Innovative antifungal strategies: enhanced biofilm inhibition of Candida albicans by a modified tea tree oil formulation

Abstract

Introduction: Candida albicans is a significant human pathogen with the ability to form biofilms, a critical factor in its resistance to antifungal treatments. This study aims to evaluate the antifungal activity and biofilm inhibition potential of Tea Tree Oil (TTO) derived from Melaleuca alternifolia cultivated in Vietnam.

Methods: The antifungal activity of TTO was assessed by determining the Minimum Inhibitory Concentration (MIC), Minimum Fungicidal Concentration (MFC), Minimum Biofilm Inhibitory Concentration (MBIC), and Minimum Biofilm Eradication Concentration (MBEC) using broth dilution methods. The experiments were conducted on C. albicans in both planktonic and biofilm states across concentrations ranging from 0.1 μL/mL to 10 μL/mL.

Results: TTO demonstrated significant antifungal efficacy, with a MIC of 0.1 μL/mL (∼91.217 μg/mL) and an MFC of 10 μL/mL (∼9121.7 μg/mL). It effectively inhibited biofilm formation with a recorded MBIC of 2 μL/mL (∼1824.34 μg/mL). However, MBEC values were not determinable as the concentrations tested did not achieve the eradication of more than 50% of mature biofilm within the experimental conditions.

Discussion: These findings highlight TTO as a promising natural antifungal agent with strong biofilm-inhibitory properties. However, its limited efficacy in eradicating mature biofilms underscores the need for further studies, potentially involving higher concentrations or synergistic combinations with conventional antifungal agents.

Keywords: Candida albicans; Minimum Biofilm Inhibitory Concentration (MBIC); Minimum Fungicidal Concentration (MFC); antifungal; biofilm; biofilm inhibition; modified tea tree oil formulation; tea tree oil.

FIGURE 1

Microscopic morphology of Candida albicans using a simple staining method.

FIGURE 2

Antifungal activity against Candida albicans of the antifungal agent AMB and TTO at various test concentrations. (A) MFC results are presented as follows: Images labeled a), b), c), d), e) show samples treated with the antifungal agent AMB at concentrations of 0.1, 1, 2, 5, and 10 μg/mL, respectively; and f), g), h), i), k) display samples treated with TTO at concentrations of 0.1, 1, 2, 5, and 10 μL/mL (equivalent to approximately 91.217, 912.17, 1824.34, 4560.85, and 9121.7 μg/mL, respectively) respectively. (B) Graphical representation of concentration thresholds inhibiting fungal growth.

FIGURE 3

Scanning Electron Microscopy (SEM) image of Candida albicans biofilm in the experimental condition of Hansen medium + 2.5% glucose. The marked region (green circle) highlights the biofilm formation.

FIGURE 4

Suppression of Candida albicans biofilm formation by AMB and TTO at different investigated concentrations (following MBIC). For AMB, statistically significant differences (p < 0.05) compared to the control included concentrations (μg/mL) of 0.1, 1, 2, 5, and 10; For TTO, statistically significant differences (p < 0.05) included concentrations (μL/mL) of 2, 5, and 10 (equivalent to approximately 1824.34, 4560.85, and 9121.7 μg/mL, respectively).

FIGURE 5

Eradication of Candida albicans biofilm formation by AMB and TTO at different test concentrations (following MBEC). For AMB, the statistically significant difference (p < 0.05) compared to the control had a threshold of only 10; For TTO, all concentrations were not statistically significant (p > 0.05).