scholarly journals Paradoxical Effect of Caspofungin: Reduced Activity against Candida albicans at High Drug Concentrations

2004 ◽  
Vol 48 (9) ◽  
pp. 3407-3411 ◽  
Author(s):  
David A. Stevens ◽  
Marife Espiritu ◽  
Rachana Parmar
Keyword(s):  
Candida Albicans ◽  
Resistance Mechanisms ◽  
Paradoxical Effect ◽  
Fungal Cell ◽  
High Drug ◽  
Minimum Fungicidal Concentration ◽  
Drug Concentrations ◽  
High Concentrations ◽  
Glucan Synthesis

ABSTRACT Resistance problems with caspofungin, an echinocandin inhibitor of fungal cell wall glucan synthesis, have been rare. We noted paradoxical turbid growth of Candida albicans isolates in broth in some high (supra-MIC) concentrations. Among isolates submitted for susceptibility testing and screened at drug concentrations up to 12.5 μg/ml, the frequency was 16%. Analysis of the turbid growth indicated slowing of growth in the presence of drug but with numbers of CFU up to 72% those of drug-free controls. Clearing of growth again by the highest drug concentrations produced a quadriphasic pattern in a tube dilution series. Cells growing at high drug concentrations were not resistant on retesting but showed the paradoxical effect of the parent. Among a selected series of isolates tested at concentrations up to 50 μg/ml, an additional 53% showed a “mini-paradoxical effect”: no turbid growth but incomplete killing at high concentrations (supra-minimum fungicidal concentration). These effects were reproducible; medium dependent in extent; noted in macro- and microdilution, in the presence or absence of serum, and on agar containing drug (but not when drug concentrations were not constant, as in agar diffusion); not seen with other echinocandins and less commonly in other Candida species; and not due to destruction of drug in tubes showing the effect. Cooperative enhancement of inhibition by a second drug could eradicate the effect. We postulate that high drug concentrations derepress or activate resistance mechanisms. The abilities of subpopulations to survive at high drug concentrations could have in vivo consequences.

scholarly journals Role for Endosomal and Vacuolar GTPases in Candida albicans Pathogenesis

2009 ◽  
Vol 77 (6) ◽  
pp. 2343-2355 ◽  
Author(s):  
Douglas A. Johnston ◽  
Karen E. Eberle ◽  
Joy E. Sturtevant ◽  
Glen E. Palmer
Keyword(s):  
Candida Albicans ◽  
Stress Resistance ◽  
Double Mutant ◽  
Hyphal Growth ◽  
Filamentous Growth ◽  
Rab Gtpases ◽  
Fungal Cell ◽  
Cellular Stresses ◽  
Vacuole Biogenesis

ABSTRACT The vacuole has crucial roles in stress resistance and adaptation of the fungal cell. Furthermore, in Candida albicans it has been observed to undergo dramatic expansion during the initiation of hyphal growth, to produce highly “vacuolated” subapical compartments. We hypothesized that these functions may be crucial for survival within the host and tissue-invasive hyphal growth. We also considered the role of the late endosome or prevacuole compartment (PVC), a distinct organelle involved in vacuolar and endocytic trafficking. We identified two Rab GTPases, encoded by VPS21 and YPT72, required for trafficking through the PVC and vacuole biogenesis, respectively. Deletion of VPS21 or YPT72 led to mild sensitivities to some cellular stresses. However, deletion of both genes resulted in a synthetic phenotype with severe sensitivity to cellular stress and impaired growth. Both the vps21Δ and ypt72Δ mutants had defects in filamentous growth, while the double mutant was completely deficient in polarized growth. The defects in hyphal growth were not suppressed by an “active” RIM101 allele or loss of the hyphal repressor encoded by TUP1. In addition, both single mutants had significant attenuation in a mouse model of hematogenously disseminated candidiasis, while the double mutant was rapidly cleared. Histological examination confirmed that the vps21Δ and ypt72Δ mutants are deficient in hyphal growth in vivo. We suggest that the PVC and vacuole are required on two levels during C. albicans infection: (i) stress resistance functions required for survival within tissue and (ii) a role in filamentous growth which may aid host tissue invasion.

