Development Of Antifungal Drugs Research Articles

Modulation of antioxidant defence system in response to berberine in Candida albicans.

Emergence of multidrug resistant species of Candida is evolving, which advocates an urgent need for the development of new therapeutic strategies and antifungal drugs. Activation of antioxidant defence system in Candida albicans is known as forefront mechanism to escape drug toxicity. This study evaluated the role of antioxidant defence genes in the susceptibility to fluconazole in C. albicans and also determined the effect of berberine on growth, antioxidant enzymes and the expression of their genes in C. albicans isolates. Expression of major antioxidant genes was significantly increased in fluconazole-resistant isolates in comparison with the susceptible group. Antifungal susceptibility against berberine showed MIC values ranging from 125 to 500 μg/ml. Berberine treatment caused upregulation of mRNA expression and enzymatic activities of the targeted major antioxidants. Interestingly, C. albicans exhibited efficient antioxidant response at lower concentrations but could not sufficiently alleviate berberine-induced oxidative stress occurring at concentrations greater than 250 μg/ml. Therefore, berberine could serve as a potent Reactive Oxygen Species (ROS)-inducing agent, disrupting the antioxidant system especially in fluconazole-resistant C. albicans to overcome antifungal drug resistance. TAKE AWAYS: Evaluated the role of antioxidant enzymes in FLC resistance in C. albicans Studied the effect of berberine on growth of different C. albicans isolates Investigated the modulation of antioxidant enzymes by berberine in C. albicans Studied the effect of berberine on antioxidant gene expression in C. albicans.

Evaluation of possible biological control of Fusarium sp. using plant extracts and antagonistic species of microbes in vitro

Background: Fusarium species is one of the most devastating fungi responsible for fruit and vegetable crops rot worldwide. The present study was designed to find an ecofriendly control measure for pathogenic Fusarium species, using suitable bioagents. Methods: Medicinal plant extracts were evaluated or their antifungal activities against Fusarium species using the poisoned food method. Antagonistic potency of some nonpathogenic microbes was also assessed on Fusarium species using the dual culture method. Results: Highest inhibitionofgrowth of Fusariumsp. wasobservedwith 68.1% (0.389 mg per 90 mm Petri plate) of mycelia on Coccinia grandis plant leaf extract, in comparison to the control grown with 100.0% (1.22 mg/dish). The highest inhibition of radial growth was observed using Trichoderma viride on Fusarium sp. (46.01% inhibition). Conclusions: The findings of present study would be benevolent for antifungal drug development to control Fusarium sp. causing fruit and vegetable rot.

The fungal-specific subunit i/j of F1FO-ATP synthase stimulates the pathogenicity of Candida albicans independent of oxidative phosphorylation.

This study aims to investigate biological functions of fungal-specific subunit i/j and subunit k of ATP synthase in C. albicans oxidative phosphorylation and virulence potential. Our results revealed that subunit i/j, and not subunit k, is critical for C. albicans pathogenicity.

Novel 2-Aryloxazoline Compounds Exhibit an Inhibitory Effect on Candida spp., Including Antifungal-Resistant Isolates.

Because of the increased resistance to currently available antifungals, fungal infections represent a significant challenge to human health. Herein, we report the synthesis of 2-aryloxazoline derivatives from the reaction between l-threonine and derivatives of salicylic or naphthoic acid. In total, 26 compounds were obtained and tested against species of Candida, Cryptococcus, and Aspergillus. We found that all of the compounds inhibited the growth of Candida species at low concentrations (<0.25 μg/mL) and exhibited reduced hemolytic and cytotoxic activities. Additionally, compounds 4i and 9i were especially effective against antifungal-resistant isolates and the emerging fungus Candida auris. However, the compounds were less active on Cryptococcus and Aspergillus. Because of the improved in vitro antifungal efficacy and attenuated cytotoxicity, these two 2-aryloxazolines obtained from salicylic and naphthoic acid derivatives, respectively, may be considered lead molecules for the development of novel antifungal drugs.

