scholarly journals Candida albicans rvs161Δ and rvs167Δ Endocytosis Mutants Are Defective in Invasion into the Oral Cavity

mBio ◽  
2019 ◽  
Vol 10 (6) ◽  
Author(s):  
Shamoon Naseem ◽  
Lois M. Douglas ◽  
James B. Konopka
Keyword(s):  
Candida Albicans ◽  
Environmental Factors ◽  
Oral Cavity ◽  
Fungal Infections ◽  
Hyphal Growth ◽  
Antifungal Drugs ◽  
Invasive Growth ◽  
Oropharyngeal Candidiasis ◽  
Content Type ◽  
Therapeutic Value

ABSTRACT Invasive growth in tissues by the human fungal pathogen Candida albicans is promoted by a switch from budding to hyphal morphogenesis that is stimulated by multiple environmental factors that can vary at different sites of infection. To identify genes that promote invasive growth in the oral cavity to cause oropharyngeal candidiasis (OPC), we first identified C. albicans mutants that failed to invade agar medium. Analysis of nine severely defective mutants in a mouse model of OPC revealed that the strongest defects were seen for the rvs161Δ and rvs167Δ mutants, which lack amphiphysin proteins needed for endocytosis. The rvsΔ mutants initially adhered to the tongue but failed to invade efficiently and were lost from the oral cavity. Previous studies indicated that rvsΔ mutants formed filamentous hyphae in the kidney albeit with morphological abnormalities, suggesting that the rvsΔ mutants were influenced by factors that vary at different sites of infection. Consistent with this, increasing concentrations of CO2, an inducer of hyphal growth that is more abundant in internal organs than air, partially rescued the invasive-growth defects of the rvsΔ mutants in vitro. Interestingly, preinduction of the rvsΔ mutants to form hyphae prior to introduction into the oral cavity restored their ability to cause OPC, identifying a key role for endocytosis in initiating invasive hyphal growth. These results highlight the influence of distinct environmental factors in promoting invasive hyphal growth in the oral cavity and indicate that blocking endocytosis could have therapeutic value in preventing the initiation of OPC. IMPORTANCE Oropharyngeal candidiasis (OPC) is a common fungal infection that is associated with severe morbidity. Another concern is that patients at risk for developing OPC often take long courses of antifungal drugs, which can lead to the emergence of drug-resistant C. albicans strains. We therefore identified nine mutants with defects in undergoing invasive hyphal growth in the oral cavity, increasing the number of genes known to be involved in OPC by more than 30%. The two strongest mutants, rvs161Δ and rvs167Δ, have defects in endocytosis. The rvsΔ mutants appear to have a specific defect in initiating invasive growth, as preinducing the cells to form hyphae prior to infection restored their ability to cause OPC. These results indicate that blocking endocytosis could have therapeutic value in preventing the initiation of OPC without leading to development of resistance against drugs currently used to treat fungal infections.

scholarly journals Candida albicans hyphal expansion causes phagosomal membrane damage and luminal alkalinization

2018 ◽  
Author(s):  
Johannes Westman ◽  
Gary Moran ◽  
Selene Mogavero ◽  
Bernhard Hube ◽  
Sergio Grinstein
Keyword(s):  
Candida Albicans ◽  
Fungal Infections ◽  
Membrane Damage ◽  
Hyphal Growth ◽  
Antifungal Drugs ◽  
Apparent Paradox ◽  
Opportunistic Fungi ◽  
Life Threatening ◽  
Proton Leakage ◽  
Phagosomal Membrane

ABSTRACTMacrophages rely on phagosomal acidity to destroy engulfed microorganisms. To survive this hostile response, opportunistic fungi such as Candida albicans developed strategies to evade the acidic environment. C. albicans is polymorphic, able to convert from yeast to hyphae, and this transition is required to subvert the microbicidal activity of the phagosome. However, the phagosomal lumen, which is acidic and nutrient-deprived, inhibits yeast-to-hypha transition. To account for this apparent paradox, it was recently proposed that C. albicans produces ammonia that alkalinizes the phagosome, thus facilitating yeast-to-hypha transformation. We re-examined the mechanism underlying phagosomal alkalinization by applying dual-wavelength ratiometric pH measurements. The phagosomal membrane was found to be highly permeable to ammonia, which is therefore unlikely to account for the pH elevation. Instead, we find that yeast-to-hypha transition begins within acidic phagosomes, and that alkalinization is a consequence of proton leakage induced by excessive membrane distension caused by the expanding hypha.IMPORTANCEC. albicans is the most common nosocomial fungal infection, and over three million people acquire life-threatening invasive fungal infections every year. Even if antifungal drugs exist, almost half of these patients will die. Despite this, fungi remain underestimated as pathogens. Our study uses quantitative biophysical approaches to demonstrate that the yeast-to-hypha transition occurs within the nutrient deprived, acidic phagosome and that alkalinization is a consequence, as opposed to the cause of hyphal growth.

