Virulence determinants of Fusobacterium nucleatum

具核梭杆菌的毒力决定因素

基本信息

批准号：
10221250
负责人：
Hung Ton-That
金额：
$ 1.31万
依托单位：
UNIVERSITY OF CALIFORNIA LOS ANGELES
依托单位国家：
美国
项目类别：
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/10221250
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

项目摘要

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.

项目概要革兰氏阴性厌氧菌具核梭杆菌是口腔生物膜形成的关键定植菌，或牙菌斑，也因其与人类疾病的关系而闻名，包括口腔感染、不良反应妊娠结局和结直肠癌。 F. nucleatum 具有与以下物质相互作用或聚集的固有能力：许多口腔生物膜的早期和晚期殖民者。它会引起啮齿动物的炎症反应和早产感染模型，以及促进体内结直肠癌的发生。尽管它具有致病潜力，但我们对梭杆菌毒力机制和相关因素的了解有限；迄今为止只有六个梭菌因子已被报道，即 FomA、FadA、Fap2、RadD、aid1 和 FAD-I，尽管超过许多具核梭菌菌株的基因组中注释了 2,000 个开放阅读框。限制的主要障碍进展是缺乏强有力的遗传工具和系统研究。我们已经开始着手解决这个问题成功开发了一种简便的 F. nucleatum 基因删除系统，并生成了一个大型随机文库具有10倍基因组覆盖度的转座子突变体。借助这些以前无法使用的工具，我们的目标是识别使用多种互补方法（包括正向和反向）确定具核梭菌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。通过电子通过显微镜观察，我们发现 F. nucleatum 产生外膜小管 (OMT)，并确定了一个关键 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粘附和侵袭。通过转座子诱变，我们揭示了几种与生物膜相关的产生硫化氢所需的因素，硫化氢在很大程度上导致口臭或口臭。在本申请中，我们旨在阐明 OMT 生物发生机制和 OMT 介导的发病机制，揭示宿主免疫防御和宿主细胞相互作用的机制，并建立生物膜的作用口臭、多种微生物相互作用和细菌毒力的相关因素。我们的学习将大大有助于我们了解这种病原体的分子毒力机制，并显着推动随着新开发的遗传工具和先进技术的应用，该领域不断发展。结果这些研究产生的结果也可能为治疗的未来发展提供有希望的目标策略。