Effect of nickel, cobalt, and iron on methanogenesis from methanol and cometabolic conversion of 1,2-dichloroethene by Methanosarcina barkeri.

Methanogens are responsible for the last step in anaerobic digestion (AD), in which methane (a biofuel) is produced. Some methanogens can cometabolize chlorinated pollutants, contributing for their removal during AD. Methanogenic cofactors involved in cometabolic reductive dechlorination, such as F430 and cobalamin, contain metal ions (nickel, cobalt, iron) in their structure. We hypothesized that the supplementation of trace metals could improve methane production and the cometabolic dechlorination of 1,2-dichloroethene (DCE) by pure cultures of Methanosarcina barkeri. Nickel, cobalt, and iron were added to cultures of M. barkeri growing on methanol and methanol plus DCE. Metal amendment improved DCE dechlorination to vinyl chloride (VC): assays with 20 µM of Fe3+ showed the highest final concentration of VC (5× higher than in controls without Fe3+ ), but also in assays with 5.5 µM of Co2+ and 5 µM of Ni2+ VC formation was improved (3.5-4× higher than in controls without the respective metals). Dosing of metals could be useful to improve anaerobic removal of chlorinated compounds, and more importantly decrease the detrimental effect of DCE on methane production in anaerobic digesters.

Secondary Mineralization of Ferrihydrite Affects Microbial Methanogenesis in Geobacter-Methanosarcina Cocultures.

UNLABELLED: The transformation of ferrihydrite to stable iron oxides over time has important consequences for biogeochemical cycling of many metals and nutrients. The response of methanogenic activity to the presence of iron oxides depends on the type of iron mineral, but the effects of changes in iron mineralogy on methanogenesis have not been characterized. To address these issues, we constructed methanogenic cocultures of Geobacter and Methanosarcina strains with different ferrihydrite mineralization pathways. In this system, secondary mineralization products from ferrihydrite are regulated by the presence or absence of phosphate. In cultures producing magnetite as the secondary mineralization product, the rates of methanogenesis from acetate and ethanol increased by 30.2% and 135.3%, respectively, compared with a control lacking ferrihydrite. Biogenic magnetite was proposed to promote direct interspecies electron transfer between Geobacter and Methanosarcina in a manner similar to that of c-type cytochrome and thus facilitate methanogenesis. Vivianite biomineralization from ferrihydrite in the presence of phosphate did not significantly influence the methanogenesis processes. The correlation between magnetite occurrence and facilitated methanogenesis was supported by increased rates of methane production from acetate and ethanol with magnetite supplementation in the defined cocultures. Our data provide a new perspective on the important role of iron biomineralization in biogeochemical cycling of carbon in diverse anaerobic environments. IMPORTANCE: It has been found that microbial methanogenesis is affected by the presence of iron minerals, and their influences on methanogenesis are associated with the mineralogical properties of the iron minerals. However, how changes in iron mineralogy affect microbial methanogenesis has not been characterized. To address this issue, we constructed methanogenic cocultures of Geobacter and Methanosarcina strains with different ferrihydrite mineralization pathways. The experimental results led to two contributions, i.e., (i) the transformation of iron minerals might exert an important influence on methanogenesis under anaerobic conditions and (ii) both biogenic and chemical magnetite can accelerate syntrophic ethanol oxidization between Geobacter metallireducens and Methanosarcina barkeri This study sheds new light on the important role of iron biomineralization in the biogeochemical cycling of carbon in diverse anaerobic environments, particularly in iron-rich natural and agricultural wetland soils.

Genetic, Genomic, and Transcriptomic Studies of Pyruvate Metabolism in Methanosarcina barkeri Fusaro.

UNLABELLED: Pyruvate, a central intermediate in the carbon fixation pathway of methanogenic archaea, is rarely used as an energy source by these organisms. The sole exception to this rule is a genetically uncharacterized Methanosarcina barkeri mutant capable of using pyruvate as a sole energy and carbon source (the Pyr(+) phenotype). Here, we provide evidence that suggests that the Pyr(+) mutant is able to metabolize pyruvate by overexpressing pyruvate ferredoxin oxidoreductase (por) and mutating genes involved in central carbon metabolism. Genomic analysis showed that the Pyr(+) strain has two mutations localized to Mbar_A1588, the biotin protein ligase subunit of the pyruvate carboxylase (pyc) operon, and Mbar_A2165, a putative transcriptional regulator. Mutants expressing the Mbar_A1588 mutation showed no growth defect compared to the wild type (WT), yet the strains lacked pyc activity. Recreation of the Mbar_A2165 mutation resulted in a 2-fold increase of Por activity and gene expression, suggesting a role in por transcriptional regulation. Further transcriptomic analysis revealed that Pyr(+) strains also overexpress the gene encoding phosphoenolpyruvate carboxylase, indicating the presence of a previously uncharacterized route for synthesizing oxaloacetate in M. barkeri and explaining the unimpaired growth in the absence of Pyc. Surprisingly, stringent repression of the por operon was lethal, even when the media were supplemented with pyruvate and/or Casamino Acids, suggesting that por plays an unidentified essential function in M. barkeri. IMPORTANCE: The work presented here reveals a complex interaction between anabolic and catabolic pathways involving pyruvate metabolism in Methanosarcina barkeri Fusaro. Among the unexpected findings were an essential role for the enzyme pyruvate-ferredoxin oxidoreductase and an alternate pathway for synthesis of oxaloacetate. These results clarify the mechanism of methanogenic catabolism of pyruvate and expand our understanding of carbon assimilation in methanogens.