Identification of Uncultivated and Unrecognized Pathogens of Periodontitis

描述(申请人提供)：近年来，使用分子技术的研究表明，在口腔中可能检测到多达700个分类群，在龈下菌斑中可能检测到约300个或更多分类群，其中大多数是不可培养的或至少不能识别的。其中一些不可培养的类群很可能是牙周病原体。因此，特殊目标1将检验培养的或未培养的细菌种类与牙周炎有关的假设，这些细菌尚未被确认为病原体。在一项横断面研究中，从80名牙周健康受试者和120名牙周炎受试者各28颗牙齿的近中侧面采集了龈下生物膜样本。每个样本将被分开，并使用两项微生物评估进行单独检查。第一种方法是使用完整的基因组DNA探针对40种物种进行直接棋盘杂交，将为区分牙周炎和健康的已知物种提供阳性和阴性对照。广泛的经验表明，牙龈假单胞菌和连翘等物种在不同的健康和疾病中会有显着的不同，而其余的大多数物种则不会。第二个微阵列芯片分析将检测300个分类群的存在，即基本上所有经常检测到的可培养和尚未培养的牙龈下分类群。区分牙周病原体和非病原体的第二个主要标准是基于这样一个概念，即抑制病原体应导致临床改善，而不减少该物种将导致疾病进展。这一标准将用于特定目标2，该目标将检查牙周治疗对新发现的潜在牙周病原体和临床参数的影响。来自特定目标1的120例牙周炎患者将接受SRP和2周的系统给药甲硝唑和阿莫西林治疗。临床监测和微生物采样将在治疗后3个月重复进行，并用SA1中描述的两种微生物方法检查龈下菌斑样本。将寻求每个测试分类群水平的变化与临床治疗反应之间的联系。由于牙周病是感染，很难建议进行比确定疾病的病原体(S)更重要的研究。这项研究将检查未培养的物种组与牙周健康和疾病的关系，从而在区分病原菌和共生菌方面迈出关键的第一步。随着时间的推移，对这种病原体的识别应导致对疾病的更好诊断、针对个别患者的最佳治疗策略、新的治疗方法和仔细有针对性的预防方法。

DESCRIPTION (provided by applicant): In recent years, studies using molecular techniques have indicated that as many as 700 taxa may be detected in the oral cavity, about 300 or more in subgingival plaque, most of which were uncultivable or at least not recognized. It is likely that some of these uncultivable taxa may be periodontal pathogens. Thus, Specific Aim 1 will test the hypothesis that cultivated or uncultivated bacterial species, not yet recognized as pathogens, are associated with periodontitis. In a cross-sectional study, subgingival biofilm samples, taken from the mesial aspect of up to 28 teeth in each of 80 periodontally healthy and 120 periodontitis subjects. Each sample will be split and individually examined using two microbiological assessments. The first method, direct checkerboard hybridization using whole genomic DNA probes to 40 species, will provide positive and negative controls for known species that discriminate periodontitis from health. Extensive experience indicates that species such as P. gingivalis and T. forsythia will differ significantly from health to disease while most remaining species will not. The second microarray chip assay will detect the presence of 300 taxa; i.e. essentially all of the frequently detected cultivable and as yet uncultivated subgingival taxa. The second major criterion to distinguish periodontal pathogens from non pathogens is based on the notion that suppression of a pathogen should lead to clinical improvement, while failure to reduce that species would lead to disease progression. This criterion will be utilized in Specific Aim 2 which will examine the effects of periodontal therapy on newly recognized potential periodontal pathogens and clinical parameters. The 120 periodontitis subjects from Specific Aim 1 will be treated by SRP and 2 weeks of systemically administered metronidazole and amoxicillin. Clinical monitoring and microbiological sampling will be repeated 3 months post therapy and the subgingival plaque samples examined by the two microbiological methods as described in SA1. Associations will be sought between changes in levels of each test taxon and clinical response to therapy. Since periodontal diseases are infections, it would be difficult to suggest studies that are more important than defining the causative agent(s) of the diseases. This study will examine the association of the uncultivated group of species to periodontal health and disease, thus taking the critical first steps in discriminating the pathogenic from commensal organisms. The identification of such pathogens should, over time, lead to better diagnosis of disease, optimal treatment strategies for the individual patient, new therapeutic approaches and carefully targeted approaches to prevention.