scholarly journals UPC2Is Universally Essential for Azole Antifungal Resistance in Candida albicans

2014 ◽  
Vol 13 (7) ◽  
pp. 933-946 ◽  
Author(s):  
Erin M. Vasicek ◽  
Elizabeth L. Berkow ◽  
Stephanie A. Flowers ◽  
Katherine S. Barker ◽  
P. David Rogers
Keyword(s):  
Candida Albicans ◽  
Iron Homeostasis ◽  
Solid Medium ◽  
Azole Resistance ◽  
Zone Of Inhibition ◽  
Clear Zone ◽  
Content Type ◽  
Fungistatic Activity ◽  
Time Kill ◽  
Increased Susceptibility

ABSTRACTInCandida albicans, the transcription factor Upc2 is central to the regulation of ergosterol biosynthesis.UPC2-activating mutations contribute to azole resistance, whereas disruption increases azole susceptibility. In the present study, we investigated the relationship ofUPC2to fluconazole susceptibility, particularly in azole-resistant strains. In addition to the reduced fluconazole MIC previously observed withUPC2disruption, we observed a lower minimum fungicidal concentration (MFC) for aupc2Δ/Δ mutant than for its azole-susceptible parent, SC5314. Moreover, theupc2Δ/Δ mutant was unable to grow on a solid medium containing 10 μg/ml fluconazole and exhibited increased susceptibility and a clear zone of inhibition by Etest. Time-kill analysis showed higher fungistatic activity against theupc2Δ/Δ mutant than against SC5314.UPC2disruption in strains carrying specific resistance mutations also resulted in reduced MICs and MFCs.UPC2disruption in a highly azole resistant clinical isolate containing multiple resistance mechanisms likewise resulted in a reduced MIC and MFC. This mutant was unable to grow on a solid medium containing 10 μg/ml fluconazole and exhibited increased susceptibility and a clear zone of inhibition by Etest. Time-kill analysis showed increased fungistatic activity against theupc2Δ/Δ mutant in the resistant background. Microarray analysis showed attenuated induction by fluconazole of genes involved in sterol biosynthesis, iron transport, or iron homeostasis in the absence ofUPC2. Taken together, these data demonstrate that theUPC2transcriptional network is universally essential for azole resistance inC. albicansand represents an attractive target for enhancing azole antifungal activity.

scholarly journals Mitochondria InfluenceCDR1Efflux Pump Activity, Hog1-Mediated Oxidative Stress Pathway, Iron Homeostasis, and Ergosterol Levels in Candida albicans

2013 ◽  
Vol 57 (11) ◽  
pp. 5580-5599 ◽  
Author(s):  
Edwina Thomas ◽  
Elvira Roman ◽  
Steven Claypool ◽  
Nikhat Manzoor ◽  
Jesús Pla ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Candida Albicans ◽  
Cellular Response ◽  
Iron Homeostasis ◽  
Sufficient Conditions ◽  
Ergosterol Biosynthesis ◽  
Biosynthesis Pathway ◽  
Content Type ◽  
Iron Assimilation ◽  
Increased Susceptibility

ABSTRACTMitochondrial dysfunction inCandida albicansis known to be associated with drug susceptibility, cell wall integrity, phospholipid homeostasis, and virulence. In this study, we deletedCaFZO1, a key component required during biogenesis of functional mitochondria. Cells withFZO1deleted displayed fragmented mitochondria, mitochondrial genome loss, and reduced mitochondrial membrane potential and were rendered sensitive to azoles and peroxide. In order to understand the cellular response to dysfunctional mitochondria, genome-wide expression profiling offzo1Δ/Δ cells was performed. Our results show that the increased susceptibility to azoles was likely due to reduced efflux activity ofCDRefflux pumps, caused by the missorting of Cdr1p into the vacuole. In addition,fzo1Δ/Δ cells showed upregulation of genes involved in iron assimilation, in iron-sufficient conditions, characteristic of iron-starved cells. One of the consequent effects was downregulation of genes of the ergosterol biosynthesis pathway with a commensurate decrease in cellular ergosterol levels. We therefore connect deregulated iron metabolism to ergosterol biosynthesis pathway in response to dysfunctional mitochondria. Impaired activation of the Hog1 pathway in the mutant was the basis for increased susceptibility to peroxide and increase in reactive oxygen species, indicating the importance of functional mitochondria in controlling Hog1-mediated oxidative stress response. Mitochondrial phospholipid levels were also altered as indicated by an increase in phosphatidylserine and phosphatidylethanolamine and decrease in phosphatidylcholine infzo1Δ/Δ cells. Collectively, these findings reinforce the connection between functional mitochondria and azole tolerance, oxidant-mediated stress, and iron homeostasis inC. albicans.

