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Applied and Environmental Microbiology, May 2004, p. 2952-2958, Vol. 70, No. 5
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.5.2952-2958.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Impact of Temperature on the Physiological Status of a Potential Bioremediation Inoculant, Arthrobacter chlorophenolicus A6

Agneta Backman,1,2 Ninwe Maraha,1,2 and Janet K. Jansson1,3*

Section for Natural Sciences, Södertörn University College, Huddinge,1 Department of Laboratory Medicine, Karolinska Institute, Huddinge University Hospital, Stockholm,2 Department of Microbiology, Swedish University of Agricultural Sciences, Uppsala, Sweden3

Received 3 October 2003/ Accepted 13 February 2004

Arthrobacter chlorophenolicus A6 (A6) can degrade large amounts of 4-chlorophenol in soil at 5 and 28°C. In this study, we investigated the effects of temperature on the physiological status of this bacterium in pure culture and in soil. A derivative of A6 tagged with the gfp gene (encoding green fluorescent protein [GFP]) was used to specifically quantify A6 cells in soil. In addition, cyano-ditolyl-tetrazoliumchloride was used to stain GFP-fluorescent cells with an active electron transfer system ("viable cells") whereas propidium iodide (PI) was used to stain cells with damaged membranes ("dead cells"). Another derivative of the strain (tagged with the firefly luciferase gene [luc]) was used to monitor the metabolic activity of the cell population, since the bioluminescence phenotype is dependent on cellular energy reserves. When the cells were incubated in soil at 28°C, the majority were stained with PI, indicating that they had lost their cell integrity. In addition, there was a corresponding decline in metabolic activity and in the ability to be grown in cultures on agar plates after incubation in soil at 28°C, indicating that the cells were dying under those conditions. When the cells were incubated in soil at 5°C, by contrast, the majority of the cells remained intact and a large fraction of the population remained metabolically active. A similar trend towards better cell survival at lower temperatures was found in pure-culture experiments. These results make A. chlorophenolicus A6 a good candidate for the treatment of chlorophenol-contaminated soil in cold climates.

* Corresponding author. Mailing address: Department of Microbiology, Swedish University of Agricultural Sciences, Box 7025, 750 07 Uppsala, Sweden. Phone: 46 18673201. Fax: 46 18673392. E-mail: Janet.Jansson{at}mikrob.slu.se.

Applied and Environmental Microbiology, May 2004, p. 2952-2958, Vol. 70, No. 5
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.5.2952-2958.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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