| Understanding of microorganisms and pathways involved in anaerobic benzene degradation is limited. Stable isotope probing of DNA was used to identify key members of a previously characterized, sulfate-reducing benzene degrading consortium. DNA extracts of cultures incubated with [13C 6]- or [12C6]benzene were separated into 13C- and 12C-labeled fractions by CsCl density gradient centrifugation. Sequencing and Terminal Restriction Fragment Length Polymorphism (T-RFLP) analysis of the 16S rRNA gene identified TRF 270 (bp), a Desulfobacterium like phylotype, which was first to derive the bulk of the 13C label for DNA synthesis, and is thus likely involved in activation of benzene degradation.;To understand the pathway of anaerobic benzene metabolism, degradation and inhibition tests were used. Based on these tests toluene was eliminated, and benzoate was identified as a possible intermediate. Metabolites detected in cultures amended with [13C6]benzene or [ 13C6]phenol indicate that in this consortium there are 2 different pathways of benzoate formation, one forms universally labeled ([13C-UL]benzoate), and the other forms ring labeled benzoate. Pathway that forms [13C-UL]benzoate is dominant during benzene degradation in which the benzene ring is carboxylated by a carbon derived from another benzene ring. This pathway is different from the proposed pathway of benzene degradation via phenol, as the labeling pattern of 13C-labeled benzoate formed from [13C6]benzene or [13 C6]phenol is not identical. In conclusion, a novel pathway that activates one benzene ring through its reaction with products of another benzene ring likely exists in this consortium.;Groundwater impacted by a manufacturing gas plant site was used for detection and quantification of metabolic intermediates of polycyclic aromatic hydrocarbons and gene analogues encoding alpha subunit of benzylsuccinate synthase ( bssA), as evidence for natural attenuation. Highest concentrations of metabolic intermediates of anaerobic naphthalene and 2-methylnaphthalene degradation were detected in an impacted monitoring well (MW)-24, near the source. Quantitative analysis of 16S rRNA gene indicated that bacterial population was enriched in the impacted wells, while bssA gene containing bacterial community was enriched in MW-24. Detection of not one, but two different indicators specific to the presence and activity of microorganisms provides strong evidence for in situ anaerobic microbial processes. |
