Nadkarni Mangala A, Caldon C Elizabeth, Chhour Kim-Ly, Fisher Ilana P, Martin F Elizabeth, Jacques Nicholas A, Hunter Neil
Institute of Dental Research, Westmead Millenium Institute, Wentworthville, New South Wales, Australia.
J Clin Microbiol. 2004 Nov;42(11):5238-44. doi: 10.1128/JCM.42.11.5238-5244.2004.
Previous analysis of the microbiology of advanced caries by culture and real-time PCR emphasized the high incidence and abundance of gram-negative anaerobic species, particularly Prevotella-like bacteria. The diversity of Prevotella-like bacteria was further explored by analyzing pooled bacterial DNA from lesions of carious dentine. This was achieved by amplification of a region of the 16S ribosomal DNA with a Prevotella genus-specific forward primer and a universal bacterial reverse primer, followed by cloning and sequencing. Cultured Prevotella species commonly associated with oral tissues constituted only 12% of the Prevotella clones isolated from advanced carious lesions. The remaining 88% consisted of a diverse range of phylotypes. These included five clusters of previously recognized but uncultured oral Prevotella spp. and a major cluster containing Prevotella-like bacteria most closely related to uncharacterized rumen bacteria. Cluster-specific primers were designed, and the numbers of bacteria within clusters were quantified by real-time PCR, confirming the abundance of these organisms. The data indicated that advanced dental caries provides a unique environment for a complex array of novel and uncultured Prevotella and Prevotella-like bacteria which, in some cases, may dominate the diverse polymicrobial community associated with the disease.
先前通过培养和实时聚合酶链反应（PCR）对重度龋齿微生物学的分析强调了革兰氏阴性厌氧菌，尤其是普氏菌属样细菌的高发生率和丰度。通过分析龋损牙本质病变处的混合细菌DNA，进一步探究了普氏菌属样细菌的多样性。这是通过使用普氏菌属特异性正向引物和通用细菌反向引物扩增16S核糖体DNA区域，随后进行克隆和测序来实现的。通常与口腔组织相关的培养普氏菌种仅占从重度龋损病变中分离出的普氏菌克隆的12%。其余88%由多种不同的系统发育型组成。这些包括五组先前已识别但未培养的口腔普氏菌种，以及一个主要的簇，其中包含与未鉴定的瘤胃细菌关系最密切的普氏菌属样细菌。设计了簇特异性引物，并通过实时PCR对簇内细菌数量进行定量，证实了这些微生物的丰度。数据表明，重度龋齿为一系列复杂的新型和未培养的普氏菌及普氏菌属样细菌提供了独特的环境，在某些情况下，这些细菌可能在与该疾病相关的多种微生物群落中占主导地位。
