Sensitive detection of a novel class of toluene-degrading denitrifiers, Azoarcus tolulyticus, with small-subunit rRNA primers and probes.

Applied and Environmental Microbiology
J ZhouJ M Tiedje

Abstract

Azoarcus tolulyticus is a new class of widely distributed toluene-degrading denitrifiers of potential importance in remediating benzene, toluene, ethylbenzene, and xylene (BTEX)-contaminated environments. To detect these organisms in the environment, 16S rRNA gene-based phylogenetic probes were developed. Two sets of specific PCR amplification primers and two oligonucleotide hybridization probes were designed and tested against both closely and distantly related environmental isolates. All of these primers and probes were specific to the species A. tolulyticus. The sensitivity of the PCR amplification primer sets was evaluated with DNA isolated from A. tolulyticus Tol-4 pure culture and from sterile soils seeded with a known number of Tol-4 and Escherichia coli cells. These primer sets were able to detect 1 fg to 1 pg of template DNA from the pure culture and 1.11 x 10(2) to 1.1 x 10(8) Tol-4 cells per g of soil in the presence of 1.56 x 10(10) E. coli cells. These two PCR amplification primers were also successfully tested at two field sites. The primers identified the A. tolulyticus strains among the toluene-degrading bacteria isolated from a low-O2-high-NO(3)- aquifer at Moffett Field, Calif. Also, the presence of A. tolulyt...Continue Reading

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Citations

Jul 1, 2004·FEMS Microbiology Ecology·Rainer U MeckenstockChristian Griebler
Jan 9, 2004·Applied and Environmental Microbiology·Blaz StresJames M Tiedje
Dec 14, 2011·Applied and Environmental Microbiology·Weimin Sun, Alison M Cupples

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