RRC ID 5154
Author Hong PY, Wu JH, Liu WT.
Title Relative abundance of Bacteroides spp. in stools and wastewaters as determined by hierarchical oligonucleotide primer extension.
Journal Appl. Environ. Microbiol.
Abstract A molecular method, termed hierarchical oligonucleotide primer extension (HOPE), was used to determine the relative abundances of predominant Bacteroides spp. present in fecal microbiota and wastewaters. For this analysis, genomic DNA in feces of healthy human adults, bovines, and swine and in wastewaters was extracted and total bacterial 16S rRNA genes were PCR amplified and used as the DNA templates for HOPE. Nineteen oligonucleotide primers were designed to detect 14 Bacteroides spp. at different hierarchical levels (domain, order, cluster, and species) and were arranged into and used in six multiplex HOPE reaction mixtures. Results showed that species like B. vulgatus, B. thetaiotaomicron, B. caccae, B. uniformis, B. fragilis, B. eggerthii, and B. massiliensis could be individually detected in human feces at abundances corresponding to as little as 0.1% of PCR-amplified 16S rRNA genes. Minor species like B. pyogenes, B. salyersiae, and B. nordii were detected only collectively using a primer that targeted the B. fragilis subgroup (corresponding to approximately 0.2% of PCR-amplified 16S rRNA genes). Furthermore, Bac303-related targets (i.e., most Bacteroidales) were observed to account for 28 to 44% of PCR-amplified 16S rRNA genes from human fecal microbiota, and their abundances were higher than those detected in the bovine and swine fecal microbiota and in wastewaters by factors of five and two, respectively. These results were comparable to those obtained by quantitative PCR and to those reported previously from studies using whole-cell fluorescence hybridization and 16S rRNA clone library methods, supporting the conclusion that HOPE can be a sensitive, specific, and rapid method to determine the relative abundances of Bacteroides spp. predominant in fecal samples.
Volume 74(9)
Pages 2882-93
Published 2008-5
DOI 10.1128/AEM.02568-07
PII AEM.02568-07
PMID 18344347
PMC PMC2394900
MeSH Adult Animals Bacteroidetes / classification Bacteroidetes / genetics* Bacteroidetes / isolation & purification* Cattle Colony Count, Microbial / methods DNA Primers / genetics DNA, Bacterial / chemistry DNA, Bacterial / genetics DNA, Ribosomal / chemistry DNA, Ribosomal / genetics Feces / microbiology* Humans Molecular Sequence Data Nucleic Acid Amplification Techniques / methods* Phylogeny RNA, Ribosomal, 16S / genetics Sensitivity and Specificity Sequence Analysis, DNA Sewage / microbiology* Swine
IF 3.633
Times Cited 27
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