Facultative Sterol Uptake in an Ergosterol-Deficient Clinical Isolate of Candida glabrata Harboring a Missense Mutation inERG11and Exhibiting Cross-Resistance to Azoles and Amphotericin B

ABSTRACTWe identified a clinical isolate ofCandida glabrata(CG156) exhibiting flocculent growth and cross-resistance to fluconazole (FLC), voriconazole (VRC), and amphotericin B (AMB), with MICs of >256, >256, and 32 μg ml−1, respectively. Sterol analysis using gas chromatography-mass spectrometry (GC-MS) revealed that CG156 was a sterol 14α-demethylase (Erg11p) mutant, wherein 14α-methylated intermediates (lanosterol was >80% of the total) were the only detectable sterols.ERG11sequencing indicated that CG156 harbored a single-amino-acid substitution (G315D) which nullified the function of native Erg11p. In heterologous expression studies using a doxycycline-regulatableSaccharomyces cerevisiae erg11strain, wild-typeC. glabrataErg11p fully complemented the function ofS. cerevisiaesterol 14α-demethylase, restoring growth and ergosterol synthesis in recombinant yeast; mutated CG156 Erg11p did not. CG156 was culturable using sterol-free, glucose-containing yeast minimal medium (glcYM). However, when grown on sterol-supplementedglcYM (with ergosta 7,22-dienol, ergosterol, cholestanol, cholesterol, Δ7-cholestenol, or desmosterol), CG156 cultures exhibited shorter lag phases, reached higher cell densities, and showed alterations in cellular sterol composition. Unlike comparator isolates (harboring wild-typeERG11) that became less sensitive to FLC and VRC when cultured on sterol-supplementedglcYM, facultative sterol uptake by CG156 did not affect its azole-resistant phenotype. Conversely, CG156 grown usingglcYM with ergosterol (or with ergosta 7,22-dienol) showed increased sensitivity to AMB; CG156 grown usingglcYM with cholesterol (or with cholestanol) became more resistant (MICs of 2 and >64 μg AMB ml−1, respectively). Our results provide insights into the consequences of sterol uptake and metabolism on growth and antifungal resistance inC. glabrata.

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CgMED3 Changes Membrane Sterol Composition To Help Candida glabrata Tolerate Low-pH Stress

Applied and Environmental Microbiology ◽

10.1128/aem.00972-17 ◽

2017 ◽

Vol 83 (17) ◽

Cited By ~ 12

Author(s):

Xiaobao Lin ◽

Yanli Qi ◽

Dongni Yan ◽

Hui Liu ◽

Xiulai Chen ◽

...

Keyword(s):

Acid Production ◽

Candida Glabrata ◽

Sterol Composition ◽

Low Ph ◽

Mediator Complex ◽

Wild Type ◽

Organic Acid Production ◽

Content Type ◽

Ph Stress

ABSTRACT Candida glabrata is a promising microorganism for organic acid production. The present study aimed to investigate the role of C. glabrata Mediator complex subunit 3 (CgMed3p) in protecting C. glabrata under low-pH conditions. To this end, genes CgMED3A and CgMED3B were deleted, resulting in the double-deletion Cgmed3ABΔ strain. The final biomass and cell viability levels of Cgmed3ABΔ decreased by 64.5% and 35.8%, respectively, compared to the wild-type strain results at pH 2.0. In addition, lack of CgMed3ABp resulted in selective repression of a subset of genes in the lipid biosynthesis and metabolism pathways. Furthermore, C18:1, lanosterol, zymosterol, fecosterol, and ergosterol were 13.2%, 80.4%, 40.4%, 78.1%, and 70.4% less abundant, respectively, in the Cgmed3ABΔ strain. In contrast, the concentration of squalene increased by about 44.6-fold. As a result, membrane integrity, rigidity, and H+-ATPase activity in the Cgmed3ABΔ strain were reduced by 62.7%, 13.0%, and 50.3%, respectively. In contrast, overexpression of CgMED3AB increased the levels of C18:0, C18:1, and ergosterol by 113.2%, 5.9%, and 26.4%, respectively. Moreover, compared to the wild-type results, dry cell weight and pyruvate production increased, irrespective of pH buffering. These results suggest that CgMED3AB regulates membrane composition, which in turn enables cells to tolerate low-pH stress. We propose that regulation of CgMed3ABp may provide a novel strategy for enhancing low-pH tolerance and increasing organic acid production by C. glabrata. IMPORTANCE The objective of this study was to investigate the role of Candida glabrata Mediator complex subunit 3 (CgMed3ABp) and its regulation of gene expression at low pH in C. glabrata. We found that CgMed3ABp was critical for cellular survival and pyruvate production during low-pH stress. Measures of the levels of plasma membrane fatty acids and sterol composition indicated that CgMed3ABp could play an important role in regulating homeostasis in C. glabrata. We propose that controlling membrane lipid composition may enhance the robustness of C. glabrata for the production of organic acids.