The Transcription Factor SomA Synchronously Regulates Biofilm Formation and Cell Wall Homeostasis in Aspergillus fumigatus.

Polysaccharides are key components of both the fungal cell wall and biofilm matrix. Despite having distinct assembly and regulation pathways, matrix exopolysaccharide and cell wall polysaccharides share common substrates and intermediates in their biosynthetic pathways. It is not clear, however, if the biosynthetic pathways governing the production of these polysaccharides are cooperatively regulated. Here, we demonstrate that cell wall stress promotes production of the exopolysaccharide galactosaminogalactan (GAG)-depend biofilm formation in the major fungal pathogen of humans Aspergillus fumigatus and that the transcription factor SomA plays a crucial role in mediating this process. A core set of SomA target genes were identified by transcriptome sequencing and chromatin immunoprecipitation coupled to sequencing (ChIP-Seq). We identified a novel SomA-binding site in the promoter regions of GAG biosynthetic genes agd3 and ega3, as well as its regulators medA and stuA Strikingly, this SomA-binding site was also found in the upstream regions of genes encoding the cell wall stress sensors, chitin synthases, and β-1,3-glucan synthase. Thus, SomA plays a direct regulation of both GAG and cell wall polysaccharide biosynthesis. Consistent with these findings, SomA is required for the maintenance of normal cell wall architecture and compositions in addition to its function in biofilm development. Moreover, SomA was found to globally regulate glucose uptake and utilization, as well as amino sugar and nucleotide sugar metabolism, which provides precursors for polysaccharide synthesis. Collectively, our work provides insight into fungal adaptive mechanisms in response to cell wall stress where biofilm formation and cell wall homeostasis were synchronously regulated.IMPORTANCE The cell wall is essential for fungal viability and is absent from human hosts; thus, drugs disrupting cell wall biosynthesis have gained more attention. Caspofungin is a member of a new class of clinically approved echinocandin drugs to treat invasive aspergillosis by blocking β-1,3-glucan synthase, thus damaging the fungal cell wall. Here, we demonstrate that caspofungin and other cell wall stressors can induce galactosaminogalactan (GAG)-dependent biofilm formation in the human pathogen Aspergillus fumigatus We further identified SomA as a master transcription factor playing a dual role in both biofilm formation and cell wall homeostasis. SomA plays this dual role by direct binding to a conserved motif upstream of GAG biosynthetic genes and genes involved in cell wall stress sensors, chitin synthases, and β-1,3-glucan synthase. Collectively, these findings reveal a transcriptional control pathway that integrates biofilm formation and cell wall homeostasis and suggest SomA as an attractive target for antifungal drug development.

Emerging roles of Wss1 in the survival of Candida albicans under genotoxic stresses.

This perspective aims to discuss the potential physiological roles and regulation mechanisms of the recently identified Candida albicans Wss1 protease important in DNA-protein crosslink (DPC) tolerance and repair. DPC is a bulky DNA lesion that blocks essential DNA transactions; thus, it poses a significant threat to genome integrity if left unrepaired. Discoveries of Wss1 in Saccharomyces cerevisiae and SPRTN in human as DPC proteases have demonstrated the importance of protease function in DPC repair. Our recent study revealed that Wss1 in C. albicans, an opportunistic pathogen that can cause life-threatening infection in immunocompromised individuals, also promotes DPC tolerance similarly to both S. cerevisiae Wss1 and human SPRTN. However, its molecular mechanism and regulation are still poorly understood. Here, we briefly discuss the recent insights into C. albicans Wss1 based on the information from S. cerevisiae, as well as outline the aspect of this protein that could make it a potential target for antifungal drug development.

Host Defense Peptides as Templates for Antifungal Drug Development.

