Enrichment and Characterization of Anaerobic Benzene-Degrading Microbial Cultures

Abstract

<p> Biodegradation of benzene, a common groundwater contaminant, occurs readily in the presence of oxygen; however, at contaminated sites, aerobic bacteria often deplete the available oxygen, resulting in anaerobic conditions. Field and laboratory studies have shown that the anaerobic biodegradation of other aromatic hydrocarbons such as toluene occurs readily, while anaerobic benzene biodegradation has only been documented in a handful of studies. Despite these difficulties, benzene biodegradation has been shown to occur under iron-reducing, sulphate-reducing and methanogenic conditions, but not under nitrate-reducing conditions.</p> <p> The goal of this thesis research was to enrich and characterize the benzene-degrading microbial populations in microcosms and transfer cultures derived from soil from four different sites. Cultures were amended with potential exogenous electron acceptors (nitrate, sulphate, ferric iron) and the rates of biodegradation under different terminal electron accepting processes were determined. Sustained, anaerobic benzene biodegradation was obtained in transfer cultures containing less than 1% of the original soil inoculum. The rate of benzene degradation was variable, ranging from 1 μM/d to more than 75 μM/d. Growth of bacteria was linked to benzene degradation under sulphate-reducing and nitrate-reducing conditions. Growth was very slow, with doubling times of 9-30 days estimated by modelling benzene depletion curves to the Monod kinetic equation. The rate of benzene degradation was influenced most by biomass concentration and much less by the terminal electron accepting process.</p> <p> The ratio of moles of electron acceptor depleted to moles of benzene degraded was calculated and compared to the theoretically predicted ratios to confirm putative terminal electron acceptors. Anaerobic benzene degradation linked to iron reduction, sulphate reduction and methanogenesis was observed in enrichment cultures, corroborating results from previous studies. In addition, in some enrichment cultures, benzene degradation was linked to nitrate reduction. This is the first report demonstrating benzene degradation linked to nitrate reduction.</p>