Diversity of vinylchloride functional genes and quantification of Dehalococcoides sp. DNA in a groundwater aquifer during a field trial

Pollution with chlorinated ethenes is a major problem in groundwater aquifers. A field trial with a closed groundwater loop recirculation system on a site polluted with chlorinated ethenes in Odense, Denmark, was established with the addition of lactate and a commercial microbial mixed culture (KB-1™). Groundwater samples were collected eight times at 4-week intervals, and DNA was extracted using a modified DNA extraction method. Dehalococcoides sp. was quantified using a previously published 16S rRNAgene targeted primer set. New primers targeting the functional genes vcrA and bvcA were developed. They proved superior to published primers in Real-Time PCR efficiency. A high degree of correlation was found between quantifications by 16S rRNA and the two functional genes, indicating that the identified degradation genes were found in Dehalococcoides sp. analysis (denaturing gradient gel electrophoresis) of the diversity of the functional genes revealed a low diversity of vinyl chloride degraders responding to the primer set on the site. During the field experiment, we found an initial increase from 8x104 Dehalococcoides sp. cells pr 100 ml groundwater to 8x106 cells pr. 100 ml. This number did not increase when KB-1™ was added, but the culture addition was followed by an increased dechlorination rate, in particular, the conversion of vinyl chloride to ethene.