Virulence determinants of Fusobacterium nucleatum

具核梭杆菌的毒力决定因素

• 批准号: 9982064
• 负责人: Hung Ton-That
• 金额: $ 44.39万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9982064
• 关键词: Adherence; Affect; Anaerobic Bacteria; Anti-Infective Agents; Antibiotic Resistance; Attenuated; Bacteria; Biochemical; Biogenesis; Biological Process; Cell Communication; Cell Proliferation; Cell physiology; Cells; Code; Colorectal Cancer; Complex; Cryo-electron tomography; Defect; Dental Plaque; Development; Electron Microscopy; Endothelial Cells; Epithelial Cells; Functional disorder; Fusobacterium nucleatum; Future; Gene Deletion; Generations; Genes; Genetic; Genome; Gingiva; Goals; Growth; Halitosis; Hydrogen Peroxide; Hydrogen Sulfide; Immune; Immune response; Immunomodulators; Impairment; Individual; Infection; Inflammasome; Inflammation; Inflammatory; Inflammatory Response; Investigation; Knowledge; Laboratories; Libraries; Link; Mass Spectrum Analysis; Mediating; Membrane; Methods; Microbial Biofilms; Microbiology; Molecular; Mutagenesis; Names; Odontogenesis; Open Reading Frames; Oxidative Stress; Pathogenesis; Pathogenicity; Pathway interactions; Periodontitis; Porphyromonas gingivalis; Premature Birth; Production; Proteins; Reporting; Rodent Model; Role; Saliva; Science; Signal Transduction; Streptococcus mutans; System; Technology; Time; Tube; Virulence; Virulence Factors; adverse pregnancy outcome; antimicrobial; cell growth regulation; colon carcinogenesis; cytokine; disorder prevention; fitness; forward genetics; human disease; in vivo; methionine sulfoxide reductase; mouse model; mutant; novel; oral biofilm; oral infection; oral pathogen; oxidative damage; pathogen; public health relevance; repaired; response; reverse genetics; therapeutic development; tool

PROJECT SUMMARY The Gram-negative anaerobe Fusobacterium nucleatum is a key colonizer in the development of oral biofilms, or dental plaque, and also known for its association with human diseases including oral infections, adverse pregnancy outcomes, and colorectal cancer. F. nucleatum has an inherent ability to interact or aggregate with many early and late colonizers of the oral biofilms. It induces inflammatory responses and preterm birth in rodent models of infection, as well as promoting colorectal carcinogenesis in vivo. Despite its pathogenic potential, we have limited knowledge about the mechanisms of fusobacterial virulence and associated factors; to date only six fusobacterial factors have been reported, i.e. FomA, FadA, Fap2, RadD, aid1, and FAD-I, although more than 2,000 open reading frames are annotated in the genome of many F. nucleatum strains. A major obstacle limiting progress is the lack of robust genetic tools and systematic investigations. We have begun to tackle this problem, successfully developing a facile gene deletion system for F. nucleatum, and generating a large library of random transposon mutants with 10-fold genome coverage. With these previously unavailable tools, we aim to identify virulence determinants of F. nucleatum using multiple complementary approaches including forward and reverse genetics, cryo-electron tomography, biochemical methods, and rodent models of infection. By electron microscopy, we discovered that F. nucleatum produces outer membrane tubules (OMTs) and identified a key factor required for OMT formation. By gene deletion, we found pathways that mediate oxidative stress defense and host cell adherence and invasion. By transposon mutagenesis, we revealed several biofilm-associated factors that are required for production of hydrogen sulfide, which largely contributes to bad breath, or halitosis. In this application, we aim to elucidate the mechanism of OMT biogenesis and OMT-mediated pathogenesis, reveal the mechanisms of host immune defense and host cell interactions, and establish the role of biofilm- associated factors in halitosis, polymicrobial interactions, and bacterial virulence. Our studies will greatly contribute to our understanding of the molecular virulence mechanisms of this pathogen and significantly move the field forward with the application of newly developed genetic tools and advanced technology. The results generated from these studies may also provide promising targets for the future development of therapeutic strategies.

项目摘要 革兰氏阴性肌杆菌核细菌是口服生物膜发展的关键菌落，， 或牙菌斑，也因与人类疾病（包括口腔感染）的关联而闻名 怀孕结局和结直肠癌。 F. nutleatum具有与 口腔生物膜的许多早期和晚期殖民者。它诱导啮齿动物的炎症反应和早产 感染模型，并在体内促进结直肠癌。尽管具有致病性，我们 关于梭菌毒力和相关因素的机制知识有限；迄今仅六个 已经报道了梭菌因子，即FOMA，FADA，FAP2，RADD，AID1和FAD-I，尽管超过 在许多F核菌株的基因组中注释了2,000个开放式阅读框。限制主要障碍 进展是缺乏强大的遗传工具和系统研究。我们已经开始解决这个问题， 成功地开发了用于F. nuteatum的轻松基因缺失系统，并生成一个大的随机库 具有10倍基因组覆盖率的转座子突变体。使用这些以前无法使用的工具，我们旨在确定 使用多种互补方法（包括前进和反向）的毒力决定因素 遗传学，冷冻电子断层扫描，生化方法和啮齿动物感染模型。由电子 显微镜，我们发现F. nutleatum产生外膜小管（OMT）并鉴定了一个钥匙 OMT形成所需的因素。通过基因删除，我们发现了介导氧化应激防御的途径 以及宿主细胞的依从性和入侵。通过转座子诱变，我们揭示了几个生物膜相关的 生产硫化氢所需的因素，这些因素很大程度上导致口臭或盐症。 在此应用中，我们旨在阐明OMT生物发生和OMT介导的发病机理的机制， 揭示宿主免疫防御和宿主细胞相互作用的机制，并确定生物膜的作用 哈特病，多数菌相互作用和细菌毒力的相关因素。我们的研究将大大 有助于我们理解该病原体的分子毒力机制，并显着移动 该领域随着新开发的遗传工具和先进技术的应用。结果 这些研究产生的可能还为未来治疗发展提供了有希望的目标 策略。