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Development and validation of a colorimetric antifungal susceptibility testing method for the dimorphic fungus Talaromyces marneffei
Antifungal drug resistance is an emerging cause of treatment failure in invasive fungal infections, and antifungal susceptibility testing (AFST) may inform treatment decisions. Currently, there are no established AFST guidelines for Talaromyces marneffei (Tm) or other dimorphic fungi. We developed a...
Autores principales: | , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Oxford University Press
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10681740/ https://www.ncbi.nlm.nih.gov/pubmed/37994652 http://dx.doi.org/10.1093/mmy/myad111 |
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author | Thu, Nguyen Thi Mai Borda, Hannah Vitsupakorn, Shawin Reddy, Kaushik Sreerama Kasmani, Navsin Barwatt, Joseph Schwartz, Ilan S Giamberardino, Charles Perfect, John R Hoa, Ngo Thi Le, Thuy |
author_facet | Thu, Nguyen Thi Mai Borda, Hannah Vitsupakorn, Shawin Reddy, Kaushik Sreerama Kasmani, Navsin Barwatt, Joseph Schwartz, Ilan S Giamberardino, Charles Perfect, John R Hoa, Ngo Thi Le, Thuy |
author_sort | Thu, Nguyen Thi Mai |
collection | PubMed |
description | Antifungal drug resistance is an emerging cause of treatment failure in invasive fungal infections, and antifungal susceptibility testing (AFST) may inform treatment decisions. Currently, there are no established AFST guidelines for Talaromyces marneffei (Tm) or other dimorphic fungi. We developed a colorimetric AFST method using a fluorescent redox indicator alamarBlue, which changes from blue to pink in proportion to cellular metabolic activity. We determined the optimal time for alamarBlue addition to be 24 h post-inoculation and for MIC reading to be 72 h post-inoculation. Our method allows three ways to determine minimum inhibitory concentration (MIC): visual inspection of color change, optical density, and fluorescence intensity. We validated the assay by determining the MICs for seven antifungals against 32 Tm clinical isolates and assessed the essential agreement (EA) and inter-rater reliability between our alamarBlue and the Clinical Laboratory Standard Institute (CLSI) broth microdilution methods. The MIC ranges (from low to high) were: 0.008–0.025 μg/ml for itraconazole, 0.004–0.13 μg/ml for voriconazole, 0.03–0.13 μg/ml for posaconazole, 0.06–0.5 µg/ml for flucytosine, 0.5–1 µg/ml for amphotericin B, 0.5–4 µg/ml for caspofungin, and 0.5–16 µg/ml for fluconazole. The EAs were 100% between all three MIC readouts of the alamarBlue method, and 94%–100% between the alamarBlue and CLSI methods. Our alamarBlue method had substantially higher inter-rater agreement and offers a more reliable method that can be standardized across laboratories in both high- and low-resource settings compared to the established CLSI methodology. |
format | Online Article Text |
id | pubmed-10681740 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | Oxford University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-106817402023-11-22 Development and validation of a colorimetric antifungal susceptibility testing method for the dimorphic fungus Talaromyces marneffei Thu, Nguyen Thi Mai Borda, Hannah Vitsupakorn, Shawin Reddy, Kaushik Sreerama Kasmani, Navsin Barwatt, Joseph Schwartz, Ilan S Giamberardino, Charles Perfect, John R Hoa, Ngo Thi Le, Thuy Med Mycol Original Article Antifungal drug resistance is an emerging cause of treatment failure in invasive fungal infections, and antifungal susceptibility testing (AFST) may inform treatment decisions. Currently, there are no established AFST guidelines for Talaromyces marneffei (Tm) or other dimorphic fungi. We developed a colorimetric AFST method using a fluorescent redox indicator alamarBlue, which changes from blue to pink in proportion to cellular metabolic activity. We determined the optimal time for alamarBlue addition to be 24 h post-inoculation and for MIC reading to be 72 h post-inoculation. Our method allows three ways to determine minimum inhibitory concentration (MIC): visual inspection of color change, optical density, and fluorescence intensity. We validated the assay by determining the MICs for seven antifungals against 32 Tm clinical isolates and assessed the essential agreement (EA) and inter-rater reliability between our alamarBlue and the Clinical Laboratory Standard Institute (CLSI) broth microdilution methods. The MIC ranges (from low to high) were: 0.008–0.025 μg/ml for itraconazole, 0.004–0.13 μg/ml for voriconazole, 0.03–0.13 μg/ml for posaconazole, 0.06–0.5 µg/ml for flucytosine, 0.5–1 µg/ml for amphotericin B, 0.5–4 µg/ml for caspofungin, and 0.5–16 µg/ml for fluconazole. The EAs were 100% between all three MIC readouts of the alamarBlue method, and 94%–100% between the alamarBlue and CLSI methods. Our alamarBlue method had substantially higher inter-rater agreement and offers a more reliable method that can be standardized across laboratories in both high- and low-resource settings compared to the established CLSI methodology. Oxford University Press 2023-11-22 /pmc/articles/PMC10681740/ /pubmed/37994652 http://dx.doi.org/10.1093/mmy/myad111 Text en © The Author(s) 2023. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. https://creativecommons.org/licenses/by-nc/4.0/This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (https://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com |
spellingShingle | Original Article Thu, Nguyen Thi Mai Borda, Hannah Vitsupakorn, Shawin Reddy, Kaushik Sreerama Kasmani, Navsin Barwatt, Joseph Schwartz, Ilan S Giamberardino, Charles Perfect, John R Hoa, Ngo Thi Le, Thuy Development and validation of a colorimetric antifungal susceptibility testing method for the dimorphic fungus Talaromyces marneffei |
title | Development and validation of a colorimetric antifungal susceptibility testing method for the dimorphic fungus Talaromyces marneffei |
title_full | Development and validation of a colorimetric antifungal susceptibility testing method for the dimorphic fungus Talaromyces marneffei |
title_fullStr | Development and validation of a colorimetric antifungal susceptibility testing method for the dimorphic fungus Talaromyces marneffei |
title_full_unstemmed | Development and validation of a colorimetric antifungal susceptibility testing method for the dimorphic fungus Talaromyces marneffei |
title_short | Development and validation of a colorimetric antifungal susceptibility testing method for the dimorphic fungus Talaromyces marneffei |
title_sort | development and validation of a colorimetric antifungal susceptibility testing method for the dimorphic fungus talaromyces marneffei |
topic | Original Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10681740/ https://www.ncbi.nlm.nih.gov/pubmed/37994652 http://dx.doi.org/10.1093/mmy/myad111 |
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