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Evaluation of a Novel FKS1 R1354H Mutation Associated with Caspofungin Resistance in Candida auris Using the CRISPR-Cas9 System

Outbreaks of invasive infections, with high mortality rates, caused by multidrug-resistant Candida auris have been reported worldwide. Although hotspot mutations in FKS1 are an established cause of echinocandin resistance, the actual contribution of these mutations to echinocandin resistance remains...

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Autores principales: Kiyohara, Maiko, Miyazaki, Taiga, Okamoto, Michiyo, Hirayama, Tatsuro, Makimura, Koichi, Chibana, Hiroji, Nakada, Nana, Ito, Yuya, Sumiyoshi, Makoto, Ashizawa, Nobuyuki, Takeda, Kazuaki, Iwanaga, Naoki, Takazono, Takahiro, Izumikawa, Koichi, Yanagihara, Katsunori, Kohno, Shigeru, Mukae, Hiroshi
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10219442/
https://www.ncbi.nlm.nih.gov/pubmed/37233240
http://dx.doi.org/10.3390/jof9050529
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author Kiyohara, Maiko
Miyazaki, Taiga
Okamoto, Michiyo
Hirayama, Tatsuro
Makimura, Koichi
Chibana, Hiroji
Nakada, Nana
Ito, Yuya
Sumiyoshi, Makoto
Ashizawa, Nobuyuki
Takeda, Kazuaki
Iwanaga, Naoki
Takazono, Takahiro
Izumikawa, Koichi
Yanagihara, Katsunori
Kohno, Shigeru
Mukae, Hiroshi
author_facet Kiyohara, Maiko
Miyazaki, Taiga
Okamoto, Michiyo
Hirayama, Tatsuro
Makimura, Koichi
Chibana, Hiroji
Nakada, Nana
Ito, Yuya
Sumiyoshi, Makoto
Ashizawa, Nobuyuki
Takeda, Kazuaki
Iwanaga, Naoki
Takazono, Takahiro
Izumikawa, Koichi
Yanagihara, Katsunori
Kohno, Shigeru
Mukae, Hiroshi
author_sort Kiyohara, Maiko
collection PubMed
description Outbreaks of invasive infections, with high mortality rates, caused by multidrug-resistant Candida auris have been reported worldwide. Although hotspot mutations in FKS1 are an established cause of echinocandin resistance, the actual contribution of these mutations to echinocandin resistance remains unknown. Here, we sequenced the FKS1 gene of a caspofungin-resistant clinical isolate (clade I) and identified a novel resistance mutation (G4061A inducing R1354H). We applied the clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 system to generate a recovered strain (H1354R) in which only this single nucleotide mutation was reverted to its wild-type sequence. We also generated mutant strains with only the R1354H mutation introduced into C. auris wild-type strains (clade I and II) and analyzed their antifungal susceptibility. Compared to their parental strains, the R1354H mutants exhibited a 4- to 16-fold increase in caspofungin minimum inhibitory concentration (MIC) while the H1354R reverted strain exhibited a 4-fold decrease in caspofungin MIC. In a mouse model of disseminated candidiasis, the in vivo therapeutic effect of caspofungin was more closely related to the FKS1 R1354H mutation and the virulence of the strain than its in vitro MIC. The CRISPR-Cas9 system could thus aid in elucidating the mechanism underlying drug resistance in C. auris.
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spelling pubmed-102194422023-05-27 Evaluation of a Novel FKS1 R1354H Mutation Associated with Caspofungin Resistance in Candida auris Using the CRISPR-Cas9 System Kiyohara, Maiko Miyazaki, Taiga Okamoto, Michiyo Hirayama, Tatsuro Makimura, Koichi Chibana, Hiroji Nakada, Nana Ito, Yuya Sumiyoshi, Makoto Ashizawa, Nobuyuki Takeda, Kazuaki Iwanaga, Naoki Takazono, Takahiro Izumikawa, Koichi Yanagihara, Katsunori Kohno, Shigeru Mukae, Hiroshi J Fungi (Basel) Article Outbreaks of invasive infections, with high mortality rates, caused by multidrug-resistant Candida auris have been reported worldwide. Although hotspot mutations in FKS1 are an established cause of echinocandin resistance, the actual contribution of these mutations to echinocandin resistance remains unknown. Here, we sequenced the FKS1 gene of a caspofungin-resistant clinical isolate (clade I) and identified a novel resistance mutation (G4061A inducing R1354H). We applied the clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 system to generate a recovered strain (H1354R) in which only this single nucleotide mutation was reverted to its wild-type sequence. We also generated mutant strains with only the R1354H mutation introduced into C. auris wild-type strains (clade I and II) and analyzed their antifungal susceptibility. Compared to their parental strains, the R1354H mutants exhibited a 4- to 16-fold increase in caspofungin minimum inhibitory concentration (MIC) while the H1354R reverted strain exhibited a 4-fold decrease in caspofungin MIC. In a mouse model of disseminated candidiasis, the in vivo therapeutic effect of caspofungin was more closely related to the FKS1 R1354H mutation and the virulence of the strain than its in vitro MIC. The CRISPR-Cas9 system could thus aid in elucidating the mechanism underlying drug resistance in C. auris. MDPI 2023-04-29 /pmc/articles/PMC10219442/ /pubmed/37233240 http://dx.doi.org/10.3390/jof9050529 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Kiyohara, Maiko
Miyazaki, Taiga
Okamoto, Michiyo
Hirayama, Tatsuro
Makimura, Koichi
Chibana, Hiroji
Nakada, Nana
Ito, Yuya
Sumiyoshi, Makoto
Ashizawa, Nobuyuki
Takeda, Kazuaki
Iwanaga, Naoki
Takazono, Takahiro
Izumikawa, Koichi
Yanagihara, Katsunori
Kohno, Shigeru
Mukae, Hiroshi
Evaluation of a Novel FKS1 R1354H Mutation Associated with Caspofungin Resistance in Candida auris Using the CRISPR-Cas9 System
title Evaluation of a Novel FKS1 R1354H Mutation Associated with Caspofungin Resistance in Candida auris Using the CRISPR-Cas9 System
title_full Evaluation of a Novel FKS1 R1354H Mutation Associated with Caspofungin Resistance in Candida auris Using the CRISPR-Cas9 System
title_fullStr Evaluation of a Novel FKS1 R1354H Mutation Associated with Caspofungin Resistance in Candida auris Using the CRISPR-Cas9 System
title_full_unstemmed Evaluation of a Novel FKS1 R1354H Mutation Associated with Caspofungin Resistance in Candida auris Using the CRISPR-Cas9 System
title_short Evaluation of a Novel FKS1 R1354H Mutation Associated with Caspofungin Resistance in Candida auris Using the CRISPR-Cas9 System
title_sort evaluation of a novel fks1 r1354h mutation associated with caspofungin resistance in candida auris using the crispr-cas9 system
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10219442/
https://www.ncbi.nlm.nih.gov/pubmed/37233240
http://dx.doi.org/10.3390/jof9050529
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