Pathways of Carbon Assimilation and Ammonia Oxidation Suggested by Environmental Genomic Analyses of Marine Crenarchaeota

Marine Crenarchaeota represent an abundant component of oceanic microbiota with potential to significantly influence biogeochemical cycling in marine ecosystems. Prior studies using specific archaeal lipid biomarkers and isotopic analyses indicated that planktonic Crenarchaeota have the capacity for autotrophic growth, and more recent cultivation studies support an ammonia-based chemolithoautotrophic energy metabolism. We report here analysis of fosmid sequences derived from the uncultivated marine crenarchaeote, Cenarchaeum symbiosum, focused on the reconstruction of carbon and energy metabolism. Genes predicted to encode multiple components of a modified 3-hydroxypropionate cycle of autotrophic carbon assimilation were identified, consistent with utilization of carbon dioxide as a carbon source. Additionally, genes predicted to encode a near complete oxidative tricarboxylic acid cycle were also identified, consistent with the consumption of organic carbon and in the production of intermediates for amino acid and cofactor biosynthesis. Therefore, C. symbiosum has the potential to function either as a strict autotroph, or as a mixotroph utilizing both carbon dioxide and organic material as carbon sources. From the standpoint of energy metabolism, genes predicted to encode ammonia monooxygenase subunits, ammonia permease, urease, and urea transporters were identified, consistent with the use of reduced nitrogen compounds as energy sources fueling autotrophic metabolism. Homologues of these genes, recovered from ocean waters worldwide, demonstrate the conservation and ubiquity of crenarchaeal pathways for carbon assimilation and ammonia oxidation. These findings further substantiate the likely global metabolic importance of Crenarchaeota with respect to key steps in the biogeochemical transformation of carbon and nitrogen in marine ecosystems.     

Population ecology of nitrifying Archaea and Bacteria in the Southern California Bight

Marine Crenarchaeota are among the most abundant microbial groups in the ocean, and although relatively little is currently known about their biogeochemical roles in marine ecosystems, recognition that Crenarchaeota posses ammonia monooxygenase (amoA) genes and may act as ammonia‐oxidizing archaea (AOA) offers another means of probing the ecology of these microorganisms. Here we use a time series approach combining quantification of archaeal and bacterial ammonia oxidizers with bacterial community fingerprints and biogeochemistry, to explore the population and community ecology of nitrification. At multiple depths (150, 500 and 890 m) in the Southern California Bight sampled monthly from 2003 to 2006, AOA were enumerated via quantitative PCR of archaeal amoA and marine group 1 Crenarchaeota 16S rRNA genes. Based on amoA genes, AOA were highly variable in time – a consistent feature of marine Crenarchaeota– however, average values were similar at different depths and ranged from 2.20 to 2.76 × 10⁴amoA copies ml^?1. Archaeal amoA genes were correlated with Crenarchaeota 16S rRNA genes (r² = 0.79) and the slope of this relationship was 1.02, demonstrating that the majority of marine group 1 Crenarchaeota present over the dates and depths sampled possessed amoA. Two AOA clades were specifically quantified and compared with betaproteobacterial ammonia‐oxidizing bacteria (β‐AOB) amoA genes at 150 m; these AOA groups were found to strongly co‐vary in time (r² = 0.70, P < 0.001) whereas AOA : β‐AOB ratios ranged from 13 to 5630. Increases in the AOA : β‐AOB ratio correlated with the accumulation of nitrite (r² = 0.87, P < 0.001), and may be indicative of differences in substrate affinities and activities leading to periodic decoupling between ammonia and nitrite oxidation. These data capture a dynamic nitrogen cycle in which multiple microbial groups appear to be active participants.     

