APPLICATION OF MOLECULAR METHODS AS A BIOMARKER IN BIOREMEDIATION STUDIES

GUERMOUCHE M.A.¹*, BENSALAH F.², GRAY N.^3
1Faculté des Sciences de la Nature et de la Vie. Département de Biotechnologie, Laboratoire de Génétique Microbienne (LGM). Université Es-Sénia- Oran 31000, Algérie.
²Faculté des Sciences de la Nature et de la Vie. Département de Biologie, Laboratoire de Génétique Microbienne (LGM), Université Es-Sénia- Oran 31000, Algérie.
³School of Civil Engineering and Geosciences, University of Newcastle, Newcastle upon Tyne, NE17RU, UK.
* Corresponding Author : amel_guermouche@yahoo.fr

Received : 26-10-2013     Accepted : 21-11-2013     Published : 30-12-2013
Volume : 5     Issue : 1       Pages : 147 - 154
Int J Biotechnol Appl 5.1 (2013):147-154
DOI : http://dx.doi.org/10.9735/0975-2943.5.1.147-154

Keywords : Bioremediation, petroleum-hydrocarbon, microbial consortium, 16S rRNA, Pseudomonas, TLC/FID, alkB genes
Conflict of Interest : None declared
Acknowledgements/Funding : This work was supported by the School of Civil Engineering and Geosciences, University of Newcastle, UK

In contaminated soils, the efficiency of natural attenuation or engineered bioremediation largely depends on the biodegradation capacities of the total microflorae. In the present study, the biodegradation capacities of various bacteria towards petroleum-hydrocarbons were determined under laboratory conditions. The purpose of the study was to isolate and characterize petroleum-degrading bacteria from contaminated soil obtained from a refinery in Arzew, Algeria. A collection of 15 bacterial isolates were obtained by enrichment cultivation from oil-contaminated soil and an indigenous microbial consortium was developed by assembling four species of bacteria which could degrade different fractions of the petroleum hydrocarbons. 16S rRNA gene analysis was used to identify members of the consortium and oil biodegradability was analyzed by Thin Layer Chromatography (TLC) with Flame Ionization Detection (FID) that performs quantitative compositional analysis of oil samples. The Iatroscan TLC/FID system measured the relative percentages of the four major fractions of petroleum i.e. saturates aromatics, resins and asphaltenes. Results indicated that the constructed consortium which comprised the genera Pseudomona, Shewanella, Enterobacter and Serratia used the hydrocarbons as sole sources of carbon where biodegradation was defined by an initial rapid decrease in the saturate and aromatic fractions from 56.44% and 34.72% to 51.77% and 27.77% respectively, coinciding with an increase in the asphaltene fraction. The resin content remained relatively constant throughout the project. Alkane hydroxylase genes (alkB) were positively amplified in the Pseudomonas isolate by the polymerase chain reaction (PCR) method using degenerate primers. This functional gene was used as a marker to assess the catabolic potential of the bacteria for alkane pollutant biodegradation. The selected bacterial consortium looks promising for its application in bioremediation technologies.

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