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Applied and Environmental Microbiology, August 2004, p. 4911-4920, Vol. 70, No. 8
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.8.4911-4920.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Change in Bacterial Community Structure during In Situ Biostimulation of Subsurface Sediment Cocontaminated with Uranium and Nitrate

Nadia N. North,1 Sherry L. Dollhopf,1 Lainie Petrie,1 Jonathan D. Istok,2 David L. Balkwill,3 and Joel E. Kostka1*

Department of Oceanography,1 Department of Biomedical Sciences, College of Medicine, Florida State University, Tallahassee, Florida,3 Department of Civil Engineering, Oregon State University, Corvallis, Oregon2

Received 8 January 2004/ Accepted 5 May 2004

Previous studies have demonstrated that metal-reducing microorganisms can effectively promote the precipitation and removal of uranium from contaminated groundwater. Microbial communities were stimulated in the acidic subsurface by pH neutralization and addition of an electron donor to wells. In single-well push-pull tests at a number of treated sites, nitrate, Fe(III), and uranium were extensively reduced and electron donors (glucose, ethanol) were consumed. Examination of sediment chemistry in cores sampled immediately adjacent to treated wells 3.5 months after treatment revealed that sediment pH increased substantially (by 1 to 2 pH units) while nitrate was largely depleted. A large diversity of 16S rRNA gene sequences were retrieved from subsurface sediments, including species from the {alpha}, ß, {delta}, and {gamma} subdivisions of the class Proteobacteria, as well as low- and high-G+C gram-positive species. Following in situ biostimulation of microbial communities within contaminated sediments, sequences related to previously cultured metal-reducing {delta}-Proteobacteria increased from 5% to nearly 40% of the clone libraries. Quantitative PCR revealed that Geobacter-type 16S rRNA gene sequences increased in biostimulated sediments by 1 to 2 orders of magnitude at two of the four sites tested. Evidence from the quantitative PCR analysis corroborated information obtained from 16S rRNA gene clone libraries, indicating that members of the {delta}-Proteobacteria subdivision, including Anaeromyxobacter dehalogenans-related and Geobacter-related sequences, are important metal-reducing organisms in acidic subsurface sediments. This study provides the first cultivation-independent analysis of the change in metal-reducing microbial communities in subsurface sediments during an in situ bioremediation experiment.

* Corresponding author. Mailing address: Department of Oceanography, Florida State University, Tallahassee, FL 32306. Phone: (850) 645-3334. Fax: (850) 644-2581. E-mail: jkostka{at}ocean.fsu.edu.

Applied and Environmental Microbiology, August 2004, p. 4911-4920, Vol. 70, No. 8
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.8.4911-4920.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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