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Methane (CH4) emissions in mangrove ecosystems may complicate the ecosystem's potential carbon offset for climate change mitigation. Microbial processes and the mass balance of CH4 in mangrove sediment are responsible for the emissions from the ecosystems. This is the follow up of our previous research which found the super saturation of CH4 in the pore water of mangrove sediment compared to atmospheric CH4 and the lack of a correlation between pore water sulphate and CH4 concentrations. This study is going to investigate methane production pathways in the sediment of overwashed mangrove forests. Two approaches were used to study methanogens here: (1) the spread plate count method and the quantitative polymerase chain reaction (qPCR) method, and (2) laboratory experiments with additional methanogenic substrates (methanol, acetate, and hydrogen) to determine which substrates are more conducive to methane production. According to the qPCR method, methanogen abundance ranged from 72 to 6 × 105 CFU g−1 sediment, while SRB abundance ranged from 2 × 102 to 2 × 105 CFU g−1 sediment. According to the plate count method, the abundance of methylotrophic methanogens (the only group of methanogens capable of competing with SRBs) ranged from 8.3 × 102 to 5.1 × 104 CFU g−1, which is higher than the abundance of the other group of methanogens (0 to 7.7 × 102 CFU g−1). The addition of methanol to the sediment slurry, a substrate for methylotropic methanogens, resulted in a massive production of CH4 (up to 9 × 104 ppm) and intriguingly the control treatments with autoclaving did not kill methanogens. These findings suggested that mangrove ecosystems in the marine environment provide favourable conditions for methanogens and further characterisation of the methanogen involved in the process is required. As a result, future research in this ecosystem should include methane production in carbon offset calculations, particularly due to methylotropic methanogenesis. [ABSTRACT FROM AUTHOR]

Microbial consortia under anaerobic conditions are involved in oxidizing organic matter in the sludge to produce methane gas. However, in developing countries like Kenya, these microbes have not been fully identified to target them for the efficient harnessing of biofuel. This study collected wet sludge from two anaerobic digestion lagoons 1 and 2 that were operational during sampling at Kangemi Sewage Treatment Plant, in Nyeri County, Kenya. DNA was extracted from samples using commercially available ZymoBIOMICS™ DNA Miniprep Kit and sequenced using Shotgun metagenomics. Samples were analyzed using MG-RAST software (Project ID: mgp100988), which allowed for identifying microorganisms directly involved in various stages of methanogenesis pathways. The study found hydrogenotrophic methanogens, such as Methanospirillum (32%), Methanobacterium (27%), Methanobrevibacter (27%), and Methanosarcina (32%), being predominant in the lagoon communities, whereas acetoclastic microorganisms such as the Methanoregula (22%) and the acetate oxidazing bacteria such as Clostridia (68%) were the key microbes for that pathway in the sewage digester sludge. Furthermore, Methanothermobacter (18%), Methanosarcina (21%), Methanosaeta (15%), and Methanospirillum (13%) carried out the methylotrophic pathway. In contrast, Methanosarcina (23%),Methanoregula (14%), methanosaeta (13%), and methnanoprevibacter (13%) seemed to play an important role in the final step of methane release. This study concluded that the sludge produced from the Nyeri-Kangemi WWTP harbors microbes with significant potential for biogas production. The study recommends a pilot study to investigate the efficiency of the identified microbes for biogas production.

