Flavones, Flavonols, and Glycosylated Derivatives—Impact on Candida albicans Growth and Virulence, Expression of CDR1 and ERG11, Cytotoxicity

Due to the high incidence of fungal infections worldwide, there is an increasing demand for the development of novel therapeutic approaches. A wide range of natural products has been extensively studied, with considerable focus on flavonoids. The antifungal capacity of selected flavones (luteolin, apigenin), flavonols (quercetin), and their glycosylated derivatives (quercitrin, isoquercitrin, rutin, and apigetrin) along with their impact on genes encoding efflux pumps (CDR1) and ergosterol biosynthesis enzyme (ERG11) has been the subject of this study. Cytotoxicity of flavonoids towards primary liver cells has also been addressed. Luteolin, quercitrin, isoquercitrin, and rutin inhibited growth of Candida albicans with the minimal inhibitory concentration of 37.5 µg/mL. The application of isoquercitrin has reduced C. albicans biofilm establishing capacities for 76%, and hyphal formation by yeast. In vitro treatment with apigenin, apigetrin, and quercitrin has downregulated CDR1. Contrary to rutin and apigenin, isoquercitrin has upregulated ERG11. Except apigetrin and quercitrin (90 µg/mL and 73 µg/mL, respectively inhibited 50% of the net cell growth), the examined flavonoids did not exhibit cytotoxicity. The reduction of both fungal virulence and expression of antifungal resistance-linked genes was the most pronounced for apigenin and apigetrin; these results indicate flavonoids’ indispensable capacity for further development as part of an anticandidal therapy or prevention strategy.

Download Full-text

Cold atmospheric plasma inhibits the growth of Candida albicans by affecting ergosterol biosynthesis and suppresses the fungal virulence factors in vitro

Photodiagnosis and Photodynamic Therapy ◽

10.1016/j.pdpdt.2015.12.007 ◽

2016 ◽

Vol 13 ◽

pp. 66-72 ◽

Cited By ~ 12

Author(s):

Neda Rahimi-Verki ◽

Atena Shapoorzadeh ◽

Mehdi Razzaghi-Abyaneh ◽

Seyed-Mohammad Atyabi ◽

Masoomeh Shams-Ghahfarokhi ◽

...

Keyword(s):

Candida Albicans ◽

Virulence Factors ◽

Atmospheric Plasma ◽

Ergosterol Biosynthesis ◽

Fungal Virulence ◽

Cold Atmospheric Plasma

Download Full-text

Genome-Wide Expression Profiling of the Response to Azole, Polyene, Echinocandin, and Pyrimidine Antifungal Agents in Candida albicans

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.49.6.2226-2236.2005 ◽

2005 ◽

Vol 49 (6) ◽

pp. 2226-2236 ◽

Cited By ~ 217

Author(s):

Teresa T. Liu ◽

Robin E. B. Lee ◽

Katherine S. Barker ◽

Richard E. Lee ◽

Lai Wei ◽

...

Keyword(s):

Candida Albicans ◽

Antifungal Agents ◽

Fungal Infections ◽

Ergosterol Biosynthesis ◽

Reverse Transcription Pcr ◽

Expression Of Genes ◽

Lipid Fatty Acid ◽

Genes Encoding ◽

Molecule Transport ◽

Small Molecule Transport

ABSTRACT Antifungal agents exert their activity through a variety of mechanisms, some of which are poorly understood. We examined changes in the gene expression profile of Candida albicans following exposure to representatives of the four currently available classes of antifungal agents used in the treatment of systemic fungal infections. Ketoconazole exposure increased expression of genes involved in lipid, fatty acid, and sterol metabolism, including NCP1, MCR1, CYB5, ERG2, ERG3, ERG10, ERG25, ERG251, and that encoding the azole target, ERG11. Ketoconazole also increased expression of several genes associated with azole resistance, including CDR1, CDR2, IFD4, DDR48, and RTA3. Amphotericin B produced changes in the expression of genes involved in small-molecule transport (ENA21), and in cell stress (YHB1, CTA1, AOX1, and SOD2). Also observed was decreased expression of genes involved in ergosterol biosynthesis, including ERG3 and ERG11. Caspofungin produced changes in expression of genes encoding cell wall maintenance proteins, including the β-1,3-glucan synthase subunit GSL22, as well as PHR1, ECM21, ECM33, and FEN12. Flucytosine increased the expression of proteins involved in purine and pyrimidine biosynthesis, including YNK1, FUR1, and that encoding its target, CDC21. Real-time reverse transcription-PCR was used to confirm microarray results. Genes responding similarly to two or more drugs were also identified. These data shed new light on the effects of these classes of antifungal agents on C. albicans.

