Methanolobus mangrovi sp. nov. and Methanolobus sediminis sp. nov, two novel methylotrophic methanogens isolated from mangrove sediments in China.

Two methylotrophic methanogens, designated strains FTZ2T and FTZ6T, were isolated from mangrove sediment sampled in Futian Mangrove Nature Reserve in Shenzhen, PR China. Cells of strains FTZ2T and FTZ6T were cocci, with diameters of 0.6-1.0 µm and 0.6-0.9 µm, respectively. Both strains grew on methanol, methylamine, dimethylamine and trimethylamine, but not on acetate, formate, H2/CO2, choline, betaine or dimethyl sulphide. Strain FTZ2T grew at 10-37 °C (optimally at 33 °C), pH 5.5-8.0 (optimally at pH 7.0) and 0-1.03 M NaCl (optimally at 0.17 M NaCl). In contrast, strain FTZ6T grew at 15-42 °C (optimally at 37 °C), pH 5.0-7.5 (optimally pH 6.5) and 0-1.03 M NaCl (optimally at 0.17 M NaCl). Both strains required magnesium for growth and were susceptible to sodium dodecyl sulphate. Biotin was required for the growth of strain FTZ2T but not of strain FTZ6T. The genomic G+C contents of strains FTZ2T and FTZ6T were 41.6 and 40.9 mol%, respectively. Phylogenetic analyses revealed that strain FTZ2T was mostly related to Methanolobus psychrotolerans YSF-03T, with 16S rRNA gene similarity of 98.6 %, an average nucleotide identity (ANI) of 82.5 %, and a digital DNA-DNA hybridization (dDDH) of 24.6 %. While strain FTZ6T was mostly related to Methanolobus vulcani PL-12/MT, with 16S rRNA gene similarity of 99.4 %, an ANI of 88.6% and a dDDH of 34.6 %. Based on phenotypic, phylogenetic and genotypic evidence, two novel species of the genus Methanolobus, Methanolobus mangrovi sp. nov. and Methanolobus sediminis sp. nov., are proposed. The type strain of M. mangrovi sp. nov. is FTZ2T (=CCAM 1276T=JCM 39396T) and the type strain of M. sediminis sp. nov. is FTZ6T (=CCAM 1277T=JCM 39397T).

Desulfovibrio mangrovi sp. nov., a sulfate-reducing bacterium isolated from mangrove sediments: a member of the proposed genus "Psychrodesulfovibrio".

"Psychrodesulfovibrio", a proposed genus within the family Desulfovibrionaceae, is a group of sulfate-reducing bacteria with biogeochemical significance but restricted child taxa availability. In this study, a strictly anaerobic bacterium, designed strain FT415T, was isolated from mangrove sediments in Futian Mangrove Nature Reserve in Shenzhen, China. The strain was Gram-stain-negative, motile, and vibrio-shaped with a single polar flagellum, which grew at the temperature range of 15-42 °C (optimum 37 °C), pH range of 6.0-7.5 (optimum 6.8), and in the presence of 0-36 g l-1 NaCl (optimum 6 g l-1 NaCl). In the presence of sulfate, electron donors including lactate, ethanol, pyruvate, malate, fumarate, succinate, cysteine, and glycerol were incompletely oxidized to acetate, and H2 and formate were used as electron donors with acetate as the carbon source by strain FT415T. Sulfate, thiosulfate, sulfide, and anthraquinone-2,6-disulfonate were reduced in the presence of lactate. Fe(III) oxide was reduced without cell growth. Fermentative growth was observed with pyruvate and cysteine. Vitamins were not required for growth. The major cellular fatty acids (> 10%) were C16:0, summed feature 10 (C18:1 c11/t9/t6 and/or unknown ECL 17.834), C16:1 cis 9, and C18:0. The major polar lipids were phosphatidylethanolamine, phospholipids, and aminolipids. The predominant menaquinone was MK-6(H2). The genomic DNA G+C content was 56.7%. Phylogenetic analysis showed that strain FT415T shared a 98.1% similarity in 16S rRNA gene sequence, an average nucleotide identity value of 84.0%, an average amino-acid identity value of 85.4%, and a digital DNA-DNA hybridization value of 25.7% with its closest relative Desulfovibrio subterraneus HN2T, which has been proposed to be transferred to the genus "Psychrodesulfovibrio". Based on phenotypic, phylogenetic, and genotypic evidence, a new species of the family Desulfovibrionaceae, Desulfovibrio mangrovi sp. nov. was proposed with the type strain FT415T (=GDMCC 1.3410T=KCTC 25525T).