Hydrogenotrophic methanogens play a key role in methane (CH4) production during various wastes' anaerobic digestion (AD). The metabolism of hydrogenotrophic methanogens involves Wolfe-cycle to reduce carbon dioxide (CO2) to CH4 using hydrogen as feed. Hydrogenotrophic methanogenesis is a potential process for up-gradation of biogas and renewable power intermittency. This review focuses on enhancing hydrogenotrophic methanogenesis for the up-gradation of biogas and, more importantly, a hybrid system that couples both in-situ and ex-situ approaches in one unit. Upgradation of biogas by hydrogenotrophic methanogenesis is advantageous for achieving a higher concentration of CH4 and removing trace elements and CO2 from biogas. This approach helps to decrease the cost of upgrading biogas to natural gas quality. Using microbial electrolysis cells coupled with AD could be a potential alternative to increase the CH4 production yield and remove CO2 from biogas. The exploitation of the hydrogen gas and integration of the available technologies on a large scale could be explored in future work and help in realizing the production of clean CH4 in an industrial set-up. This review summarizes the metabolic background of hydrogenotrophic methanogens and their efficient utilization towards methane production. Limited works are available in public domain covering this aspect and thus a review article is important for the readers to update their understanding and research work in light of recent findings. [ABSTRACT FROM AUTHOR]

Marine benthic environments may be shaped by anthropogenic and other localized events, leading to changes in microbial community composition evident decades after a disturbance. Marine sediments in particular harbor exceptional taxonomic diversity and can shed light on distinctive evolutionary strategies. Genetic code expansion is a strategy that increases the structural and functional diversity of proteins in cells, by repurposing stop codons to encode non-canonical amino acids: pyrrolysine (Pyl) and selenocysteine (Sec). Here, we report both a study of the microbiome at a deep sea industrial waste dumpsite and an unanticipated discovery of codon reassignment in its most abundant member, with potential ramifications for interpreting microbial interactions with ocean-dumped wastes. The genomes of abundant Deltaproteobacteria from the sediments of a deep-ocean chemical waste dump site have undergone genetic code expansion. Pyl and Sec in these organisms appear to augment trimethylamine (TMA) and one-carbon metabolism, representing an increased metabolic versatility. The inferred metabolism of these sulfate-reducing bacteria places them in competition with methylotrophic methanogens for TMA, a contention further supported by earlier isotope tracer studies and reanalysis of metatranscriptomic studies. A survey of genomic data further reveals a broad geographic distribution of a niche group of similarly specialized Deltaproteobacteria, including at sulfidic sites in the Atlantic Ocean, Gulf of Mexico, Guaymas Basin, and North Sea, as well as in terrestrial and estuarine environments. These findings reveal an important biogeochemical role for specialized Deltaproteobacteria at the interface of the carbon, nitrogen, selenium, and sulfur cycles, with their niche adaptation and ecological success potentially augmented by genetic code expansion.

Marine benthic environments may be shaped by anthropogenic and other localized events, leading to changes in microbial community composition evident decades after a disturbance. Marine sediments in particular harbor exceptional taxonomic diversity and can shed light on distinctive evolutionary strategies. Genetic code expansion is a strategy that increases the structural and functional diversity of proteins in cells, by repurposing stop codons to encode non-canonical amino acids: pyrrolysine (Pyl) and selenocysteine (Sec). Here, we report both a study of the microbiome at a deep sea industrial waste dumpsite and an unanticipated discovery of codon reassignment in its most abundant member, with potential ramifications for interpreting microbial interactions with ocean-dumped wastes. The genomes of abundant Deltaproteobacteria from the sediments of a deep-ocean chemical waste dump site have undergone genetic code expansion. Pyl and Sec in these organisms appear to augment trimethylamine (TMA) and one-carbon metabolism, representing an increased metabolic versatility. The inferred metabolism of these sulfate-reducing bacteria places them in competition with methylotrophic methanogens for TMA, a contention further supported by earlier isotope tracer studies and reanalysis of metatranscriptomic studies. A survey of genomic data further reveals a broad geographic distribution of a niche group of similarly specialized Deltaproteobacteria, including at sulfidic sites in the Atlantic Ocean, Gulf of Mexico, Guaymas Basin, and North Sea, as well as in terrestrial and estuarine environments. These findings reveal an important biogeochemical role for specialized Deltaproteobacteria at the interface of the carbon, nitrogen, selenium, and sulfur cycles, with their niche adaptation and ecological success potentially augmented by genetic code expansion. [ABSTRACT FROM AUTHOR]