Contribution of crenarchaeal autotrophic ammonia oxidizers to the dark primary production in Tyrrhenian deep waters (Central Mediterranean Sea)

Mesophilic Crenarchaeota have recently been thought to be significant contributors to nitrogen (N) and carbon (C) cycling. In this study, we examined the vertical distribution of ammonia-oxidizing Crenarchaeota at offshore site in Southern Tyrrhenian Sea. The median value of the crenachaeal cell to amoA gene ratio was close to one suggesting that virtually all deep-sea Crenarchaeota possess the capacity to oxidize ammonia. Crenarchaea-specific genes, nirK and ureC, for nitrite reductase and urease were identified and their affiliation demonstrated the presence of ‘deep-sea'' clades distinct from ‘shallow'' representatives. Measured deep-sea dark CO₂ fixation estimates were comparable to the median value of photosynthetic biomass production calculated for this area of Tyrrhenian Sea, pointing to the significance of this process in the C cycle of aphotic marine ecosystems. To elucidate the pivotal organisms in this process, we targeted known marine crenarchaeal autotrophy-related genes, coding for acetyl-CoA carboxylase (accA) and 4-hydroxybutyryl-CoA dehydratase (4-hbd). As in case of nirK and ureC, these genes are grouped with deep-sea sequences being distantly related to those retrieved from the epipelagic zone. To pair the molecular data with specific functional attributes we performed [¹⁴C]HCO₃ incorporation experiments followed by analyses of radiolabeled proteins using shotgun proteomics approach. More than 100 oligopeptides were attributed to 40 marine crenarchaeal-specific proteins that are involved in 10 different metabolic processes, including autotrophy. Obtained results provided a clear proof of chemolithoautotrophic physiology of bathypelagic crenarchaeota and indicated that this numerically predominant group of microorganisms facilitate a hitherto unrecognized sink for inorganic C of a global importance.     

Identification of a Thiomicrospira denitrificans-Like Epsilonproteobacterium as a Catalyst for Autotrophic Denitrification in the Central Baltic Sea

Identification and functional analysis of key members of bacterial communities in marine and estuarine environments are major challenges for obtaining a mechanistic understanding of biogeochemical processes. In the Baltic Sea basins, as in many other marine environments with anoxic bodies of water, the oxic-anoxic interface is considered a layer of high bacterial turnover of sulfur, nitrogen, and carbon compounds that has a great impact on matter balances in the whole ecosystem. We focused on autotrophic denitrification by oxidation of reduced sulfur compounds as a biogeochemically important process mediating concomitant turnover of sulfur, nitrogen, and carbon. We used a newly developed approach consisting of molecular analyses in stimulation experiments and in situ abundance. The molecular approach was based on single-strand conformational polymorphism (SSCP) analysis of the bacterial community RNA, which allowed identification of potential denitrifiers based on the sequences of enhanced SSCP bands and monitoring of the overall bacterial community during the experiments. Sequences of the SSCP bands of interest were used to design highly specific primers that enabled (i) generation of almost complete 16S rRNA gene sequences using experimental and environmental DNA as templates and (ii) quantification of the bacteria of interest by real-time PCR. By using this approach we identified the bacteria responsible for autotrophic denitrification as a single taxon, an epsilonproteobacterium related to the autotrophic denitrifier Thiomicrospira denitrificans. This finding was confirmed by material balances in the experiments that were consistent with those obtained with continuous cultures of T. denitrificans. The presence and activity of a bacterium that is phylogenetically and physiologically closely related to T. denitrificans could be relevant for the carbon budget of the central Baltic Sea because T. denitrificans exhibits only one-half the efficiency for carbon dioxide fixation per mol of sulfide oxidized and mol of nitrate reduced of Thiobacillus denitrificans hypothesized previously for this function.     

Satellite remote sensing data can be used to model marine microbial metabolite turnover

