Towards bioremediation of toxic unresolved complex mixtures of hydrocarbons: identification of bacteria capable of rapid degradation of alkyltetralins

Abstract

Unresolved complex mixtures (UCMs) of aromatic hydrocarbons are widespread, but often overlooked, environmental contaminants. Since UCMs are generally rather resistant to bacterial degradation, bioremediation of UCM-contaminated sites by bacteria is a challenging goal. Branched chain alkyltetralins are amongst the individual classes of components of aromatic UCMs which have been identified in hydrocarbon-contaminated sediments and a number of synthetic alkyltetralins have proved toxic in laboratory studies. Thus, alkyltetralins should perhaps be amongst the targets for UCM bioremediation strategies. The slow degradation of several alkyltetralins by a microbial consortium has been reported previously; however, the bacteria involved remain unidentified and no single strain capable of alkyltetralin biodegradation has been isolated. The present project therefore aimed to enrich and identify bacterial consortia and single strains of bacteria from a naturally hydrocarbon-contaminated site (Whitley Bay, UK), which were capable of the degradation of two synthetic alkyltetralins (6-cyclohexyltetralin (CHT) and 1-(3’-methylbutyl)-7-cyclohexyltetralin (MBCHT)). Bacteria were enriched from sediment collected from Whitley Bay, UK by culturing with CHT and MBCHT for a period of 4 months. Biodegradation experiments were then established and degradation of model compounds monitored by gas chromatography–mass spectrometry. Internal standards allowed the generation of quantitative data. 16S rRNA gene clone libraries were constructed from individual enrichments to allow assessment of microbial community structure. Selective media containing MBCHT were used to isolate single bacterial strains. These strains were then tested in liquid culture for their ability to degrade MBCHT. The consortia obtained through enrichment culture were able to degrade 87% of CHT and 76% of MBCHT after only 46 days compared with abiotic controls. The 16S ribosomal RNA gene clone libraries of these bacteria were dominated by sequences of Rhodococcus spp. Using selective media, a strain of Rhodococcus was then isolated that was also able to biodegrade 63% of MBCHT in only 21 days. The present report describes the isolation of a single bacterial strain able to degrade the resistant MBCHT. Although significant losses of MBCHT were observed, putative metabolites were not detectable. Rhodococcus sp. have been reported previously to be able to biodegrade a range of hydrocarbon compounds. Due to their environmental persistence and toxicity, aromatic UCMs require bioremediation. The culturing and identification of such bacteria capable of rapid degradation of alkyltetralins may be an important step toward the development of bioremediation strategies for sites contaminated with toxic UCMs.