scholarly journals Macrophage Autophagy in Immunity to Cryptococcus neoformans and Candida albicans

2012 ◽  
Vol 80 (9) ◽  
pp. 3065-3076 ◽  
Author(s):  
André Moraes Nicola ◽  
Patrícia Albuquerque ◽  
Luis R. Martinez ◽  
Rafael Antonio Dal-Rosso ◽  
Carolyn Saylor ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Cryptococcus Neoformans ◽  
Fungal Pathogens ◽  
Cytokine Signaling ◽  
Alternatively Activated Macrophages ◽  
Content Type ◽  
Material Recycling ◽  
Fungistatic Activity ◽  
Increased Susceptibility

ABSTRACTAutophagy is used by eukaryotes in bulk cellular material recycling and in immunity to intracellular pathogens. We evaluated the role of macrophage autophagy in the response toCryptococcus neoformansandCandida albicans, two important opportunistic fungal pathogens. The autophagosome marker LC3 (microtubule-associated protein 1 light chain 3 alpha) was present in most macrophage vacuoles containingC. albicans. In contrast, LC3 was found in only a few vacuoles containingC. neoformanspreviously opsonized with antibody but never after complement-mediated phagocytosis. Disruption of host autophagyin vitroby RNA interference against ATG5 (autophagy-related 5) decreased the phagocytosis ofC. albicansand the fungistatic activity of J774.16 macrophage-like cells against both fungi, independent of the opsonin used. ATG5-knockout bone marrow-derived macrophages (BMMs) also had decreased fungistatic activity againstC. neoformanswhen activated. In contrast, nonactivated ATG5-knockout BMMs actually restrictedC. neoformansgrowth more efficiently, suggesting that macrophage autophagy plays different roles againstC. neoformans, depending on the macrophage type and activation. Interference with autophagy in J774.16 cells also decreased nonlytic exocytosis ofC. neoformans, increased interleukin-6 secretion, and decreased gamma interferon-induced protein 10 secretion. Mice with a conditionally knocked out ATG5 gene in myeloid cells showed increased susceptibility to intravenousC. albicansinfection. In contrast, these mice manifested no increased susceptibility toC. neoformans, as measured by survival, but had fewer alternatively activated macrophages and less inflammation in the lungs after intratracheal infection than control mice. These results demonstrate the complex roles of macrophage autophagy in restricting intracellular parasitism by fungi and reveal connections with nonlytic exocytosis, humoral immunity, and cytokine signaling.

scholarly journals Head-to-Head Comparison of Inhibitory and Fungicidal Activities of Fluconazole, Itraconazole, Voriconazole, Posaconazole, and Isavuconazole against Clinical Isolates of Trichosporon asahii

2013 ◽  
Vol 57 (10) ◽  
pp. 4841-4847 ◽  
Author(s):  
Gulsen Hazirolan ◽  
Emilia Canton ◽  
Selma Sahin ◽  
Sevtap Arikan-Akdagli
Keyword(s):  
Rank Order ◽  
Growth Control ◽  
Clinical Isolates ◽  
Content Type ◽  
Trichosporon Asahii ◽  
Killing Activity ◽  
Fungistatic Activity ◽  
Microdilution Method ◽  
Inhibition Of Growth ◽  
Time Kill

