DETECTION OF HYDROCARBON DEGRADINGTHERMOPHILIC BACTERIA USING MOLECULARBIOLOGY TECHNIQUES AND POSSIBLE USES INACCELERATED HYDROCARBON BIOREMEDIATION

摘要

During the past two years we have established that thermophilicrnbacteria belonging to the genus Geobacillus exist in large numbersrnin otherwise cool soil environments. These thermophiles have arngrowth range of 40-80℃ and can be found in soils in high numbersrn(104 colony forming units (CFU)/g). These thermophilicrngeobacilli have a rapid growth rate with doubling times betweenrn20-30 minutes at 65-70℃ on nutrient broth and most have a widernrang ability to degrade hydrocarbons including aromatic molecules.rnThis suggested that they could play a role in the attenuationrnof hydrocarbon-contaminated soils. To assess their activity in situ,rnmolecular biology techniques were employed firstly though designingrna taxon specific forward PCR primer (THERM) to detectrnthe geobacilli and secondly through targeting the alkane monooxygenasern(alkB) gene expression. A reverse transcriptase PCRrnexperiment established that the alkane monooxygenase activity inrnpure cultures of Geobacillus are only inducible when they arerngrown at temperatures > 40℃ in the presence of alkane. Similarlyrnsoil microcosms spiked with alkane only gave the alkB RT PCRrnproduct when incubated at temperatures above 40℃. Mixedrnsequences of the alkB gene fragment were detected in soil andrnappear to belong to numerous thermophilic rather than mesophilicrnbacteria. We believe these thermophilic geobacilli may have arnsignificant role in hydrocarbon biodegradation at elevated temperaturesrnand may be utilized for bioremediation of hydrocarbon-rnhydrocarboncontaminatedrnsoils.