Alternative primer sets for PCR detection of genotypes involved in bacterial aerobic BTEX degradation: Distribution of the genes in BTEX degrading isolates and in subsurface soils of a BTEX contaminated industrial site

Eight new primer sets were designed for PCR detection of (i) mono-oxygenase and dioxygenase gene sequences involved in initial attack of bacterial aerobic BTEX degradation and of (ii) catechol 2,3-dioxygenase gene sequences responsible for meta-cleavage of the aromatic ring. The new primer sets allo...

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Published in:	Journal of microbiological methods Vol. 64; no. 2; pp. 250 - 265
Main Authors:	Hendrickx, Barbara, Junca, Howard, Vosahlova, Jolana, Lindner, Antje, Rüegg, Irene, Bucheli-Witschel, Margarete, Faber, Folkert, Egli, Thomas, Mau, Margit, Schlömann, Michael, Brennerova, Maria, Brenner, VLadimir, Pieper, Dietmar H., Top, Eva M., Dejonghe, Winnie, Bastiaens, Leen, Springael, Dirk
Format:	Journal Article
Language:	English
Published:	Shannon Elsevier B.V 01.02.2006 Elsevier Science
Subjects:	Actinobacteria Actinobacteria - genetics Actinobacteria - metabolism Aerobic BTEX biodegradation Bacteria, Aerobic - genetics Bacteria, Aerobic - metabolism Bacterial Proteins Bacteriology Biodegradation, Environmental Biological and medical sciences BTEX contaminated site BTEX degrading isolates Carbohydrate Epimerases Catabolic gene distribution DNA Primers Environmental Pollution Fundamental and applied biological sciences. Psychology Gene Transfer, Horizontal Genes, Bacterial - physiology Genetics Hydrocarbons - metabolism Industrial Waste Microbiology PCR detection Polymerase Chain Reaction - methods Proteobacteria Proteobacteria - genetics Proteobacteria - metabolism Pseudomonas Soil Microbiology Soil Pollutants - metabolism Species Specificity Substrate Specificity PCR detection Aerobic BTEX biodegradation BTEX contaminated site Catabolic gene distribution BTEX degrading isolates Pseudomonadales Biodegradation Eubacteria Toluene Pseudomonas Polymerase chain reaction Gene Benzene Xylene Bacteria Pseudomonadaceae Actinobacteria Catabolism Detection
ISSN:	0167-7012, 1872-8359
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Summary:	Eight new primer sets were designed for PCR detection of (i) mono-oxygenase and dioxygenase gene sequences involved in initial attack of bacterial aerobic BTEX degradation and of (ii) catechol 2,3-dioxygenase gene sequences responsible for meta-cleavage of the aromatic ring. The new primer sets allowed detection of the corresponding genotypes in soil with a detection limit of 10 3–10 4 or 10 5–10 6 gene copies g − 1 soil, assuming one copy of the gene per cell. The primer sets were used in PCR to assess the distribution of the catabolic genes in BTEX degrading bacterial strains and DNA extracts isolated from soils sampled from different locations and depths (vadose, capillary fringe and saturated zone) within a BTEX contaminated site. In both soil DNA and the isolates, tmoA-, xylM- and xylE1-like genes were the most frequently recovered BTEX catabolic genes. xylM and xylE1 were only recovered from material from the contaminated samples while tmoA was detected in material from both the contaminated and non-contaminated samples. The isolates, mainly obtained from the contaminated locations, belonged to the Actinobacteria or Proteobacteria (mainly Pseudomonas). The ability to degrade benzene was the most common BTEX degradation phenotype among them and its distribution was largely congruent with the distribution of the tmoA-like genotype. The presence of tmoA and xylM genes in phylogenetically distant strains indicated the occurrence of horizontal transfer of BTEX catabolic genes in the aquifer. Overall, these results show spatial variation in the composition of the BTEX degradation genes and hence in the type of BTEX degradation activity and pathway, at the examined site. They indicate that bacteria carrying specific pathways and primarily carrying tmoA/ xylM/ xylE1 genotypes, are being selected upon BTEX contamination.
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ISSN:	0167-7012 1872-8359
DOI:	10.1016/j.mimet.2005.04.018