Genus Methanothermobacter Research Articles

Comparison of hyper-thermophilic–mesophilic two-stage with single-stage mesophilic anaerobic digestion of waste activated sludge: Process performance and microbial community analysis

The conventional mesophilic digestion of waste activated sludge (WAS) is challenged by the updated wastewater treatment process. A hyper-thermophilic (70°C)–mesophilic (35°C) temperature-phased two-stage systems without and with recirculation were employed to upgrade the single-stage mesophilic digestion of waste activated sludge in this study. The solid reduction increased by more than 10% in the two-stage systems. Similar methane production was observed in the single-stage system and two-stage systems, with a methane yield of 0.24L/g VS added. In the case of two-stage system without recirculation, hyper-thermophilic stage played an important role in COD solubilization, with 34.4% of the WAS solubilized in that stage, while mesophilic stage contributed more in methane production, with 36.2% of COD in the WAS produced as methane in the mesophilic stage. Firmicutes became the most dominant bacteria phylum in the mesophilic stages of two-stage systems as compared to the dominant phylum Proteobacteria in single-stage system, with the genus Methanothermobacter cardinal in the hyper-thermophilic stage. The recirculation in the two-stage system increased the microbial community diversity in the hyper-thermophilic stage, which was one factor to result in improved methane production in the hyper-thermophilic stage.

Roles of thermophilic thiosulfate-reducing bacteria and methanogenic archaea in the biocorrosion of oil pipelines.

Thermophilic sulfide-producing microorganisms from an oil pipeline network were enumerated with different sulfur oxyanions as electron acceptors at 55°C. Most-probable number (MPN) analysis showed that thiosulfate-reducing bacteria were the most numerous sulfidogenic microorganisms in pipeline inspection gauge (PIG) scrapings. Thiosulfate-reducing and methanogenic enrichments were obtained from the MPN cultures that were able to use yeast extract as the electron donor. Molecular analysis revealed that both enrichments harbored the same dominant bacterium, which belonged to the genus Anaerobaculum. The dominant archaeon in the methanogenic enrichment was affiliated with the genus Methanothermobacter. With yeast extract as the electron donor, the general corrosion rate by the thiosulfate-reducing enrichment (8.43 ± 1.40 milli-inch per year, abbreviated as mpy) was about 5.5 times greater than the abiotic control (1.49 ± 0.15 mpy), while the comparable measures for the methanogenic culture were 2.03 ± 0.49 mpy and 0.62 ± 0.07 mpy, respectively. Total iron analysis in the cultures largely accounted for the mass loss of iron measured in the weight loss determinations. Profilometry analysis of polished steel coupons incubated in the presence of the thiosulfate-reducing enrichment revealed 59 pits over an area of 71.16 mm2, while only 6 pits were evident in the corresponding methanogenic incubations. The results show the importance of thiosulfate-utilizing, sulfide-producing fermentative bacteria such as Anaerobaculum sp. in the corrosion of carbon steel, but also suggest that Anaerobaculum sp. are of far less concern when growing syntrophically with methanogens.

Evaluation of microbial community composition in thermophilic methane-producing incubation of production water from a high-temperature oil reservoir

Investigation of petroleum microbes is fundamental for the development and utilization of oil reservoirs’ microbial resources, and also provides great opportunities for research and development of bio-energy. Production water from a high-temperature oil reservoir was incubated anaerobically at 55°C for more than 400 days without amendment of any nutrients. Over the time of incubation, about 1.6 mmol of methane and up to 107 μ mol of hydrogen (H2) were detected in the headspace. Methane formation indicated that methanogenesis was likely the predominant process in spite of the presence of 23.4 mM in the production water. Microbial community composition of the incubation was characterized by means of 16S rRNA gene clone libraries construction. Bacterial composition changed from Pseudomonales as the dominant population initially to Hydrogenophilales-related microorganisms affiliated to Petrobacter spp. closely. After 400 days of incubation, other bacterial members detected were related to Anareolineales, β-, γ-, and δ-Proteobacteria. The archaeal composition of the original production water was essentially composed of obligate acetoclastic methanogens of the genus Methanosaeta, but the incubation was predominantly composed of CO2-reducing methanogens of the genus Methanothermobacter and Crenarchaeotes-related microorganisms. Our results suggest that methanogenesis could be more active than expected in oil reservoir environments and methane formation from CO2-reduction played a significant role in the methanogenic community. This conclusion is consistent with the predominant role played by H2-oxidizing methanogens in the methanogenic conversion of organic matter in high-temperature petroleum reservoirs.