Sampling ecosystems, even at a local scale, at the temporal and spatial resolution necessary to capture natural variability in microbial communities are prohibitively expensive. We extrapolated marine surface microbial community structure and metabolic potential from 72 16S rRNA amplicon and 8 metagenomic observations using remotely sensed environmental parameters to create a system-scale model of marine microbial metabolism for 5904 grid cells (49 km²) in the Western English Chanel, across 3 years of weekly averages. Thirteen environmental variables predicted the relative abundance of 24 bacterial Orders and 1715 unique enzyme-encoding genes that encode turnover of 2893 metabolites. The genes'' predicted relative abundance was highly correlated (Pearson Correlation 0.72, P-value <10⁻⁶) with their observed relative abundance in sequenced metagenomes. Predictions of the relative turnover (synthesis or consumption) of CO₂ were significantly correlated with observed surface CO₂ fugacity. The spatial and temporal variation in the predicted relative abundances of genes coding for cyanase, carbon monoxide and malate dehydrogenase were investigated along with the predicted inter-annual variation in relative consumption or production of ∼3000 metabolites forming six significant temporal clusters. These spatiotemporal distributions could possibly be explained by the co-occurrence of anaerobic and aerobic metabolisms associated with localized plankton blooms or sediment resuspension, which facilitate the presence of anaerobic micro-niches. This predictive model provides a general framework for focusing future sampling and experimental design to relate biogeochemical turnover to microbial ecology.     

Unravelling the Carbon and Sulphur Metabolism in Coastal Soil Ecosystems Using Comparative Cultivation-Independent Genome-Level Characterisation of Microbial Communities

Bacterial autotrophy contributes significantly to the overall carbon balance, which stabilises atmospheric CO₂ concentration and decelerates global warming. Little attention has been paid to different modes of carbon/sulphur metabolism mediated by autotrophic bacterial communities in terrestrial soil ecosystems. We studied these pathways by analysing the distribution and abundance of the diagnostic metabolic marker genes cbbM, apsA and soxB, which encode for ribulose-1,5-bisphosphate carboxylase/oxygenase, adenosine phosphosulphate reductase and sulphate thiohydrolase, respectively, among different contrasting soil types. Additionally, the abundance of community members was assessed by quantifying the gene copy numbers for 16S rRNA, cbbL, cbbM, apsA and soxB. Distinct compositional differences were observed among the clone libraries, which revealed a dominance of phylotypes associated with carbon and sulphur cycling, such as Gammaproteobacteria (Thiohalomonas, Allochromatium, Chromatium, Thiomicrospira) and Alphaproteobacteria (Rhodopseudomonas, Rhodovulum, Paracoccus). The rhizosphere soil was devoid of sulphur metabolism, as the soxB and apsA genes were not observed in the rhizosphere metagenome, which suggests the absence or inadequate representation of sulphur-oxidising bacteria. We hypothesise that the novel Gammaproteobacteria sulphur oxidisers might be actively involved in sulphur oxidation and inorganic carbon fixation, particularly in barren saline soil ecosystems, suggesting their significant putative ecological role and contribution to the soil carbon pool.     

Carbon assimilating fungi from surface ocean to subseafloor revealed by coupled phylogenetic and stable isotope analysis

Fungi are ubiquitous in the ocean and hypothesized to be important members of marine ecosystems, but their roles in the marine carbon cycle are poorly understood. Here, we use ¹³C DNA stable isotope probing coupled with phylogenetic analyses to investigate carbon assimilation within diverse communities of planktonic and benthic fungi in the Benguela Upwelling System (Namibia). Across the redox stratified water column and in the underlying sediments, assimilation of ¹³C-labeled carbon from diatom extracellular polymeric substances (¹³C-dEPS) by fungi correlated with the expression of fungal genes encoding carbohydrate-active enzymes. Phylogenetic analysis of genes from ¹³C-labeled metagenomes revealed saprotrophic lineages related to the facultative yeast Malassezia were the main fungal foragers of pelagic dEPS. In contrast, fungi living in the underlying sulfidic sediments assimilated more ¹³C-labeled carbon from chemosynthetic bacteria compared to dEPS. This coincided with a unique seafloor fungal community and dissolved organic matter composition compared to the water column, and a 100-fold increased fungal abundance within the subseafloor sulfide-nitrate transition zone. The subseafloor fungi feeding on ¹³C-labeled chemolithoautotrophs under anoxic conditions were affiliated with Chytridiomycota and Mucoromycota that encode cellulolytic and proteolytic enzymes, revealing polysaccharide and protein-degrading fungi that can anaerobically decompose chemosynthetic necromass. These subseafloor fungi, therefore, appear to be specialized in organic matter that is produced in the sediments. Our findings reveal that the phylogenetic diversity of fungi across redox stratified marine ecosystems translates into functionally relevant mechanisms helping to structure carbon flow from primary producers in marine microbiomes from the surface ocean to the subseafloor.Subject terms: Microbial ecology, Fungal ecology, Microbiome, Biogeochemistry     

