Phylogenetic analysis of the human gut microbiota using 16S rDNA clone libraries and strictly anaerobic culture-based methods
Phylogenetic analysis of the human gut microbiota using 16S rDNA clone libraries and strictly anaerobic culture-based methods
复制标题
DOI：
10.1111/j.1348-0421.2002.tb02731.x
复制
发表时间：
2002-01-01
影响因子：
2.6
通讯作者：
Benno, Y
中科院分区：
文献类型：
--
作者：
Hayashi, H;Sakamoto, M;Benno, Y
通过16S rDNA文库序列分析和基于培养的方法比较了三名健康受试者的肠道微生物区系。直接计数为1.9×10(11)-4.0×10(11)个/g(湿重)，平板计数为6.6×10(10)-1.2×10(11)cfu/g(湿重)。人类肠道中60%到70%的细菌不能用目前可用的方法培养。3个受试者的16S rDNA文库均以肠道总群落DNA为模板，用通用引物构建。随机选择的克隆被部分测序。从琼脂平板表面检测到的所有纯化菌落用于16S rDNA的部分测序。在序列相似性的基础上，将克隆和菌落划分为若干簇，对应于域细菌的主要门。在获得的744个克隆中，约有25%属于31个已知物种。剩下的克隆中约有75%是新的“系统型”(克隆序列至少有98%的相似性)。根据本研究的序列数据，肠道优势微生物群落由130个种或系统型组成。16S rDNA文库和菌落包括类杆菌群、链球菌群、双歧杆菌群和梭状芽孢杆菌rRNA簇IV、IX、XIVa和XVIII。此外，在克隆文库和克隆中还发现了几种以前未鉴定和未培养的微生物。我们的结果还显示，肠道微生物区系的组成存在显著的个体差异。
The human gut microbiota from three healthy subjects were compared by the use of a sequence analysis of 16S rDNA libraries and a culture-based method. Direct counts ranged from 1.9 X 10(11) to 4.0 X 10(11) cells/g (wet weight), and plate counts totaled 6.6 X 10(10) to 1.2 X 10(11) CFU/g (wet weight). Sixty to seventy percent of the bacteria in the human intestinal tract cannot be cultured with currently available methods. The 16S rDNA libraries from three subjects were generated from total community DNA in the intestinal tract with universal primer sets. Randomly selected clones were partially sequenced. All purified colonies detected from the surface of the agar plate were used for a partial sequencing of 16S rDNA. On the basis of sequence similarities, the clones and colonies were classified into several clusters corresponding to the major phylum of the domain Bacteria. Among a total of 744 clones obtained, approximately 25% of them belonged to 31 known species. About 75% of the remaining clones were novel "phylotypes" (at least 98% similarity of clone sequence). The predominant intestinal microbial community consisted of 130 species or phylotypes according to the sequence data in this study. The 16S rDNA libraries and colonies included the Bacteroides group, Streptococcus group, Bifidobacterium group, and Clostridium rRNA clusters IV, IX, XIVa, and XVIII. Moreover, several previously uncharacterized and uncultured microorganisms were recognized in clone libraries and colonies. Our results also showed marked individual differences in the composition of intestinal microbiota.