USE OF MICROBIAL RESTRICTION FRAGMENT PROFILES AS MAKERs OF GROUNDWATER SOURCE

Karst groundwater constitutes a critical source of municipal and private water in West Virginia and throughout the Appalachian region. Identifying the sources of groundwater flows in karst systems is therefore critical for the assessment of sustainable water resources in the Appalachians. However characterization of groundwater flow source is challenging and traditionally performed by the use of tracers or analyses of specific chemicals.

In the present research, we propose an innovative approach for tracking the origin of groundwater based on the specific structure of the microbial community using terminal-restriction fragment length polymorphism (T-RFLP) analysis. The DNA pool of a complex microbial community, such as found in most ecosystems, including groundwater, provides very specific restriction fragment profiles that can be used as the fingerprint of a body of water. The objective of this study is to validate the use of T-RFLP profiles obtained from different spring waters as a marker of the groundwater source. T-RFLP results are then compared with other geochemical indicators in water, such as elemental analyses.

Water samples were collected from cold and warm springs in Huntingdon, PA, Great Valley (Martinsburg), WV, and Berkeley Springs, WV. Three liters of water samples were filtered in the field through 0.2 micrometer. Total DNA was directly extracted from the filters and used for the PCR-amplification of full-size16S ribosomal DNA (rDNA) genes. 16S rDNA pools were characterized by restriction fragment analysis (T-RFLP). In addition, genotypic identification of individual bacterial species inhabiting the groundwater was conducted by screening a 16S rDNA gene library. The composition of the bacterial community at the species level provides information about the microbial metabolic activities in groundwater. Identifying trophic groups in groundwater helps determine the geochemistry and, therefore, the origin of water flows in karst systems. Results obtained at the completion of this research are expected to pave the way for more extensive use of complex microbiological profiles as indicators of the groundwater origin.