Monitored natural attenuation (MNA) and enhanced in situ bioremediation (EISB) remedies have the potential to reduce costs associated with cleanup of Department of Defense (DoD) sites impacted by chlorinated solvents. However, there are many DoD sites where tetrachloroethene (PCE) and trichloroethene (TCE) are undergoing only partial dechlorination to cis-1,2-dichloroethene (cDCE) even when sufficient electron donor is present, either because of the absence of required bacteria (Dehalococcoides) or aerobic conditions. Under SERDP project ER-1168, a novel aerobic bacterium (Polaromonas sp. strain JS666) that uses cDCE as a sole carbon and energy source was isolated and characterized. Since it requires no exotic growth factors, JS666 is a promising bioaugmentation culture for aerobic sites where cDCE is recalcitrant. The microorganism grows and thrives where oxygen and cDCE are co-located, and JS666 degrades 1,2-dichloroethane (DCA) and cometabolizes TCE and vinyl chloride (VC). Ideal groundwater conditions for JS666 include dissolved oxygen (DO) levels between 0.01 mg/L and 8 mg/L, low ionic strength (conductivity <15 milliSiemens per centimeter [mS/cm]), a pH of 6.5 to 8, and relatively low concentrations of TCE, 1,2-DCA, and VC (<500 μg/L).

The demonstration was conducted at Site 21, St. Julien’s Creek Annex (SJCA) in Chesapeake, Virginia. This site had appropriate site conditions and a suitable on-site support network, as well as several relatively well-characterized groundwater plumes of chlorinated volatile organic compounds (VOCs); primarily cDCE, TCE, and VC. In the vicinity of the pilot test area (PTA), groundwater flowed towards the west and shallow groundwater typically ranged from 2 to 7 ft below ground surface (bgs). Estimates of the hydraulic gradient and groundwater velocity for the Columbia aquifer were 0.004-0.01 ft/ft and 72 ft/yr, respectively. Preliminary baseline sampling indicated that the groundwater pH was in the 6 to 6.3 range and that buffering would be required.


The objective of this field demonstration was to evaluate the effectiveness of JS666 in competing with indigenous organisms and biodegrading cDCE and other chlorinated ethenes and ethanes in situ. Also assessed was the use of molecular markers to detect the spread of JS666 in groundwater and the effectiveness of isotopes for detecting and quantifying cDCE biodegradation. The project also sought to provide technical data relevant to field-scale aerobic biotreatment using JS666, including documenting benefits of the technology in terms of expected reduction in the duration and cost of remediation of sites where cDCE persists in groundwater.

Demonstration Results

Greater cDCE reductions were observed in many of the wells in the bioaugmented plots compared to the control plots, as evidenced by analysis of VOCs and carbon stable isotopes. However, cDCE biodegradation in the bioaugmentated plots was likely limited by lack of oxygen and inhibited by high levels of TCE in some areas. Reductions in average cDCE concentrations of up to 44% were observed in the bioaugmentation plot receiving oxygen and buffer and up to 25% in the bioaugmentation plot receiving only buffer. qPCR and microcosm results demonstrated the spread, in-situ survival, and sustained activity of the JS666 organisms in the bioaugmented plots. However, it was difficult to tell whether growth was occurring, as bacterial densities did not consistently increase over time.

Addition of the aerobic culture via injection wells was straightforward; aeration of the test plots using the Waterloo Emitter was not difficult, but not effective in distributing oxygen beyond the injection wells. Injection of buffer also was not difficult, but was time consuming and required reapplication due to the soluble nature of the buffer employed. The cost assessment showed a 47% cost savings compared to pump and treat, assuming no aeration or buffering is required and sufficient oxygen is present in the groundwater naturally.

Implementation Issues

At full scale, an underground injection control (UIC) permit may be required for the injection of bacteria, buffer amendments (if needed), and extraction and re-injection of contaminated groundwater (if needed). Buffer addition will be required if the pH of the groundwater is low (pH<6.5), and the amendment process may be time-consuming and require repetition if a soluble buffer is used. Aeration also may be required if the ambient dissolved oxygen is not sufficient to support biodegradation. However, JS666 does not tolerate oxygen concentrations above 10 mg/L; thus, care must be taken not to achieve concentrations above this level. JS666 can degrade cDCE metabolically and TCE and VC cometabolically. As the concentration of TCE increases, the rate of cDCE degradation decreases due to competitive inhibition. Therefore, JS666 will perform better when there are lower concentrations of TCE (<500 µg/L) in groundwater. To mitigate the effects of competitive inhibition due to high TCE concentrations to some extent, higher densities of JS666 can be employed.