Serial passaging of Aspergillus fumigatus on Galleria extract agar produces strains with altered response of oxidative stress

Aspergillus fumigatus is an environmental saprophyte and opportunistic fungal pathogen of the human lung. Serially passaging of A. fumigatus on an agar produced from Galleria mellonella larvae produced strains showing alterations in virulence, anti-fungal susceptibility and in protein abundances that may indicate a degree of adaptation or selection after 25 passages on the artificial agar. While passaged strains demonstrated reduced virulence in G. mellonella larvae, they demonstrated increased tolerance to haemocyte mediated killing, hydrogen peroxide, itraconazole and amphotericin B. Label-free proteomic analysis of control and passaged A. fumigatus strains revealed a total of 3329 proteins of which, 1902 remained following filtration and 32 proteins were statistically significant and differentially abundant. Proteins that play a role in the oxidative stress response were altered in abundance in the passaged strain and included (S)-S-oxide reductase (+2.63 fold), developmental regulator FlbA (+2.27 fold) and histone H2A.Z (-1.82 fold). The results presented here indicate that prolonged culturing of A. fumigatus on Galleria extract agar results in alterations in susceptibility to oxidative stress and in the abundance of proteins associated with the oxidative stress response. The phenomenon may be a result of selection for strains suitable for survival in the adverse conditions imposed by the innate immune response.烟曲霉（Aspergillus fumigatus）是一种环境腐生菌，同时也是人类肺部的机会致病性真菌病原体。研究团队将烟曲霉在大蜡螟（Galleria mellonella）幼虫所制备的琼脂上连续传代后，获得的菌株在毒力、抗真菌药物敏感性以及蛋白质丰度上均出现改变，提示其在该人工琼脂上传代25次后发生了一定程度的适应性演化或筛选。尽管传代菌株在大蜡螟幼虫体内的毒力有所降低，但它们对血细胞介导的杀伤作用、过氧化氢（hydrogen peroxide）、伊曲康唑（itraconazole）以及两性霉素B（amphotericin B）的耐受性显著提升。对对照菌株与传代烟曲霉菌株开展无标记蛋白质组学分析（Label-free proteomic analysis），共鉴定到3329种蛋白质，经过滤后保留1902种，其中32种蛋白质的丰度具有统计学显著性差异。传代菌株中参与氧化应激反应的蛋白质丰度发生改变，包括(S)-S-氧化物还原酶（表达量上调2.63倍）、发育调节因子FlbA（表达量上调2.27倍）以及组蛋白H2A.Z（表达量下调1.82倍）。本研究结果表明，烟曲霉在大蜡螟提取物琼脂上长期培养，会导致其对氧化应激的敏感性以及与氧化应激反应相关的蛋白质丰度发生改变，该现象可能是固有免疫应答所施加的不利环境中，适用于存活的菌株被筛选保留所导致的。