Potential for mercury methylation by Asgard archaea in mangrove sediments.

Methylmercury (MeHg) is a potent neurotoxin that bioaccumulates along food chains. The conversion of MeHg from mercury (Hg) is mediated by a variety of anaerobic microorganisms carrying hgcAB genes. Mangrove sediments are potential hotspots of microbial Hg methylation; however, the microorganisms responsible for Hg methylation are poorly understood. Here, we conducted metagenomic and metatranscriptomic analyses to investigate the diversity and distribution of putative microbial Hg-methylators in mangrove ecosystems. The highest hgcA abundance and expression occurred in surface sediments in Shenzhen, where the highest MeHg concentration was also observed. We reconstructed 157 metagenome-assembled genomes (MAGs) carrying hgcA and identified several putative novel Hg-methylators, including one Asgard archaea (Lokiarchaeota). Further analysis of MAGs revealed that Deltaproteobacteria, Euryarchaeota, Bacteroidetes, Chloroflexi, and Lokiarchaeota were the most abundant and active Hg-methylating groups, implying their crucial role in MeHg production. By screening publicly available MAGs, 104 additional Asgard MAGs carrying hgcA genes were identified from a wide range of coast, marine, permafrost, and lake sediments. Protein homology modelling predicts that Lokiarchaeota HgcAB proteins contained the highly conserved amino acid sequences and folding structures required for Hg methylation. Phylogenetic tree revealed that hgcA genes from Asgard clustered with fused hgcAB genes, indicating a transitional stage of Asgard hgcA genes. Our findings thus suggest that Asgard archaea are potential novel Hg-methylating microorganisms and play an important role in hgcA evolution.

Non-syntrophic methanogenic hydrocarbon degradation by an archaeal species.

The methanogenic degradation of oil hydrocarbons can proceed through syntrophic partnerships of hydrocarbon-degrading bacteria and methanogenic archaea1-3. However, recent culture-independent studies have suggested that the archaeon 'Candidatus Methanoliparum' alone can combine the degradation of long-chain alkanes with methanogenesis4,5. Here we cultured Ca. Methanoliparum from a subsurface oil reservoir. Molecular analyses revealed that Ca. Methanoliparum contains and overexpresses genes encoding alkyl-coenzyme M reductases and methyl-coenzyme M reductases, the marker genes for archaeal multicarbon alkane and methane metabolism. Incubation experiments with different substrates and mass spectrometric detection of coenzyme-M-bound intermediates confirm that Ca. Methanoliparum thrives not only on a variety of long-chain alkanes, but also on n-alkylcyclohexanes and n-alkylbenzenes with long n-alkyl (C≥13) moieties. By contrast, short-chain alkanes (such as ethane to octane) or aromatics with short alkyl chains (C≤12) were not consumed. The wide distribution of Ca. Methanoliparum4-6 in oil-rich environments indicates that this alkylotrophic methanogen may have a crucial role in the transformation of hydrocarbons into methane.

Comparative genomic analysis reveals metabolic flexibility of Woesearchaeota.

The archaeal phylum Woesearchaeota, within the DPANN superphylum, includes phylogenetically diverse microorganisms that inhabit various environments. Their biology is poorly understood due to the lack of cultured isolates. Here, we analyze datasets of Woesearchaeota 16S rRNA gene sequences and metagenome-assembled genomes to infer global distribution patterns, ecological preferences and metabolic capabilities. Phylogenomic analyses indicate that the phylum can be classified into ten subgroups, termed A-J. While a symbiotic lifestyle is predicted for most, some members of subgroup J might be host-independent. The genomes of several Woesearchaeota, including subgroup J, encode putative [FeFe] hydrogenases (known to be important for fermentation in other organisms), suggesting that these archaea might be anaerobic fermentative heterotrophs.

Expanded diversity of Asgard archaea and their relationships with eukaryotes.

