Phylogenetic Analysis of 16S Ribosomal RNA Gene Sequences from Human Fecal Microbiota and Improved Utility of Terminal Restriction Fragment Length Polymorphism Profiling

Abstract

We have developed a terminal restriction fragment length polymorphism (T-RFLP) method for determining the structure and dynamics of the microbial gut community. In this paper, the improved T-RFLP method in combination with an analysis of the fecal 16S rDNA (ribosomal RNA gene) clone libraries from six individuals is described. A total of 418 different partial sequences of 16S rDNA were determined and subjected to a phylogenetic analysis and homology examination. We found that the sequences were roughly divided into six phylogenetic groups containing seven subgroups and were related to 71 known species with over 90% similarity. With the exception of a few cases, we found that in silica BstI-digestioa of the sequences belonging to the same bacterial group or subgroup generated terminal restriction fragments of similar lengths. We concluded that human intestinal microbiota predominantly consists of the members of approximately ten phylogenetic bacterial groups and that these bacterial groups are effectively distinguished by our T-RFLP system.