In vitro and in vivo Effect of Exogenous Farnesol Exposure Against Candida auris

Abstract

The spreading of multidrug-resistant Candida auris is considered as an emerging global health threat. The number of effective therapeutic regimens is strongly limited; therefore, development of novel strategies is needed. Farnesol is a quorum-sensing molecule with a potential antifungal and/or adjuvant effect; it may be a promising candidate in alternative treatment against Candida species including C. auris. To examine the effect of farnesol on C. auris, we performed experiments focusing on growth, biofilm production ability, production of enzymes related to oxidative stress, triazole susceptibility and virulence. Concentrations ranging from 100 to 300 μM farnesol caused a significant growth inhibition against C. auris planktonic cells for 24 h (p < 0.01-0.05). Farnesol treatment showed a concentration dependent inhibition in terms of biofilm forming ability of C. auris; however, it did not inhibit significantly the biofilm development at 24 h. Nevertheless, the metabolic activity of adhered farnesol pre-exposed cells (75 μM) was significantly diminished at 24 h depending on farnesol treatment during biofilm formation (p < 0.001-0.05). Moreover, 300 μM farnesol exerted a marked decrease in metabolic activity against one-day-old biofilms between 2 and 24 h (p < 0.001). Farnesol increased the production of reactive species remarkably, as revealed by 2',7'-dichlorofluorescein (DCF) assay {3.96 ± 0.89 [nmol DCF (OD₆₄₀)^-1] and 23.54 ± 4.51 [nmol DCF (OD₆₄₀)^-1] for untreated cells and farnesol exposed cells, respectively; p < 0.001}. This was in line with increased superoxide dismutase level {85.69 ± 5.42 [munit (mg protein)^-1] and 170.11 ± 17.37 [munit (mg protein)^-1] for untreated cells and farnesol exposed cells, respectively; p < 0.001}, but the catalase level remained statistically comparable between treated and untreated cells (p > 0.05). Concerning virulence-related enzymes, exposure to 75 μM farnesol did not influence phospholipase or aspartic proteinase activity (p > 0.05). The interaction between fluconazole, itraconazole, voriconazole, posaconazole, isavuconazole and farnesol showed clear synergism (FICI ranges from 0.038 to 0.375) against one-day-old biofilms. Regarding in vivo experiments, daily 75 μM farnesol treatment decreased the fungal burden in an immunocompromised murine model of disseminated candidiasis, especially in case of inocula pre-exposed to farnesol (p < 0.01). In summary, farnesol shows a promising therapeutic or adjuvant potential in traditional or alternative therapies such as catheter lock therapy.

Keywords: biofilm; in vivo; oxidative stress; quorum-sensing; synergy; therapy; triazoles; virulence.

FIGURE 1

Time-kill curves of farnesol against Candida auris (A,B) and Candida albicans (C,D) isolates in RPMI-1640 for farnesol unexposed (A,C) and farnesol pre-exposed (B,D) cells (75 μM), respectively. Each timepoint represents mean ± SEM (standard error of mean) of cell count derived from isolates.

FIGURE 2

Metabolic activity changes over time in case of biofilm formation in the presence of given farnesol concentrations (10–300 μM) for C. auris (A) and C. albicans (D), respectively. Metabolic activity changes over time in case of biofilm formation by farnesol pre-exposed cells (75 μM) in the presence of given farnesol concentrations (10–300 μM) for C. auris (B) and C. albicans (E), respectively. Metabolic activity changes over time for one-day-old preformed biofilms in the presence of given farnesol concentrations (10–300 μM) for C. auris (C) and C. albicans (F), respectively. Each time-point represents mean ± SEM (standard error of mean) of metabolic activity of clinical isolates (three independent experiments per isolate).

FIGURE 3

The kidney burden of Candida auris in a systemically infected mouse model. The bars represent the means ± SEM (standard error of mean) of kidney tissue burdens of BALB/c mice. Significant differences between CFU numbers were determined based on comparison with the untreated controls. Levels of significant differences are indicated (**p < 0.01). Histological changes in kidney tissue from mice suffering from systemic candidiasis with or without farnesol treatment in the presence or absence of farnesol pre-exposure were examined by Periodic acid-Schiff staining.

FIGURE 4

The kidney burden of Candida albicans in a systemically infected mouse model. The bars represent the means ± SEM (standard error of mean) of kidney tissue burdens of BALB/c mice. Significant differences between CFU numbers were determined based on comparison with the untreated controls. Histological changes in kidney tissue from mice suffering from systemic candidiasis with or without farnesol treatment in the presence or absence of farnesol pre-exposure were examined by Periodic acid-Schiff staining.