Download Full-text

Efficacy of Novel Schiff base Derivatives as Antifungal Compounds in Combination with Approved Drugs Against Candida Albicans

Medicinal Chemistry ◽

10.2174/1573406415666181203115957 ◽

2019 ◽

Vol 15 (6) ◽

pp. 648-658 ◽

Cited By ~ 1

Author(s):

Manzoor Ahmad Malik ◽

Shabir Ahmad Lone ◽

Parveez Gull ◽

Ovas Ahmad Dar ◽

Mohmmad Younus Wani ◽

...

Keyword(s):

Schiff Base ◽

Candida Albicans ◽

Antifungal Activity ◽

Fungal Infections ◽

Total Sterol ◽

Antifungal Drugs ◽

New Drugs ◽

Ergosterol Biosynthesis ◽

Dependent Manner ◽

Schiff Base Derivatives

Background: The increasing incidence of fungal infections, especially caused by Candida albicans, and their increasing drug resistance has drastically increased in recent years. Therefore, not only new drugs but also alternative treatment strategies are promptly required. Methods: We previously reported on the synergistic interaction of some azole and non-azole compounds with fluconazole for combination antifungal therapy. In this study, we synthesized some non-azole Schiff-base derivatives and evaluated their antifungal activity profile alone and in combination with the most commonly used antifungal drugs- fluconazole (FLC) and amphotericin B (AmB) against four drug susceptible, three FLC resistant and three AmB resistant clinically isolated Candida albicans strains. To further analyze the mechanism of antifungal action of these compounds, we quantified total sterol contents in FLC-susceptible and resistant C. albicans isolates. Results: A pyrimidine ring-containing derivative SB5 showed the most potent antifungal activity against all the tested strains. After combining these compounds with FLC and AmB, 76% combinations were either synergistic or additive while as the rest of the combinations were indifferent. Interestingly, none of the combinations was antagonistic, either with FLC or AmB. Results interpreted from fractional inhibitory concentration index (FICI) and isobolograms revealed 4-10-fold reduction in MIC values for synergistic combinations. These compounds also inhibit ergosterol biosynthesis in a concentration-dependent manner, supported by the results from docking studies. Conclusion: The results of the studies conducted advocate the potential of these compounds as new antifungal drugs. However, further studies are required to understand the other mechanisms and in vivo efficacy and toxicity of these compounds.

Download Full-text

Camphor and Eucalyptol—Anticandidal Spectrum, Antivirulence Effect, Efflux Pumps Interference and Cytotoxicity

International Journal of Molecular Sciences ◽

10.3390/ijms22020483 ◽

2021 ◽

Vol 22 (2) ◽

pp. 483

Author(s):

Marija Ivanov ◽

Abhilash Kannan ◽

Dejan S. Stojković ◽

Jasmina Glamočlija ◽

Ricardo C. Calhelha ◽

...

Keyword(s):

Fungal Pathogens ◽

Efflux Pump ◽

Efflux Pumps ◽

Liver Cells ◽

Ergosterol Biosynthesis ◽

Antifungal Compound ◽

Porcine Liver ◽

Fungal Virulence ◽

Genes Encoding ◽

Plant Constituents

Candidaalbicans represents one of the most common fungal pathogens. Due to its increasing incidence and the poor efficacy of available antifungals, finding novel antifungal molecules is of great importance. Camphor and eucalyptol are bioactive terpenoid plant constituents and their antifungal properties have been explored previously. In this study, we examined their ability to inhibit the growth of different Candida species in suspension and biofilm, to block hyphal transition along with their impact on genes encoding for efflux pumps (CDR1 and CDR2), ergosterol biosynthesis (ERG11), and cytotoxicity to primary liver cells. Camphor showed excellent antifungal activity with a minimal inhibitory concentration of 0.125–0.35 mg/mL while eucalyptol was active in the range of 2–23 mg/mL. The results showed camphor’s potential to reduce fungal virulence traits, that is, biofilm establishment and hyphae formation. On the other hand, camphor and eucalyptol treatments upregulated CDR1;CDR2 was positively regulated after eucalyptol application while camphor downregulated it. Neither had an impact on ERG11 expression. The beneficial antifungal activities of camphor were achieved with an amount that was non-toxic to porcine liver cells, making it a promising antifungal compound for future development. The antifungal concentration of eucalyptol caused cytotoxic effects and increased expression of efflux pump genes, which suggests that it is an unsuitable antifungal candidate.

