Evolved Heterogeneity Contributes to Chronic Fungal Lung Infections

慢性真菌感染通常不愿接受治疗。慢性感染的顽固性不佳 可以理解，但可能涉及多种因素，包括生物膜的形成和生长缓慢。此外，多样化 对从细菌感染到人类肿瘤的不同系统的研究发现，人群可以 甚至在治疗开始之前，药物敏感性就不同，导致治疗失败。 真菌病原体种群内的异质性还没有得到很好的研究。我们已经确定了慢性 三名囊性纤维化(CF)无关受试者的高负担卢氏弧菌肺部感染。卢西塔尼亚种具有 以前曾在CF肺部感染中记录过，但以卢西坦氏弧菌为主的感染并不广泛 据报道。在所有三个受试者中，每个种群内的卢西坦氏杆菌分离株在抗真菌方面表现出很大的差异。 敏感性，以最低抑菌浓度(MIC)衡量，一些分离物稳定耐药。我们 进一步描述了来自一个对象的分离物，并通过全基因组测序发现 临床耐药相关基因MRR1等位基因异质性与MIC值的关系 在其他念珠菌属中。暴露在毒品中。单个群体中MRR1等位基因的变异令人惊讶，因为 这名受试者在前一年没有开过抗真菌药。由于易于操作， Lusitaniae，一种单倍体，遗传易驯化，交配能力强的酵母，我们建议利用这些菌株 收集研究耐药性的基础，促进选择增加耐药性的因素， 以及促进肺部异质性人口结构的因素。在目标1中，我们将检验假设 MRr1中央调控结构域的替换和mr1C-末端的截断导致增加 活性(1.1)，共同和不同的基因受不同类别的MRr1变体(1.2)控制，以及 差异表达的MRr1调控基因与氟康唑耐药有关(1.3)。在目标2中，我们将测试 假设特定的SNPs影响代谢多样性(2.1)，高MRr1活性促进健康 糖酵解率高的菌株(2.2)，代谢多样化促进药物的异质性 营养复杂环境中的抵抗力(2.3)。因为卢西塔尼亚种有发展的倾向 对两性霉素B等强效药物的耐药性，通常是最后的抗真菌药物，与 在卢氏假丝酵母菌和最近引起关注的多重耐药真菌病原体金黄色念珠菌之间，这些 研究与发现更好地了解慢性感染的组成的方法高度相关 限制抗真菌耐药性的发展，并在耐药菌株形成后与之抗争。

Chronic fungal infections are often recalcitrant to treatment. The recalcitrance of chronic infections is not well understood, but likely involves multiple factors including biofilm formation, and slow growth. In addition, diverse studies in different systems from bacterial infections to human tumors, found that populations can have heterogeneous drug susceptibilities even before treatment is initiated, leading to treatment failures. Heterogeneity within populations of fungal pathogens has not been well studied. We have identified chronic high burden C. lusitaniae lung infections in three unrelated subjects with cystic fibrosis (CF). C. lusitaniae has been documented in CF lung infections previously, but infections dominated by C. lusitaniae are not widely reported. In all three subjects, C. lusitaniae isolates within each population showed high variance in antifungal sensitivity, measured as the minimum inhibitory concentration (MIC), with some isolates stably resistant. We characterized the isolates from one subject further and discovered, through whole genome sequencing, that variable MICs correlated with heterogeneity in alleles of MRR1, a gene associated with clinical drug resistance in other Candida spp. exposed to drugs. Variation in MRR1 alleles within a single population was surprising as this subject had not been prescribed antifungals in the preceding year. Because of the ease of manipulation of C. lusitaniae, a haploid, genetically tractable, mating competent, yeast, we propose to leverage these isolate collections to study the basis for drug resistance, factors that promote selection for increased drug resistance, and factors that promote heterogeneous population structures in the lung. In Aim 1, we will test the hypothesis that substitutions in the Mrr1 central regulatory domain and Mrr1 C-terminal truncations lead to increased activity (1.1), that common and distinct genes are controlled by different classes of Mrr1 variants (1.2), and that differentially expressed Mrr1-regulated genes contribute to fluconazole resistance (1.3). In Aim 2, we will test the hypothesis that specific SNPs impact metabolic diversity (2.1), high Mrr1 activity promotes fitness in isolates with high rates of glycolysis (2.2), and that metabolic diversification promotes heterogeneity in drug resistance in nutritionally complex environments (2.3). Because of the propensity of C. lusitaniae to develop resistance to potent drugs like amphotericin B, often the antifungal of last resort, and the close relationship between C. lusitaniae and a recent multi-drug resistant fungal pathogen of concern, Candida auris, these studies are highly relevant to discovering ways to better understand the composition of chronic infections, to limit the development of antifungal resistance, and to combat resistant isolates once they develop.