自养微生物同化CO_2的分子生态研究及同化碳在土壤中的转化

大气中CO2浓度持续升高和全球气候变暖是亟待解决的重大环境问题。自养微生物在环境中广泛分布,能直接参与CO2的同化,因此研究自养微生物同化CO2的分子生态学机制具有重大的科学意义。以往对自养微生物的研究多针对基因组DNA,从DNA水平揭示了不同生态系统中碳同化自养微生物的种群结构和多样性,但这些微生物在生态系统中的具体功能有待进一步的研究。近年来,随着转录组学研究技术和稳定同位素探针技术(SIP)的发展,自养微生物同化CO2的生态机理研究不断深入,这些研究明确揭示了碳同化自养微生物是河流、湖泊和海洋生态系统中CO2固定作用的驱动者,并新发现了一些具有CO2同化功能的微生物群落。基于国内外有关研究进展,从DNA和RNA水平上对自养微生物同化CO2的分子机理以及稳定同位素探针技术(SIP)在碳同化微生物研究中的应用进行了分析和总结,初步展望了RNA-SIP技术在陆地生态系统碳同化微生物分子生态学研究中的前景。同时,探讨了陆地生态系统同化碳的转化和稳定性机理,以期为深入了解生态系统碳循环过程和应对气候变化提供理论依据。     

海洋奇古菌门认知的拓展：从新类群到新功能

奇古菌门是全球海洋中的重要微生物类群,在海洋原核浮游生物中的比例可达20%–40%。作为一类化能无机自养微生物,奇古菌门成员可通过氧化氨获得能量,实现不依赖光照的无机碳固定,在碳、氮等元素的地球化学循环中起关键作用。奇古菌门是海洋中氨氧化反应的主要执行者,其化能合成的有机质是海洋特别是深海环境中微生物的重要能量来源。随着研究的逐步深入,有关该类群生理代谢特性的认知不断被拓展,包括奇古菌门异养代谢的揭示、不具氨氧化能力类群在深海中的发现,以及最新报道的奇古菌门在厌氧条件下介导氧气、氧化亚氮和氮气的产生等。这些研究揭示了奇古菌门参与海洋生物地球化学循环和气候变化调节的新机制,为围绕该类群的深入探究和培养提供了新的思路和方向。本文从群落组成、环境适应、生态功能、进化历史和培养现状等方面综述了近年来有关海洋奇古菌门的新发现和新认识,以期增进对该类群的了解。     

Interactions between Thaumarchaea,Nitrospira and methanotrophs modulate autotrophic nitrification in volcanic grassland soil

Ammonium/ammonia is the sole energy substrate of ammonia oxidizers, and is also an essential nitrogen source for other microorganisms. Ammonia oxidizers therefore must compete with other soil microorganisms such as methane-oxidizing bacteria (MOB) in terrestrial ecosystems when ammonium concentrations are limiting. Here we report on the interactions between nitrifying communities dominated by ammonia-oxidizing archaea (AOA) and Nitrospira-like nitrite-oxidizing bacteria (NOB), and communities of MOB in controlled microcosm experiments with two levels of ammonium and methane availability. We observed strong stimulatory effects of elevated ammonium concentration on the processes of nitrification and methane oxidation as well as on the abundances of autotrophically growing nitrifiers. However, the key players in nitrification and methane oxidation, identified by stable-isotope labeling using ¹³CO₂ and ¹³CH₄, were the same under both ammonium levels, namely type 1.1a AOA, sublineage I and II Nitrospira-like NOB and Methylomicrobium-/Methylosarcina-like MOB, respectively. Ammonia-oxidizing bacteria were nearly absent, and ammonia oxidation could almost exclusively be attributed to AOA. Interestingly, although AOA functional gene abundance increased 10-fold during incubation, there was very limited evidence of autotrophic growth, suggesting a partly mixotrophic lifestyle. Furthermore, autotrophic growth of AOA and NOB was inhibited by active MOB at both ammonium levels. Our results suggest the existence of a previously overlooked competition for nitrogen between nitrifiers and methane oxidizers in soil, thus linking two of the most important biogeochemical cycles in nature.     