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ERG6 and ERG2 Are Major Targets Conferring Reduced Susceptibility to Amphotericin B in Clinical Candida glabrata Isolates in Kuwait

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01900-18 ◽

2018 ◽

Vol 63 (2) ◽

pp. e01900-18 ◽

Cited By ~ 7

Author(s):

Suhail Ahmad ◽

Leena Joseph ◽

Josie E. Parker ◽

Mohammad Asadzadeh ◽

Steven L. Kelly ◽

...

Keyword(s):

Amphotericin B ◽

Candida Glabrata ◽

Molecular Mechanisms ◽

Total Sterol ◽

Resistant Isolate ◽

Ergosterol Biosynthesis ◽

Sterol Content ◽

Wild Type ◽

Content Type ◽

Synonymous Mutations

ABSTRACT Candida glabrata is intrinsically less susceptible to azoles, and resistance to echinocandins and reduced susceptibility (RS) to amphotericin B (AMB) have also been detected. The molecular mechanisms of RS to AMB were investigated in C. glabrata strains in Kuwait by sequence analyses of genes involved in ergosterol biosynthesis. A total of 1,646 C. glabrata isolates were tested by Etest, and results for 12 selected isolates were confirmed by reference broth microdilution. PCR sequencing of three genes (ERG2, ERG6, and ERG11) was performed for all isolates with RS to AMB (RS-AMB isolates) and 5 selected wild-type C. glabrata isolates by using gene-specific primers. The total cell sterol content was analyzed by gas chromatography-mass spectrometry. The phylogenetic relationship among the isolates was investigated by multilocus sequence typing. Wild-type isolates contained only synonymous mutations in ERG2, ERG6, or ERG11, and the total sterol content was similar to that of the reference strains. A nonsynonymous ERG6 mutation (AGA48AAA, R48K) was found in both RS-AMB and wild-type isolates. Four RS-AMB isolates contained novel nonsense mutations at Trp286, Tyr192, and Leu341, and 2 isolates contained a nonsynonymous mutation in ERG6 (V126F or C198F); and the sterol content of these isolates was consistent with ERG6 deficiency. Two other RS-AMB isolates contained a novel nonsynonymous ERG2 mutation (G119S or G122S), and their sterol content was consistent with ERG2 deficiency. Of 8 RS-AMB isolates, 1 fluconazole-resistant isolate also contained nonsynonymous Y141H plus L381M mutations, while 7 isolates contained only synonymous mutations in ERG11. All isolates with ERG6, ERG2, and ERG11 mutations were genotypically distinct strains. Our data show that ERG6 and ERG2 are major targets conferring RS-AMB in clinical C. glabrata isolates.

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Rezafungin In Vitro Activity against Contemporary Nordic Clinical Candida Isolates and Candida auris Determined by the EUCAST Reference Method

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02438-19 ◽

2020 ◽

Vol 64 (4) ◽

Cited By ~ 3

Author(s):

Marie Helleberg ◽

Karin Meinike Jørgensen ◽

Rasmus Krøger Hare ◽

Raluca Datcu ◽

Anuradha Chowdhary ◽

...