Current treatment for invasive fungal diseases is limited to three classes of antifungal drugs: azoles, polyenes, and echinocandins. The most recently introduced antifungal class, the echinocandins, was first approved nearly 30 years ago. The limited antifungal drug portfolio is rapidly losing its clinical utility due to the inexorable rise in the incidence of invasive fungal infections and the emergence of multidrug resistant (MDR) fungal pathogens. New antifungal therapeutic agents and novel approaches are desperately needed. Here, we detail attempts to exploit the antifungal and immunoregulatory properties of host defense peptides (HDPs) in the design and evaluation of new antifungal therapeutics and discuss historical limitations and recent advances in this quest.

Antifungal activity of farnesol incorporated in liposomes and associated with fluconazole

Candida infections represent a threat to human health. Candida albicans is the main causative agent of invasive candidiasis, especially in immunosuppressed patients. The emergence of resistant strains has required the development of new therapeutic strategies. In this context, the use of liposomes as drug carrier systems is a promising alternative in drug development. Thus, considering the evidence demonstrating that sesquiterpene farnesol is a bioactive compound with antifungal properties, this study evaluated the activity farnesol-containing liposomes against different Candida strains. The IC50 of farnesol and its liposomal formulation was assessed in vitro using cultures of Candida albicans, Candida tropicalis, and Candida krusei. The Minimum Fungicidal Concentration (MFC) was established by subculture in solid medium. The occurrence of fungal dimorphism was analyzed using optical microscopy. The effects on antifungal resistance to fluconazole were assessed by evaluating the impact of combined therapy on the growth of Candida strains. The characterization of liposomes was carried out considering their vesicular size, polydispersion index, and zeta medium potential, in addition to electron microscopy analysis. Farnesol exerted an antifungal activity that might be associated with the inhibition of fungal dimorphism, especially in Candida albicans. The incorporation of farnesol into liposomes significantly increased its antifungal activity against C. albicans, C. tropicalis, and C. krusei. In addition, liposomal farnesol potentiated the action of fluconazole against C. albicans and C. tropicalis. On the other hand, the association of unconjugated farnesol with fluconazole resulted in antagonistic effects. In conclusion, farnesol-containing liposomes have the potential to be used in antifungal drug development. However, further research is required to investigate how the antifungal properties of farnesol are affected by the interaction with liposomes, contributing to the modulation of antifungal resistance to conventional drugs.

Fungal Infections in Burn Patients.

Background: Fungal burn wound infections are among the most devastating complications in patients who are severely burned. Increasing incidence of burn wound infections caused by fungi led to new challenges in diagnostic and therapeutic approaches. The wide use of broad-spectrum antibiotic agents, an increased prevalence of molds and non-Candida albicans spp., and the variety of available antifungal agents underline the importance of identifying the causative species, to initiate adequate therapy within an adequate timeframe. Methods: Review of the pertinent English and German literature. Results: Fungal burn wound infections go along with a delay of identifying the causative fungus species and can be mistaken for early bacterial burn wound infection. Recently, an increase of uncommon fungal pathogens and fungi resistance against antifungal agents has been reported. Amphotericin B and voriconazole remain the antifungal drugs used most commonly. Conclusions: Adequate therapy remains challenging. Early radical debridement and wound closure play an imperative part, particularly in preventing infections caused by yeasts and molds or any other agent. Prophylactic empiric pharmacologic treatment is reserved for those highly at risk for invasive burn wound infection only. Because of the emergence of drug-resistant fungi, the development of new antifungal drugs is essential for the battle against fungal burn wound infections.

COVID-19-Associated Candidiasis (CAC): An Underestimated Complication in the Absence of Immunological Predispositions?

The recent global pandemic of COVID-19 has predisposed a relatively high number of patients to acute respiratory distress syndrome (ARDS), which carries a risk of developing super-infections. Candida species are major constituents of the human mycobiome and the main cause of invasive fungal infections, with a high mortality rate. Invasive yeast infections (IYIs) are increasingly recognized as s complication of severe COVID-19. Despite the marked immune dysregulation in COVID-19, no prominent defects have been reported in immune cells that are critically required for immunity to Candida. This suggests that relevant clinical factors, including prolonged ICU stays, central venous catheters, and broad-spectrum antibiotic use, may be key factors causing COVID-19 patients to develop IYIs. Although data on the comparative performance of diagnostic tools are often lacking in COVID-19 patients, a combination of serological and molecular techniques may present a promising option for the identification of IYIs. Clinical awareness and screening are needed, as IYIs are difficult to diagnose, particularly in the setting of severe COVID-19. Echinocandins and azoles are the primary antifungal used to treat IYIs, yet the therapeutic failures exerted by multidrug-resistant Candida spp. such as C. auris and C. glabrata call for the development of new antifungal drugs with novel mechanisms of action.