Genome of the Epsilonproteobacterial Chemolithoautotroph Sulfurimonas denitrificans

Sulfur-oxidizing epsilonproteobacteria are common in a variety of sulfidogenic environments. These autotrophic and mixotrophic sulfur-oxidizing bacteria are believed to contribute substantially to the oxidative portion of the global sulfur cycle. In order to better understand the ecology and roles of sulfur-oxidizing epsilonproteobacteria, in particular those of the widespread genus Sulfurimonas, in biogeochemical cycles, the genome of Sulfurimonas denitrificans DSM1251 was sequenced. This genome has many features, including a larger size (2.2 Mbp), that suggest a greater degree of metabolic versatility or responsiveness to the environment than seen for most of the other sequenced epsilonproteobacteria. A branched electron transport chain is apparent, with genes encoding complexes for the oxidation of hydrogen, reduced sulfur compounds, and formate and the reduction of nitrate and oxygen. Genes are present for a complete, autotrophic reductive citric acid cycle. Many genes are present that could facilitate growth in the spatially and temporally heterogeneous sediment habitat from where Sulfurimonas denitrificans was originally isolated. Many resistance-nodulation-development family transporter genes (10 total) are present; of these, several are predicted to encode heavy metal efflux transporters. An elaborate arsenal of sensory and regulatory protein-encoding genes is in place, as are genes necessary to prevent and respond to oxidative stress.     

Metagenomic and 14C tracing evidence for autotrophic microbial CO2 fixation in paddy soils

Autotrophic carbon dioxide (CO₂) fixation by microbes is ubiquitous in the environment and potentially contributes to the soil organic carbon (SOC) pool. However, the multiple autotrophic pathways of microbial carbon assimilation and fixation in paddy soils remain poorly characterized. In this study, we combine metagenomic analysis with ¹⁴C-labelling to investigate all known autotrophic pathways and CO₂ assimilation mechanisms in five typical paddy soils from southern China. Marker genes of six autotrophic pathways are detected in all soil samples, which are dominated by the cbbL genes (67%–82%) coding the ribulose-bisphosphate carboxylase large chain in the Calvin cycle. These marker genes are associated with a broad range of phototrophic and chemotrophic genera. Significant amounts of ¹⁴C-CO₂ are assimilated into SOC (74.3–175.8 mg ¹⁴C kg⁻¹) and microbial biomass (5.2–24.1 mg ¹⁴C kg⁻¹) after 45 days incubation, where more than 70% of ¹⁴C-SOC was concentrated in the relatively stable humin fractions. These results show that paddy soil microbes contain the genetic potential for autotrophic carbon fixation spreading over broad taxonomic ranges, and can incorporate atmospheric carbon into organic components, which ultimately contribute to the stable SOC pool.     