scholarly journals Genetic Analysis of Sirtuin Deacetylases in Hyphal Growth of Candida albicans

mSphere ◽  
2021 ◽  
Vol 6 (3) ◽  
Author(s):  
Guolei Zhao ◽  
Laura N. Rusche
Keyword(s):  
Candida Albicans ◽  
Environmental Changes ◽  
Fungal Infections ◽  
Hyphal Growth ◽  
Morphological Plasticity ◽  
The United States ◽  
Environmental Cues ◽  
Growth Forms ◽  
Content Type ◽  
Hypha Formation

ABSTRACT Candida albicans is a major human fungal pathogen that encounters varied host environments during infection. In response to environmental cues, C. albicans switches between ovoid yeast and elongated hyphal growth forms, and this morphological plasticity contributes to virulence. Environmental changes that alter the cell’s metabolic state could be sensed by sirtuins, which are NAD+-dependent deacetylases. Here, we studied the roles of three sirtuin deacetylases—Sir2, Hst1, and Hst2—in the hyphal growth of C. albicans. We made single, double, and triple sirtuin knockout strains and tested their ability to switch from yeast to hyphae. We found that true hypha formation was significantly reduced by the deletion of SIR2 but not HST1 or HST2. Moreover, the expression of hypha-specific genes HWP1, ALS3, and ECE1 decreased in the sir2Δ/Δ mutant compared to the wild type. This regulation of hypha formation was likely dependent on the deacetylase activity of Sir2, as a similar defect in hypha formation was observed when an asparagine known to be required for deacetylation was mutated. Finally, we found that Sir2 and Hst1 were localized to the nucleus, with Sir2 specifically focused in the nucleolus. This nuclear localization suggests a role for Sir2 and Hst1 in regulating gene expression. In contrast, Hst2 was localized to the cytoplasm. In conclusion, our results suggest that Sir2 plays a critical and nonredundant role in hyphal growth of C. albicans. IMPORTANCE Candida albicans is one of the most common causes of hospital-acquired systemic fungal infections in the United States. It can switch between ovoid yeast and elongated hyphal growth forms in response to environmental cues. This morphological transition is essential for its survival in the host. Thus, identifying regulators involved in this process can lead to new therapies. In this study, we examined the contribution of three regulators called sirtuins (Sir2, Hst1, and Hst2) to the yeast-to-hypha transition of C. albicans. We found that loss of Sir2 but not Hst1 or Hst2 hampered hypha formation. Moreover, the defect was caused by the loss of the catalytic activity of Sir2. Our study may lay the groundwork for discovering novel targets for antifungal therapies.

scholarly journals In Situ Imaging of Candida albicans Hyphal Growth via Atomic Force Microscopy

mSphere ◽  
2020 ◽  
Vol 5 (6) ◽  
Author(s):  
Arzu Çolak ◽  
Mélanie A. C. Ikeh ◽  
Clarissa J. Nobile ◽  
Mehmet Z. Baykara
Keyword(s):  
Atomic Force Microscopy ◽  
Candida Albicans ◽  
Adhesion Force ◽  
Hyphal Growth ◽  
Antifungal Drugs ◽  
Content Type ◽  
Force Microscopy ◽  
Atomic Force ◽  
Host Tissues

ABSTRACT Candida albicans is an opportunistic fungal pathogen of humans known for its ability to cause a wide range of infections. One major virulence factor of C. albicans is its ability to form hyphae that can invade host tissues and cause disseminated infections. Here, we introduce a method based on atomic force microscopy to investigate C. albicans hyphae in situ on silicone elastomer substrates, focusing on the effects of temperature and antifungal drugs. Hyphal growth rates differ significantly for measurements performed at different physiologically relevant temperatures. Furthermore, it is found that fluconazole is more effective than caspofungin in suppressing hyphal growth. We also investigate the effects of antifungal drugs on the mechanical properties of hyphal cells. An increase in Young’s modulus and a decrease in adhesion force are observed in hyphal cells subjected to caspofungin treatment. Young’s moduli are not significantly affected following treatment with fluconazole; the adhesion force, however, increases. Overall, our results provide a direct means of observing the effects of environmental factors and antifungal drugs on C. albicans hyphal growth and mechanics with high spatial resolution. IMPORTANCE Candida albicans is one of the most common pathogens of humans. One important virulence factor of C. albicans is its ability to form elongated hyphae that can invade host tissues and cause disseminated infections. Here, we show the effect of different physiologically relevant temperatures and common antifungal drugs on the growth and mechanical properties of C. albicans hyphae using atomic force microscopy. We demonstrate that minor temperature fluctuations within the normal range can have profound effects on hyphal cell growth and that different antifungal drugs impact hyphal cell stiffness and adhesion in different ways.

