Molecular Characterization and Sterol Profiles Identify Nonsynonymous Mutations in ERG2 as a Major Mechanism Conferring Reduced Susceptibility to Amphotericin B in Candida kefyr
ABSTRACT The molecular basis of reduced susceptibility to amphotericin B (rs-AMB) among any yeasts is poorly defined. Genetic alterations in genes involved in ergosterol biosynthesis and total cell sterols were investigated among clinical Candida kefyr isolates. C. kefyr isolates (n = 81) obtained f...
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| Format: | Article |
| Language: | English |
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American Society for Microbiology
2023-08-01
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| Series: | Microbiology Spectrum |
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| Online Access: | https://journals.asm.org/doi/10.1128/spectrum.01474-23 |
| _version_ | 1797741710875295744 |
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| author | Mohammad Asadzadeh Wadha Alfouzan Josie E. Parker Jacques F. Meis Steven L. Kelly Leena Joseph Suhail Ahmad |
| author_facet | Mohammad Asadzadeh Wadha Alfouzan Josie E. Parker Jacques F. Meis Steven L. Kelly Leena Joseph Suhail Ahmad |
| author_sort | Mohammad Asadzadeh |
| collection | DOAJ |
| description | ABSTRACT The molecular basis of reduced susceptibility to amphotericin B (rs-AMB) among any yeasts is poorly defined. Genetic alterations in genes involved in ergosterol biosynthesis and total cell sterols were investigated among clinical Candida kefyr isolates. C. kefyr isolates (n = 81) obtained from 74 patients in Kuwait and identified by phenotypic and molecular methods were analyzed. An Etest was initially used to identify isolates with rs-AMB. Specific mutations in ERG2 and ERG6 involved in ergosterol biosynthesis were detected by PCR sequencing. Twelve selected isolates were also tested by the SensiTitre Yeast One (SYO), and total cell sterols were evaluated by gas chromatography-mass spectrometry and ERG3 and ERG11 sequencing. Eight isolates from 8 patients showed rs-AMB by Etest, including 2 isolates with additional resistance to fluconazole or to all three antifungals. SYO correctly identified 8 of 8 rs-AMB isolates. A nonsynonymous mutation in ERG2 was detected in 6 of 8 rs-AMB isolates but also in 3 of 73 isolates with a wild-type AMB pattern. One rs-AMB isolate contained a deletion (frameshift) mutation in ERG2. One or more nonsynonymous mutations was detected in ERG6 in 11 of 81 isolates with the rs-AMB or wild-type AMB pattern. Among 12 selected isolates, 2 and 2 isolates contained a nonsynonymous mutation(s) in ERG3 and ERG11, respectively. Ergosterol was undetectable in 7 of 8 rs-AMB isolates, and the total cell sterol profiles were consistent with loss of ERG2 function in 6 rs-AMB isolates and loss of ERG3 activity in another rs-AMB isolate. Our data showed that ERG2 is a major target conferring rs-AMB in clinical C. kefyr isolates. IMPORTANCE Some yeast species exhibit intrinsic resistance or rapidly acquire resistance to azole antifungals. Despite >50 years of clinical use, resistance to amphotericin B (AMB) among yeast species has been extremely rarely reported until recently. Reduced susceptibility to AMB (rs-AMB) among yeast species is, therefore, a matter of serious concern due to the availability of only four classes of antifungal drugs. Recent studies in Candida glabrata, Candida lusitaniae, and Candida auris have identified ERG genes involved in ergosterol biosynthesis as the major targets conferring rs-AMB. The results of this study also show that nonsynonymous mutations in ERG2 impair its function, abolish ergosterol from C. kefyr, and confer rs-AMB. Thus, rapid detection of rs-AMB among clinical isolates will help in proper management of invasive C. kefyr infections. |
| first_indexed | 2024-03-12T14:30:39Z |
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| id | doaj.art-baf9d8e20cba41319bdbbd3ae0696368 |
| institution | Directory Open Access Journal |
| issn | 2165-0497 |
| language | English |
| last_indexed | 2024-03-12T14:30:39Z |
| publishDate | 2023-08-01 |
| publisher | American Society for Microbiology |