Asgard is a recently discovered superphylum of archaea that appears to include the closest archaeal relatives of eukaryotes1-5. Debate continues as to whether the archaeal ancestor of eukaryotes belongs within the Asgard superphylum or whether this ancestor is a sister group to all other archaea (that is, a two-domain versus a three-domain tree of life)6-8. Here we present a comparative analysis of 162 complete or nearly complete genomes of Asgard archaea, including 75 metagenome-assembled genomes that-to our knowledge-have not previously been reported. Our results substantially expand the phylogenetic diversity of Asgard and lead us to propose six additional phyla that include a deep branch that we have provisionally named Wukongarchaeota. Our phylogenomic analysis does not resolve unequivocally the evolutionary relationship between eukaryotes and Asgard archaea, but instead-depending on the choice of species and conserved genes used to build the phylogeny-supports either the origin of eukaryotes from within Asgard (as a sister group to the expanded Heimdallarchaeota-Wukongarchaeota branch) or a deeper branch for the eukaryote ancestor within archaea. Our comprehensive protein domain analysis using the 162 Asgard genomes results in a major expansion of the set of eukaryotic signature proteins. The Asgard eukaryotic signature proteins show variable phyletic distributions and domain architectures, which is suggestive of dynamic evolution through horizontal gene transfer, gene loss, gene duplication and domain shuffling. The phylogenomics of the Asgard archaea points to the accumulation of the components of the mobile archaeal 'eukaryome' in the archaeal ancestor of eukaryotes (within or outside Asgard) through extensive horizontal gene transfer.

Diversity, metabolism and cultivation of archaea in mangrove ecosystems.

Mangroves comprise a globally significant intertidal ecosystem that contains a high diversity of microorganisms, including fungi, bacteria and archaea. Archaea is a major domain of life that plays important roles in biogeochemical cycles in these ecosystems. In this review, the potential roles of archaea in mangroves are briefly highlighted. Then, the diversity and metabolism of archaeal community of mangrove ecosystems across the world are summarized and Bathyarchaeota, Euryarchaeota, Thaumarchaeota, Woesearchaeota, and Lokiarchaeota are confirmed as the most abundant and ubiquitous archaeal groups. The metabolic potential of these archaeal groups indicates their important ecological function in carbon, nitrogen and sulfur cycling. Finally, some cultivation strategies that could be applied to uncultivated archaeal lineages from mangrove wetlands are suggested, including refinements to traditional cultivation methods based on genomic and transcriptomic information, and numerous innovative cultivation techniques such as single-cell isolation and high-throughput culturing (HTC). These cultivation strategies provide more opportunities to obtain previously uncultured archaea.

Global distribution and environmental drivers of methylmercury production in sediments.

Neurotoxic methylmercury (MeHg) in environments poses substantial risks to human health. Saturated sediments are basic sources of MeHg in food chains; however, distribution patterns and environmental drivers of MeHg at a global scale remain largely unexplored. Here, we characterized global patterns of MeHg distribution and environmental drivers of MeHg production based on 495 sediment samples across five typical ecosystems from the literature (1995-2018) and our own field survey. Our results showed the MeHg concentration ranged from 0.009 to 55.7 µg kg-1 across the different ecosystems, and the highest MeHg concentration and Hg methylation potential were from the sediments of paddy and marine environments, respectively. Further, using combined analysis of random forest and structural equation modeling, we identified temperature and precipitation as important regulators of MeHg production after accounting for the well-known drivers including Hg availability and sediment geochemistry. More importantly, we found increased MeHg production in sediments with elevated mean annual Hg precipitation, and warmer temperature could also accelerate MeHg production by facilitating activities of microbial methylators. Together, this work advances our understanding of global MeHg distribution in sediments and environmental drivers, which are fundamental to the prediction and management of MeHg production and its potential health risk globally.

Genomic and transcriptomic insights into methanogenesis potential of novel methanogens from mangrove sediments.

