Use of 16S rRNA gene-targeted group-specific primers for real-time PCR analysis of predominant bacteria in human feces.
16S rRNA gene-targeted group-specific primers were designed and validated for specific detection and quantification of the Clostridium leptum subgroup and the Atopobium cluster. To monitor the predominant bacteria in human feces by real-time PCR, we used these specific primers together with four sets of group-specific primers for the Clostridium coccoides group, the Bacteroides fragilis group, Bifidobacterium, and Prevotella developed in a previous study (T. Matsuki, K. Watanabe, J. Fujimoto, Y. Miyamoto, T. Takada, K. Matsumoto, H. Oyaizu, and R. Tanaka, Appl. Environ. Microbiol. 68:5445-5451, 2002). Examination of DNA extracted from the feces of 46 healthy adults showed that the C. coccoides group was present in the greatest numbers (log10 10.3 +/- 0.3 cells per g [wet weight] [average +/- standard deviation]), followed by the C. leptum subgroup (log10 9.9 +/- 0.7 cells per g [wet weight]), the B. fragilis group (log10 9.9 +/- 0.3 cells per g [wet weight]), Bifidobacterium (log10 9.4 +/- 0.7 cells per g [wet weight]), and the Atopobium cluster (log10 9.3 +/- 0.7 cells per g [wet weight]). These five bacterial groups were detected in all 46 volunteers. Prevotella was found in only 46% of the subjects at a level of log10 9.7 +/- 0.8 cells per g (wet weight). Examination of changes in the population and the composition of the intestinal flora for six healthy adults over an 8-month period revealed that the composition of the flora of each volunteer remained stable throughout the test period.
Matsuki
Takahiro
T
Yakult Central Institute for Microbiological Research, Kunitachi, Tokyo, Japan. takahiro-matsuki@yakult.co.jp
Watanabe
Koichi
K
Fujimoto
Junji
J
Takada
Toshihiko
T
Tanaka
Ryuichiro
R
D000328:Adult; D001419:Bacteria / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D001441:Bacteroides fragilis / Q000235:genetics / Q000302:isolation & purification; D001644:Bifidobacterium / Q000254:growth & development / Q000302:isolation & purification; D003013:Clostridium / Q000235:genetics / Q000302:isolation & purification; D017931:DNA Primers; D005243:Feces / Q000382:microbiology*; D020459:Genes, rRNA; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D018720:Prevotella / Q000235:genetics / Q000302:isolation & purification; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D013045:Species Specificity
D017931:DNA Primers; D012336:RNA, Ribosomal, 16S