ABSTRACTTreatment of disseminatedTrichosporoninfections still remains difficult. Amphotericin B frequently displays inadequate fungicidal activity and echinocandins have no meaningful antifungal effect against this genus. Triazoles are currently the drugs of choice for the treatment ofTrichosporoninfections. This study evaluates the inhibitory and fungicidal activities of five triazoles against 90 clinical isolates ofTrichosporon asahii. MICs (μg/ml) were determined according to Clinical and Laboratory Standards Institute microdilution method M27-A3 at 24 and 48 h using two endpoints, MIC-2 and MIC-0 (the lowest concentrations that inhibited ∼50 and 100% of growth, respectively). Minimum fungicidal concentrations (MFCs; μg/ml) were determined by seeding 100 μl of all clear MIC wells (using an inoculum of 104CFU/ml) onto Sabouraud dextrose agar. Time-kill curves were assayed against four clinicalT. asahiiisolates and theT. asahiiATCC 201110 strain. The MIC-2 (∼50% reduction in turbidity compared to the growth control well)/MIC-0 (complete inhibition of growth)/MFC values that inhibited 90% of isolates at 48 h were, respectively, 8/32/64 μg/ml for fluconazole, 1/2/8 μg/ml for itraconazole, 0.12/0.5/2 μg/ml for voriconazole, 0.5/2/4 μg/ml for posaconazole, and 0.25/1/4 μg/ml for isavuconazole. The MIC-0 endpoints yielded more consistent MIC results, which remained mostly unchanged when extending the incubation to 48 h (98 to 100% agreement with 24-h values) and are easier to interpret. Based on the time-kill experiments, none of the drugs reached the fungicidal endpoint (99.9% killing), killing activity being shown but at concentrations not reached in serum. Statistical analysis revealed that killing rates are dose and antifungal dependent. The lowest concentration at which killing activity begins was for voriconazole, and the highest was for fluconazole. These results suggest that azoles display fungistatic activity and lack fungicidal effect againstT. asahii. By rank order, the most active triazole is voriconazole, followed by itraconazole ∼ posaconazole ∼ isavuconazole > fluconazole.

scholarly journals Disruption of the Transcriptional Regulator Cas5 Results in Enhanced Killing of Candida albicans by Fluconazole

2014 ◽  
Vol 58 (11) ◽  
pp. 6807-6818 ◽  
Author(s):  
Erin M. Vasicek ◽  
Elizabeth L. Berkow ◽  
Vincent M. Bruno ◽  
Aaron P. Mitchell ◽  
Nathan P. Wiederhold ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Resistance Mechanisms ◽  
Transcriptional Network ◽  
Transcriptional Analysis ◽  
Resistance Mutations ◽  
Content Type ◽  
Minimum Fungicidal Concentration ◽  
Fungistatic Activity ◽  
Yeast Extract Peptone Dextrose ◽  
Genes Encoding

ABSTRACTAzole antifungal agents such as fluconazole exhibit fungistatic activity againstCandida albicans. Strategies to enhance azole antifungal activity would be therapeutically appealing. In an effort to identify transcriptional pathways that influence the killing activity of fluconazole, we sought to identify transcription factors (TFs) involved in this process. From a collection ofC. albicansstrains disrupted for genes encoding TFs (O. R. Homann, J. Dea, S. M. Noble, and A. D. Johnson, PLoS Genet. 5:e1000783, 2009,http://dx.doi.org/10.1371/journal.pgen.1000783), four strains exhibited marked reductions in minimum fungicidal concentration (MFCs) in both RPMI and yeast extract-peptone-dextrose (YPD) media. One of these genes,UPC2, was previously characterized with regard to its role in azole susceptibility. Of mutants representing the three remaining TF genes of interest, one (CAS5) was unable to recover from fluconazole exposure at concentrations as low as 2 μg/ml after 72 h in YPD medium. This mutant also showed reduced susceptibility and a clear zone of inhibition by Etest, was unable to grow on solid medium containing 10 μg/ml fluconazole, and exhibited increased susceptibility by time-kill analysis.CAS5disruption in highly azole-resistant clinical isolates exhibiting multiple resistance mechanisms did not alter susceptibility. However,CAS5disruption in strains with specific resistance mutations resulted in moderate reductions in MICs and MFCs. Genome-wide transcriptional analysis was performed in the presence of fluconazole and was consistent with the suggested role ofCAS5in cell wall organization while also suggesting a role in iron transport and homeostasis. These findings suggest that Cas5 regulates a transcriptional network that influences the response ofC. albicansto fluconazole. Further delineation of this transcriptional network may identify targets for potential cotherapeutic strategies to enhance the activity of the azole class of antifungals.

scholarly journals Potent In Vitro Synergism of Fluconazole and Berberine Chloride against Clinical Isolates of Candida albicans Resistant to Fluconazole