Molecular analysis of the microbial community structures in water-flooding petroleum reservoirs with different temperatures

Abstract. Analyses of microbial communities from six water-flooding petroleum reservoirs at temperatures from 21 to 63 °C by 16S rRNA gene clone libraries indicates the presence of physiologically diverse and temperature-dependent microorganisms in these subterrestrial ecosystems. In samples originating from high-temperature petroleum reservoirs, most of the archaeal sequences belong to thermophiles affiliated with members of the genera Thermococcus, Methanothermobacter and the order Thermoplasmatales, whereas bacterial sequences predominantly belong to the phyla Firmicutes, Thermotogae and Thermodesulfobacteria. In contrast to high-temperature petroleum reservoirs, microorganisms belonging to the Proteobacteria, Methanobacteriales and Methanomicrobiales were the most encountered in samples collected from low-temperature petroleum reservoirs. Canonical correspondence analysis (CCA) revealed that temperature, mineralization, ionic type as well as volatile fatty acids showed correlation with the microbial community structures, in particular members of the Firmicutes and the genus Methanothermobacter showed positive correlation with temperature and the concentration of acetate. Overall, these data indicate the large occurrence of hydrogenotrophic methanogens in petroleum reservoirs and imply that acetate metabolism via syntrophic oxidation may represent the main methanogenic pathway in high-temperature petroleum reservoirs.

Involvement of thermophilic archaea in the biocorrosion of oil pipelines

Two thermophilic archaea, strain PK and strain MG, were isolated from a culture enriched at 80°C from the inner surface material of a hot oil pipeline. Strain PK could ferment complex organic nitrogen sources (e.g. yeast extract, peptone, tryptone) and was able to reduce elemental sulfur (S°), Fe(3+) and Mn(4+) . Phylogenetic analysis revealed that the organism belonged to the order Thermococcales. Incubations of this strain with elemental iron (Fe°) resulted in the abiotic formation of ferrous iron and the accumulation of volatile fatty acids during yeast extract fermentation. The other isolate, strain MG, was a H(2) :CO(2) -utilizing methanogen, phylogenetically affiliated with the genus Methanothermobacter family. Co-cultures of the strains grew as aggregates that produced CH(4) without exogenous H(2) amendment. The co-culture produced the same suite but greater concentrations of fatty acids from yeast extract than did strain PK alone. Thus, the physiological characteristics of organisms both alone and in combination could conceivably contribute to pipeline corrosion. The Thermococcus strain PK could reduce elemental sulfur to sulfide, produce fatty acids and reduce ferric iron. The hydrogenotrophic methanogen strain MG enhanced fatty acid production by fermentative organisms but could not couple the dissolution Fe° with the consumption of water-derived H(2) like other methanogens.

Thermophilic two-stage dry anaerobic digestion of model garbage with ammonia stripping

To avoid the inhibition of methane production by ammonia that occurs during the degradation of garbage, anaerobic digestion with prior ammonia production and subsequent stripping was investigated. In the ammonia production phase, the maximum ammonia concentration was approximately 2800 mg N/kg of total wet sludge in the range of 4 days of sludge retention time, indicating that only 43% of total nitrogen in the model garbage was converted to ammonia. The model garbage from which ammonia was produced and stripped was subjected to semi-continuous thermophilic dry anaerobic digestion over 180 days. The gas yield was in the range of 0.68 to 0.75 Nm(3)/kg volatile solid, and it decreased with the decrease of the sludge retention time. The ammonia-nitrogen concentration in the sludge was kept below 3000 mg N/kg total wet sludge. Microbial community structure analysis revealed that the phylum Firmicutes dominated in the ammonia production, but the community structure changed at different sludge retention times. In dry anaerobic digestion, the dominant bacteria shifted from the phylum Thermotogae to Firmicutes. The dominant archaeon was the genus Methanothermobacter, but the ratio of Methanosarcina increased during the process of dry anaerobic digestion.