Emissions of microbially produced methane (CH 4 ) from lake sediments are a major source of this potent greenhouse gas to the atmosphere. The rates of CH 4 production and emission are believed to be influenced by electron acceptor distributions and organic carbon contents, which in turn are affected by anthropogenic inputs of nutrients leading to eutrophication. Here, we investigate how eutrophication influences the abundance and community structure of CH 4 producing Archaea and methanogenesis pathways across time-resolved sedimentary records of five Swiss lakes with well-characterized trophic histories. Despite higher CH 4 concentrations which suggest higher methanogenic activity in sediments of eutrophic lakes, abundances of methanogens were highest in oligotrophic lake sediments. Moreover, while the methanogenic community composition differed significantly at the lowest taxonomic levels (OTU), depending on whether sediment layers had been deposited under oligotrophic or eutrophic conditions, it showed no clear trend in relation to in situ distributions of electron acceptors. Remarkably, even though methanogenesis from CO 2 -reduction was the dominant pathway in all sediments based on carbon isotope fractionation values, taxonomic identities, and genomes of resident methanogens, CO 2 -reduction with hydrogen (H 2 ) was thermodynamically unfavorable based on measured reactant and product concentrations. Instead, strong correlations between genomic abundances of CO 2 -reducing methanogens and anaerobic bacteria with potential for extracellular electron transfer suggest that methanogenic CO 2 -reduction in lake sediments is largely powered by direct electron transfer from syntrophic bacteria without involvement of H 2 as an electron shuttle., Competing Interests: None declared., (© The Author(s) 2024. Published by Oxford University Press on behalf of the International Society for Microbial Ecology.)

The aim is to conduct the biodiagnosis of soil sampled in the area of the underground storage of natural gas (UGS) (Stepnoye village, Saratov region) with the use of microbiological analysis.Methods. In the course of the work, using Koch’s solid medium method were estimated the following: the total number of heterotrophic microorganisms on meat peptone agar, amount of methylotrophic, hydrocarbon oxidizing, ironoxidizing and sulfur-oxidizing bacteria in the respective selective mediums; as well as the hydrogen index, oxidation-reduction potential and magnetic susceptibility of the soil.Results. Microbiological analysis of the soil in the Stepnovsky UGS area has shown changes in soil biocenoses. A reduced content of heterotrophic microorganisms was detected in comparison with background samples. In a number of samples, we also discovered an increased content of hydrocarbon-oxidizing and methylotrophic microorganisms, including obligate ones, which suggest the methane entering the upper layers of the soil. An increased content of sulfur-oxidizing bacteria was also detected; the presence of iron-oxidizing microorganisms in the soil was shown.Conclusions. The revealed features in the content of indicator microorganisms in the soil sampled in the area of the underground storage of natural gas, indicating the development of specialized soil microorganisms, confirmed the presence of contaminants in the soil which are hydrocarbons and sulfur compounds. The use of microbiological indicator values seems promising for environmental monitoring of soils in the areas of gas storage and early detection of methane leakage.

Although it is generally assumed that CH 4 production in peatlands is dominated by the acetoclastic and hydrogenotrophic methanogenic pathways, we found evidence that methylated substrates (including methanol, methylamines, and dimethylsulfide) support methanogenesis in these organic-rich environments. The role of methylotrophic methanogenesis was investigated in three northern Minnesota peatlands by amending surface soils with 13 C-labeled substrates. Labeled methanol was converted to enriched δ 13 C-CH 4 at all three sites. Additionally, dimethylsulfide and monomethylamine were processed through methanogenic pathways, although this was not evident at every site. The addition of 13 C labeled methanol and monomethylamine also resulted in the enrichment of δ 13 C-CO 2 , further suggesting that either methanogens or non-methanogenic microbes were processing methylated substrates. Results demonstrate the potential for methylotrophic methanogenesis in northern peatlands and suggest that further studies are warranted to quantify the amount of CH 4 produced by this process in comparison to traditional pathways of methanogenesis. [ABSTRACT FROM AUTHOR]

