Moving Beyond Diversity by Revealing Biological Functions of Uncultured Bacteria

通过揭示未培养细菌的生物学功能超越多样性

• 批准号: 8197607
• 负责人: Jeffrey Scott McLean
• 金额: $ 41.37万
• 依托单位: J. CRAIG VENTER INSTITUTE, INC.
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-12-01 至 2014-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8197607
• 关键词: Acids; Adult; Bacteria; Behavior; Bioinformatics; Biological; Biological Assay; Biological Models; Biological Process; Carbohydrates; Cells; Child; Clinical; Clinical Management; Collaborations; Communicable Diseases; Communities; Complex; Cryoultramicrotomy; Data; Data Set; Dental; Dental Plaque; Dental caries; Development; Disease; Disease Progression; Dominant Genes; Ecology; Environment; Fluorescent Antibody Technique; Fluorescent Probes; Fluorescent in Situ Hybridization; Freezing; Future; Gene Cluster; Gene Expression; Genes; Genetic; Genome; Genomics; Growth; Health; Human; In Vitro; Incubated; Individual; Infection; Institutes; Isotopes; Knowledge; Label; Laboratories; Maps; Messenger RNA; Metabolic; Metabolic Pathway; Metabolism; Metagenomics; Methods; Microbe; Microbial Biofilms; Modeling; Molecular Profiling; Monitor; Nature; Oral; Organism; Pathogenesis; Pathway interactions; Patients; Pattern; Physiological; Physiology; Population; Positioning Attribute; Prevalence; Process; Production; Property; Proteins; Public Health; RNA; Recombinant DNA; Relative (related person); Research; Resources; Reverse Transcriptase Polymerase Chain Reaction; Role; Saliva; Sampling; Sequence Analysis; Solutions; Spatial Distribution; Technology; Thinking; Transcript; United States; Virulence; Work; analytical method; base; combat; environmental change; genome sequencing; human disease; in vivo; insight; interest; laser capture microdissection; member; microbial; microbial community; microorganism; novel; novel strategies; novel therapeutic intervention; pathogen; public health relevance; response; stable isotope; success; tool; treatment strategy

DESCRIPTION (provided by applicant): Many human diseases are polymicrobial in nature and therefore require thinking beyond the traditional one organism-one disease concept to find solutions to combating them. Of fundamental importance is the need for an understanding of how these communities evolve from a healthy to a diseased state at a community, species and gene level. Although many model species found associated with healthy or disease conditions have been characterized in vitro, it is difficult to determine if their physiology in pure cultures are maintained in vivo in the presence of a mixed microbial community. In addition, most host-associated microbes remain uncultured (estimated at ~ 50% of those identified) therefore little is known about the uncultured species except for their 16S rDNA sequence. Understanding the physiology of uncultured species has become a major limiting step in studying the ecology of these polymicrobial infections. With the advent of new approaches in high throughput sequencing, single cell genomics as well as new tools in microbial ecology, we can now achieve a deeper and more detailed understanding of the physiological and ecological principals that govern the behavior of host-associated microbial communities. In doing so, we can also reveal the functions and biological role of currently uncultured members. Specifically in this application we will investigate oral microbial communities and seek to; 1) identify the active species, that are highly correlated with high and low pH microbial processes through Stable Isotope Probing (SIP); 2) identify the genes and dominant pathway(s) that are expressed and also shared between these active species through metatranscriptomics; 3) investigate the spatial patterns and relationships of uncultured species with the other community members using Laser Capture Microdissection and 4) sequence the genomes of key suspected pathogens that are currently uncultured, through whole genome amplification from captured single cells. The success of this study would greatly expand our knowledge of oral microbial pathogenesis as it will help us to understand the virulence properties of uncultured species as well as known pathogens, not only in pure culture, but also in multi-species dental biofilms. We are particularly focused on revealing more about how these currently uncultivated bacteria may contribute to heath and disease processes. The present lack of such information presents a major barrier to understanding polymicrobial diseases. This will have a great impact on the future clinical management of dental caries and provide a comprehensive approach for characterizing the function of species and their interactions for other host-associated communities. PUBLIC HEALTH RELEVANCE: Many widespread human diseases, such as tooth decay (caries), result from the interactions of complex communities of microorganisms (polymicrobial diseases). This complexity severely complicates treatment strategies. Caries, for example, remains a major health issue in the United States and worldwide with a prevalence of more than 50% in young children, increasing to about 85% in the adult population. The majority of bacteria associated with polymicrobial diseases have not been well studied due to the fact that they cannot be grown in the laboratory. We propose to use new analytical methods to investigate the biological functions of these uncultured bacteria and their roles in human health. These approaches include generating the genomes and monitoring the physiology of these species in their natural environment. This new approach will provide a deeper understanding of disease progression and allow for the subsequent development of novel therapeutic approaches to battle these diseases.

描述（由申请人提供）：许多人类疾病本质上是多微生物的，因此需要超越传统的一种微生物一种疾病的概念来寻找解决方案。至关重要的是，需要了解这些群落是如何在群落、物种和基因水平上从健康状态进化到患病状态的。尽管许多与健康或疾病状况相关的模式物种已经在体外进行了表征，但很难确定它们在纯培养物中是否能在体内混合微生物群落中维持生理机能。此外，大多数宿主相关微生物仍未培养（估计约占已鉴定微生物的50%），因此除了对未培养的物种的16S rDNA序列知之甚少。了解未培养物种的生理学已成为研究这些多微生物感染生态学的主要限制步骤。随着高通量测序新方法、单细胞基因组学以及微生物生态学新工具的出现，我们现在可以更深入、更详细地了解控制宿主相关微生物群落行为的生理和生态原理。通过这样做，我们还可以揭示目前未培养成员的功能和生物学作用。具体而言，在本应用中，我们将调查口腔微生物群落并寻求；1)通过稳定同位素探测（SIP）鉴定出与高、低pH微生物过程高度相关的活性种；2)通过亚转录组学鉴定这些活性物种之间表达和共享的基因和显性途径；3)利用激光捕获显微解剖技术研究未培养物种与其他群落成员的空间格局和关系；4)通过捕获的单细胞全基因组扩增，对目前未培养的关键可疑病原体进行基因组测序。这项研究的成功将极大地扩展我们对口腔微生物发病机制的认识，因为它将帮助我们了解未培养物种和已知病原体的毒力特性，不仅在纯培养中，而且在多物种口腔生物膜中。我们特别专注于揭示更多关于这些目前未培养的细菌如何有助于健康和疾病过程的信息。目前缺乏这类信息是了解多微生物疾病的一个主要障碍。这将对未来龋齿的临床管理产生重大影响，并为表征物种功能及其与其他宿主相关群落的相互作用提供全面的方法。