Premium
Molecular Analysis of Fecal Microbiota in Elderly Individuals Using 16S rDNA Library and T‐RFLP
Author(s) -
Hayashi Hidenori,
Sakamoto Mitsuo,
Kitahara Maki,
Benno Yoshimi
Publication year - 2003
Publication title -
microbiology and immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.664
H-Index - 70
eISSN - 1348-0421
pISSN - 0385-5600
DOI - 10.1111/j.1348-0421.2003.tb03418.x
Subject(s) - phylotype , biology , restriction fragment length polymorphism , gammaproteobacteria , terminal restriction fragment length polymorphism , 16s ribosomal rna , genetics , microbiology and biotechnology , bacteroides , ribosomal rna , clostridium , genotype , gene , bacteria
Fecal microbiota in six elderly individuals were characterized by the 16S rDNA libraries and terminal restriction fragment length polymorphism (T‐RFLP) analysis. Random clones of 16S rRNA gene sequences were isolated after PCR amplification with universal primer sets from total genomic DNA extracted from feces of three elderly individuals. These clones were partially sequenced (about 500 bp). T‐RFLP analysis was performed using 16S rDNA amplified from six subjects. The lengths of the terminal restriction fragment (T‐RF) were analyzed after digestion by Hha I and Msp I. Among 240 clones obtained, approximately 46% belonged to 27 known species. About 54% of the other clones were 56 novel “phylotypes” (at least 98% homology of clone sequence). These libraries included 83 species or phylotypes. In addition, about 13% (30 phylotypes) of these phylotypes were newly discovered in these libraries. A large number of species that are not yet known exist in the feces of elderly individuals. 16S rDNA libraries and T‐RFLP analysis revealed that the majority of bacteria were Bacteroides and relatives, Clostridium rRNA cluster IV, IX, Clostridium rRNA subcluster XIVa, and “ Gammaproteobacteria ”. The proportion of Clostridium rRNA subcluster XIVa was lower than in healthy adults. In addition, although Ruminococcus obeum and its closely related phylotypes were detected in high frequency in healthy young subjects, hardly any were detected in our elderly individuals. “ Gammaproteobacteria ” were detected at high frequency.