Comprehensive detection of periodontal pocket bacterial flora and quantitation of disease-specific bacterial species

背景：人类牙周袋中已发现近400种细菌。其中一半被认为至今尚未培养。牙周感染的微生物学诊断需要进一步研究未培养菌群的生态学特征。为了解慢性牙周炎患者龈下菌群生态，为牙周微生物生态学的诊断提供依据，本研究采用序列分析法对4例慢性牙周炎患者的龈下菌斑进行16 S rRNA基因文库的构建。结果：该物种丰富度包括一组丰富的共有种（17个亚种型）和一组个体多样的稀有种（58个亚种型）。两组均由约25%的已描述的和约75%的尚未培养的大肠杆菌类型组成。这些细菌类型被分配到拟杆菌门、梭杆菌门、变形菌门、厚壁菌门、螺旋菌门、放线菌门和TM 7门。以前报道的牙周相关的物种和牙周类型中发现的一些样本，但不是所有共享。序列相似性大于99%的各种亚基类型呈偏态分布。结论：龈下菌群中未培养菌的物种丰富度和丰度分别为70-80%和70-80%。然而，大多数这些细菌类型在遗传学上接近已被充分描述的牙周相关细菌。个体差异被发现在斑块的物种内尚未培养，以及良好的描述物种的微生物区系。这些个别不同的斑片状分布是预期为“量身定制”的牙周治疗微生物诊断。

Background : Nearly 400 species of bacteria have been found in the human periodontal pockets. Half of those are considered to remain not-yet-cultured to this date. Further characterization of the ecology of those not-yet-cultured species in the microflora should be done for establishment of microbiological diagnosis of periodontal infection. The purpose of this study is to know ecology of subgingival microflora of chronic periodontitis patients by sequence-based method for an approach to diagnosis in human periodontal microbial ecology.Methods : Four 16S rRNA gene libraries were constructed from subgingival plaque samples taken from all of diseased sites of 4 chronic periodontitis patients. The 480 clones from each libraries were analyzed by phylotyping.Results : The species richness contained a group of abundant shared species (17 phylotypes) and a group of individually diverse rare species (58〜83 phylotypes). Both groups consisted of〜25% of well-described and〜75% of not-yet-cultured phylotypes. The phylotypes were assigned to the Bacteroidetes, Fusobacteria, Proteobacteria, Firmicutes, Spirochetes, Actinobacteria, and TM7 phyla. Previously reported periodontally associated species and phylotypes were found in some samples but not to be shared by all. Diverse sub-phylotypes showing more than 99% of sequence similarity showed individually skewed distribution.Conclusions : The species richness and abundance of the not-yet-cultured species seemed to be 70-80% and 70-80% respectively in subgingival microflora. However most of those phylotypes were phylogenetically close to well-described periodontally associated bacteria. Individual differences were found in patchness of the species within not-yet-cultured as well as well-described species of the microflora. These individually distinct patchy distributions are expectable to serve microbial diagnosis for 'tailor-made' periodontal treatment.