Keyword(s):

Amphotericin B ◽

Target Genes ◽

Specific Activity ◽

Significant Proportion ◽

Reference Method ◽

Cross Resistance ◽

Wild Type ◽

Candida Auris ◽

Content Type

ABSTRACT Rezafungin (formerly CD101) is a novel echinocandin in clinical development. EUCAST epidemiological cutoff values (ECOFFs) have not yet been established. We determined the in vitro activity of rezafungin and comparators against 1,293 Nordic yeast isolates and 122 Indian Candida auris isolates and established single-center wild-type upper limits (WT-UL). The isolates (19 Candida spp. and 13 other yeast species) were identified using Chromagar; matrix-assisted laser desorption ionization–time of flight (MALDI-TOF); and, when needed, internal transcribed spacer sequencing. EUCAST E.Def 7.3.1 susceptibility testing included rezafungin, anidulafungin, micafungin, amphotericin B, and fluconazole. WT-UL were established following EUCAST principles for visual and statistical ECOFF setting. fks target genes were sequenced for rezafungin non-wild-type isolates. EUCAST clinical breakpoints for fungi version 9.0 were adopted for susceptibility classification. Rezafungin had species-specific activity similar to that of anidulafungin and micafungin. On a milligram-per-liter basis, rezafungin was overall less active than anidulafungin and micafungin but equally or more active than fluconazole and amphotericin B against the most common Candida species, except C. parapsilosis. We identified 37 (3.1%) rezafungin non-wild-type isolates of C. albicans (1.9%), C. glabrata (3.0%), C. tropicalis (2.7%), C. dubliniensis (2.9%), C. krusei (1.2%), and C. auris (14.8%). Alterations in Fks hot spots were found in 26/26 Nordic and 8/18 non-wild-type C. auris isolates. Rezafungin displayed broad in vitro activity against Candida spp., including C. auris. Adopting WT-UL established here, few Nordic strains, but a significant proportion of C. auris isolates, had elevated MICs with mutations in fks target genes that conferred echinocandin cross-resistance. fks1 mutations raised rezafungin MICs notably less than anidulafungin and micafungin MICs in C. auris.

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Molecular Mechanisms of Drug Resistance in Clinical Candida Species Isolated from Tunisian Hospitals

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00555-13 ◽

2013 ◽

Vol 57 (7) ◽

pp. 3182-3193 ◽

Cited By ~ 67

Author(s):

Jamel Eddouzi ◽

Josie E. Parker ◽

Luis A. Vale-Silva ◽

Alix Coste ◽

Françoise Ischer ◽

...

Keyword(s):

