Use of Fixed-Film Bioreactors, in Situ Microcosms, and Molecular Biological Analyses to Evaluate Bioremediation of Chlorinated Benzenes By Indigenous Bacteria and a Bioaugmented Dechlorinating Consortium

Evaluation of bioremediation is complicated by contaminant mixtures, high concentrations, variable site conditions, and multiple possible degradation pathways. In this study, fixed-film bioreactor experiments, in situ microcosms, and microbial analyses were utilized to evaluate both anaerobic and aerobic biodegradation processes for tri- and dichlorobenzene isomers, monochlorobenzene, and benzene in a wetland. Biofilm-based bioreactors provide a robust assessment tool because of their typically high degree of stability, even with major and repeated perturbations. Two bioreactor units seeded with an anaerobic dechlorinating consortium (WBC-2) and one unit seeded only with bacteria indigenous to the site were operated under flow-through conditions to compare biougmentation and natural attenuation. Electron donor levels were varied to fluctuate between anaerobic and aerobic conditions, and inflow concentrations of total chlorobenzenes were transitioned from 1-10 mg/L to 50-100 mg/L. Biodegradation resulted in removal efficiencies of 80 to 99 percent for the different compounds and inflow concentrations. Degradation efficiency in the native bioreactor was not impacted by cycling between anaerobic and aerobic conditions, although removal rates for monochlorobenzene and benzene increased under aerobic conditions. In situ microcosms were incubated below the wetland surface in sets of 3 treatments—unamended, biostimulated (lactate addition), and bioaugmented (WBC-2 and lactate). Additional treatment sets contained 13C-labeled contaminants to monitor for production of 13C-containing carbon dioxide and cellular material. Microcosm results verified that WBC-2 bioaugmentation can enhance biodegradation, with complete mineralization of chlorobenzene and benzene in bioaugmented and native treatments. Microbial analyses using QuantArrayTM for functional and taxonomic genes indicated potential for co-occurrence of anaerobic and aerobic biodegradation. Compared to the unamended in situ microcosms, the WBC-2 microcosm contained two to five orders of magnitude higher quantities of targeted microbial populations that are associated with degradation of chlorinated and petroleum compounds through both anaerobic and aerobic pathways.