A New Antifungal Agent (4-phenyl-1, 3-thiazol-2-yl) Hydrazine Induces Oxidative Damage in Candida albicans.

A gradual rise in immunocompromised patients over past years has led to the increasing incidence of invasive fungal infections. Development of effective fungicides can not only provide new means for clinical treatment, but also reduce the occurrence of fungal resistance. We identified a new antifungal agent (4-phenyl-1, 3-thiazol-2-yl), hydrazine (numbered as 31C) which showed high-efficiency, broad-spectrum and specific activities. The minimum inhibitory concentration of 31C against pathogenic fungi was between 0.0625-4 μg/ml in vitro, while 31C had no obvious cytotoxicity to human umbilical vein endothelial cells with the concentration of 4 μg/ml. In addition, 31C of 0.5 μg/ml could exhibit significant fungicidal activity and inhibit the biofilm formation of C. albicans. In vivo fungal infection model showed that 31C of 10 mg/kg significantly increased the survival rate of Galleria mellonella. Further study revealed that 31C-treatment increased the reactive oxygen species (ROS) in C. albicans and elevated the expression of some genes related to anti-oxidative stress response, including CAP1, CTA1, TRR1, and SODs. Consistently, 31C-induced high levels of intracellular ROS resulted in considerable DNA damage, which played a critical role in antifungal-induced cellular death. The addition of ROS scavengers, such as glutathione (GSH), N-Acetyl-L-cysteine (NAC) or oligomeric proanthocyanidins (OPC), dramatically reduced the antifungal activities of 31C and rescued the 31C-induced filamentation defect. Collectively, these results showed that 31C exhibited strong antifungal activity and induced obvious oxidative damage, which indicated that compounds with a structure similar to 31C may provide new sight for antifungal drug development.

Precise Expression of Afmed15 Is Crucial for Asexual Development, Virulence, and Survival of Aspergillus fumigatus.

The rise of drug resistance in fungal pathogens is becoming a serious problem owing to the limited number of antifungal drugs available. Identifying and targeting factors essential for virulence or development unique to fungal pathogens is one approach to develop novel treatments for fungal infections. In this study, we present the identification and functional characterization of a novel developmental regulator in Aspergillus fumigatus, AfMed15, which contained a conserved Med15_fungal domain, as determined by screening of a mutant library that contained more than 2,000 hygromycin-resistant A. fumigatus transformants. Downregulating the expression of Afmed15 abolished the conidiation and decreased the fungal virulence in an insect model. Strikingly, the overexpression of Afmed15 caused fungal death accompanied by intensive autophagy. RNA sequencing of an Afmed15 overexpression strain revealed that altered gene expression patterns were associated with carbon metabolism, energy metabolism, and translation. Interestingly, the addition of metal ions could partially rescue fungal death caused by the overexpression of Afmed15, indicating that disordered ion homeostasis is a potential reason for the fungal death caused by the overexpression of Afmed15 Considering that the precise expression of Afmed15 is crucial for fungal development, virulence, and survival and that no ortholog was found in humans, Afmed15 is an ideal target for antifungal-drug development.IMPORTANCE The identification and characterization of regulators essential for virulence or development constitute one approach for antifungal drug development. In this study, we screened and functionally characterized Afmed15, a novel developmental regulator in A. fumigatus We demonstrate that the precise transcriptional expression of Afmed15 is crucial for fungal asexual development, virulence, and survival. Downregulating the expression of Afmed15 abolished the conidiation and decreased the fungal virulence in an insect model. In contrast, the overexpression of Afmed15 caused fungal death accompanied by intensive autophagy. Our study provides a foundation for further studies to identify compounds perturbing the expression of Afmed15 that may be used for the prevention of invasive A. fumigatus infections.

