DESIGN AND EVALUATION OF A 16S RIBOSOMAL-RNA-TARGETED OLIGONUCLEOTIDEPROBE FOR SPECIFIC DETECTION AND QUANTITATION OF HUMAN FECAL BACTEROIDES POPULATIONS

Colonic Bacteroides include several species which, by their population level and activities, are significant contributers to the metabolic a ctivity and health of man and animals. Yet, the understanding of their ecology has been hampered by the lack of highly specific and reliable enumeration techniques. Based on 16S rRNA sequence comparisons within the available database, we have designed an 18-mer oligonucleotide th at targets a region common to and specific for the Bacteroides-Porphyr omonas-Prevotella group. We have tested the specificity of the probe a nd its usefulness for studies of human faecal samples. Under experimen tally optimized hybridization conditions, the probe was shown to simil arly recognize the rDNA obtained from 40 strains representing 8 specie s of the Bacteroides-Porphyromonas-Prevotella group. Importantly, it d id not recognize 31 strains of microorganisms representing 8 genera of the dominant human faecal microbiota. Among selected colonies of domi nant microorganisms of the faecal flora of two human individuals, stra ins identified as B. vulgatus by immunoblots using a species-specific monoclonal antibody were ail detected by the probe. Colony hybridizati on was used to enumerate total Bacteroides-group microorganisms in fae cal specimen from children and adults. The probe described therein was further used in quantitative RNA blots to monitor fluctuations of the Bacteroides-group versus Bifidobacterium genus in frozen faecal sampl es from a child between 85 and 125 days of age. It will be applicable to similar investigations of other anaerobic environments.