Assessment of the diversity and stability of faecal bacteria from healthy adults using molecular methods

Abstract

The dominant members of the human colonic microbiota play an important role in colonic health through their activity on dietary substrates. Different molecular methods have been used to describe colonic populations in healthy humans; however, the comparability of these different methodologies is not fully understood. Dominant members of the microbiota were studied in faecal specimens from eight healthy adults using 16S rRNA gene PCR, denaturing gradient gel electrophoresis (DGGE), 16S rRNA gene clone library construction, DNA sequencing and phylogenetic analysis. Bacterial populations were stable over time for each volunteer, but clear inter-individual differences in population composition were observed. Comparative analysis of dominant faecal phylotypes by DGGE and 16S rRNA gene clone library techniques revealed significant differences in the dominant phylotypes detected despite numerous closely related sequences arising from the different techniques. Sequences encompassed a number of clades within the Bacteroidales and Clostridiales . Lineages, containing phylotypes present in the majority of individuals, formed two groups within the Clostridial cluster IV, one group within the, as yet undescribed, ‘cluster B’ and one uncultivated clade within the Bacteroidales . Phylotypes from these groups demonstrated poor homology with previously cultivated bacteria, despite their apparent abundance in the colon. Key words: DGGE, faecal bacteria, 16S rRNA, bowel health