BACKGROUND: Methanogens are crucial to global methane budget and carbon cycling. Methanogens from the phylum Euryarchaeota are currently classified into one class and seven orders, including two novel methanogen taxa, Methanofastidiosa and Methanomassiliicoccales. The relative importance of the novel methanogens to methane production in the natural environment is poorly understood. RESULTS: Here, we used a combined metagenomic and metatranscriptomic approach to investigate the metabolic activity of methanogens in mangrove sediments in Futian Nature Reserve, Shenzhen. We obtained 13 metagenome-assembled genomes (MAGs) representing one class (Methanofastidiosa) and five orders (Methanomassiliicoccales, Methanomicrobiales, Methanobacteriales, Methanocellales, and Methanosarcinales) of methanogens, including the two novel methanogens. Comprehensive annotation indicated the presence of an H2-dependent methylotrophic methanogenesis pathway in Methanofastidiosa and Methanomassiliicoccales. Based on the functional gene analysis, hydrogenotrophic and methylotrophic methanogenesis are the dominant pathways in mangrove sediments. MAG mapping revealed that hydrogenotrophic Methanomicrobiales were the most abundant methanogens and that methylotrophic Methanomassiliicoccales were the most active methanogens in the analyzed sediment profile, suggesting their important roles in methane production. CONCLUSIONS: Partial or near-complete genomes of two novel methanogen taxa, Methanofastidiosa and Methanomassiliicoccales, in natural environments were recovered and analyzed here for the first time. The presented findings highlight the ecological importance of the two novel methanogens and complement knowledge of how methane is produced in mangrove ecosystem. This study implies that two novel methanogens play a vital role in carbon cycle. Video Abstract.

Spatial and seasonal variation of methanogenic community in a river-bay system in South China.

River-bay system is a transitional zone connecting land and ocean and an important natural source for methane emission. Methanogens play important roles in the global greenhouse gas budget and carbon cycle since they produce methane. The abundance and community assemblage of methanogens in such a dynamic system are not well understood. Here, we used quantitative PCR and high-throughput sequencing of the mcrA gene to investigate the abundance and community composition of methanogens in the Shenzhen River-Bay system, a typical subtropical river-bay system in Southern of China, during the wet and dry seasons. Results showed that mcrA gene abundance was significantly higher in the sediments of river than those of estuary, and was higher in wet season than dry season. Sequences of mcrA gene were mostly assigned to three orders, including Methanosarcinales, Methanomicrobiales, and Methanobacteriales. Specifically, Methanosarcina, Methanosaeta, and Methanobacterium were the most abundant and ubiquitous genera. Methanogenic communities generally clustered according to habitat (river vs. estuary), and salinity was the major factor driving the methanogenic community assemblage. Furthermore, the indicator groups for two habitats were identified. For example, Methanococcoides, Methanoculleus, and Methanogenium preferentially existed in estuarine sediments, whereas Methanomethylovorans, Methanolinea, Methanoregula, and Methanomassiliicoccales were more abundant in riverine sediments, indicating distinct ecological niches. Overall, these findings reveal the distribution patterns of methanogens and expand our understanding of methanogenic community assemblage in the river-bay system. Key Points • Abundance of methanogens was relatively higher in riverine sediments. • Methanogenic community in estuarine habitat separated from that in riverine habitat. • Salinity played a vital role in regulating methanogenic community assemblage.

Prokaryotic Diversity in Mangrove Sediments across Southeastern China Fundamentally Differs from That in Other Biomes.

Mangroves, as a blue carbon reservoir, provide an environment for a variety of microorganisms. Mangroves lie in special locations connecting coastal and estuarine areas and experience fluctuating conditions, which are expected to intensify with climate change, creating a need to better understand the relative roles of stochastic and deterministic processes in shaping microbial community assembly. Here, a study of microbial communities inhabiting mangrove sediments across southeastern China, spanning mangroves in six nature reserves, was conducted. We performed high-throughput DNA sequencing of these samples and compared them with data of 1,370 sediment samples collected from the Earth Microbiome Project (EMP) to compare the microbial diversity of mangroves with that of other biomes. Our results showed that prokaryotic alpha diversity in mangroves was significantly higher than that in other biomes and that microbial beta diversity generally clustered according to biome types. The core operational taxonomic units (OTUs) in mangroves were mostly assigned to Gammaproteobacteria, Deltaproteobacteria, Chloroflexi, and Euryarchaeota The majority of beta nearest-taxon index values were higher than 2, indicating that community assembly in mangroves was better explained through a deterministic process than through a stochastic process. Mean annual precipitation (MAP) and total organic carbon (TOC) were main deterministic factors explaining variation in the microbial community. This study fills a gap in addressing the unique microbial diversity of mangrove ecosystems and their microbial community assembly mechanisms.IMPORTANCE Understanding the underlying mechanisms of microbial community assembly patterns is a vital issue in microbial ecology. Mangroves, as an important and special ecosystem, provide a unique environment for examining the relative importance of stochastic and deterministic processes. We made the first global-scale comparison and found that microbial diversity was significantly different in mangrove sediments compared to that of other biomes. Furthermore, our results suggest that a deterministic process is more important in shaping microbial community assembly in mangroves.

