Environmental Uncultivable Bacteria Model for Understanding Human Periodontitis

描述（由申请人提供）：I025是一种新的不可培养的细菌群，与牙周病有关，在工业化国家，牙周病影响18%的成年人口。这一群体的天然水库和完全多样性都不为人所知。I025属于候选门TM7，它首先在包括海水，土壤和活性污泥在内的各种自然环境中发现，并且完全由不可培养的物种组成。最近的独立调查已经在健康和患病个体的人类口腔微生物群落中检测到TM7核糖体DNA（rDNA）基因序列，而I025主要在患病个体中检测到。环境和人类TM7菌株之间的rDNA序列比较表明，有些几乎是相同的，这表明它们是近亲，如果不是同一物种。我们假设环境中的I025样细菌可以作为更好地理解I025在人类牙周炎中的作用的模型。我们的目标是建立这样一个I025模型。然而，对栽培的抗性使TM7物种除了它们的rDNA序列和很少的形态特征之外没有被描述，尽管它们在人类口腔斑块中长达70 μ m的明显细丝。SJSU的本科生和硕士生将应用现代和高度敏感的非培养分子技术来研究I025样细菌在其自然栖息地中的功能。 该项目的具体目标是： 目标1：通过16S rDNA基因的PCR扩增和序列分析，确定TM7的生态位点，并对与人类牙周病相关的环境样品中TM7的多样性进行分析。 目标二：通过实时定量PCR和荧光原位杂交技术对与牙周炎相关的TM7（I025）密切相关的环境TM7进行定量。 目标3：通过联合收割机显微放射自显影和原位杂交相结合的创新方法来表征TM7细胞在其自然环境中的代谢功能，以测量单个细胞的营养吸收，而无需培养。 生态环境是许多人类病原体的储存库。I025是一种潜在的新兴人类病原体，一旦确定了它的环境来源并了解了它的营养需求，我们就可以着手建立一个模型，以更好地了解它在人类中的作用。

DESCRIPTION (provided by applicant): I025, a novel uncultivable bacteria group, has been associated with periodontal disease, which affects 18% of the adult population in the industrialized world. Neither the natural reservoir nor the complete diversity of this group is known. I025 belongs to the candidate phylum TM7, which was first discovered in diverse natural environments including seawater, soil, and activated sludge, and is made entirely of uncultivable species. Recent independent investigations have detected TM7 ribosomal DNA (rDNA) gene sequences in the human oral microbial community of both healthy and diseased individuals, whereas I025 was detected primarily in diseased individuals. rDNA sequence comparison between environmental and human TM7 strains indicate some are nearly identical, suggesting they are close relatives, if not the same species. We hypothesize that an environmental I025-like bacteria can serve as model for better understanding the I025 role in human periodontitis. Our goal is to establish such an I025 model. Resistance to cultivation, however, has kept TM7 species undescribed beyond their rDNA sequence and few morphological features, despite their conspicuous filaments of up to 70 ¿m in length in the human oral plaque. Undergraduate and Master's students at SJSU will apply modern and highly sensitive culture-independent molecular techniques to investigate the function of I025-like bacteria in their natural habitat. The specific aims of this project are: Aim 1: Identify ecological sites with TM7 and then characterize the diversity of TM7 in those environmental samples related to human periodontal-associated TM7 species through PCR amplification of 16S rDNA gene and sequence analysis. Aim 2: Quantify environmental TM7 closely related to the periodontitis-associated TM7 (I025) through real-time quantitative PCR and fluorescence in situ hybridization. Aim 3: Characterize the metabolic function of TM7 cells in their natural environment through innovative methods that combine microautoradiography and in situ hybridization to measure nutrient uptake by single cells, without the need of cultivation. Ecological environments serve as the reservoir for many human pathogens. I025 is a potential emerging human pathogen, and once its environmental source is defined and the nutrient requirements understood, we can proceed to establish a model for better understanding its role in humans.