Conventional cultivation method relies on the use of nutrient-rich medium, which might limit the exploration of oligotrophic bacteria from soil ecosystems. In the present study attempts were made to isolate oligotrophic bacteria from non-rhizospheric samples collected from mountain roadside, beach, bay and wetland using 1000-fold diluted nutrient agar. Phylogenetic analysis and biogeographic studies of these isolates were performed based on their 16S rDNA sequences. Besides, tests for the utilization of methanol or agar as their carbon source (methylotrophic or agarolytic), light as their energy source (photoheterotrophic), fixation of nitrogen or solubilization of tricalcium phosphate were conducted. A total of 74 isolates with facultatively oligotrophic behavior were obtained, since they can proliferate on both 1000-fold diluted and conventional nutrient agar. 16S rDNA sequence analyses assigned them mainly to class Actinobacteria and α-Proteobacteria. All these isolates belonged to 24 genera which encompassed 35 species, demonstrating that these facultatively oligotrophic bacteria occupied a wide range of bacterial lineages. Fifty-seven out of 74 isolates formed colonies on methanol-containing agar, while only 21 showed better growth in methanol-containing medium. Nine isolates were verified to utilize agar instead of methanol for growth. A total of 4 isolates showed photoheterotrophic trait and the pufLM gene was successfully PCR amplified and sequenced. The methylotrophic or photoheterotrophic behaviors might provide advantages for them to inhabit oligotrophic environments. Members affiliated with many genera were first demonstrated to fix nitrogen or solubilize phosphate, and isolates with profound activities have potentials to be developed as bioinoculants used to promote plant growth. In addition, the use of diluted nutrient agar helped to explore several unrecognized species from non-rhizospheric soils. All these isolates may provide opportunities for studying their oligotrophic growth, metabolism and interaction with plants in a near future. [ABSTRACT FROM AUTHOR]

Advances in synthetic genomics are now well underway in yeasts due to the low cost of synthetic DNA. These new capabilities also bring greater need for quantitating the presence, loss and rearrangement of loci within synthetic yeast genomes. Methods for achieving this will ideally; i) be robust to industrial settings, ii) adhere to a global standard and iii) be sufficiently rapid to enable at-line monitoring during cell growth. The methylotrophic yeast Pichia pastoris ( P. pastoris ) is increasingly used for industrial production of biotherapeutic proteins so we sought to answer the following questions for this particular yeast species. Is time-consuming DNA purification necessary to obtain accurate end-point polymerase chain reaction (e-pPCR) and quantitative PCR (qPCR) data? Can the novel linear regression of efficiency qPCR method (LRE qPCR), which has properties desirable in a synthetic biology standard, match the accuracy of conventional qPCR? Does cell cultivation scale influence PCR performance? To answer these questions we performed e-pPCR and qPCR in the presence and absence of cellular material disrupted by a mild 30s sonication procedure. The e-pPCR limit of detection (LOD) for a genomic target locus was 50 pg (4.91 × 10 3 copies) of purified genomic DNA (gDNA) but the presence of cellular material reduced this sensitivity sixfold to 300 pg gDNA (2.95 × 10 4 copies). LRE qPCR matched the accuracy of a conventional standard curve qPCR method. The presence of material from bioreactor cultivation of up to OD 600 = 80 did not significantly compromise the accuracy of LRE qPCR. We conclude that a simple and rapid cell disruption step is sufficient to render P. pastoris samples of up to OD 600 = 80 amenable to analysis using LRE qPCR which we propose as a synthetic biology standard. [ABSTRACT FROM AUTHOR]

The methylotrophic thermo tolerant actinomycete strain 381, was isolated from a compost sample, assimilated methanol during growth. Methanol dehydrogenase could not be detected in cell extracts using dye-linked assays in the absence of NAD+. Dichlorophenol-indophenol-linked methanol oxidation was detected in broken cell suspensions in the presence of NAD+ and the absence of cyanide. Taxonomic studies showed that actinomycete strain 381 was distinct from Nocardia species 239, which has also been reported to contain an NAD+– dependent methanol dehydrogenase. The organism can also utilizes pyruvate, casamino acid, glucose, glycerol and many other carbon sources as described before. Preliminary enzyme studies suggested that the cell extracts contained phosphoribulokinase and (RUBP) ribulose bisphosphate carboxylase (the two key enzymes for Calvin cycle) which–compounds indicating that actinomycete strain 381, assimilates C1by the means of Serine pathway.