2006 ◽  
Vol 50 (3) ◽  
pp. 1096-1099 ◽  
Author(s):  
Hua Quan ◽  
Ying-Ying Cao ◽  
Zheng Xu ◽  
Jing-Xia Zhao ◽  
Ping-Hui Gao ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Clinical Isolates ◽  
Antagonistic Action ◽  
Berberine Chloride ◽  
Agar Diffusion ◽  
Fungistatic Activity ◽  
Fluconazole Resistant ◽  
Time Kill ◽  
In Vitro Interaction

ABSTRACT In vitro interaction of fluconazole and berberine chloride was investigated against 40 fluconazole-resistant clinical isolates of Candida albicans. Synergism in fungistatic activity was found with the checkerboard microdilution assay. The findings of agar diffusion tests and time-kill curves confirmed the synergistic interaction, but no antagonistic action was observed.

scholarly journals Fungemia Surveillance in Denmark Demonstrates Emergence of Non-albicans Candida Species and Higher Antifungal Usage and Resistance Rates than in Other Nations

2018 ◽  
Vol 56 (4) ◽  
pp. e01907-17 ◽  
Author(s):  
Mariana Castanheira
Keyword(s):  
Candida Albicans ◽  
Candida Species ◽  
Susceptibility Testing ◽  
Antifungal Susceptibility ◽  
Fungal Species ◽  
Azole Resistance ◽  
Content Type ◽  
Fungal Organisms ◽  
Echinocandin Resistance ◽  
Resistance Rates

ABSTRACT Recent changes in the occurrence of fungal species and the difficulties in performing reference antifungal susceptibility testing highlight the importance of surveillance of fungal organisms and antifungal resistance rates. K. M. T. Astvad et al. report results from recent (2012 to 2015) fungemia surveillance in Denmark and compare the results to previous data (2004 to 2011), showing a decrease in Candida albicans infections accompanied by an increase in C. glabrata and C. dubliniensis infections (J Clin Microbiol 56:e01564-17, 2018, https://doi.org/10.1128/JCM.01564-17). Azole resistance among C. tropicalis and C. parapsilosis isolates and echinocandin resistance in C. krusei isolates were higher in Denmark than in other regions. Interestingly, the usage of antifungals is higher in Denmark than in other Nordic countries.

scholarly journals Antifungal pharmacodynamic characteristics of fluconazole and amphotericin B tested against Candida albicans.

1997 ◽  
Vol 41 (6) ◽  
pp. 1392-1395 ◽  
Author(s):  
M E Klepser ◽  
E J Wolfe ◽  
R N Jones ◽  
C H Nightingale ◽  
M A Pfaller
Keyword(s):  
Candida Albicans ◽  
Amphotericin B ◽  
Fungicidal Activity ◽  
Narrow Range ◽  
Growth Effects ◽  
Fungistatic Activity ◽  
Time Kill

Time-kill curves were determined for three isolates of Candida albicans tested against fluconazole and amphotericin B at multiples of the MIC. Fluconazole produced fungistatic activity, with concentration-related growth effects observed over a narrow range of concentrations. Amphotericin B exhibited fungicidal activity, with enhancement of activity over a broader range of concentrations.

scholarly journals Activity of Isavuconazole and Other Azoles against Candida Clinical Isolates and Yeast Model Systems with Known Azole Resistance Mechanisms

2015 ◽  
Vol 60 (1) ◽  
pp. 229-238 ◽  
Author(s):  
Dominique Sanglard ◽  
Alix T. Coste
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Candida Albicans ◽  
Abc Transporters ◽  
Resistance Mechanisms ◽  
Model Systems ◽  
Clinical Isolates ◽  
Azole Resistance ◽  
Limited Effect ◽  
Content Type ◽  
Major Facilitator