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| spelling | doaj.art-baf9d8e20cba41319bdbbd3ae06963682023-08-17T13:04:15ZengAmerican Society for MicrobiologyMicrobiology Spectrum2165-04972023-08-0111410.1128/spectrum.01474-23Molecular Characterization and Sterol Profiles Identify Nonsynonymous Mutations in ERG2 as a Major Mechanism Conferring Reduced Susceptibility to Amphotericin B in Candida kefyrMohammad Asadzadeh0Wadha Alfouzan1Josie E. Parker2Jacques F. Meis3Steven L. Kelly4Leena Joseph5Suhail Ahmad6Department of Microbiology, Faculty of Medicine, Kuwait University, Safat, KuwaitDepartment of Microbiology, Faculty of Medicine, Kuwait University, Safat, KuwaitMolecular Biosciences Division, School of Biosciences, Cardiff University, Cardiff, United KingdomDepartment of Medical Microbiology and Infectious Diseases, Canisius-Wilhelmina Hospital, Nijmegen, the NetherlandsInstitute of Life Science, Faculty of Health, Medicine and Life Sciences, Swansea University, Swansea, Wales, United KingdomDepartment of Microbiology, Faculty of Medicine, Kuwait University, Safat, KuwaitDepartment of Microbiology, Faculty of Medicine, Kuwait University, Safat, KuwaitABSTRACT The molecular basis of reduced susceptibility to amphotericin B (rs-AMB) among any yeasts is poorly defined. Genetic alterations in genes involved in ergosterol biosynthesis and total cell sterols were investigated among clinical Candida kefyr isolates. C. kefyr isolates (n = 81) obtained from 74 patients in Kuwait and identified by phenotypic and molecular methods were analyzed. An Etest was initially used to identify isolates with rs-AMB. Specific mutations in ERG2 and ERG6 involved in ergosterol biosynthesis were detected by PCR sequencing. Twelve selected isolates were also tested by the SensiTitre Yeast One (SYO), and total cell sterols were evaluated by gas chromatography-mass spectrometry and ERG3 and ERG11 sequencing. Eight isolates from 8 patients showed rs-AMB by Etest, including 2 isolates with additional resistance to fluconazole or to all three antifungals. SYO correctly identified 8 of 8 rs-AMB isolates. A nonsynonymous mutation in ERG2 was detected in 6 of 8 rs-AMB isolates but also in 3 of 73 isolates with a wild-type AMB pattern. One rs-AMB isolate contained a deletion (frameshift) mutation in ERG2. One or more nonsynonymous mutations was detected in ERG6 in 11 of 81 isolates with the rs-AMB or wild-type AMB pattern. Among 12 selected isolates, 2 and 2 isolates contained a nonsynonymous mutation(s) in ERG3 and ERG11, respectively. Ergosterol was undetectable in 7 of 8 rs-AMB isolates, and the total cell sterol profiles were consistent with loss of ERG2 function in 6 rs-AMB isolates and loss of ERG3 activity in another rs-AMB isolate. Our data showed that ERG2 is a major target conferring rs-AMB in clinical C. kefyr isolates. IMPORTANCE Some yeast species exhibit intrinsic resistance or rapidly acquire resistance to azole antifungals. Despite >50 years of clinical use, resistance to amphotericin B (AMB) among yeast species has been extremely rarely reported until recently. Reduced susceptibility to AMB (rs-AMB) among yeast species is, therefore, a matter of serious concern due to the availability of only four classes of antifungal drugs. Recent studies in Candida glabrata, Candida lusitaniae, and Candida auris have identified ERG genes involved in ergosterol biosynthesis as the major targets conferring rs-AMB. The results of this study also show that nonsynonymous mutations in ERG2 impair its function, abolish ergosterol from C. kefyr, and confer rs-AMB. Thus, rapid detection of rs-AMB among clinical isolates will help in proper management of invasive C. kefyr infections.https://journals.asm.org/doi/10.1128/spectrum.01474-23Candida kefyrERG2nonsynonymous mutationsreduced susceptibilityamphotericin B |
| spellingShingle | Mohammad Asadzadeh Wadha Alfouzan Josie E. Parker Jacques F. Meis Steven L. Kelly Leena Joseph Suhail Ahmad Molecular Characterization and Sterol Profiles Identify Nonsynonymous Mutations in ERG2 as a Major Mechanism Conferring Reduced Susceptibility to Amphotericin B in Candida kefyr Microbiology Spectrum Candida kefyr ERG2 nonsynonymous mutations reduced susceptibility amphotericin B |
| title | Molecular Characterization and Sterol Profiles Identify Nonsynonymous Mutations in ERG2 as a Major Mechanism Conferring Reduced Susceptibility to Amphotericin B in Candida kefyr |
| title_full | Molecular Characterization and Sterol Profiles Identify Nonsynonymous Mutations in ERG2 as a Major Mechanism Conferring Reduced Susceptibility to Amphotericin B in Candida kefyr |
| title_fullStr | Molecular Characterization and Sterol Profiles Identify Nonsynonymous Mutations in ERG2 as a Major Mechanism Conferring Reduced Susceptibility to Amphotericin B in Candida kefyr |
| title_full_unstemmed | Molecular Characterization and Sterol Profiles Identify Nonsynonymous Mutations in ERG2 as a Major Mechanism Conferring Reduced Susceptibility to Amphotericin B in Candida kefyr |
| title_short | Molecular Characterization and Sterol Profiles Identify Nonsynonymous Mutations in ERG2 as a Major Mechanism Conferring Reduced Susceptibility to Amphotericin B in Candida kefyr |
| title_sort | molecular characterization and sterol profiles identify nonsynonymous mutations in erg2 as a major mechanism conferring reduced susceptibility to amphotericin b in candida kefyr |
| topic | Candida kefyr ERG2 nonsynonymous mutations reduced susceptibility amphotericin B |
| url | https://journals.asm.org/doi/10.1128/spectrum.01474-23 |
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