Intraspecies variation in a widely distributed tree species regulates the responses of soil microbiome to different temperature regimes.

Plant characteristics in different provenances within a single species may vary in response to climate change, which might alter soil microbial communities and ecosystem functions. We conducted a glasshouse experiment and grew seedlings of three provenances (temperate, subtropical and tropical origins) of a tree species (i.e., Eucalyptus tereticornis) at different growth temperatures (18, 21.5, 25, 28.5, 32 and 35.5°C) for 54 days. At the end of the experiment, bacterial and fungal community composition, diversity and abundance were characterized. Measured soil functions included surrogates of microbial respiration, enzyme activities and nutrient cycling. Using Permutation multivariate analysis of variance (PerMANOVA) and network analysis, we found that the identity of tree provenances regulated both structure and function of soil microbiomes. In some cases, tree provenances substantially affected the response of microbial communities to the temperature treatments. For example, we found significant interactions of temperature and tree provenance on bacterial community and relative abundances of Chloroflexi and Zygomycota, and inorganic nitrogen. Microbial abundance was altered in response to increasing temperature, but was not affected by tree provenances. Our study provides novel evidence that even a small variation in biotic components (i.e., intraspecies tree variation) can significantly influence the response of soil microbial community composition and specific soil functions to global warming.

Impacts of Projected Climate Warming and Wetting on Soil Microbial Communities in Alpine Grassland Ecosystems of the Tibetan Plateau.

Climate change is projected to have impacts on precipitation and temperature regimes in drylands of high elevation regions, with especially large effects in the Qinghai-Tibetan Plateau. However, there was limited information about how the projected climate change will impact on the soil microbial community and their activity in the region. Here, we present results from a study conducted across 72 soil samples from 24 different sites along a temperature and precipitation gradient (substituted by aridity index ranging from 0.079 to 0.89) of the Plateau, to assess how changes in aridity affect the abundance, community composition, and diversity of bacteria, ammonia-oxidizers, and denitrifers (nirK/S and nosZ genes-containing communities) as well as nitrogen (N) turnover enzyme activities. We found V-shaped or inverted V-shaped relationships between the aridity index (AI) and soil microbial parameters (gene abundance, community structures, microbial diversity, and N turnover enzyme activities) with a threshold at AI = 0.27. The increasing or decreasing rates of the microbial parameters were higher in areas with AI < 0.27 (alpine steppes) than in mesic areas with 0.27 < AI < 0.89 (alpine meadow and swamp meadow). The results indicated that the projected warming and wetting have a strong impact on soil microbial communities in the alpine steppes.

Interactive effects of multiple climate change factors on ammonia oxidizers and denitrifiers in a temperate steppe.

Global climate change could have profound effects on belowground microbial communities and subsequently affect soil biogeochemical processes. The interactive effects of multiple co-occurring climate change factors on microbially mediated processes are not well understood. A four-factorial field experiment with elevated CO2, watering, nitrogen (N) addition and night warming was conducted in a temperate steppe of northern China. Real-time polymerase chain reaction and terminal-restriction fragment length polymorphism, combined with clone library techniques, were applied to examine the effects of those climate change factors on N-related microbial abundance and community composition. Only the abundance of ammonia-oxidizing bacteria significantly increased by nitrogen addition and decreased by watering. The interactions of watering × warming on the bacterial amoA community and warming × nitrogen addition on the nosZ community were found. Redundancy analysis indicated that the ammonia-oxidizing archaeal community was affected by total N and total carbon, while the community of bacterial amoA and nosZ were significantly affected by soil pH. According to a structural equation modeling analysis, climate change influenced net primary production indirectly by altering microbial abundance and activities. These results indicated that microbial responses to the combination of chronic global change tend to be smaller than expected from single-factor global change manipulations.