Drug Resistance ◽

Molecular Mechanisms ◽

Major Facilitator Superfamily ◽

Single Amino Acid ◽

Cross Resistance ◽

Clinical Samples ◽

Ergosterol Biosynthesis ◽

Toxic Metabolite ◽

Wild Type ◽

Content Type

ABSTRACTAntifungal resistance ofCandidaspecies is a clinical problem in the management of diseases caused by these pathogens. In this study we identified from a collection of 423 clinical samples taken from Tunisian hospitals two clinicalCandidaspecies (Candida albicansJEY355 andCandida tropicalisJEY162) with decreased susceptibility to azoles and polyenes. For JEY355, the fluconazole (FLC) MIC was 8 μg/ml. Azole resistance inC. albicansJEY355 was mainly caused by overexpression of a multidrug efflux pump of the major facilitator superfamily, Mdr1. The regulator of Mdr1,MRR1, contained a yet-unknown gain-of-function mutation (V877F) causingMDR1overexpression. TheC. tropicalisJEY162 isolate demonstrated cross-resistance between FLC (MIC > 128 μg/ml), voriconazole (MIC > 16 μg/ml), and amphotericin B (MIC > 32 μg/ml). Sterol analysis using gas chromatography-mass spectrometry revealed that ergosterol was undetectable in JEY162 and that it accumulated 14α-methyl fecosterol, thus indicating a perturbation in the function of at least two main ergosterol biosynthesis proteins (Erg11 and Erg3). Sequence analyses ofC. tropicalis ERG11(CtERG11) andCtERG3from JEY162 revealed a deletion of 132 nucleotides and a single amino acid substitution (S258F), respectively. These two alleles were demonstrated to be nonfunctional and thus are consistent with previous studies showing thatERG11mutants can only survive in combination with otherERG3mutations.CtERG3andCtERG11wild-type alleles were replaced by the defective genes in a wild-typeC. tropicalisstrain, resulting in a drug resistance phenotype identical to that of JEY162. This genetic evidence demonstrated thatCtERG3andCtERG11mutations participated in drug resistance. During reconstitution of the drug resistance inC. tropicalis, a strain was obtained harboring only defectiveCterg11allele and containing as a major sterol the toxic metabolite 14α-methyl-ergosta-8,24(28)-dien-3α,6β-diol, suggesting thatERG3was still functional. This strain therefore challenged the current belief thatERG11mutations cannot be viable unless accompanied by compensatory mutations. In conclusion, this study, in addition to identifying a novelMRR1mutation inC. albicans, constitutes the first report on a clinicalC. tropicaliswith defective activity of sterol 14α-demethylase and sterol Δ5,6-desaturase leading to azole-polyene cross-resistance.

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Antifungal Activity of SCY-078 and Standard Antifungal Agents against 178 Clinical Isolates of Resistant and Susceptible Candida Species

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01102-17 ◽

2017 ◽

Vol 61 (11) ◽

Cited By ~ 20

Author(s):

Wiley A. Schell ◽

A. M. Jones ◽

Katyna Borroto-Esoda ◽

Barbara D. Alexander

Keyword(s):

Candida Glabrata ◽

Antifungal Agents ◽

Candida Parapsilosis ◽

Candida Krusei ◽

Wild Type ◽

Content Type ◽

Clinical Resistance ◽

Excellent Activity ◽

Candida Lusitaniae

ABSTRACT SCY-078 in vitro activity was determined for 178 isolates of resistant or susceptible Candida albicans, Candida dubliniensis, Candida glabrata, Candida krusei, Candida lusitaniae, and Candida parapsilosis, including 44 Candida isolates with known genotypic (FKS1 or FKS2 mutations), phenotypic, or clinical resistance to echinocandins. Results were compared to those for anidulafungin, caspofungin, micafungin, fluconazole, and voriconazole. SCY-078 was shown to have excellent activity against both wild-type isolates and echinocandin- and azole-resistant isolates of Candida species.

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Characterization of the Proliferating Cell Nuclear Antigen of Leishmania donovani Clinical Isolates and Its Association with Antimony Resistance

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01847-13 ◽

2014 ◽

Vol 58 (6) ◽

pp. 2997-3007 ◽

Cited By ~ 9

Author(s):

Rati Tandon ◽

Sharat Chandra ◽

Rajendra Kumar Baharia ◽

Sanchita Das ◽

Pragya Misra ◽

...

Keyword(s):