Evidence for syntrophic acetate oxidation coupled to hydrogenotrophic methanogenesis in the high‐temperature petroleum reservoir of Yabase oil field (Japan)

The methanogenic communities and pathways in a high-temperature petroleum reservoir were investigated through incubations of the production water and crude oil, combined with radiotracer experiments and molecular biological analyses. The incubations were conducted without any substrate amendment and under high-temperature and pressurized conditions that mimicked the in situ environment (55°C, 5 MPa). Changes in methane and acetate concentrations during the incubations indicated stoichiometric production of methane from acetate. Rates of hydrogenotrophic methanogenesis measured using [(14)C]-bicarbonate were 42-68 times those of acetoclastic methanogenesis measured using [2-(14) C]-acetate, implying the dominance of methane production by syntrophic acetate oxidation coupled to hydrogenotrophic methanogenesis in the environment. 16S rRNA gene sequence analyses of the incubated production water showed bacterial communities dominated by the genus Thermacetogenium, known as a thermophilic syntrophic acetate-oxidizing bacterium, and archaeal communities dominated by thermophilic hydrogenotrophic methanogens belonging to the genus Methanothermobacter. Furthermore, group-specific real-time PCR assays revealed that 16S rRNA gene copy numbers of the hydrogenotrophic methanogens affiliated with the order Methanobacteriales were almost identical to those of archaeal 16S rRNA genes. This study demonstrates that syntrophic acetate oxidation is the main methanogenic pathway in a high-temperature petroleum reservoir.

Recombination between elongation factor 1alpha genes from distantly related archaeal lineages.

Homologous recombination (HR) and lateral gene transfer are major processes in genome evolution. The combination of the two processes, HR between genes in different species, has been documented but is thought to be restricted to very similar sequences in relatively closely related organisms. Here we report two cases of interspecific HR in the gene encoding the core translational protein translation elongation factor 1alpha (EF-1alpha) between distantly related archaeal groups. Maximum-likelihood sliding window analyses indicate that a fragment of the EF-1alpha gene from the archaeal lineage represented by Methanopyrus kandleri was recombined into the orthologous gene in a common ancestor of the Thermococcales. A second recombination event appears to have occurred between the EF-1alpha gene of the genus Methanothermobacter and its ortholog in a common ancestor of the Methanosarcinales, a distantly related euryarchaeal lineage. These findings suggest that HR occurs across a much larger evolutionary distance than generally accepted and affects highly conserved essential "informational" genes. Although difficult to detect by standard whole-gene phylogenetic analyses, interspecific HR in highly conserved genes may occur at an appreciable frequency, potentially confounding deep phylogenetic inference and hypothesis testing.

Phylogenetic analysis of 18 thermophilic Methanobacterium isolates supports the proposals to create a new genus, Methanothermobacter gen. nov., and to reclassify several isolates in three species, Methanothermobacter thermautotrophicus comb. nov., Methanothermobacter wolfeii comb. nov., and Methanothermobacter marburgensis sp. nov.

Using a combination of 16S rRNA analysis and antigenic fingerprinting consisting of new and published data, the phylogenetic position of 18 thermophilic isolates currently classified as Methanobacterium species was reinvestigated. The results were verified by independent methods, including, where applicable, plasmid and phage typing. Comparative analysis of 16S rRNA data for 30 strains belonging to the order Methanobacteriales strongly suggested that mesophilic and thermophilic Methanobacterium isolates are distantly related and should be assigned to separate genera. For the thermophilic strains the genus Methanothermobacter was initially proposed by Boone, Whitman and Rouvière. Furthermore, the results support a reclassification of 15 isolates in three species within the proposed genus: (i) Methanothermobacter thermautotrophicus comb. nov., containing eight isolates, six of which are able to utilize formate (type strain deltaHT); (ii) Methanothermobacter wolfeii comb. nov., containing four formate-utilizing isolates (type strain DSM 2970T); (iii) Methanothermobacter marburgensis sp. nov., containing three obligately autotrophic isolates (type strain MarburgT). Of the nine isolates formerly referred to as Methanobacterium thermoformicicum, six were reclassified as Methanothermobacter thermautotrophicus and three as Methanothermobacter wolfeii.