scholarly journals CO2 Signaling through the Ptc2-Ssn3 Axis Governs Sustained Hyphal Development of Candida albicans by Reducing Ume6 Phosphorylation and Degradation

mBio ◽  
2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Yang Lu ◽  
Chang Su ◽  
Shatarupa Ray ◽  
Yuncong Yuan ◽  
Haoping Liu
Keyword(s):  
Carbon Dioxide ◽  
Candida Albicans ◽  
Signaling Pathway ◽  
Fungal Infections ◽  
Pathogenic Fungus ◽  
Hyphal Growth ◽  
Dependent Manner ◽  
Content Type ◽  
Hyphal Development ◽  
Carbon Dioxide Levels

ABSTRACT Candida albicans is the most common cause of invasive fungal infections in humans. Its ability to sense and adapt to changing carbon dioxide levels is crucial for its pathogenesis. Carbon dioxide promotes hyphal development. The hypha-specific transcription factor Ume6 is rapidly degraded in air, but is stable under physiological CO2 and hypoxia to sustain hyphal elongation. Here, we show that Ume6 stability is regulated by two parallel E3 ubiquitin ligases, SCFGrr1 and Ubr1, in response to CO2 and O2, respectively. To uncover the CO2 signaling pathway that regulates Ume6 stability, we performed genetic screens for mutants unable to respond to CO2 for sustained filamentation. We find that the type 2C protein phosphatase Ptc2 is specifically required for CO2-induced stabilization of Ume6 and hyphal elongation. In contrast, the cyclin-dependent kinase Ssn3 is found to be required for Ume6 phosphorylation and degradation in atmospheric CO2. Furthermore, we find that Ssn3 is dephosphorylated in 5% CO2 in a Ptc2-dependent manner, whereas deletion of PTC2 has no effect on Ssn3 phosphorylation in air. Our study uncovers the Ptc2-Ssn3 axis as a new CO2 signaling pathway that controls hyphal elongation by regulating Ume6 stability in C. albicans. IMPORTANCE The capacity to sense and adapt to changing carbon dioxide levels is crucial for all organisms. In fungi, CO2 is a key determinant involved in fundamental biological processes, including growth, morphology, and virulence. In the pathogenic fungus Candida albicans, high CO2 is directly sensed by adenylyl cyclase to promote hyphal growth. However, little is known about the mechanism by which hyphal development is maintained in response to physiological levels of CO2. Here we report that a signal transduction system mediated by a phosphatase-kinase pair controls CO2-responsive Ume6 phosphorylation and stability that in turn dictate hyphal elongation. Our results unravel a new regulatory mechanism of CO2 signaling in fungi.

scholarly journals High-Throughput Screening of a Collection of Known Pharmacologically Active Small Compounds for Identification of Candida albicans Biofilm Inhibitors

2013 ◽  
Vol 57 (8) ◽  
pp. 3681-3687 ◽  
Author(s):  
Samuel A. Siles ◽  
Anand Srinivasan ◽  
Christopher G. Pierce ◽  
José L. Lopez-Ribot ◽  
Anand K. Ramasubramanian
Keyword(s):  
Candida Albicans ◽  
Antifungal Activity ◽  
Biofilm Formation ◽  
High Throughput Screening ◽  
Fungal Infections ◽  
Rapid Development ◽  
Unmet Need ◽  
Antifungal Drugs ◽  
Content Type ◽  
Pharmacologically Active

