Mutations in hmg1, Challenging the Paradigm of Clinical Triazole Resistance in Aspergillus fumigatus
Mutations in hmg1, Challenging the Paradigm of Clinical Triazole Resistance in Aspergillus fumigatus
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DOI：
10.1128/mbio.00437-19
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发表时间：
2019-03-01
期刊：
影响因子：
6.4
通讯作者：
Fortwendel, Jarrod R.
中科院分区：
文献类型：
--
作者：
Rybak, Jeffrey M.;Ge, Wenbo;Fortwendel, Jarrod R.
烟曲霉是侵袭性曲霉病的主要病原体，这种疾病每年导致超过 200,000 例危及生命的感染。三唑类抗真菌药在临床上对于治疗侵袭性曲霉病至关重要，既可以作为一线治疗，也可以作为挽救治疗。不幸的是，现在世界范围内越来越多地报道烟曲霉分离株对三唑类药物的耐药性，并且这种耐药性的很大一部分仍然无法解释。在这项工作中，我们在大量高度三唑耐药的临床烟曲霉分离株中，描述了先前确定的三唑耐药机制的贡献，包括甾醇脱甲基酶编码基因 cyp51A 的突变、甾醇脱甲基酶基因的过度表达以及外排泵编码基因 abcC 的过度表达。在揭示这些机制本身不能证实该集合所表现出的大多数三唑耐药性后，我们随后描述了三唑耐药性的新型遗传决定因素的鉴定和表征。在我们收集的大多数三唑耐药临床分离株中发现了 3-羟基-3-甲基-戊二酰辅酶 A (HMG-CoA) 还原酶编码基因 hmg1 的突变。引入三种不同的 hmg1 突变，预计编码 Hmg1 保守甾醇感应结构域中的残基改变，导致对三唑类药物的耐药性显着增加。此外，用新型 Cas9 核糖核蛋白介导的系统纠正烟曲霉的泛三唑耐药临床分离株中的 hmg1 突变，可以恢复对所有三唑药物的临床敏感性。通过甾醇分析，hmg1 突变还被证明会导致麦角甾醇前体（如依布立考）的积累，同时不会改变甾醇脱甲基酶基因的表达。 重要性 烟曲霉是侵袭性曲霉菌病的主要病原体，这种疾病每年会导致超过 200,000 例危及生命的感染。三唑类抗真菌药在临床上对于治疗侵袭性曲霉菌病至关重要。不幸的是，现在世界范围内越来越多地报道了烟曲霉分离株对三唑类的耐药性。在这项工作中，我们挑战了目前烟曲霉临床三唑耐药性的范式，首先证明先前表征的耐药机制对三唑敏感性具有名义影响，随后确定了一种对临床三唑敏感性具有深远影响的新耐药机制。我们证明，HMG-CoA 还原酶基因 hmg1 的突变在耐药临床分离株中很常见，并且 hmg1 突变赋予对所有临床可用的三唑抗真菌药物的耐药性。
Aspergillus fumigatus is the predominant pathogen of invasive aspergillosis, a disease state credited with over 200,000 life-threatening infections each year. The triazole class of antifungals are clinically essential to the treatment of invasive aspergillosis, both as frontline and as salvage therapy. Unfortunately, resistance to the triazoles among A. fumigatus isolates is now increasingly reported worldwide, and a large proportion of this resistance remains unexplained. In this work, we characterize the contributions of previously identified mechanisms of triazole resistance, including mutations in the sterol-demethylase-encoding gene cyp51A, overexpression of sterol-demethylase genes, and overexpression of the efflux pump-encoding gene abcC, among a large collection of highly triazole-resistant clinical A. fumigatus isolates. Upon revealing that these mechanisms alone cannot substantiate the majority of triazole resistance exhibited by this collection, we subsequently describe the identification and characterization of a novel genetic determinant of triazole resistance. Mutations in the 3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase-encoding gene, hmg1, were identified in a majority of triazole-resistant clinical isolates in our collection. Introduction of three different hmg1 mutations, predicted to encode residue alterations in the conserved sterol sensing domain of Hmg1, resulted in significantly increased resistance to the triazole class of agents. Additionally, correction of a hmg1 mutation in a pan-triazole-resistant clinical isolate of A. fumigatus with a novel Cas9-ribonucleoprotein-mediated system was shown to restore clinical susceptibility to all triazole agents. Mutations in hmg1 were also shown to lead to the accumulation of ergosterol precursors, such as eburicol, by sterol profiling, while not altering the expression of sterol-demethylase genes.IMPORTANCE Aspergillus fumigatus is the predominant pathogen of invasive aspergillosis, a disease state credited with over 200,000 life-threatening infections annually. The triazole class of antifungals are clinically essential to the treatment of invasive aspergillosis. Unfortunately, resistance to the triazoles among A. fumigatus isolates is now increasingly reported worldwide. In this work, we challenge the current paradigm of clinical triazole resistance in A. fumigatus, by first demonstrating that previously characterized mechanisms of resistance have nominal impact on triazole susceptibility and subsequently identifying a novel mechanism of resistance with a profound impact on clinical triazole susceptibility. We demonstrate that mutations in the HMG-CoA reductase gene, hmg1, are common among resistant clinical isolates and that hmg1 mutations confer resistance to all clinically available triazole antifungals.