Zheng Lijun, Xu Yi, Wang Chen, Guo Liangsheng
Department of Ultrasound Medicine, The Second Affiliated Hospital of Soochow University, Suzhou, China.
Department of Pharmacy, The 960th Hospital of PLA, Jinan, China.
Front Microbiol. 2024 Jul 30;15:1450557. doi: 10.3389/fmicb.2024.1450557. eCollection 2024.
The emergence of tolerance to antifungal agents in complicates the treatment of fungal infections. Understanding the mechanisms underlying this tolerance is crucial for developing effective therapeutic strategies.
This study aims to elucidate the genetic and molecular basis of ketoconazole tolerance in , focusing on the roles of chromosomal aneuploidy, Hsp90, and calcineurin.
The wild-type strain SC5314 was exposed to increasing concentrations of ketoconazole (0.015-32 μg/mL) to select for tolerant adaptors. Disk diffusion and spot assays were used to assess tolerance. Whole-genome sequencing identified chromosomal changes in the adaptors. The roles of Hsp90 and calcineurin in maintaining and developing ketoconazole tolerance were investigated using specific inhibitors and knockout strains.
Adaptors exhibited tolerance to ketoconazole concentrations up to 16 μg/mL, a significant increase from the parent strain's inhibition at 0.015 μg/mL. All tolerant adaptors showed amplification of chromosome R, with 29 adaptors having trisomy and one having tetrasomy. This aneuploidy was unstable, reverting to euploidy and losing tolerance in drug-free conditions. Both Hsp90 and calcineurin were essential for maintaining and developing ketoconazole tolerance. Inhibition of these proteins resulted in loss of tolerance. The efflux gene was not required for the development of tolerance. Chromosome R trisomy and tetrasomy induce cross-tolerance to other azole antifungal agents, including clotrimazole and miconazole, but not to other antifungal classes, such as echinocandins and pyrimidines, exemplified by caspofungin and 5-flucytosine.
Ketoconazole tolerance in is mediated by chromosomal aneuploidy, specifically chromosome R amplification, and requires Hsp90 and calcineurin. These findings highlight potential targets for therapeutic intervention to combat antifungal tolerance and improve treatment outcomes.
抗真菌药物耐受性的出现使真菌感染的治疗变得复杂。了解这种耐受性背后的机制对于制定有效的治疗策略至关重要。
本研究旨在阐明白色念珠菌中酮康唑耐受性的遗传和分子基础，重点关注染色体非整倍性、热休克蛋白90（Hsp90）和钙调神经磷酸酶的作用。
将野生型白色念珠菌菌株SC5314暴露于浓度不断增加的酮康唑（0.015 - 32μg/mL）中，以筛选出耐受性适应株。采用纸片扩散法和点试验评估耐受性。全基因组测序确定了适应株中的染色体变化。使用特异性抑制剂和基因敲除菌株研究Hsp90和钙调神经磷酸酶在维持和发展酮康唑耐受性中的作用。
适应株对高达16μg/mL的酮康唑浓度表现出耐受性，与亲本菌株在0.015μg/mL时的抑制情况相比有显著提高。所有耐受性适应株均显示染色体R扩增，其中29个适应株为三体，1个为四体。这种非整倍性不稳定，在无药物条件下会恢复为整倍体并失去耐受性。Hsp90和钙调神经磷酸酶对于维持和发展酮康唑耐受性均至关重要。抑制这些蛋白会导致耐受性丧失。耐受性的发展不需要外排基因。染色体R三体和四体诱导对其他唑类抗真菌药物（包括克霉唑和咪康唑）的交叉耐受性，但对其他抗真菌类别（如棘白菌素类和嘧啶类，以卡泊芬净和5-氟胞嘧啶为例）则无交叉耐受性。
白色念珠菌中的酮康唑耐受性由染色体非整倍性介导，特别是染色体R扩增，并且需要Hsp90和钙调神经磷酸酶。这些发现突出了对抗真菌耐受性和改善治疗结果的治疗干预潜在靶点。
