Anaerobic microbial communities and their potential for bioenergy production in heavily biodegraded petroleum reservoirs.

Most of the oil in low temperature, non-uplifted reservoirs is biodegraded due to millions of years of microbial activity, including via methanogenesis from crude oil. To evaluate stimulating additional methanogenesis in already heavily biodegraded oil reservoirs, oil sands samples were amended with nutrients and electron acceptors, but oil sands bitumen was the only organic substrate. Methane production was monitored for over 3000 days. Methanogenesis was observed in duplicate microcosms that were unamended, amended with sulfate or that were initially oxic, however methanogenesis was not observed in nitrate-amended controls. The highest rate of methane production was 0.15 µmol CH4 g-1 oil d-1 , orders of magnitude lower than other reports of methanogenesis from lighter crude oils. Methanogenic Archaea and several potential syntrophic bacterial partners were detected following the incubations. GC-MS and FTICR-MS revealed no significant bitumen alteration for any specific compound or compound class, suggesting that the very slow methanogenesis observed was coupled to bitumen biodegradation in an unspecific manner. After 3000 days, methanogenic communities were amended with benzoate resulting in methanogenesis rates that were 110-fold greater. This suggests that oil-to-methane conversion is limited by the recalcitrant nature of oil sands bitumen, not the microbial communities resident in heavy oil reservoirs.

Volatile hydrocarbons inhibit methanogenic crude oil degradation.

Methanogenic degradation of crude oil in subsurface sediments occurs slowly, but without the need for exogenous electron acceptors, is sustained for long periods and has enormous economic and environmental consequences. Here we show that volatile hydrocarbons are inhibitory to methanogenic oil biodegradation by comparing degradation of an artificially weathered crude oil with volatile hydrocarbons removed, with the same oil that was not weathered. Volatile hydrocarbons (nC5-nC10, methylcyclohexane, benzene, toluene, and xylenes) were quantified in the headspace of microcosms. Aliphatic (n-alkanes nC12-nC34) and aromatic hydrocarbons (4-methylbiphenyl, 3-methylbiphenyl, 2-methylnaphthalene, 1-methylnaphthalene) were quantified in the total hydrocarbon fraction extracted from the microcosms. 16S rRNA genes from key microorganisms known to play an important role in methanogenic alkane degradation (Smithella and Methanomicrobiales) were quantified by quantitative PCR. Methane production from degradation of weathered oil in microcosms was rapid (1.1 ± 0.1 µmol CH4/g sediment/day) with stoichiometric yields consistent with degradation of heavier n-alkanes (nC12-nC34). For non-weathered oil, degradation rates in microcosms were significantly lower (0.4 ± 0.3 µmol CH4/g sediment/day). This indicated that volatile hydrocarbons present in the non-weathered oil inhibit, but do not completely halt, methanogenic alkane biodegradation. These findings are significant with respect to rates of biodegradation of crude oils with abundant volatile hydrocarbons in anoxic, sulphate-depleted subsurface environments, such as contaminated marine sediments which have been entrained below the sulfate-reduction zone, as well as crude oil biodegradation in petroleum reservoirs and contaminated aquifers.

Anaerobic hydrocarbon biodegradation in deep subsurface oil reservoirs.

Biodegradation of crude oil in subsurface petroleum reservoirs is an important alteration process with major economic consequences. Aerobic degradation of petroleum hydrocarbons at the surface is well documented and it has long been thought that the flow of oxygen- and nutrient-bearing meteoric waters into reservoirs was necessary for in-reservoir petroleum biodegradation. The occurrence of biodegraded oils in reservoirs where aerobic conditions are unlikely, together with the identification of several anaerobic microorganisms in oil fields and the discovery of anaerobic hydrocarbon biodegradation mechanisms, suggests that anaerobic degradation processes could also be responsible. The extent of anaerobic hydrocarbon degradation processes in the world's deep petroleum reservoirs, however, remains strongly contested. Moreover, no organism has yet been isolated that has been shown to degrade hydrocarbons under the conditions found in deep petroleum reservoirs. Here we report the isolation of metabolites indicative of anaerobic hydrocarbon degradation from a large fraction of 77 degraded oil samples from both marine and lacustrine sources from around the world, including the volumetrically important Canadian tar sands. Our results therefore suggest that anaerobic hydrocarbon degradation is a common process in biodegraded subsurface oil reservoirs.