 vs. nontreated controls. Potential mechanism underlying the effects of 3,2′-DHF and flavone in C. albicans (B).

Fig. 6

Chemical toxicities of 3,2′-dihydroxyflavone (3,2′-DHF) and flavone. B. rapa seed germination assay was performed with or without 3,2′-DHF or flavone at 25 °C for 2 days (A). Plant total lengths were measured on days 1, 2, 3, 4, and 5 (B and C), and C. elegans survival was assessed in the presence or absence of 3,2′-DHF or flavone for 10 days (D). *, p < 0.05 vs. nontreated controls. Nontreated controls (None).