ABSTRACTIsavuconazole is a novel, broad-spectrum, antifungal azole. In order to evaluate its interactions with known azole resistance mechanisms, isavuconazole susceptibility among different yeast models and clinical isolates expressing characterized azole resistance mechanisms was tested and compared to those of fluconazole, itraconazole, posaconazole, and voriconazole.Saccharomyces cerevisiaeexpressing theCandida albicansandC. glabrataATP binding cassette (ABC) transporters (CDR1,CDR2, andCgCDR1), major facilitator (MDR1), and lanosterol 14-α-sterol-demethylase (ERG11) alleles with mutations were used. In addition, pairs ofC. albicansandC. glabratastrains from matched clinical isolates with known azole resistance mechanisms were investigated. The expression of ABC transporters increased all azole MICs, suggesting that all azoles tested were substrates of ABC transporters. The expression ofMDR1did not increase posaconazole, itraconazole, and isavuconazole MICs. Relative increases of azole MICs (from 4- to 32-fold) were observed for fluconazole, voriconazole, and isavuconazole when at least two mutations were present in the sameERG11allele. Upon MIC testing of azoles with clinicalC. albicansandC. glabrataisolates with known resistance mechanisms, the MIC90s ofC. albicansfor fluconazole, voriconazole, itraconazole, posaconazole, and isavuconazole were 128, 2, 1, 0.5, and 2 μg/ml, respectively, while inC. glabratathey were 128, 2, 4, 4, and 16 μg/ml, respectively. In conclusion, the effects of azole resistance mechanisms on isavuconazole did not differ significantly from those of other azoles. Resistance mechanisms in yeasts involving ABC transporters andERG11decreased the activity of isavuconazole, whileMDR1had limited effect.

scholarly journals Occidiofungin's Chemical Stability andIn VitroPotency against Candida Species

2011 ◽  
Vol 56 (2) ◽  
pp. 765-769 ◽  
Author(s):  
Dayna Ellis ◽  
Jiten Gosai ◽  
Charles Emrick ◽  
Rachel Heintz ◽  
Lanette Romans ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Chemical Stability ◽  
Extreme Temperatures ◽  
Content Type ◽  
Burkholderia Contaminans ◽  
Time Kill ◽  
Postantifungal Effect ◽  
Minimal Lethal Concentration ◽  
Stable Molecule

ABSTRACTOccidiofungin is a cyclic glyco-lipopeptide produced byBurkholderia contaminans. MICs againstCandidaspecies were between 0.5 and 2.0 μg/ml. Occidiofungin retains itsin vitropotency in the presence of 5% and 50% human serum with a minimal lethal concentration (MLC) of 2 and 4 μg/ml, respectively. Time-kill and postantifungal effect (PAFE) experiments of occidiofungin againstCandida albicanswere performed. The results demonstrate that occidiofungin is fungicidal. Occidiofungin was also found to be a very stable molecule. It is resistant to extreme temperatures and pH and maintains its activity following exposure to gastric proteases.

scholarly journals In VitroInteractions between Aspirin and Amphotericin B against Planktonic Cells and Biofilm Cells of Candida albicans and C. parapsilosis

2012 ◽  
Vol 56 (6) ◽  
pp. 3250-3260 ◽  
Author(s):  
Yabin Zhou ◽  
Ganggang Wang ◽  
Yutang Li ◽  
Yang Liu ◽  
Yu Song ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Amphotericin B ◽  
Antimicrobial Agents ◽  
Positive Interactions ◽  
Antibiofilm Activity ◽  
Data Generation ◽  
Planktonic Cells ◽  
Content Type ◽  
Time Kill

ABSTRACTThe increase in drug resistance and invasion caused by biofilm formation brings enormous challenges to the management ofCandidainfection. Aspirin's antibiofilm activityin vitrowas discovered recently. The spectrophotometric method and the XTT {2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino)carbonyl]-2H-tetrazolium hydroxide} reduction assay used for data generation make it possible to evaluate fungal biofilm growth accurately. The combined use of the most commonly used methods, the fractional inhibitory concentration index (FICI) and a newly developed method, the ΔEmodel, which uses the concentration-effect relationship over the whole concentration range instead of using the MIC index alone, makes the interpretation of results more reliable. As an attractive tool for studying the pharmacodynamics of antimicrobial agents, time-kill curves can provide detailed information about antimicrobial efficacy as a function of both time and concentration. In the present study,in vitrointeractions between aspirin (acetylsalicylic acid [ASA]) and amphotericin B (AMB) against planktonic cells and biofilm cells ofCandida albicansandC. parapsilosiswere evaluated by the checkerboard microdilution method and the time-kill test. Synergistic and indifferent effects were found for the combination of ASA and AMB against planktonic cells, while strong synergy was found against biofilm cells analyzed by FICI. The ΔEmodel gave more consistent results with FICI. The positive interactions in concentration were also confirmed by the time-kill test. Moreover, this approach also revealed the pharmacodynamics changes of ASA and synergistic action on time. Our findings suggest a potential clinical use for combination therapy with ASA and AMB to augment activity against biofilm-associated infections.

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