Genetic Techniques for the Verification and Monitoring of Dihaloethane Biodegradation in New Mexico Aquifers

The dihaloethanes 1,2-dibromoethane (EDB) and 1,2-dichloroethane (EDC) are used in industrial applications. Both are carcinogenic and cytotoxic. The primary source of dihaloethane contamination is associated with petroleum refining industries and fuel dispensing systems. In New Mexico, approximately 175 locations have dihaloethane-contaminated soil and groundwater. The objective was to determine the potential application of molecular biological tools to monitor biodegradation potential of contaminated aquifers.

Sites for preliminary experiments were in Ribera and Socorro, New Mexico. Methods for isolation of microbes from aquifer samples included centrifugation and micro-filtration. Both were adequate, but micro-filtration on-site allowed the collection of larger sample volumes and eliminated the need to transport water to the lab. Once isolated and concentrated, the samples were divided for DNA and protein isolation. Polymerase chain reaction (PCR) was used to amplify the16SrRNA gene from the DNA. The PCR product was cloned and sequenced. Bacterial species were determined by sequence comparison to GenBank. Attempts to amplify the gene for dehaloalkane dehalogenase (dhlA) from the DNA proved inconclusive. However, enzyme activity was detected in protein extracts from contaminated aquifers. The ability to quantify enzyme activity directly from groundwater provides a rapid method for estimation of biodegradation potential.