Candida aurisにCRISPR-Cas9法を応用した、カスポファンギン耐性に関与する新たなFKS1 R1354H変異の評価

• Main Author: 清原, 舞子
• Format: Other/Unknown Material
• Language: English
• Published: MDPI 2023
• Subjects: Candida auris; echinocandin; caspofungin; antifungal resistance; FKS1; CRISPR-Cas9; taiga
• Online Access: https://nagasaki-u.repo.nii.ac.jp/record/2000366/files/ISYK1546_Kiyohara.pdf

Nagasaki University (長崎大学) 博士(医学) 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. 長崎大学学位論文 学位記番号:博(医歯薬)甲第1546号 学位授与年月日:令和5年9月6日 Author: Maiko Kiyohara, Taiga Miyazaki, Michiyo Okamoto, Tatsuro Hirayama, Koichi Makimura, Hiroji Chibana, Nana Nakada, Yuya Ito, Makoto Sumiyoshi, Nobuyuki Ashizawa, Kazuaki Takeda, Naoki Iwanaga, Takahiro Takazono, Koichi Izumikawa, Katsunori Yanagihara, Shigeru Kohno and Hiroshi Mukae Citation: Journal of Fungi, 9, art. no. 529; 2023 Nagasaki University (長崎大学), 博士(医学) (2023-09-06) doctoral thesis