Basic Studies On Aspergillus Species

曲霉属的基础研究

• 批准号: 9354725
• 负责人: Kyung Kwon-Chung
• 金额: $ 38.42万
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9354725
• 关键词: Adrenal Cortex Hormones; Affect; Aspergillosis; Aspergillus; Aspergillus fumigatus; Assimilations; Azoles; Biological; Breathing; Catabolism; Cells; Characteristics; Childhood; Chromosomes; Chronic Granulomatous Disease; Color; Copper; Disease; Drug resistance; Enabling Factors; Environment; Exposure to; Fatty Acids; Frequencies; Fungal Drug Resistance; Genes; Genetic; Genetic Recombination; Genetic study; Goals; Growth; Heating; Host Defense; Human; Hyphae; Immune; Immunocompromised Host; Individual; Infection; Interferons; Iron; Life; Mating Types; Meiosis; Messenger RNA; Metabolism; Methods; Mitotic; Modeling; Molecular; Mus; Mutate; Mutation; Neutropenia; Partner in relationship; Patients; Phagocytes; Phenotype; Polyenes; Production; Proteins; Reactive Oxygen Species; Refractory; Reporting; Reproduction; Reproduction spores; Resistance; Sister; System; Systemic disease; Systemic infection; Virulence; Virulence Factors; Virulent; Vitamin K 3; differential expression; fungus; genome sequencing; glyoxylate; interest; mouse model; mutant; neutrophil; novel therapeutics; pathogen; peroxisome; resistance gene; response; tool; trait; ward; whole genome

Aspergillus is one of the most common fungal pathogens affecting neutropenic patients and other types of immunocompromised individuals such as those with Chronic Granulomatous Disease(CGD) of Childhood. Among a dozen species of Aspergillus reported to cause infection in humans, A. fumigatus is the most common species reported to cause invasive aspergillosis. All Aspergillus species propagate by conidia (spores), which humans encounter daily through inhalation. During 2007-2008, we investigated the Aspergillus response to host immune cells by studying the genes differentially expressed in conidia vs hyphae when challenged with neutrophils from healthy donors as well as from those with chronic granulomatous disease (CGD). To our knowledge, this is the first study that investigated the genes differentially expressed in conidia vs hyphae of A. fumigatus in response to neutrophils from healthy donors as well as from those with CGD which are defective in the production of reactive oxygen species. Upon exposure to either normal or CGD neutrophils, 244 genes were up-regulated in conidia but not in hyphae. Several of these genes are involved in the degradation of fatty acids, peroxisome function and the glyoxylate cycle which suggests that conidia exposed to neutrophils reprogram their metabolism to adjust to the host environment. In addition, the mRNA levels of four genes encoding proteins putatively involved in iron/copper assimilation were found to be higher in conidia and hyphae exposed to normal neutrophils compared to those exposed to CGD neutrophils. Deletants of several of these differentially expressed genes showed phenotypes related to the proposed functions, i.e. deletants of genes involved in fatty acid catabolism showed defective growth on fatty acids and the deletants of iron/copper assimilation showed a higher sensitivity to the oxidative agent menadione. None of these deletants, however, showed reduced resistance to neutrophil attack. During 2008-2009, we characterized the differences in disease manifestations and the manner of dissemination beween A. fumigatus and its sister species, A. udagawae. Both A. udagawae (Neosartorya udagawae) and A. fumigatus belong to Section Fumigatii in the genus Aspergillus and they can not be differentiated by their morphological characteristics but can be differentiated by genetic methods. We also characterized the biological differences between the two species.During 2009-2010, we focused on the identification of MAT-1 and MAT-2 strains that could be used as the foundational strains for the construction of an isogenic pair for the genetic study of virulence factors and other traits of pathobiological importance. We found two super maters of the opposite mating type and decided to use the AFB62 strain that originated from a case of invasive aspergillosis. During 2010-2011, we characterized the sexual reproduction by the super mater pair which was chosen for the construction of the MAT-1 and MAT-2 isogenic set. The meiotic product (ascospores) of the pair could be readily purified from contaminating mitotic spores by heat treatment for 30 minutes at 70 C. High recombination frequencies obtained by mating two conidial color mutants derived from the super mater pair:MAT-1 carrying abr2 mutation, and MAT-2 carrying alb1 mutation and the mutated genes were located 19 kb apart on the same chromosome, indicated that the pair can be used as a genetic tool for recombinational analysis. During 2011-2012, we discovered a new pathogenic species of Aspergillus phylogenetically unrelated to A. fumigatus that causes invasive aspergillosis refractory to currently available antimycotic agents. The new species was described as A. tanneri belonging to the Aspergillus section Circumdati.The species was innately resistant to polyenes, azoles and echinocandins and caused fatal infections in CGD patients.During 2012-2013, we compared the virulence of A.tanneri with that of A. fumigatus in various host systems including CGD mice, corticosteroid treated mice and Galleria mellonela and found that the new species is significantly more virulent for Galleria but less virulent for the mice models. During 2013-2014, we discovered three new species A. fumigatus-look alike species that are genetically and pathobiologically different from A. fumigatus. During 2014-2015, we have identified new drug resistance gene using the whole genome sequence of the isogenic mating pair combined with creating drug resistant mutants. During 2015-2016, we found that exogenous Type 1 IFN protects mice infected with A. fumigatus as well as the Aspergillus species resistant to antifungals such as A. tennari, A. nidulans.