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 The Paradoxical Effect of Echinocandins in Aspergillus fumigatus Relies on Recovery of the β-1,3-Glucan Synthase Fks1

2016 ◽  
Vol 61 (2) ◽  
Author(s):  
Veronika Loiko ◽  
Johannes Wagener
Keyword(s):  
Cell Wall ◽  
Aspergillus Fumigatus ◽  
Minor Importance ◽  
Initial Growth ◽  
Paradoxical Effect ◽  
Fungal Cell ◽  
Glucan Synthase ◽  
Content Type ◽  
Drug Concentrations ◽  
Very High

ABSTRACT Echinocandins target the fungal cell wall by inhibiting biosynthesis of the cell wall carbohydrate β-1,3-glucan. This antifungal drug class exhibits a paradoxical effect that is characterized by the resumption of growth of otherwise susceptible strains at higher drug concentrations (approximately 4 to 32 μg/ml). The nature of this phenomenon is still unknown. In this study, we analyzed the paradoxical effect of the echinocandin caspofungin on the pathogenic mold Aspergillus fumigatus. Using a conditional fks1 mutant, we show that very high caspofungin concentrations exert an additional antifungal activity besides inhibition of the β-1,3-glucan synthase. This activity could explain the suppression of paradoxical growth at very high caspofungin concentrations. Additionally, we found that exposure to inhibitory caspofungin concentrations always causes initial growth deprivation independently of the capability of the drug concentration to induce the paradoxical effect. Paradoxically growing hyphae emerge from microcolonies essentially devoid of β-1,3-glucan. However, these hyphae expose β-1,3-glucan again, suggesting that β-1,3-glucan synthesis is restored. In agreement with this hypothesis, we found that expression of the β-1,3-glucan synthase Fks1 is an essential requirement for the paradoxical effect. Surprisingly, overexpression of fks1 renders A. fumigatus more susceptible, whereas reduced expression leads to hyphae that are more resistant to the growth-inhibitory and limited fungicidal activity of caspofungin. Upregulation of chitin synthesis appears to be of minor importance for the paradoxical effect, since paradoxically growing hyphae exhibit significantly less chitin than the growth-deprived parental microcolonies. Our results argue for a model where the paradoxical effect primarily relies on recovery of β-1,3-glucan synthase activity.

scholarly journals Patients with Long-Term Oral Carriage Harbor High-Persister Mutants of Candida albicans

2009 ◽  
Vol 54 (1) ◽  
pp. 39-44 ◽  
Author(s):  
Michael D. LaFleur ◽  
Qingguo Qi ◽  
Kim Lewis
Keyword(s):  
Candida Albicans ◽  
Amphotericin B ◽  
Antimicrobial Agents ◽  
Oral Candidiasis ◽  
Wild Type ◽  
Persister Cells ◽  
Microtiter Plates ◽  
High Concentrations

ABSTRACT Fungal biofilms produce a small number of persister cells which can tolerate high concentrations of fungicidal agents. Persisters form upon attachment to a surface, an important step in the pathogenesis of Candida strains. The periodic application of antimicrobial agents may select for strains with increased levels of persister cells. In order to test this possibility, 150 isolates of Candida albicans and C. glabrata were obtained from cancer patients who were at high risk for the development of oral candidiasis and who had been treated with topical chlorhexidine once a day. Persister levels were measured by exposing biofilms growing in the wells of microtiter plates to high concentrations of amphotericin B and plating for survivors. The persister levels of the isolates varied from 0.2 to 9%, and strains isolated from patients with long-term carriage had high levels of persisters. High-persister strains were isolated from every patient with Candida carriage of more than 8 consecutive weeks but from no patients with transient carriage. All of the high-persister isolates had an amphotericin B MIC that was the same as that for the wild type, indicating that these strains were drug-tolerant rather than drug-resistant mutants. Biofilms of the majority of high-persister strains also showed an increased tolerance to chlorhexidine and had the same MIC for this antimicrobial as the wild type. This study suggests that persister cells are clinically relevant, and antimicrobial therapy selects for high-persister strains in vivo. The drug tolerance of persisters may be a critical but overlooked component responsible for antimicrobial drug failure and relapsing infections.