Clinical Isolate ◽

Proliferating Cell Nuclear Antigen ◽

Leishmania Donovani ◽

Clinical Isolates ◽

Resistant Isolate ◽

Nuclear Antigen ◽

Wild Type ◽

Content Type ◽

Sensitive Isolate ◽

Proliferating Cell

ABSTRACTPreviously, through a proteomic analysis, proliferating cell nuclear antigen (PCNA) was found to be overexpressed in the sodium antimony gluconate (SAG)-resistant clinical isolate compared to that in the SAG-sensitive clinical isolate ofLeishmania donovani. The present study was designed to explore the potential role of the PCNA protein in SAG resistance inL. donovani. For this purpose, the protein was cloned, overexpressed, purified, and modeled. Western blot (WB) and real-time PCR (RT-PCR) analyses confirmed that PCNA was overexpressed by ≥3-fold in the log phase, stationary phase, and peanut agglutinin isolated procyclic and metacyclic stages of the promastigote form and by ∼5-fold in the amastigote form of the SAG-resistant isolate compared to that in the SAG-sensitive isolate.L. donovaniPCNA (LdPCNA) was overexpressed as a green fluorescent protein (GFP) fusion protein in a SAG-sensitive clinical isolate ofL. donovani, and modulation of the sensitivities of the transfectants to pentavalent antimonial (SbV) and trivalent antimonial (SbIII) drugs was assessedin vitroagainst promastigotes and intracellular (J774A.1 cell line) amastigotes, respectively. Overexpression of LdPCNA in the SAG-sensitive isolate resulted in an increase in the 50% inhibitory concentrations (IC50) of SbV(from 41.2 ± 0.6 μg/ml to 66.5 ± 3.9 μg/ml) and SbIII(from 24.0 ± 0.3 μg/ml to 43.4 ± 1.8 μg/ml). Moreover, PCNA-overexpressing promastigote transfectants exhibited less DNA fragmentation compared to that of wild-type SAG-sensitive parasites upon SbIIItreatment. In addition, SAG-induced nitric oxide (NO) production was found to be significantly inhibited in the macrophages infected with the transfectants compared with that in wild-type SAG-sensitive parasites. Consequently, we infer that LdPCNA has a significant role in SAG resistance inL. donovaniclinical isolates, which warrants detailed investigations regarding its mechanism.

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Single Amino Acid Substitutions in HXT2.4 from Scheffersomyces stipitis Lead to Improved Cellobiose Fermentation by Engineered Saccharomyces cerevisiae

Applied and Environmental Microbiology ◽

10.1128/aem.03253-12 ◽

2012 ◽

Vol 79 (5) ◽

pp. 1500-1507 ◽

Cited By ~ 25

Author(s):

Suk-Jin Ha ◽

Heejin Kim ◽

Yuping Lin ◽

Myoung-Uoon Jang ◽

Jonathan M. Galazka ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Amino Acids ◽

Single Amino Acid ◽

Kinetic Properties ◽

Wild Type ◽

Scheffersomyces Stipitis ◽

Content Type ◽

Charged Amino Acids ◽

Cellodextrin Transporter ◽

Engineered Strain

ABSTRACTSaccharomyces cerevisiaecannot utilize cellobiose, but this yeast can be engineered to ferment cellobiose by introducing both cellodextrin transporter (cdt-1) and intracellular β-glucosidase (gh1-1) genes fromNeurospora crassa. Here, we report that an engineeredS. cerevisiaestrain expressing the putative hexose transporter geneHXT2.4fromScheffersomyces stipitisandgh1-1can also ferment cellobiose. This result suggests that HXT2.4p may function as a cellobiose transporter whenHXT2.4is overexpressed inS. cerevisiae. However, cellobiose fermentation by the engineered strain expressingHXT2.4andgh1-1was much slower and less efficient than that by an engineered strain that initially expressedcdt-1andgh1-1. The rate of cellobiose fermentation by theHXT2.4-expressing strain increased drastically after serial subcultures on cellobiose. Sequencing and retransformation of the isolated plasmids from a single colony of the fast cellobiose-fermenting culture led to the identification of a mutation (A291D) in HXT2.4 that is responsible for improved cellobiose fermentation by the evolvedS. cerevisiaestrain. Substitutions for alanine (A291) of negatively charged amino acids (A291E and A291D) or positively charged amino acids (A291K and A291R) significantly improved cellobiose fermentation. The mutant HXT2.4(A291D) exhibited 1.5-fold higherKmand 4-fold higherVmaxvalues than those from wild-type HXT2.4, whereas the expression levels were the same. These results suggest that the kinetic properties of wild-type HXT2.4 expressed inS. cerevisiaeare suboptimal, and mutations of A291 into bulky charged amino acids might transform HXT2.4p into an efficient transporter, enabling rapid cellobiose fermentation by engineeredS. cerevisiaestrains.