[Responses of Soil Ammonia Oxidizers to Simulated Warming and Increased Precipitation in a Temperate Steppe of Inner Mongolia].

Soil ammonia oxidizers, as key players for the ammonia oxidation process in soil N cycling, could respond, adapt, and give feedback to global change. In this research, soil samples were collected from a long-term field experiment with increased precipitation and warming in a temperate steppe of Inner Mongolia. We analyzed the responses of the abundance, diversity, and community structure of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) to warming and increased precipitation using quantitative real-time PCR, terminal restriction fragment length polymorphism (T-RFLP), and clone library. The results showed that increased precipitation significantly stimulated soil pH and warming significantly reduced soil respiration (SR). No significant difference was detected regarding the abundances of amoA genes across all treatments, whereas increased precipitation significantly affected the community structure of soil AOB. However, the interactive effect between warming and increased precipitation had no significant influence on the community structure of soil ammonia oxidizers. The result of the structural equation model indicated that the plant diversity and community structures of soil ammonia oxidizers were significantly correlated, suggesting that there were certain relationships among climate change, microbes, and plants. In conclusion, this study confirmed that soil microorganisms had the ability to adapt to climate change, which could provide important information for predicting future changes in ecosystems.

Primary Succession of Nitrogen Cycling Microbial Communities Along the Deglaciated Forelands of Tianshan Mountain, China.

Structural succession and its driving factors for nitrogen (N) cycling microbial communities during the early stages of soil development (0-44 years) were studied along a chronosequence in the glacial forelands of the Tianshan Mountain No.1 glacier in the arid and semi-arid region of central Asia. We assessed the abundance and population of functional genes affiliated with N-fixation (nifH), nitrification (bacterial and archaeal amoA), and denitrification (nirK/S and nosZ) in a glacier foreland using molecular methods. The abundance of functional genes significantly increased with soil development. N cycling community compositions were also significantly shifted within 44 years and were structured by successional age. Cyanobacterial nifH gene sequences were the most dominant N fixing bacteria and its relative abundance increased from 56.8-93.2% along the chronosequence. Ammonia-oxidizing communities shifted from the Nitrososphaera cluster (AOA-amoA) and the Nitrosospira cluster ME (AOB-aomA) in younger soils (0 and 5 years) to communities dominated by soil and sediment 1 (AOA-amoA) and Nitrosospira Cluster 2 Related (AOB-aomA) in older soils (≥17 years). Most of the denitrifers closest relatives were potential aerobic denitrifying bacteria, and some other types of denitrifying bacteria (like autotrophic nitrate-reducing, sulfide-oxidizing bacteria and denitrifying phosphorus removing bacteria) were also detected in all soil samples. The regression analysis showed that N cycling microbial communities were dominant in younger soils (0-5 years) and significantly correlated with soil total carbon, while communities that were most abundant in older soils were significantly correlated with soil total nitrogen. These results suggested that the shift of soil C and N contents during the glacial retreat significantly influenced the abundance, composition and diversity of N cycling microbial communities.

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In recent decades, global climate change is one of the main concerns around the world. Land use change and the high demand for fossil fuel have caused severe consequences of climate change, such as elevated greenhouse gases, warming, and altering precipitation pattern. These combined factors have substantial impacts on ecosystem processes, especially carbon and nitrogen cycles in terrestrial ecosystems. Since the 1970s, a series of field manipulative experiments had been set up to stimulate the influences of monofactorial and/or multifactorial climate changes, improving our understanding of ecosystem response and feedback to global change. In this review, we summarized the development history of global change experiments, and discussed the main issues of using field manipulative experiments in simulating global change. The application of multifactorial experiments, such as CO2 enrichment, warming, precipitation and nitrogen deposition, were highlighted in the research of soil microbial ecology. Moreover, the response and feedback of soil biota as well as the biogeochemical processes that they mediated were further addressed. We also proposed the prospects of their application in global change research to explore the impact of global change on terrestrial ecosystems.