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Applied and Environmental Microbiology, March 2003, p. 1372-1376, Vol. 69, No. 3
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.3.1372-1376.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Expression of Chlorocatechol 1,2-Dioxygenase and Chlorocatechol 2,3-Dioxygenase Genes in Chlorobenzene-Contaminated Subsurface Samples

Albin Alfreider,* Carsten Vogt, and Wolfgang Babel

Department of Environmental Microbiology, UFZ Centre for Environmental Research, Leipzig 04318, Germany

Received 9 August 2002/ Accepted 10 December 2002

In order to evaluate the in situ degradative capabilities of microorganisms in an underground reactor facility housing two flowthrough columns filled with aquifer soil, we examined the distribution and phylogeny of gene transcripts encoding enzymes capable of catalyzing the cleavage of the chlorinated aromatic ring during transformation of the main pollutant, chlorobenzene. Initial biostimulation of the autochthonous bacteria in the originally anaerobic reactor columns was achieved by injecting nitrate and oxygen in the form of H2O2. Two broad-range primer pairs were used for reverse transcriptase PCR (RT-PCR) of partial subunit genes of chlorocatechol 1,2-dioxygenase and catechol 2,3-dioxygenase from RNA directly extracted from different groundwater and aquifer samples. Samples retrieved from the lowermost sections of the reactor columns, which were operated in upflow mode, were positive for the presence of chlorocatechol 1,2-dioxygenase and catechol 2,3-dioxygenase mRNA. On the other hand, chlorocatechol 1,2-dioxygenase RT-PCR products were detected in a larger part of each reactor column, up to a zone 5.5 m above the bottom. Phylogenetic analyses of these chlorocatechol 1,2-dioxygenase sequences clearly separated them into two main clusters, one of which was closely affiliated with the broad-spectrum chlorocatechol 1,2-dioxygenase from Pseudomonas chlororaphis RW71. Analysis of sequences obtained from RT-PCR products amplified with catechol 2,3-dioxygenase primers revealed that their closest relative was the chlorocatechol 2,3-dioxygenase gene cbzE from Pseudomonas putida GJ31 (A. E. Mars, J. Kingma, S. R. Kaschabek, W. Reineke, and D. B. Janssen, J. Bacteriol. 181:1309-1318, 1999), with sequence similarities between 97.8 and 99.0%.

* Corresponding author. Mailing address: Department of Environmental Microbiology, UFZ Centre for Environmental Research, Leipzig 04318, Germany. Phone: 49-341-235-2367. Fax: 49-341-235-2247. E-mail: alfreid{at}umb.ufz.de.

Applied and Environmental Microbiology, March 2003, p. 1372-1376, Vol. 69, No. 3
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.3.1372-1376.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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  • Sharkey, F. H., Banat, I. M., Marchant, R. (2004). Detection and Quantification of Gene Expression in Environmental Bacteriology. Appl. Environ. Microbiol. 70: 3795-3806 [Full Text]  


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