This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Kuritza, A P Articles by Salyers, A A Search for Related Content PubMed PubMed Citation Articles by Kuritza, A P Articles by Salyers, A A Agricola Articles by Kuritza, A P Articles by Salyers, A A

 Previous Article  |  Next Article 

Appl Environ Microbiol. 1985 October; 50(4): 958-964

Use of a species-specific DNA hybridization probe for enumerating Bacteroides vulgatus in human feces.

ABSTRACT

pBV-1, a recombinant plasmid that contains a chromosomal DNA fragment from Bacteroides vulgatus, hybridized to DNA from B. vulgatus but not to DNA from other colonic Bacteroides species. This plasmid was used as a DNA probe to detect and enumerate B. vulgatus in pure culture, in mixed cultures, and in a bacterial fraction from human feces. Bacteria in a pure or mixed culture were lysed by heating the culture in NaOH. The DNA in the disrupted cell suspension was then trapped on nitrocellulose paper by vacuum filtration. If fecal samples were used instead of pure or mixed cultures, it was first necessary to partially purify the DNA by low-speed centrifugation (2,000 X g) and phenol-chloroform extraction before filtering. When 32P-labeled pBV-1 was incubated with filters containg B. vulgatus DNA, the amount of radioactivity that bound to the filters was proportional to the number of B. vulgatus filtered as long as the filtering capacity of the nitrocellulose was not exceeded. Using this procedure, we obtained a value for the concentration of B. vulgatus in human feces (2 X 10(10) to 3 X 10(10) per g of dry weight) that is similar to values obtained by other investigators using conventional bacteriological techniques (3 X 10(10) to 6 X 10(10) per g of dry weight). The advantage of the DNA hybridization method over conventional techniques is that it is not necessary to isolate pure cultures of bacteria from complex specimens such as feces. Furthermore, our method bypasses the cumbersome set of biochemical tests normally used to identify anaerobic bacteria. The major limitation of our method is its sensitivity.(ABSTRACT TRUNCATED AT 250 WORDS)