Iron Uptake and Virulence of Burkholderia pseudomallei

鼻疽伯克霍尔德菌的铁吸收和毒力

• 批准号: 6816714
• 负责人: JORGE H CROSA
• 金额: $ 30.2万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-15 至 2006-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6816714
• 关键词: Burkholderia; bacteria infection mechanism; bacterial cytopathogenic effect; bacterial disease; bacterial genetics; biological transport; bioterrorism /chemical warfare; chemical structure; chemical structure function; gene expression; gene targeting; high performance liquid chromatography; host organism interaction; intracellular; iron; iron metabolism; laboratory mouse; macrophage; mass spectrometry; microarray technology; mutant; nuclear magnetic resonance spectroscopy; septicemia; siderophores; virulence; zoonosis

DESCRIPTION (provided by applicant): Melioidosis and glanders are life threatening infectious diseases with similar patho-physiology caused respectively by the select agents Burkholderia pseudomallei and B. mallei, with dramatic repercussions in the field of bio-defense. Despite major recent advances including the elucidation of the entire genome sequence of B. pseudomallei K96243 and the ongoing sequencing of the B. mallei genome, little is known about their mechanisms of virulence. Our long-range goal is to elucidate the pathogenesis of these diseases. Although many factors can contribute to bacterial virulence, one important nonspecific mechanism of defense pathogenic bacteria must overcome to establish infection, is the ability of human and animal hosts to withhold iron. Thus, a competitive advantage for microorganisms is the possession of genetic determinants encoding products that allow them to utilize otherwise unavailable iron. The immediate goal of our research is to use a combination of genetic and biochemical approaches to unveil specific mechanisms of iron uptake employed by these pathogens during both the septicemic and the intracellular phases of disease. To accomplish these goals we will perform: 1. Microarray and mutational analysis. We will use microarray chips to examine the differential gene expression of B. pseudomallei K96243, from cells grown in vitro under conditions of iron repletion and limitation. We will also examine patterns of gene expression induced in B. pseudomallei during infection of macrophages. In conjunction with microarray analysis we will generate knock-out mutants of important iron uptake genes in B. pseudomallei, identified from the available genome sequencing project. Selected iron uptake mutants will be assessed for their subsequent ability to invade and survive intracellularly in macrophages and for virulence in animal models. 2. The characterization and structural analysis of siderophores from B. pseudomallei and B. mallei. We will characterize the structure of siderophores produced by wild type and mutant B. pseudomallei (K96243) and B. mallei (ATCC 23344). These compounds will be purified by high performance liquid chromatography and their structural details determined by nuclear magnetic resonance and mass spectrometry. Knowledge gained from our investigation will contribute to the development of measures to control the diseases caused by these pathogens.

描述(申请人提供)：类鼻疽病和扁桃体病是由选定的假鼻疽伯克霍尔德氏菌和马来伯克霍尔德氏菌分别引起的具有相似病理生理的危及生命的传染病，在生物防御领域产生巨大影响。尽管最近取得了重大进展，包括阐明了假单胞菌K96243的全基因组序列，以及正在进行的马来双胞菌基因组测序，但对它们的毒力机制知之甚少。我们的长期目标是阐明这些疾病的发病机制。虽然许多因素都可能导致细菌的毒力，但病原菌必须克服一个重要的非特异性防御机制来建立感染，那就是人类和动物宿主抑制铁的能力。因此，微生物的一个竞争优势是拥有编码产品的基因决定因素，这些产品允许它们利用原本无法获得的铁。我们研究的直接目标是使用遗传和生化方法的组合来揭示这些病原体在败血症和细胞内疾病阶段吸收铁的特定机制。为了实现这些目标，我们将进行：1.微阵列和突变分析。我们将使用微阵列芯片来检测在铁补充和限制条件下体外培养的假鼻疽杆菌K96243的差异基因表达。我们还将研究在巨噬细胞感染期间诱导假鼻疽杆菌基因表达的模式。结合微阵列分析，我们将从现有的基因组测序项目中鉴定出假鼻疽杆菌中重要的铁摄取基因的敲除突变。选定的铁摄取突变体将被评估其随后在巨噬细胞内入侵和存活的能力以及动物模型中的毒力。2.假马利氏杆菌和马利氏杆菌铁载体的性质和结构分析我们将研究野生型和突变型假单胞菌(K96243)和马利氏杆菌(ATCC 23344)产生的铁载体的结构。这些化合物将通过高效液相色谱进行纯化，并通过核磁共振和质谱学确定其结构细节。我们从调查中获得的知识将有助于制定控制这些病原体引起的疾病的措施。