scholarly journals Novel Inhibitor of Plasmodium Histone Deacetylase That Cures P. berghei-Infected Mice

2009 ◽  
Vol 53 (5) ◽  
pp. 1727-1734 ◽  
Author(s):  
S. Agbor-Enoh ◽  
C. Seudieu ◽  
E. Davidson ◽  
A. Dritschilo ◽  
M. Jung
Keyword(s):  
Histone Deacetylases ◽  
Mammalian Cells ◽  
Malaria Parasite ◽  
Antimalarial Activity ◽  
Hdac Inhibitors ◽  
High Drug ◽  
Lead Compounds ◽  
Drug Concentrations

ABSTRACT Histone deacetylases (HDAC) are potential targets for the development of new antimalarial drugs. The growth of Plasmodium falciparum and other apicomplexans can be suppressed in the presence of potent HDAC inhibitors in vitro and in vivo; however, in vivo parasite suppression is generally incomplete or reversible after the discontinuation of drug treatment. Furthermore, most established HDAC inhibitors concurrently show broad toxicities against parasites and human cells and high drug concentrations are required for effective antimalarial activity. Here, we report on HDAC inhibitors that are potent against P. falciparum at subnanomolar concentrations and that have high selectivities; the lead compounds have mean 50% inhibitory concentrations for the killing of the malaria parasite up to 950 times lower than those for the killing of mammalian cells. These potential drugs improved survival and completely and irreversibly suppressed parasitemia in P. berghei-infected mice.

scholarly journals Assessment of the Paradoxical Effect of Caspofungin in Therapy of Candidiasis

2006 ◽  
Vol 50 (4) ◽  
pp. 1293-1297 ◽  
Author(s):  
Karl V. Clemons ◽  
Marife Espiritu ◽  
Rachana Parmar ◽  
David A. Stevens
Keyword(s):  
Candida Albicans ◽  
Clinical Isolates ◽  
Paradoxical Effect ◽  
Cure Rate ◽  
Systemic Model ◽  
Paradoxical Growth ◽  
Intravenous Inoculation

ABSTRACT Paradoxical growth of some Candida albicans isolates in the presence of caspofungin (CAS) in vitro has been demonstrated previously. We sought to determine whether a similar phenomenon occurred in vivo. A systemic model of candidiasis was studied in CD-1 mice by intravenous inoculation of different isolates of C. albicans. Infected animals were treated with CAS at various dosages (0.01 to 20 mg/kg) and CFU remaining in the kidneys determined. Four clinical isolates that showed paradoxical growth in vitro and one that did not were tested. Recovery of CFU from the kidneys showed that dosages of CAS at 0.1 mg/kg and above were efficacious in the reduction of C. albicans, but were not curative. Against isolates that show paradoxical growth in vitro, CAS was efficacious, but lacked dose responsiveness above 0.5 mg/kg against three of the four. One isolate, 95-68, showed paradoxical growth in vivo with significantly higher CFU recovered from mice given CAS at 20 mg/kg than those given CAS at 5 mg/kg, but the effect was not reproducible in a subsequent experiment. When CAS was given prophylactically and therapeutically, improved efficacy and cure rate were observed. Overall, these data indicate that CAS is highly efficacious against systemic murine candidiasis and a paradoxical effect was not reproducibly demonstrated in vivo.

scholarly journals Lon Protease Is Essential for Paradoxical Survival of Escherichia coli Exposed to High Concentrations of Quinolone

2009 ◽  
Vol 53 (7) ◽  
pp. 3103-3105 ◽  
Author(s):  
Muhammad Malik ◽  
Joseph Capecci ◽  
Karl Drlica
Keyword(s):  
Escherichia Coli ◽  
Dna Lesions ◽  
Lon Protease ◽  
High Drug ◽  
Cell Lysate ◽  
Drug Concentrations ◽  
Parallel Increase ◽  
High Concentrations ◽  
Dna Size ◽  
Very High

ABSTRACT A deficiency of the Escherichia coli Lon protease blocked paradoxical survival occurring at very high nalidixic acid concentrations. The absence of Lon also blocked a parallel increase in cell lysate viscosity likely to reflect DNA size. Thus, Lon may participate in repairing quinolone-mediated DNA lesions formed at high drug concentrations.

