Bacterial cultures preferentially removing singly flanked chlorine substituents from chlorobenzenes

The wide though not ubiquitous distribution of chlorobenzene-dechlorinating bacteria in anaerobic sludge from German sewage plants is demonstrated. The model substrates 1,2,3- and 1,2,4-trichlorobenzene (TCB) were dechlorinated to dichlorobenzenes (DCBs) and monochlorobenzene (MCB) via distinct pathways. For easy visualization and differentiation of the pathways, a novel plotting method was developed. While many of the cultures showed a dechlorination pattern similar to that previously found for Dehalococcoides species, removing doubly flanked rather than singly flanked chlorine substituents from TCBs, some cultures formed 1,2-DCB from 1,2,3-TCB and/or 1,3-DCB from 1,2,4-TCB. Stable cultures preferentially catalyzing the removal of singly flanked chlorines were obtained by repeated subcultivation in sediment-free synthetic medium. This dechlorination pattern is potentially of great benefit for remediation as the accumulation of persistent intermediates such as 1,3,5-TCB from highly chlorinated compounds can be avoided. In addition, the cultures dechlorinated 1,3,5-TCB, pentachlorobenzene (PeCB), and hexachlorobenzene (HCB). Nested PCR demonstrated the presence of low numbers of Dehalococcoides species. However, the observed insensitivity of the dechlorinating bacteria in our cultures to oxygen and sensitivity to vancomycin is not in accordance with the reported properties of Dehalococcoides species, suggesting that other bacteria than Dehalococcoides catalyzed the removal of singly flanked chlorines from TCB.