Synthesis, structural characterization, and prospects for new cobalt (II) complexes with thiocarbamoyl-pyrazoline ligands as promising antifungal agents

Candida spp. cause invasive fungal infections. One species, Candida glabrata, may present intrinsic resistance to conventional antifungal agents, thereby increasing mortality rates in hospitalized patients. In this context, metal complexes present an alternative for the development of new antifungal drugs owing to their biological and pharmacological activities demonstrated in studies in the last decades. Accordingly, in this study we have synthesized and characterized two new Co(II) complexes with thiocarbamoyl-pyrazoline ligands to assess their antimicrobial, mutagenic, and cytotoxic potential. For antimicrobial activity, the broth microdilution method was performed against ATCC strains of Candida spp. and fluconazole dose-dependent isolates of C. glabrata obtained from urine samples. The Ames test was used to assess mutagenic potential. The reduction method of the MTS reagent (3 [4,5-dimethylthiazol-2-yl]-5-[3-carboxymethoxyphenyl]-2-[4-sulfophenyl]-2H-tetrazolium) was performed with HeLa, SiHa, and Vero cells to determine cytotoxicity. Both complexes exhibited fungistatic and fungicidal activity for the yeasts used in the study, demonstrating greater potential for C. glabrata ATCC 2001 and the C. glabrata CG66 isolate with a Minimum Inhibitory Concentration MIC from 3.90 to 7.81 μg mL−1 and fungicidal action from 7.81 to 15.62 μg mL−1. The complexes inhibited and degraded biofilms by up to 90% and did not present mutagenic and cytotoxic potential at the concentrations evaluated for MIC. Thus, the complexes examined herein suggest promising alternatives for the development of new antifungal drugs.

Azidothymidine (AZT) Inhibits Proliferation of Human Ovarian Cancer Cells by Regulating Cell Cycle Progression.

Drug resistance is a significant cause of high mortality in ovarian cancer (OC) patients. The reverse transcriptase inhibitor azidothymidine (AZT) has been utilized as a treatment for tumors, but its role in OC treatment has not been revealed. The aim of the present in vitro study was to examine the influence of AZT on the growth of human OC cells and the involved proteins. The proliferation, cell cycle distribution, extent of apoptosis, mitotic index, and terminal restriction fragment length were examined in three OC cell lines, CaOV3, TOV112D, and TOV21G, treated with AZT. AZT inhibited growth of the TOV21G and CaOV3 cell lines by regulating cell cycle distribution. Specifically, AZT caused G2/M phase arrest on TOV21G cells and S phase arrest on CaOV3 cells. In addition, AZT treatment induced up-regulation of p21 and p16 in the TOV21G and CaOV3 cell line, respectively. AZT inhibited cell proliferation in serous and clear cell OC via the regulation of cell cycle distribution.

Antifungal Drug Development: Targeting the Fungal Sphingolipid Pathway.

Fungal infections are becoming more prevalent and problematic due to the continual rise of immune deficient patients as well as the progressive development of drug resistance towards currently available antifungal drugs. There has been a significant increase in the development of antifungal compounds with a similar mechanism of action of current drugs. In contrast, there has been very little progress in developing compounds inhibiting totally new fungal targets or/and fungal pathways. This review focuses on novel compounds recently discovered to target the fungal sphingolipids and their metabolizing enzymes.

