Anaerobic degradation of cyclohexane by sulfate-reducing bacteria from hydrocarbon-contaminated marine sediments.

Ulrike Jaekel,Florin Musat,Heinz Wilkes,Johannes Zedelius

doi:10.3389/fmicb.2015.00116

Abstract

The fate of cyclohexane, often used as a model compound for the biodegradation of cyclic alkanes due to its abundance in crude oils, in anoxic marine sediments has been poorly investigated. In the present study, we obtained an enrichment culture of cyclohexane-degrading sulfate-reducing bacteria from hydrocarbon-contaminated intertidal marine sediments. Microscopic analyses showed an apparent dominance by oval cells of 1.5 × 0.8 μm. Analysis of a 16S rRNA gene library, followed by whole-cell hybridization with group- and sequence-specific oligonucleotide probes showed that these cells belonged to a single phylotype, and were accounting for more than 80% of the total cell number. The dominant phylotype, affiliated with the Desulfosarcina-Desulfococcus cluster of the Deltaproteobacteria, is proposed to be responsible for the degradation of cyclohexane. Quantitative growth experiments showed that cyclohexane degradation was coupled with the stoichiometric reduction of sulfate to sulfide. Substrate response tests corroborated with hybridization with a sequence-specific oligonucleotide probe suggested that the dominant phylotype apparently was able to degrade other cyclic and n-alkanes, including the gaseous alkane n-butane. Based on GC-MS analyses of culture extracts cyclohexylsuccinate was identified as a metabolite, indicating an activation of cyclohexane by addition to fumarate. Other metabolites detected were 3-cyclohexylpropionate and cyclohexanecarboxylate providing evidence that the overall degradation pathway of cyclohexane under anoxic conditions is analogous to that of n-alkanes.

Highlights

Cycloalkanes are major constituents of crude oils, accounting for 20–40% of the total hydrocarbon fractions (Tissot and Welte, 1984), and are common in refined petroleum products
The enrichment culture obtained from intertidal hydrocarboncontaminated marine sediments was dominated by a single phylotype affiliated with the Desulfosarcina-Desulfococcus cluster of the Deltaproteobacteria
Due to its abundance in the enrichment culture, we propose that this phylotype was responsible for the degradation of cyclohexane

Summary

Introduction

Cycloalkanes are major constituents of crude oils, accounting for 20–40% of the total hydrocarbon fractions (Tissot and Welte, 1984), and are common in refined petroleum products Cyclopentane, cyclohexane and their alkylated derivatives, especially methylcyclopentane and methylcyclohexane are most abundant (Tissot and Welte, 1984). A relatively low number of aerobic bacterial strains able to degrade cyclohexane have been isolated. These were affiliated with the Actinobacteria (Stirling et al, 1977), or with the Proteobacteria (Anderson et al, 1980; Trower et al, 1985; Rouviere and Chen, 2003). Genes coding for the cyclohexane degradation pathway were found in metagenome and metatranscriptome libraries, indicating that cyclohexane-degrading bacteria were abundant and active in the crude oil contaminated waters (Mason et al, 2012)

Methods

Results

Conclusion