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Molecular Analysis of Resistance and Detection of Non-Wild-Type Strains Using Etest Epidemiological Cutoff Values for Amphotericin B and Echinocandins for Bloodstream Candida Infections from a Tertiary Hospital in Qatar

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00214-18 ◽

2018 ◽

Vol 62 (9) ◽

Cited By ~ 4

Author(s):

Saad J. Taj-Aldeen ◽

Husam Salah ◽

Winder B. Perez ◽

Muna Almaslamani ◽

Mary Motyl ◽

...

Keyword(s):

Amphotericin B ◽

Hot Spot ◽

Antifungal Susceptibility ◽

Tertiary Hospital ◽

Wild Type ◽

Content Type ◽

Cutoff Values ◽

Echinocandin Resistance ◽

Candida Infections ◽

Type Strains

ABSTRACT A total of 301 Candida bloodstream isolates collected from 289 patients over 5 years at a tertiary hospital in Qatar were evaluated. Out of all Candida infections, 53% were diagnosed in patients admitted to the intensive care units. Steady increases in non-albicans Candida species were reported from 2009 to 2014 (30.2% for Candida albicans versus 69.8% for the other Candida species). Etest antifungal susceptibility testing was performed on all recovered clinical isolates to determine echinocandin (micafungin and anidulafungin) and amphotericin B susceptibilities and assess non-wild-type (non-WT) strains (strains for which MICs were above the epidemiological cutoff values). DNA sequence analysis was performed on all isolates to assess the presence of FKS mutations, which confer echinocandin resistance in Candida species. A total of 3.9% of isolates (12/301) among strains of C. albicans and C. orthopsilosis contained FKS hot spot mutations, including heterozygous mutations in FKS1. For C. tropicalis, the Etest appeared to overestimate strains non-WT for micafungin, anidulafungin, and amphotericin B, as 14%, 11%, and 35% of strains, respectively, had values above the epidemiological cutoff value. However, no FKS mutations were identified in this species. For all other species, micafungin best reported the echinocandin non-WT strains relative to the FKS genotype, as anidulafungin tended to overestimate non-wild-type strains. Besides C. tropicalis, few strains were classified as non-WT for amphotericin B.

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In Vivo Efficacy of Liposomal Amphotericin B against Wild-Type and Azole-Resistant Aspergillus fumigatus Isolates in Two Different Immunosuppression Models of Invasive Aspergillosis

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02479-16 ◽

2017 ◽

Vol 61 (6) ◽

Cited By ~ 3

Author(s):

Seyedmojtaba Seyedmousavi ◽

Johan W. Mouton ◽

Willem J. G. Melchers ◽

Paul E. Verweij

Keyword(s):