Download Full-text

Identification of Putative Virulence Genes by DNA Methylation Studies in the Cereal Pathogen Fusarium graminearum

Cells ◽

10.3390/cells10051192 ◽

2021 ◽

Vol 10 (5) ◽

pp. 1192

Author(s):

Francesco Tini ◽

Giovanni Beccari ◽

Gianpiero Marconi ◽

Andrea Porceddu ◽

Micheal Sulyok ◽

...

Keyword(s):

Dna Methylation ◽

Fusarium Graminearum ◽

Environmental Changes ◽

Virulent Strain ◽

Fungal Virulence ◽

Wheat Seedlings ◽

Wide Range ◽

Conidia Production ◽

Natural Hosts

DNA methylation mediates organisms’ adaptations to environmental changes in a wide range of species. We investigated if a such a strategy is also adopted by Fusarium graminearum in regulating virulence toward its natural hosts. A virulent strain of this fungus was consecutively sub-cultured for 50 times (once a week) on potato dextrose agar. To assess the effect of subculturing on virulence, wheat seedlings and heads (cv. A416) were inoculated with subcultures (SC) 1, 23, and 50. SC50 was also used to re-infect (three times) wheat heads (SC50×3) to restore virulence. In vitro conidia production, colonies growth and secondary metabolites production were also determined for SC1, SC23, SC50, and SC50×3. Seedling stem base and head assays revealed a virulence decline of all subcultures, whereas virulence was restored in SC50×3. The same trend was observed in conidia production. The DNA isolated from SC50 and SC50×3 was subject to a methylation content-sensitive enzyme and double-digest, restriction-site-associated DNA technique (ddRAD-MCSeEd). DNA methylation analysis indicated 1024 genes, whose methylation levels changed in response to the inoculation on a healthy host after subculturing. Several of these genes are already known to be involved in virulence by functional analysis. These results demonstrate that the physiological shifts following sub-culturing have an impact on genomic DNA methylation levels and suggest that the ddRAD-MCSeEd approach can be an important tool for detecting genes potentially related to fungal virulence.

Download Full-text

Eap1p, an Adhesin That Mediates Candida albicans Biofilm Formation In Vitro and In Vivo

Eukaryotic Cell ◽

10.1128/ec.00049-07 ◽

2007 ◽

Vol 6 (6) ◽

pp. 931-939 ◽

Cited By ~ 96

Author(s):

Fang Li ◽

Michael J. Svarovsky ◽

Amy J. Karlsson ◽

Joel P. Wagner ◽

Karen Marchillo ◽

...

Keyword(s):

Candida Albicans ◽

Cell Adhesion ◽

Biofilm Formation ◽

Fungal Infections ◽

Gene Dosage ◽

Venous Catheter ◽

Flow Chamber ◽

Dependent Manner

ABSTRACT Candida albicans is the leading cause of systemic fungal infections in immunocompromised humans. The ability to form biofilms on surfaces in the host or on implanted medical devices enhances C. albicans virulence, leading to antimicrobial resistance and providing a reservoir for infection. Biofilm formation is a complex multicellular process consisting of cell adhesion, cell growth, morphogenic switching between yeast form and filamentous states, and quorum sensing. Here we describe the role of the C. albicans EAP1 gene, which encodes a glycosylphosphatidylinositol-anchored, glucan-cross-linked cell wall protein, in adhesion and biofilm formation in vitro and in vivo. Deleting EAP1 reduced cell adhesion to polystyrene and epithelial cells in a gene dosage-dependent manner. Furthermore, EAP1 expression was required for C. albicans biofilm formation in an in vitro parallel plate flow chamber model and in an in vivo rat central venous catheter model. EAP1 expression was upregulated in biofilm-associated cells in vitro and in vivo. Our results illustrate an association between Eap1p-mediated adhesion and biofilm formation in vitro and in vivo.