Methylotrophic bacteria which are known to utilize C1 compounds including methane. Research during past few decades increased the interest in finding out novel genera of methane degrading bacteria to efficiently utilize methane to decrease global warming effect. Moreover, evaluation of certain known plant growth promoting strains for their methane degrading potential may open up a new direction for multiple utility of such cultures. In this study, efficient methylotrophic cultures were isolated from wetland paddy fields of Gujarat. From the overall morphological, biochemical and molecular characterization studies, the isolates were identified and designated as Bacillus aerius AAU M 8; Rhizobium sp. AAU M 10; B. subtilis AAU M 14; Paenibacillus illinoisensis AAU M 17 and B. megaterium AAU M 29. Gene specific PCR analysis of the isolates, P. illinoisensis, B. aerius, Rhizobium sp. and B. subtilis showed presence of pmoA gene encoding α subunit particulate methane monooxygenase cluster. B. megaterium, P. illinoisensis, Rhizobium sp. and Methylobacterium extrorquens showed presence of mmoX gene encoding α subunit of the hydroxylase component of the soluble methane monooxygenase cluster. P. illinoisensis and Rhizobium sp. showed presence mxaF gene encoding α subunit region of methanol dehydrogenase gene cluster showing that both isolates are efficient utilizers of methane. To the best of our knowledge, this is the first time report showing presence of methane degradation enzymes and genes within the known PGPB group of organisms from wet land paddy agro-ecosystem, which is considered as one of the leading methane producer. [ABSTRACT FROM AUTHOR]

Desulfotomaculum kuznetsovii is a moderately thermophilic member of the polyphyletic spore-forming genus Desulfotomaculum in the family Peptococcaceae. This species is of interest because it originates from deep subsurface thermal mineral water at a depth of about 3,000 m. D. kuznetsovii is a rather versatile bacterium as it can grow with a large variety of organic substrates, including short-chain and long-chain fatty acids, which are degraded completely to carbon dioxide coupled to the reduction of sulfate. It can grow methylotrophically with methanol and sulfate and autotrophically with H + CO and sulfate. For growth it does not require any vitamins. Here, we describe the features of D. kuznetsovii together with the genome sequence and annotation. The chromosome has 3,601,386 bp organized in one contig. A total of 3,567 candidate protein-encoding genes and 58 RNA genes were identified. Genes of the acetyl-CoA pathway, possibly involved in heterotrophic growth with acetate and methanol, and in CO fixation during autotrophic growth are present. Genomic comparison revealed that D. kuznetsovii shows a high similarity with Pelotomaculum thermopropionicum. Genes involved in propionate metabolism of these two strains show a strong similarity. However, main differences are found in genes involved in the electron acceptor metabolism. [ABSTRACT FROM AUTHOR]