ABSTRACTCandida albicansis the most common etiologic agent of systemic fungal infections with unacceptably high mortality rates. The existing arsenal of antifungal drugs is very limited and is particularly ineffective againstC. albicansbiofilms. To address the unmet need for novel antifungals, particularly those active against biofilms, we have screened a small molecule library consisting of 1,200 off-patent drugs already approved by the Food and Drug Administration (FDA), the Prestwick Chemical Library, to identify inhibitors ofC. albicansbiofilm formation. According to their pharmacological applications that are currently known, we classified these bioactive compounds as antifungal drugs, as antimicrobials/antiseptics, or as miscellaneous drugs, which we considered to be drugs with no previously characterized antifungal activity. Using a 96-well microtiter plate-based high-content screening assay, we identified 38 pharmacologically active agents that inhibitC. albicansbiofilm formation. These drugs were subsequently tested for their potency and efficacy against preformed biofilms, and we identified three drugs with novel antifungal activity. Thus, repurposing FDA-approved drugs opens up a valuable new avenue for identification and potentially rapid development of antifungal agents, which are urgently needed.

scholarly journals Candida albicansHyphal Expansion Causes Phagosomal Membrane Damage and Luminal Alkalinization

mBio ◽  
2018 ◽  
Vol 9 (5) ◽  
Author(s):  
Johannes Westman ◽  
Gary Moran ◽  
Selene Mogavero ◽  
Bernhard Hube ◽  
Sergio Grinstein
Keyword(s):  
Fungal Infections ◽  
Membrane Damage ◽  
Hyphal Growth ◽  
Antifungal Drugs ◽  
Apparent Paradox ◽  
Opportunistic Fungi ◽  
Content Type ◽  
Life Threatening ◽  
Proton Leakage ◽  
Phagosomal Membrane

ABSTRACTMacrophages rely on phagosomal acidity to destroy engulfed microorganisms. To survive this hostile response, opportunistic fungi such asCandida albicansdeveloped strategies to evade the acidic environment.C. albicansis polymorphic and able to convert from yeast to hyphae, and this transition is required to subvert the microbicidal activity of the phagosome. However, the phagosomal lumen, which is acidic and nutrient deprived, is believed to inhibit the yeast-to-hypha transition. To account for this apparent paradox, it was recently proposed thatC. albicansproduces ammonia that alkalinizes the phagosome, thus facilitating yeast-to-hypha transition. We reexamined the mechanism underlying phagosomal alkalinization by applying dual-wavelength ratiometric pH measurements. The phagosomal membrane was found to be highly permeable to ammonia, which is therefore unlikely to account for the pH elevation. Instead, we find that yeast-to-hypha transition begins within acidic phagosomes and that alkalinization is a consequence of proton leakage induced by excessive membrane distension caused by the expanding hypha.IMPORTANCEC. albicansis the most common cause of nosocomial fungal infection, and over 3 million people acquire life-threatening invasive fungal infections every year. Even if antifungal drugs exist, almost half of these patients will die. Despite this, fungi remain underestimated as pathogens. Our study uses quantitative biophysical approaches to demonstrate that yeast-to-hypha transition occurs within the nutrient-deprived, acidic phagosome and that alkalinization is a consequence, as opposed to the cause, of hyphal growth.

scholarly journals Potential Targets for Antifungal Drug Discovery Based on Growth and Virulence in Candida albicans

2015 ◽  
Vol 59 (10) ◽  
pp. 5885-5891 ◽  
Author(s):  
Xiuyun Li ◽  
Yinglong Hou ◽  
Longtao Yue ◽  
Shuyuan Liu ◽  
Juan Du ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Drug Discovery ◽  
Fungal Infections ◽  
Current Knowledge ◽  
Antifungal Drugs ◽  
Clinical Settings ◽  
Challenging Problem ◽  
Content Type ◽  
Pathogenic Mechanisms ◽  
Novel Targets

ABSTRACTFungal infections, especially infections caused byCandida albicans, remain a challenging problem in clinical settings. Despite the development of more-effective antifungal drugs, their application is limited for various reasons. Thus, alternative treatments with drugs aimed at novel targets inC. albicansare needed. Knowledge of growth and virulence in fungal cells is essential not only to understand their pathogenic mechanisms but also to identify potential antifungal targets. This article reviews the current knowledge of the mechanisms of growth and virulence inC. albicansand examines potential targets for the development of new antifungal drugs.