Download Full-text

In Vitro Incorporation of Helicobacter pylori into Candida albicans Caused by Acidic pH Stress

Pathogens ◽

10.3390/pathogens9060489 ◽

2020 ◽

Vol 9 (6) ◽

pp. 489 ◽

Cited By ~ 1

Author(s):

Kimberly Sánchez-Alonzo ◽

Cristian Parra-Sepúlveda ◽

Samuel Vega ◽

Humberto Bernasconi ◽

Víctor L. Campos ◽

...

Keyword(s):

Helicobacter Pylori ◽

Candida Albicans ◽

Pcr Amplification ◽

Growth Curves ◽

Acidic Ph ◽

Ph Values ◽

16S Gene ◽

Wide Range ◽

H Pylori

Yeasts can adapt to a wide range of pH fluctuations (2 to 10), while Helicobacter pylori, a facultative intracellular bacterium, can adapt to a range from pH 6 to 8. This work analyzed if H. pylori J99 can protect itself from acidic pH by entering into Candida albicans ATCC 90028. Growth curves were determined for H. pylori and C. albicans at pH 3, 4, and 7. Both microorganisms were co-incubated at the same pH values, and the presence of intra-yeast bacteria was evaluated. Intra-yeast bacteria-like bodies were detected using wet mounting, and intra-yeast binding of anti-H. pylori antibodies was detected using immunofluorescence. The presence of the H. pylori rDNA 16S gene in total DNA from yeasts was demonstrated after PCR amplification. H. pylori showed larger death percentages at pH 3 and 4 than at pH 7. On the contrary, the viability of the yeast was not affected by any of the pHs evaluated. H. pylori entered into C. albicans at all the pH values assayed but to a greater extent at unfavorable pH values (pH 3 or 4, p = 0.014 and p = 0.001, respectively). In conclusion, it is possible to suggest that H. pylori can shelter itself within C. albicans under unfavorable pH conditions.

Download Full-text

Spatial Distribution of a Porphyrin-Based Photosensitizer Reveals Mechanism of Photodynamic Inactivation of Candida albicans

Frontiers in Medicine ◽

10.3389/fmed.2021.641244 ◽

2021 ◽

Vol 8 ◽

Author(s):

Thomas Voit ◽

Fabian Cieplik ◽

Johannes Regensburger ◽

Karl-Anton Hiller ◽

Anita Gollmer ◽

...

Keyword(s):

Cell Wall ◽

Candida Albicans ◽

Fungal Infections ◽

Cell Envelope ◽

Antimicrobial Photodynamic Therapy ◽

Fungal Cell ◽

Before And After ◽

Antifungal Action ◽

Complete Cell ◽

Further Development

The antimicrobial photodynamic therapy (aPDT) is a promising approach for the control of microbial and especially fungal infections such as mucosal mycosis. TMPyP [5,10,15, 20-tetrakis(1-methylpyridinium-4-yl)-porphyrin tetra p-toluenesulfonate] is an effective photosensitizer (PS) that is commonly used in aPDT. The aim of this study was to examine the localization of TMPyP in Candida albicans before and after irradiation with visible light to get information about the cellular mechanism of antifungal action of the photodynamic process using this PS. Immediately after incubation of C. albicans with TMPyP, fluorescence microscopy revealed an accumulation of the PS in the cell envelope. After irradiation with blue light the complete cell showed red fluorescence, which indicates, that aPDT is leading to a damage in the cell wall with following influx of PS into the cytosol. Incubation of C. albicans with Wheat Germ Agglutinin (WGA) could confirm the cell wall as primary binding site of TMPyP. The finding that the porphyrin accumulates in the fungal cell wall and does not enter the interior of the cell before irradiation makes it unlikely that resistances can emerge upon aPDT. The results of this study may help in further development and modification of PS in order to increase efficacy against fungal infections such as those caused by C. albicans.

Download Full-text

Systematic characterization and genetic interaction analysis of adhesins in Candida albicans virulence

10.1101/2020.10.22.350991 ◽

2020 ◽

Author(s):

Sierra Rosiana ◽

Liyang Zhang ◽

Grace H. Kim ◽

Alexey V. Revtovich ◽

Arjun Sukumaran ◽

...