A novel methylotrophic methanogen Methanococcoides orientis sp. nov. was isolated from East China Sea sediment. Type strain LMO-1 T of Methanococcoides orientis sp. nov. was irregular 1-2 µm cocci without flagella. Strain LMO-1 T could utilize a variety of methylated compounds including methanol, methylamine, dimethylamine and trimethylamine for growth and methanogenesis, while H 2 /CO 2 or acetate could not be used for growth or methanogenesis. Optimum growth temperature was 30-35 °C, optimum pH range for growth was 7.0-7.5, while the optimum salinity spectrum for growth was 1.0%-5.0% NaCl. Based on 16S rRNA gene similarity, strain LMO-1 T belongs to Methanococcoides , with the highest sequence similarity to Methanococcoides methylutens DSM 2657 T (99.8 %), Methanococcoides vulcani SLH33 T (99.4 %), followed by Methanococcoides alaskense AK-5 T (98.1 %), Methanococcoides burtonii DSM 6242 T (98.0 %). Digital DNA-DNA hybridization also showed highest similarity with Methanococcoides methylutens DSM 2657 T , with the value of 58.4 %. The average nucleotide identity between strain LMO-1 T and Methanococcoides methylutens DSM 2657 T was 94.06 %. In summary, LMO-1 T represents a novel species of the genus Methaococcoides , for which the name Methanococcoides orientis sp. nov. is proposed. The type strain is LMO-1 T (=MCCC 4K00106 T =JCM 39195 T ).

The article reports the preliminary results obtained in a pilot plant that treats a spent tetramethylammonium hydroxide (TMAH) solution, a typical chemical compound used in the microelectronics industry and massively present in wastewaters. The plant is based on three aerobic bioreactors in series, fed by a real TMAH solution effluent from the semiconductor industry. The plant ran for 14 days in continuous operating mode with a flow rate of 5 L/h; then, the plant switched to batch mode. The microbial population was analyzed to determine if microbial population changes occurred. The results indicated that TMAH degradation only occurred during the batch operation mode. The TMAH reduction was about 80% in the first reactor and complete (100%) in the second and third bioreactors. At the same time, the microorganisms increased their mass by 338%, 58%, and 1% in the first, second, and third bioreactor, respectively. The degradation intermediates confirmed the efficiency of the process and the possible reuse of water, which closes the water loop's circularity. Furthermore, a nitrate concentration of 150–250 mg/L was found in the last two bioreactors: this implies a significant presence of ammonium-oxidizing bacteria. Community genomic analysis was conducted to detect changes in the microbial community due to changes in operating conditions that could cause reactor failure if methylotrophs were washed out or outcompeted by unspecific bacteria. [ABSTRACT FROM AUTHOR]

• Methanol supply enhances the culture characteristics under H 2 /CO 2 condition. • Methanol metabolism exhibits high correspondence with H 2 /CO 2 metabolism. • Methanol addition can enhance substrate consumption during syngas fermentation. This study analyzed the effect of methanol on the metabolism of syngas components (i.e. , H 2 and CO) by the syngas fermenting acetogenic strain E. limosum KIST612. The culture characteristics and relevant proteomic expressions (as fold changes) were carefully analyzed under CO/CO 2 and H 2 /CO 2 conditions with and without methanol addition, as well as, under methanol/CO 2 conditions. The culture characteristics (specific growth rate and H 2 consumption rate) under H 2 /CO 2 conditions were greatly enhanced in the presence of methanol, by 4.0 and 2.7 times, respectively. However, the promoting effect of methanol was not significant under CO/CO 2 conditions. Proteomic fold changes in most enzyme expression levels in the Wood-Ljungdahl pathway and chemiosmotic energy conservation also exhibited high correspondence between methanol and H 2 /CO 2 but not between methanol and CO/CO 2. These findings suggest the advantages of methanol addition to H 2 /CO 2 for biomass enhancement and faster consumption of gaseous substrates during syngas fermentation. [ABSTRACT FROM AUTHOR]

