Using 'omics' approaches to study anaerobic hydrocarbon biodegradation by microbes indigenous to oil sands tailings ponds

In oil sands tailings ponds, methanogenesis is driven in part by the degradation of hydrocarbons in residual solvents used as a diluent during bitumen extraction, such as naphtha. Alkanes constitute a large proportion of these unrecovered hydrocarbons in mature fine tailings (MFT). Methanogenic degradation of alkanes has been poorly described in the literature. Fumarate addition, widely reported for activation of alkanes and monoaromatic compounds under sulfate- and nitrate-reducing conditions, has not been demonstrated conclusively for alkane degradation under methanogenic conditions because signature metabolites and key organisms have not been detected and/or isolated. In order to understand methanogenic alkane degradation by microorganisms indigenous to oil sands tailings ponds, a model alkane-degrading culture (SCADC) was established using MFT obtained from Mildred Lake Settling Basin (MLSB) tailings pond.;Data mining of metagenomes of MLSB and hydrocarbon-impacted environments recovered novel fumarate addition genes undescribed previously, indicating the overall ubiquitous nature of these genes in anoxic environments. Comparative metagenomic analysis of SCADC to two other metagenomes of methanogenic toluene- and naphtha-degrading cultures, in addition to physiological studies, suggests that fumarate addition may be the bottleneck reaction in these three cultures. The cultures have the genetic capability of degrading structurally diverse hydrocarbons and share highly conserved and streamlined functions for anaerobic respiration and methanogenesis, unlike in situ environments impacted by hydrocarbons, which are highly variable in their functional capabilities. This observation provides future prospects for development of commercial cultures for bioremediation and biomethanization.;SCADC degraded many lower molecular weight alkanes, represented by n-C6 -C10, 2-methylpentane and methylcyclopentane during year-long methanogenic incubation, but expected fumarate addition products were only detected for 2-methylpentane and methylcyclopentane. Nucleic acids isolated from SCADC were subjected to metagenomic and metatranscriptomic analysis using Illumina Hi-Seq. Metagenomic binning using multiple approaches recovered several partial genomes, including novel syntrophic Desulfotomaculum and Smithella spp. that are genetically capable of fumarate addition, which was previously unknown. Metatranscriptomic analysis further confirmed the high expression of genes encoding enzymes for alkane addition to fumarate by Desulfotomaculum but not Smithella during active methanogenesis, indicating the importance of Firmicutes in fumarate activation of low molecular weight alkanes.