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

2019 ◽  
Vol 15 (6) ◽  
pp. 648-658 ◽  
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.

scholarly journals Dectin-2-Targeted Antifungal Liposomes Exhibit Enhanced Efficacy

mSphere ◽  
2019 ◽  
Vol 4 (5) ◽  
Author(s):  
Suresh Ambati ◽  
Emma C. Ellis ◽  
Jianfeng Lin ◽  
Xiaorong Lin ◽  
Zachary A. Lewis ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Aspergillus Fumigatus ◽  
Effective Dose ◽  
Cryptococcus Neoformans ◽  
Amphotericin B ◽  
Antifungal Drugs ◽  
Extracellular Matrices ◽  
Content Type ◽  
Order Of Magnitude ◽  
Life Threatening

ABSTRACT Candida albicans, Cryptococcus neoformans, and Aspergillus fumigatus cause life-threatening candidiasis, cryptococcosis, and aspergillosis, resulting in several hundred thousand deaths annually. The patients at the greatest risk of developing these life-threatening invasive fungal infections have weakened immune systems. The vulnerable population is increasing due to rising numbers of immunocompromised individuals as a result of HIV infection or immunosuppressed individuals receiving anticancer therapies and/or stem cell or organ transplants. While patients are treated with antifungals such as amphotericin B, all antifungals have serious limitations due to lack of sufficient fungicidal effect and/or host toxicity. Even with treatment, 1-year survival rates are low. We explored methods of increasing drug effectiveness by designing fungicide-loaded liposomes specifically targeted to fungal cells. Most pathogenic fungi are encased in cell walls and exopolysaccharide matrices rich in mannans. Dectin-2 is a mammalian innate immune membrane receptor that binds as a dimer to mannans and signals fungal infection. We coated amphotericin-loaded liposomes with monomers of Dectin-2’s mannan-binding domain, sDectin-2. sDectin monomers were free to float in the lipid membrane and form dimers that bind mannan substrates. sDectin-2-coated liposomes bound orders of magnitude more efficiently to the extracellular matrices of several developmental stages of C. albicans, C. neoformans, and A. fumigatus than untargeted control liposomes. Dectin-2-coated amphotericin B-loaded liposomes reduced the growth and viability of all three species more than an order of magnitude more efficiently than untargeted control liposomes and dramatically decreased the effective dose. Future efforts focus on examining pan-antifungal targeted liposomal drugs in animal models of fungal diseases. IMPORTANCE Invasive fungal diseases caused by Candida albicans, Cryptococcus neoformans, and Aspergillus fumigatus have mortality rates ranging from 10 to 95%. Individual patient costs may exceed $100,000 in the United States. All antifungals in current use have serious limitations due to host toxicity and/or insufficient fungal cell killing that results in recurrent infections. Few new antifungal drugs have been introduced in the last 2 decades. Hence, there is a critical need for improved antifungal therapeutics. By targeting antifungal-loaded liposomes to α-mannans in the extracellular matrices secreted by these fungi, we dramatically reduced the effective dose of drug. Dectin-2-coated liposomes loaded with amphotericin B bound 50- to 150-fold more strongly to C. albicans, C. neoformans, and A. fumigatus than untargeted liposomes and killed these fungi more than an order of magnitude more efficiently. Targeting drug-loaded liposomes specifically to fungal cells has the potential to greatly enhance the efficacy of most antifungal drugs.

scholarly journals Cell Wall-Related Bionumbers and Bioestimates of Saccharomyces cerevisiae and Candida albicans

2013 ◽  
Vol 13 (1) ◽  
pp. 2-9 ◽  
Author(s):  
Frans M. Klis ◽  
Chris G. de Koster ◽  
Stanley Brul
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Wall ◽  
Candida Albicans ◽  
Cell Size ◽  
Pathogenic Fungus ◽  
Turgor Pressure ◽  
Hyphal Growth ◽  
Biological Research ◽  
Content Type ◽  
Starting Point

ABSTRACTBionumbers and bioestimates are valuable tools in biological research. Here we focus on cell wall-related bionumbers and bioestimates of the budding yeastSaccharomyces cerevisiaeand the polymorphic, pathogenic fungusCandida albicans. We discuss the linear relationship between cell size and cell ploidy, the correlation between cell size and specific growth rate, the effect of turgor pressure on cell size, and the reason why using fixed cells for measuring cellular dimensions can result in serious underestimation ofin vivovalues. We further consider the evidence that individual buds and hyphae grow linearly and that exponential growth of the population results from regular formation of new daughter cells and regular hyphal branching. Our calculations show that hyphal growth allowsC. albicansto cover much larger distances per unit of time than the yeast mode of growth and that this is accompanied by strongly increased surface expansion rates. We therefore predict that the transcript levels of genes involved in wall formation increase during hyphal growth. Interestingly, wall proteins and polysaccharides seem barely, if at all, subject to turnover and replacement. A general lesson is how strongly most bionumbers and bioestimates depend on environmental conditions and genetic background, thus reemphasizing the importance of well-defined and carefully chosen culture conditions and experimental approaches. Finally, we propose that the numbers and estimates described here offer a solid starting point for similar studies of other cell compartments and other yeast species.

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