Peatlands store close to half of the terrestrial soil carbon and release substantial amounts of the greenhouse gas methane (CH 4) to the atmosphere. While CH 4 is thought to be produced mainly by hydrogenotrophic and acetoclastic methanogenic pathways, methylotrophic methanogenesis may be a hidden, yet potentially important, source of CH 4 in these globally significant ecosystems. To explore the importance of methylotrophic methanogenesis in northern peatlands, 13C-labeled methylated substrates (methanol, dimethylsulfide, and trimethylamine) were added to soils along a depth profile and the production of 13C–CH 4 was measured. Additionally, the concentrations of methylated compounds were measured across a soil depth profile in situ. Results suggest that methylated substrates can be used by peatland microbes, resulting in CH 4 production in all sites at all depths. Methanol was converted to CH 4 in each site at all depths. In contrast, trimethylamine and dimethylsulfide were only converted to CH 4 at the surface depth regardless of site. In situ concentrations of dimethylsulfide were below detection limit in all sites and depths. Monomethylamine and methanol were present in situ with concentrations ranging from 30 to 1225 nM. For the first time in peatlands, we have estimated contributions of methylated substrates to the total CH 4 produced. Microbial processing of methylated substrates contributed a maximum of 11% of the total CH 4 produced at depth in one peatland; but, in shallow soils with higher rates of CH 4 production, the potential contribution of methylotrophic methanogenesis to overall CH 4 production was minor. • Methylotrophic methanogenesis is possible through the depth profile in three peatlands. • Utilization of methylated substrates contributes a maximum of ~11% of CH 4 production in the studied soils. • Methanol and monomethylamine are dominant methylated substrates in situ. [ABSTRACT FROM AUTHOR]

A halotolerant, psychrotolerant and methylotrophic methanogen, strain SY-01 T , was isolated from the saline Lake Tus in Siberia. Cells of strain SY-01 T were non-motile, cocci and 0.8-1.0 µm in diameter. The only methanogenic substrate utilized by strain SY-01 T was methanol. The temperature range of growth for strain SY-01 T was from 4 to 40 °C and the optimal temperature for growth was 30 °C. The pH range of growth was from pH 7.2 to 9.0, with optimal growth at pH 8.0. The NaCl range of growth was 0-1.55 M with optimal growth at 0.51 M NaCl. The G+C content of the genome of strain SY-01 T was 43.6 mol % as determined by genome sequencing. Phylogenetic analysis revealed that strain SY-01 T was most closely related to Methanolobus zinderi SD1 T (97.3 % 16S rRNA gene sequence similarity), and had 95.5-97.2 % similarities to other Methanolobus species with valid names. Genome relatedness between strain SY-01 T and DSM 21339 T was computed using average nucleotide identity and digital DNA-DNAhybridization, which yielded values of 79.7 and 21.7 %, respectively. Based on morphological, phenotypic, phylogenetic and genomic relatedness data presented here, it is evident that strain SY-01 T represents a novel species of the genus Methanolobus , and the name Methanolobus halotolerans sp. nov. is proposed. The type strain is SY-01 T (=BCRC AR10051 T =NBRC 113166 T =DSM 107642 T ).

A novel anaerobic methylotrophic halophilic methanogen strain SLHTYRO T was isolated from a deep hypersaline anoxic basin called "Tyro" located in the Eastern Mediterranean Sea. Cells of SLHTYRO T were motile cocci. The strain SLHTYRO T grew between 12 and 37 °C (optimum 30 °C), at pH between 6.5 and 8.2 (optimum pH 7.5) and salinity from 45 to 240 g L -1 NaCl (optimum 135 g L -1 ). Strain SLHTYRO T was methylotrophic methanogen able to use methylated compounds (trimethylamine, dimethylamine, monomethylamine and methanol). Strain SLHTYRO T was able to grow at in situ hydrostatic pressure and temperature conditions (35 MPa, 14 °C). Phylogenetic analysis based on 16S rRNA gene and mcrA gene sequences indicated that strain SLHTYRO T was affiliated to genus Methanohalophilus within the order Methanosarcinales. It shared >99.16% of the 16S rRNA gene sequence similarity with strains of other Methanohalophilus species. Based on ANIb, AAI and dDDH measurements, and the physiological properties of the novel isolate, we propose that strain SLHTYRO T should be classified as a representative of a novel species, for which the name Methanohalophilus profundi sp. nov. is proposed; the type strain is SLHTYRO T (=DSM 108854 = JCM 32768 = UBOCC-M-3308)., (Copyright © 2020 Elsevier GmbH. All rights reserved.)