Determination of biodegradation potential by two culture-independent methods in PAH-contaminated soils

Exiguobacterium sp. CNU020, an alkaliphile and extremohalophile bacterium, is resistant to 20 mM H2O2, a concentration that is lethal to most bacteria. Representative antioxidant genes of catalase and superoxide dismutase (SOD), obtained by PCR amplification of the genomic DNA, were characterized: the 252-bp catalase gene shared 77% similarity in the deduced amino acid sequence to that of E. oxidotolerans T-2-2(T). The 420-bp SOD gene had the closest similarity (94.3%) to the manganese-SOD of E. sibiricum 255-15. Through activity-staining analysis, stain CNU020 had at least four catalase isoforms: C1, C2, C3 and C4. Expression of each catalase isoform was dependent on the growth phase and oxidants but two catalases (C3 and C4) were always induced and expressed at a similar rate, indicating that they were constitutively expressed. RT-PCR-based expression analysis at the transcriptional level suggested that the catalase gene is strongly expressed in response to 2 mM H2O2, 0.2 mM Paraquat and 0.2 mM menadione. However, the SOD gene exhibited no observable expression pattern with 2 mM H2O2 despite its strong expression when exposed to Paraquat and menadione.This research was supported by grants from the New Jersey Hazardous Substance Management Research Center and the National Institute of Environmental Health Sciences. Additional support was provided by the KOSEF through the AEBRC at POSTECH. The authors also thank the Research Institute of Engineering Science at Seoul National University for technical assistance

Evaluation of carbazole degradation by Pseudomonas rhodesiae strain KK1 isolated from soil contaminated with coal tar

In this study, strain KK1 isolated from coal tar-contaminated soil was found to be able to mineralize carbazole as a sole source of carbon by radiorespirometric analysis. KK1 cells pregrown on phenanthrene were able to mineralize carbazole much more rapidly than cells pregrown on naphthalene, suggesting a possible close linkage between the pathways for carbazole and phenanthrene catabolism. Also, Rieske-type iron sulfur center sequence of dioxygenase from KK1 was analyzed to evaluate carbazole catabolism by KK1. A gene cloned out from KK1 using a universal dioxygenase primer set was found a dioxygenase for initial catabolism of carbazole based on deduced amino acid sequences. Northern hybridization using the putative carbazole dixoygenase gene fragment as a probe provided the information that catabolism of carbazole might be greatly activated in phenanthrene-grown cells. Analysis of PLFAs extracted from KK1 cells exposed to carbazole revealed that lipids 10 : 0 3OH, 17 : 0 cyclo, and 18 : 0 were representatives produced or significantly increased in response to carbazole. Strain KK1 was identified as Pseudomonas species with 94% confidence when BIOLOG system was applied, as Pseudomonas sp. with over 90% confidence by total cellular compositions of fatty acid, and as Pseudomonas rhodesiae with 99% confidence by 16S rRNA sequence. Accordingly, strain KK1 was identified as Pseudomonas rhodesiae based on combination of the data, and designated Pseudomonas rhodesiae KK1. The phylogenetic tree based on 16S rRNA suggested that strain KK1 was far away in the phylogenetic distance from the strains that can degrade carbazole. The global pollution of soils, rivers, lakes, marshes, etc by polycyclic aromatic hydrocarons (PAHs) such as anthracene, chrysene, fluorene, naphthalene, phenanthrene, and pyrene has been of great concern to environmental microbiologists, because PAHs are considered serious pollutants that are hard to be degraded in the environment. However, little attention has been paid to carbazole that is a nitrogen heterocyclic aromatic compound, despite its toxicity Microorganisms play a primary role in the removal of many types of chemical pollutants including carbazole from the environment. Studies on biodegradation of carbazole by microorganisms such as Pseudomonas, Ralstonia, and Sphingomonas revealed the metabolic pathway for initial carbazole methabolism as shown i