The Triazole Ring as a Privileged Scaffold for Putative Antifungals: Synthesis and Evaluation of a Series of New Analogues

The significant antifungal activity of a series of novel 1,2,4-triazole derivatives against different strains of Candida albicans, Candida krusei and Aspergillus fumigatus, compared to the commercial fungicides ketoconazole and itraconazole, is reported. Systemic mycosis and invasive fungal infections, whether from immunodeficiency or hospital-acquired infection, have been on an upward trend for several years. The 1,2,4-triazole ring substituted with other aromatic and heteroaromatic systems plays an important role in the field of antifungal drug discovery and development. Thus, an extensive series of 29 triazoles, substituted in different positions with a variety of aromatic rings, has been designed, synthesized, and evaluated for their fungicidal activity. Almost all the agents tested in vitro showed high activity against all examined fungal strains. It is noteworthy that, in the case of A. fumigatus, all the examined compounds achieved equal or higher antifungal activity than ketoconazole, but less activity than itraconazole. Among all the derivatives studied, the dichlorourea analogue and bromo-substituted triazole stand out as the most promising compounds. Quantitative structure-activity relationship (QSAR) models were built for a systematic structure-activity relationship (SAR) profile to explain and potentially explore the potency characteristics of 1,2,4-triazole analogues.

Antifungal activity of styrylpyridinium compounds against Candida albicans.

We synthesized a set of 13 new and earlier described styrylpyridinium compounds (N-alkyl styrylpyridinium salts with bromide or tosylate anions) in order to evaluate antifungal activity against C.albicans cells, to assay the possible synergism with fluconazole, and to estimate cytotoxicity to mammalian cells. All compounds were synthesized according to a well-known two-step procedure involving alkylation of γ-picoline with appropriate alkyl bromide and further condensation with substituted benzaldehyde. Compounds with long N-alkyl chains (C18 H37 -C20 H41 ) had no antifungal activity against the cells of all tested C.albicans strains. Other styrylpyridinium compounds were able to inhibit yeast growth at the concentrations of 0.06-16μg/ml. At fungicidal concentrations, the compound with the CN- group was least toxic to mammalian cells, showed the most effective synergism with fluconazole, and only slightly inhibited the respiration of C.albicans. The compound with the 4'-diethylamino group exhibited the strongest fungicidal properties and effectively blocked the respiration of C.albicans cells. However, toxicity to mammalian cells was also high. Summarizing, the results of our study indicate that styrylpyridinium compounds are promising candidates in the development of new antifungal drugs.

Current Antimycotics, New Prospects, and Future Approaches to Antifungal Therapy

Fungal infections represent an increasing threat to a growing number of immune- and medically compromised patients. Fungi are eukaryotic organisms and, as such, there is a limited number of selective targets that can be exploited for antifungal drug development. This has also resulted in a very restricted number of antifungal drugs that are clinically available for the treatment of invasive fungal infections at the present time—polyenes, azoles, echinocandins, and flucytosine. Moreover, the utility of available antifungals is limited by toxicity, drug interactions and the emergence of resistance, which contribute to high morbidity and mortality rates. This review will present a brief summary on the landscape of current antifungals and those at different stages of clinical development. We will also briefly touch upon potential new targets and opportunities for novel antifungal strategies to combat the threat of fungal infections.

An Antivirulence Approach for Preventing Cryptococcus neoformans from Crossing the Blood-Brain Barrier via Novel Natural Product Inhibitors of a Fungal Metalloprotease.