Keyword(s):

Candida Albicans ◽

Genetic Interaction ◽

Interaction Analysis ◽

Genetic Interactions ◽

Fungal Virulence ◽

C Elegans ◽

Adhesin Proteins ◽

Insight Into

AbstractCandida albicans is a microbial fungus that exists as a commensal member of the human microbiome and an opportunistic pathogen. Cell surface-associated adhesin proteins play a crucial role in C. albicans’ ability to undergo cellular morphogenesis, develop robust biofilms, colonize, and cause infection in a host. However, a comprehensive analysis of the role and relationships between these adhesins has not been explored. We previously established a CRISPR-based platform for efficient generation of single- and double-gene deletions in C. albicans, which was used to construct a library of 144 mutants, comprising 12 unique adhesin genes deleted singly, or in every possible combination of double deletions. Here, we exploit this adhesin mutant library to explore the role of adhesin proteins in C. albicans virulence. We perform a comprehensive, high-throughput screen of this library, using Caenorhabditis elegans as a simplified model host system, which identified mutants critical for virulence and significant genetic interactions. We perform follow-up analysis to assess the ability of high- and low-virulence strains to undergo cellular morphogenesis and form biofilms in vitro, as well as to colonize the C. elegans host. We further perform genetic interaction analysis to identify novel significant negative genetic interactions between adhesin mutants, whereby combinatorial perturbation of these genes significantly impairs virulence, more than expected based on virulence of the single mutant constituent strains. Together, this yields important new insight into the role of adhesins, singly and in combinations, in mediating diverse facets of virulence of this critical fungal pathogen.SummaryCandida albicans is a human fungal pathogen and cause of life-threatening systemic infections. Cell surface-associated adhesins play a central role in this pathogen’s ability to establish infection. Here, we provide a comprehensive analysis of adhesin factors, and their role in fungal virulence. Exploiting a high-throughput workflow, we screened an adhesin mutant library using C. elegans as a simple model host, and identified mutants and genetic interactions involved in virulence. We found that adhesin mutants are impaired in in vitro pathogenicity, irrespective of their virulence. Together, this work provides new insight into the role of adhesin factors in mediating fungal virulence.

Download Full-text

Synergistic Effect of Calcineurin Inhibitors and Fluconazole against Candida albicans Biofilms

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01397-07 ◽

2008 ◽

Vol 52 (3) ◽

pp. 1127-1132 ◽

Cited By ~ 145

Author(s):

Priya Uppuluri ◽

Jeniel Nett ◽

Joseph Heitman ◽

David Andes

Keyword(s):

Candida Albicans ◽

Calcineurin Inhibitors ◽

Drug Susceptibility ◽

Specific Protein ◽

Therapeutic Interventions ◽

Ergosterol Biosynthesis ◽

Drug Synergism ◽

Planktonic Cells

ABSTRACT Calcineurin is a Ca2+-calmodulin-activated serine/threonine-specific protein phosphatase that governs multiple aspects of fungal physiology, including cation homeostasis, morphogenesis, antifungal drug susceptibility, and virulence. Growth of Candida albicans planktonic cells is sensitive to the calcineurin inhibitors FK506 and cyclosporine A (CsA) in combination with the azole antifungal fluconazole. This drug synergism is attributable to two effects: first, calcineurin inhibitors render fluconazole fungicidal rather than simply fungistatic, and second, membrane perturbation by azole inhibition of ergosterol biosynthesis increases intracellular calcineurin inhibitor concentrations. C. albicans cells in biofilms are up to 1,000-fold more resistant to fluconazole than planktonic cells. In both in vitro experiments and in an in vivo rat catheter model, C. albicans cells in biofilms were resistant to individually delivered fluconazole or calcineurin inhibitors but exquisitely sensitive to the combination of FK506-fluconazole or CsA-fluconazole. C. albicans strains lacking FKBP12 or expressing a dominant FK506-resistant calcineurin mutant subunit (Cnb1-1) formed biofilms that were resistant to FK506-fluconazole but susceptible to CsA-fluconazole, demonstrating that drug synergism is mediated via direct calcineurin inhibition. These findings reveal that calcineurin contributes to fluconazole resistance of biofilms and provide evidence that synergistic drug combinations may prove efficacious as novel therapeutic interventions to treat or prevent biofilms.

Download Full-text