曲霉是影响中性粒细胞减少患者和其他类型的免疫功能低下的个体，例如儿童期慢性肉芽肿（CGD）的最常见的真菌病原体之一。在据报道会引起人类感染的十几种曲霉菌中，富马图斯是据报道会引起侵袭性曲霉病的最常见物种。所有曲霉物种都由分生孢子（孢子）传播，人类每天通过吸入而遇到。在2007年至2008年期间，我们通过研究健康供体的中性粒细胞以及慢性肉芽肿性疾病（CGD）的中性粒细胞挑战时，研究了曲霉对宿主免疫细胞的反应。据我们所知，这是第一项研究，研究了烟曲霉和富米曲霉菌丝中差异表达的基因，这些基因响应健康供体的中性粒细胞，以及来自健康供体的中性粒细胞，这些供体的中性粒细胞在产生反应性氧物种中有缺陷的CGD。 暴露于正常或CGD嗜中性粒细胞后，在分生孢子中上调了244个基因，但在菌丝中没有上调。 这些基因中的几个与脂肪酸，过氧化物酶体功能和甘酰胺循环的降解有关，这表明暴露于中性粒细胞的分生孢子重编程其代谢以适应宿主环境。 此外，与暴露于CGD嗜中性粒细胞的蛋白质相比，发现分生孢子和暴露于正常嗜中性粒细胞的菌丝的四种编码蛋白质的四个基因的mRNA水平更高。 这些差异表达基因的几种缺失物显示出与所提出功能有关的表型，即参与脂肪酸分解代谢的基因的缺失显示脂肪酸的生长有缺陷，铁/铜同化的缺点对氧化剂的氧化剂的敏感性更高。 然而，这些缺失剂都没有显示出对中性粒细胞攻击的耐药性。在2008 - 2009年期间，我们表征了疾病表现的差异以及烟曲霉及其姊妹物种A. udagawae之间的传播方式。 A. udagawae（Neosartorya udagawae）和烟曲霉均属于曲霉属中的fumigatii部分，它们无法通过其形态学特征来区分，但可以通过遗传学方法区分。我们还表征了这两个物种之间的生物学差异。在2009-2010，我们专注于鉴定MAT-1和MAT-2菌株，MAT-1和MAT-2菌株可以用作构建同基对的基础菌株，以构建毒力因子和其他病理学重要性特征的基因研究。我们发现了两个相反交配类型的超级均值，并决定使用起源于侵入性曲霉病的AFB62菌株。在2010-2011期间，我们表征了超级母校对Mat-1和Mat-2等源性集选择的性繁殖。这对的减数分裂产物（孢子孢子）可以通过热处理污染有丝分裂孢子来纯化30分钟，在70 C中30分钟C.通过配对两个来自Super Mater Pair的分生类颜色突变体获得的高重组频率：MAT-1携带ABR2突变，MAT-1携带ABR2突变，MAT-2携带Alb1突变的GenES均具有19 K进行重组分析。在2011年至2012年期间，我们发现了一种新的曲霉曲霉病原体，与烟曲霉无关，这会引起侵袭性的曲霉菌病难治性，以使其对当前可用的抗菌毛剂。新物种被描述为属于曲霉部分的坦纳里。该物种天生对多烯，阿唑，偶氮和棘齿轮蛋白具有抗性，并在CGD患者中引起致命感染。在2012- 2013年，我们将a.tanneri的病毒率与包括cgd Meters andersy的A.tanneri的病毒率相比梅洛内拉（Mellonela）发现，新物种对美术馆的毒性明显更大，但对小鼠模型的毒力较少。在2013 - 2014年期间，我们发现了三种新物种烟曲霉，它们在遗传和病原体上与烟曲霉的不同。在2014-2015期间，我们使用ISEGENIC交配对的整个基因组序列鉴定了新的耐药性基因，并结合了产生耐药性突变体。在2015-2016期间，我们发现外源性1型IFN保护感染了烟曲霉的小鼠以及对抗真菌性抗真菌性抗真菌性抗真菌性抗体的曲霉物种，例如A. tennari，A。nidulans。