Cryptococcus neoformans (Cn) is the leading cause of fungal meningitis, a deadly disease with limited therapeutic options. Dissemination to the central nervous system hinges on the ability of Cn to breach the blood-brain barrier (BBB) and is considered an attribute of Cn virulence. Targeting virulence instead of growth for antifungal drug development has not been fully exploited despite the benefits of this approach. Mpr1 is a secreted fungal metalloprotease not required for fungal growth, but rather, it functions as a virulence factor by facilitating Cn migration across the BBB. This central role for Mpr1, its extracellular location, and lack of expression in mammalian cells make Mpr1 a high-value target for an antivirulence approach aimed at developing therapeutics for cryptococcal meningitis. To test this notion, we devised a large-scale screen to identify compounds that prohibited Cn from crossing the BBB by selectively blocking Mpr1 proteolytic activity, without inhibiting the growth of Cn A phytochemical natural product-derived library was screened to identify new molecular scaffolds of prototypes unique to a Cn microecosystem. Of the 240 pure natural products examined, 3 lead compounds, abietic acid, diosgenin, and lupinine inhibited Mpr1 proteolytic activity with 50% inhibitory concentration (IC50) values of <10 μM, displayed little to no mammalian cell toxicity, and did not affect Cn growth. Notably, the lead compounds blocked Cn from crossing the BBB, without damaging the barrier integrity, suggesting the bioactive molecules had no off-target effects. We propose that these new drug scaffolds are promising candidates for the development of antivirulence therapy against cryptococcal meningitis.IMPORTANCE Fungal infections like cryptococcal meningitis are difficult to resolve because of the limited therapies available. The small arsenal of antifungal drugs reflect the difficulty in finding available targets in fungi because like mammalian cells, fungi are eukaryotes. The limited efficacy, toxicity, and rising resistance of antifungals contribute to the high morbidity and mortality of fungal infections and further underscore the dire but unmet need for new antifungal drugs. The traditional approach in antifungal drug development has been to target fungal growth, but an attractive alternative is to target mechanisms of pathogenesis. An important attribute of Cryptococcus neoformans (Cn) pathogenesis is its ability to enter the central nervous system. Here, we describe a large-scale screen that identified three natural products that prevented Cn from crossing the blood-brain barrier by inhibiting the virulence factor Mpr1 without affecting the growth of Cn We propose that compounds identified here could be further developed as antivirulence therapy that would be administered preemptively or serve as a prophylactic in patients at high risk for developing cryptococcal meningitis.

Antifungal Activity of the Natural Coumarin Scopoletin Against Planktonic Cells and Biofilms From a Multidrug-Resistant Candida tropicalis Strain.

Candida tropicalis is one the most relevant biofilm-forming fungal species increasingly associated with invasive mucosal candidiasis worldwide. The amplified antifungal resistance supports the necessity for more effective and less toxic treatment, including the use of plant-derived natural products. Scopoletin, a natural coumarin, has shown antifungal properties against plant yeast pathogens. However, the antifungal activity of this coumarin against clinically relevant fungal species such as C. tropicalis remains to be established. Here, we investigated the potential antifungal properties and mechanisms of action of scopoletin against a multidrug-resistant C. tropicalis strain (ATCC 28707). First, scopoletin was isolated by high-performance liquid chromatography from Mitracarpus frigidus, a plant species (family Rubiaceae) distributed throughout South America. Next, scopoletin was tested on C. tropicalis cultivated for 48h in both planktonic and biofilm forms. Fungal planktonic growth inhibition was analyzed by evaluating minimal inhibitory concentration (MIC), time-kill kinetics and cell density whereas the mechanisms of action were investigated with nucleotide leakage, efflux pumps and sorbitol and ergosterol bioassays. Finally, the scopoletin ability to affect C. tropicalis biofilms was evaluated through spectrophotometric and whole slide imaging approaches. In all procedures, fluconazole was used as a positive control. MIC values for scopoletin and fluconazole were 50 and 250 μg/L respectively, thus demonstrating a fungistatic activity for scopoletin. Scopoletin induced a significant decrease of C. tropicalis growth curves and cell density (91.7% reduction) compared to the growth control. Its action was related to the fungal cell wall, affecting plasma membrane sterols. When associated with fluconazole, scopoletin led to inhibition of efflux pumps at the plasma membrane. Moreover, scopoletin not only inhibited the growth rate of preformed biofilms (68.2% inhibition at MIC value) but also significantly decreased the extent of biofilms growing on the surface of coverslips, preventing the formation of elongated fungal forms. Our data demonstrate, for the first time, that scopoletin act as an effective antifungal phytocompound against a multidrug-resistant strain of C. tropicalis with properties that affect both planktonic and biofilm forms of this pathogen. Thus, the present findings support additional studies for antifungal drug development based on plant isolated-scopoletin to treat candidiasis caused by C. tropicalis.