Invasive Aspergillosis ◽

Aspergillus Fumigatus ◽

Amphotericin B ◽

Liposomal Amphotericin ◽

Treated Group ◽

Liposomal Amphotericin B ◽

Wild Type ◽

Resistance Mutations ◽

Content Type

ABSTRACT Using an immunocompetent murine model of invasive aspergillosis (IA), we previously reported that the efficacy of liposomal amphotericin B (L-AmB) (Ambisome) is not hampered by the presence of azole resistance mutations in Aspergillus fumigatus (S. Seyedmousavi, W. J. G. Melchers, J. W. Mouton, and P. E. Verweij, Antimicrob Agents Chemother 57:1866–1871, 2013, https://doi.org/10.1128/AAC.02226-12 ). We here investigated the role of immune suppression, i.e., neutropenia and steroid treatment, in L-AmB efficacy in mice infected with wild-type (WT) A. fumigatus and with azole-resistant A. fumigatus harboring a TR34/L98H mutation in the cyp-51A gene. Survival of treated animals at day 14 in both immunosuppressed models was significantly better than that of nontreated controls. A dose-response relationship was observed that was independent of the azole-resistant mechanism and the immunosuppression method used. In the neutropenic model, 100% survival was reached at an L-AmB dose of 16 mg/kg of body weight for the WT strain and the TR34/L98H isolate. In the steroid-treated group, 90.9% survival and 100% survival were achieved for the WT isolate and the TR34/L98H isolate with an L-AmB dose of 16 mg/kg, respectively. The 50% effective dose (ED50) was 1.40 mg/kg (95% confidence interval [CI], 0.66 to 3.00 mg/kg) for the WT isolate and 1.92 mg/kg (95% CI, 0.60 to 6.17 mg/kg) for the TR34/L98H isolate in the neutropenic model and was 2.40 mg/kg (95% CI, 1.93 to 2.97 mg/kg) for the WT isolate and 2.56 mg/kg (95% CI, 1.43 to 4.56 mg/kg) for the TR34/L98H isolate in the steroid-treated group. Overall, there were no significant differences between the two different immunosuppressed conditions in the efficacy of L-AmB against the wild-type and azole-resistant isolates (P > 0.9). However, the required L-AmB exposure was significantly higher than that seen in the immunocompetent model.

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Manogepix (APX001A) Displays Potent In Vitro Activity against Human Pathogenic Yeast, but with an Unexpected Correlation to Fluconazole MICs

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00429-20 ◽

2020 ◽

Vol 64 (7) ◽

Cited By ~ 4

Author(s):

Maiken Cavling Arendrup ◽

Karin Meinike Jørgensen

Keyword(s):

Amphotericin B ◽

Broad Spectrum ◽

Hot Spot ◽

Its Sequencing ◽

Wild Type ◽

Pathogenic Yeast ◽

Fluconazole Resistance ◽

Content Type ◽

Spectrum Activity ◽

Broad Spectrum Activity

ABSTRACT Manogepix (APX001A) is the active moiety of the novel drug candidate fosmanogepix (APX001). We previously reported the broad-spectrum activity of manogepix but also observed a correlation between increased manogepix and fluconazole MICs. Here, we extended this study and included isolates with acquired fluconazole resistance. Isolates (n = 835) were identified using CHROMagar, matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS), and, when needed, internal transcribed spacer (ITS) sequencing. EUCAST E.Def 7.3.1 susceptibility testing included manogepix, amphotericin B, anidulafungin, micafungin, fluconazole, and voriconazole. Manogepix wild-type-upper-limit (WT-UL) values were established following EUCAST principles for the epidemiological cutoff value (ECOFF) setting allowing wild-type/non-wild-type classification. Drug-specific MIC correlations were investigated using Pearson’s correlation. Manogepix modal MICs were low (range, 0.004 to 0.06 mg/liter against 16/20 included species). Exceptions were Candida krusei and Candida inconspicua and, to a lesser extent, Candida kefyr and Pichia kluyveri. The activity was independent of Fks echinocandin hot spot alterations (n = 17). Adopting the WT-UL established for Candida albicans, Candida dubliniensis, Candida glabrata, Candida parapsilosis, and Candida tropicalis, 14/724 (1.9%) isolates were non-wild type for manogepix. Twelve of these (85.7%) were also non-wild type for fluconazole. A statistically significant correlation was observed between manogepix and fluconazole MICs for C. albicans, C. dubliniensis, C. glabrata, C. parapsilosis, and C. tropicalis (Pearson’s r = 0.401 to 0.575) but not between manogepix and micafungin or amphotericin B MICs for any species except C. tropicalis (r = 0.519 for manogepix versus micafungin). Broad-spectrum activity was confirmed for manogepix against contemporary yeast. However, a 1 to 4 2-fold dilutions increase in manogepix MICs is observed in a subset of isolates with acquired fluconazole resistance. Further studies on the potential underlying mechanism and implication for optimal dosing are warranted.

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