ABSTRACT
BACKGROUND: Over three-fifths of the world's known crude oil cannot be recovered using state-of-the-art techniques, but microbial conversion of petroleum hydrocarbons trapped in oil reservoirs to methane is one promising path to increase the recovery of fossil fuels. The process requires cooperation between syntrophic bacteria and methanogenic archaea, which can be affected by sulfate-reducing prokaryotes (SRPs). However, the effects of sulfate on hydrocarbon degradation and methane production remain elusive, and the microbial communities involved are not well understood. RESULTS: In this study, a methanogenic hexadecane-degrading enrichment culture was treated with six different concentrations of sulfate ranging from 0.5 to 25 mM. Methane production and maximum specific methane production rate gradually decreased to 44 and 56% with sulfate concentrations up to 25 mM, respectively. There was a significant positive linear correlation between the sulfate reduction/methane production ratio and initial sulfate concentration, which remained constant during the methane production phase. The apparent methanogenesis fractionation factor (αapp) gradually increased during the methane production phase in each treatment, the αapp for the treatments with lower sulfate (0.5-4 mM) eventually plateaued at ~1.047, but that for the treatment with 10-25 mM sulfate only reached ~1.029. The relative abundance levels of Smithella and Methanoculleus increased almost in parallel with the increasing sulfate concentrations. Furthermore, the predominant sulfate reducer communities shifted from Desulfobacteraceae in the low-sulfate cultures to Desulfomonile in the high-sulfate cultures. CONCLUSION: The distribution of hexadecane carbon between methane-producing and sulfate-reducing populations is dependent on the initial sulfate added, and not affected during the methane production period. There was a relative increase in hydrogenotrophic methanogenesis activity over time for all sulfate treatments, whereas the total activity was inhibited by sulfate addition. Both Smithella and Methanoculleus, the key alkane degraders and methane producers, can adapt to sulfate stress. Specifically, different SRP populations were stimulated at various sulfate concentrations. These results could help to evaluate interactions between sulfate-reducing and methanogenic populations during anaerobic hydrocarbon degradation in oil reservoirs.
ABSTRACT
The methanogenic degradation of hydrocarbons plays an important role in hydrocarbon-contaminated environments in the absence of an external electron acceptor. Members of Syntrophaceae sublineages were previously reported to be responsible for syntrophic alkane degradation. However, limited information is currently available on their physiological capabilities in nature because it is very challenging to cultivate these as-yet uncultured microbes. We herein performed metagenomic sequencing of the methanogenic hexadecane-degrading culture M82 and recovered a nearly complete genome (2.75 Mb, estimated completeness ≥97%) belonging to Syntrophaceae sublineage II. The assembly genome was tentatively named "Candidatus Smithella cisternae strain M82_1". Genes encoding alkylsuccinate synthase for alkane activation were identified, suggesting that this organism is capable of oxidizing alkanes through fumarate addition. This capability was further supported by the detection of methyl pentadecyl succinic acid and methyl tetradecyl succinic acid in cultures amended with hexadecane and pentadecane, respectively. Genes encoding enzymes for the ß-oxidation of long-chain fatty acids and butyrate were also identified. The electron transfer flavoprotein/DUF224 complex is presumed to link electron flow from acyl-CoA dehydrogenase to a membrane hydrogenase or formate dehydrogenase. Although no indications of Rnf complexes were detected, genes encoding electron-confurcating hydrogenase and formate dehydrogenase were proposed to couple the thermodynamically favorable oxidation of ferredoxin to generate H2 and formate from NADH. Strain M82_1 synthesized ATP from acetyl-CoA by substrate-level phosphorylation or F1F0-ATP synthases. These results provide an insight into the potential metabolic traits and ecophysiological roles of the syntrophic alkane degrader Syntrophaceae.
