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Applied and Environmental Microbiology, January 2003, p. 461-467, Vol. 69, No. 1
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.1.461-467.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

In Situ Exposure to Low Herbicide Concentrations Affects Microbial Population Composition and Catabolic Gene Frequency in an Aerobic Shallow Aquifer

Julia R. de Lipthay,^1, Nina Tuxen,² Kaare Johnsen,^1,3 Lars H. Hansen,⁴ Hans-Jørgen Albrechtsen,² Poul L. Bjerg,² and Jens Aamand^1*

Department of Geochemistry, Geological Survey of Denmark and Greenland, Øster Voldgade 10, DK-1350 Copenhagen K,¹ Environment & Resources DTU, Technical University of Denmark, DK-2800 Lyngby,² Danish Veterinary Institute, DK-1790 Copenhagen V,³ Department of General Microbiology, University of Copenhagen, DK-1307 Copenhagen K, Denmark⁴

Received 13 May 2002/ Accepted 30 October 2002

The aim of this study was to evaluate how the in situ exposure of a Danish subsurface aquifer to phenoxy acid herbicides at low concentrations (<40 µg l^-1) changes the microbial community composition. Sediment and groundwater samples were collected inside and outside the herbicide-exposed area and were analyzed for the presence of general microbial populations, Pseudomonas bacteria, and specific phenoxy acid degraders. Both culture-dependent and culture-independent methods were applied. The abundance of microbial phenoxy acid degraders (10⁰ to 10⁴ g^-1 sediment) was determined by most probable number assays, and their presence was only detected in herbicide-exposed sediments. Similarly, PCR analysis showed that the 2,4-dichlorophenoxyacetic acid degradation pathway genes tfdA and tfdB (10² to 10³ gene copies g^-1 sediment) were only detected in sediments from contaminated areas of the aquifer. PCR-restriction fragment length polymorphism measurements demonstrated the presence of different populations of tfd genes, suggesting that the in situ herbicide degradation was caused by the activity of a heterogeneous population of phenoxy acid degraders. The number of Pseudomonas bacteria measured by either PCR or plating on selective agar media was higher in sediments subjected to high levels of phenoxy acid. Furthermore, high numbers of CFU compared to direct counting of 4',6-diamidino-2-phenylindole-stained cells in the microscope suggested an increased culturability of the indigenous microbial communities from acclimated sediments. The findings of this study demonstrate that continuous exposure to low herbicide concentrations can markedly change the bacterial community composition of a subsurface aquifer.

Present address: Department of General Microbiology, University of Copenhagen, Sølvgade 83H, DK-1307 Copenhagen K, Denmark.