Q-186. Diversity of Naphthalene-Degrading Bacteria in an Aged, PAH-Contaminated Soil Identified by Stable-Isotope Probing

M. D. Jones, D. R. Singleton, M. D. Aitken;
Univ. of North Carolina Sch. of Pub. Hlth., Chapel Hill, NC.

Naphthalene is a low-molecular-weight polycyclic aromatic hydrocarbon (PAH) to which humans are commonly exposed. Soil and groundwater can become contaminated with naphthalene from industrial sources of PAHs. Bioremediation of PAH contamination can be aided by identifying the indigenous organisms able to degrade the compound(s) of interest. To define the community of bacteria responsible for naphthalene degradation in an aged, PAH-contaminated soil, we performed a DNA-based stable-isotope probing (SIP) investigation. PAH-contaminated soil was obtained from a former manufactured gas plant site in Salisbury, North Carolina. Continuously mixed slurry microcosms were incubated in the dark at room temperature for 2 days. The aqueous phase was then replaced with fresh buffer and [U-13C]naphthalene and incubated in the dark at room temperature for 8 additional hours. Recovered 13C-enriched DNA was analyzed by denaturing-gradient gel electrophoresis (DGGE) and 16S rRNA gene clone libraries. The DGGE banding pattern of the enriched community was a subset of the unenriched community. The majority of the sequences (53/69) resulting from clone library analysis grouped closely with one of three genera: Pseudomonas, Sphingomonas, or Variovorax. The remaining sequences were most similar to members of the Acidovorax, Achromobacter, Alcaligenes, Pigmentiphaga, or Pseudoxanthomonas genera. SIP investigations of other PAH-contaminated soils have previously identified Pseudomonas and Variovorax as participating in naphthalene degradation. SIP has also linked Acidovorax species to phenanthrene but not naphthalene degradation. The other represented genera have all been detected in PAH-contaminated soils by various methods, but not by SIP. Our results provide evidence that these organisms participate specifically in the metabolism of naphthalene.