Studies of the paradoxical effect of caspofungin at high drug concentrations

2005 ◽  
Vol 51 (3) ◽  
pp. 173-178 ◽  
Author(s):  
David A. Stevens ◽  
Theodore C. White ◽  
David S. Perlin ◽  
Claude P. Selitrennikoff
Keyword(s):  
Paradoxical Effect ◽  
High Drug ◽  
Drug Concentrations

scholarly journals Cellular Accumulation, Localization, and Activity of a Synthetic Cyclopeptamine in Fungi

1998 ◽  
Vol 42 (2) ◽  
pp. 389-393 ◽  
Author(s):  
John O. Capobianco ◽  
Dorothy Zakula ◽  
David J. Frost ◽  
Robert C. Goldman ◽  
Leping Li ◽  
...  
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
Glucan Synthase ◽  
Cellular Accumulation ◽  
Drug Concentrations ◽  
Drug Reservoir ◽  
Membrane Bound ◽  
Glucan Synthesis ◽  
Major Drug

ABSTRACT A novel synthetic cyclopeptamine, A172013, rapidly accumulated by passive diffusion into Candida albicans CCH442. Drug influx could not be totally facilitated by the membrane-bound target, β-(1,3)-glucan synthase, since accumulation was unsaturable at drug concentrations up to 10 μg/ml (about 1.6 × 10−7molecules/cell), or 25× MIC. About 55 and 23% of the cell-incorporated drug was associated with the cell wall and protoplasts, respectively. Isolated microsomes contained 95% of the protoplast-associated drug, which was fully active against glucan synthesis in vitro. Drug (0.1 μg/ml) accumulation was rapid and complete after 5 min in several fungi tested, including a lipopeptide/cyclopeptamine-resistant strain of C. albicans(LP3-1). The compound penetrated to comparable levels in both yeast and hyphal forms of C. albicans, and accumulation inAspergillus niger was 20% that in C. albicans. These data indicated that drug-cell interactions were driven by the amphiphilic nature of the compound and that the cell wall served as a major drug reservoir.

scholarly journals Fluconazole Treatment Is Effective against a Candida albicans erg3/erg3 Mutant In Vivo Despite In Vitro Resistance

2006 ◽  
Vol 50 (2) ◽  
pp. 580-586 ◽  
Author(s):  
Taiga Miyazaki ◽  
Yoshitsugu Miyazaki ◽  
Koichi Izumikawa ◽  
Hiroshi Kakeya ◽  
Shunichi Miyakoshi ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Resistance Mechanism ◽  
Resistance Mechanisms ◽  
In Vivo Studies ◽  
Azole Resistance ◽  
In Vitro Testing ◽  
Fluconazole Resistance ◽  
Hyphal Formation

ABSTRACT Candida albicans ERG3 encodes a sterol C5,6-desaturase which is essential for synthesis of ergosterol. Defective sterol C5,6 desaturation has been considered to be one of the azole resistance mechanisms in this species. However, the clinical relevance of this resistance mechanism is still unclear. In this study, we created a C. albicans erg3/erg3 mutant by the “Ura-blaster” method and confirmed the expected azole resistance using standard in vitro testing and the presence of ergosta-7,22-dien-3β-ol instead of ergosterol. For in vivo studies, a wild-type URA3 was placed back into its native locus in the erg3 homozygote to avoid positional effects on URA3 expression. Defective hyphal formation of the erg3 homozygote was observed not only in vitro but in kidney tissues. A marked attenuation of virulence was shown by the longer survival and the lower kidney burdens of mice inoculated with the reconstituted Ura+ erg3 homozygote relative to the control. To assess fluconazole efficacy in a murine model of disseminated candidiasis, inoculum sizes of the control and the erg3 homozygote were chosen which provided a similar organ burden. Under these conditions, fluconazole was highly effective in reducing the organ burden in both groups. This study demonstrates that an ERG3 mutation causing inactivation of sterol C5,6-desaturase cannot confer fluconazole resistance in vivo by itself regardless of resistance measured by standard in vitro testing. The finding questions the clinical significance of this resistance mechanism.

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