Carbon and nitrogen metabolism in legume root nodules

The literature concerning the metabolism of carbon compounds during the reduction, assimilation and translocation of nitrogen in root nodules of leguminous plants is reviewed. The reduction of dinitrogen requires an energy source (ATP) and a reluctant which are both supplied by respiratory catabolism of carbohydrates produced by the host plant. Photosynthates are also required to generate the carbon skeletons for amino acid or urcide synthesis during the assimilation of ammonia produced by the bacteria within the nodule tissue. Competition for photosynthates occurs between the bacteroids, nodule tissue and the various vegetative and reproductive sinks in the host plant. The nature of carbon compounds involved in these processes, their routes of metabolism, the mechanisms of control and the partitioning of metabolises between the various sites of utilization are only poorly understood. It is apparent that dinitrogen is reduced to ammonia in the bacteroids. Both fast- and slow-growing strains of Rhizobium possess the Entner-Doudoroff pathway of glucose catabolism, and some, if not all, enzymes of the Emden-Meyerhof pathway. Some bacterial cultures also metabolize carbon through the ketogluconate pathway but only the fast-growing strains of cultured rhizobia possess the key enzyme of the pentose phosphate pathway (6-phosphogluconate dehydrogenase). The host cells are thought to contain the complete Emden-Meyerhof pathway and tricarboxylic acid cycle, which provides the carbon skeletons for assimilation of the ammonia, formed by the bacteroids, into α-amino acids. A pathway of anapleurotic carbon conservation, operative in the host cells, synthesizes oxaloacetic acid through β-carboxylation of phosphoenol pyruvate. This process could be important in the recapture and assimilation of respired CO₂ in the rhizosphere. The main route of assimilation of ammonia produced by the bacteroids would appear to be via the glutamine synthetase-glutamate synthase pathway in the host cells. However, glutamate dehydrogenase may also be involved in ammonia assimilation. These enzymes also occur in in vitro cultures of Rhizobium and in bacteroids where they presumably participate in the synthesis of amino acids for growth of the bacteria or bacteroids. Nitrogen assimilated into glutamine or glutamate is exported from the nodules in a variety of forms, which include asparagine, glutamine, aspartate, homoserine and allantoates, in proportions which depend on the legume species. Studies on regulation of the overall process have focussed on expression of bacteroid genes and on the control of enzyme activity, at the level of nitrogenase and enzymes of nitrogen assimilation in particular. However, due to the wide range of experimental techniques, environmental conditions and plant species which have been used, no clear conclusions can yet be drawn. The pathways of carbon flow in nitrogen metabolism, particularly in relation to the synthesis of ureides and the regulation of carbon metabolism, remain key areas for future research in symbiotic nitrogen fixation.     

Ecological Genomics of Marine Roseobacters

Bacterioplankton of the marine Roseobacter clade have genomes that reflect a dynamic environment and diverse interactions with marine plankton. Comparative genome sequence analysis of three cultured representatives suggests that cellular requirements for nitrogen are largely provided by regenerated ammonium and organic compounds (polyamines, allophanate, and urea), while typical sources of carbon include amino acids, glyoxylate, and aromatic metabolites. An unexpectedly large number of genes are predicted to encode proteins involved in the production, degradation, and efflux of toxins and metabolites. A mechanism likely involved in cell-to-cell DNA or protein transfer was also discovered: vir-related genes encoding a type IV secretion system typical of bacterial pathogens. These suggest a potential for interacting with neighboring cells and impacting the routing of organic matter into the microbial loop. Genes shared among the three roseobacters and also common in nine draft Roseobacter genomes include those for carbon monoxide oxidation, dimethylsulfoniopropionate demethylation, and aromatic compound degradation. Genes shared with other cultured marine bacteria include those for utilizing sodium gradients, transport and metabolism of sulfate, and osmoregulation.     

Millimeter-scale vertical partitioning of nitrogen cycling in hypersaline mats reveals prominence of genes encoding multi-heme and prismane proteins

Microbial mats are modern analogues of the first ecosystems on the Earth. As extant representatives of microbial communities where free oxygen may have first been available on a changing planet, they offer an ecosystem within which to study the evolution of biogeochemical cycles requiring and inhibited by oxygen. Here, we report the distribution of genes involved in nitrogen metabolism across a vertical oxygen gradient at 1 mm resolution in a microbial mat using quantitative PCR (qPCR), retro-transcribed qPCR (RT-qPCR) and metagenome sequencing. Vertical patterns in the presence and expression of nitrogen cycling genes, corresponding to oxygen requiring and non-oxygen requiring nitrogen metabolism, could be seen across gradients of dissolved oxygen and ammonium. Metagenome analysis revealed that genes annotated as hydroxylamine dehydrogenase (proper enzyme designation EC 1.7.2.6, hao) and hydroxylamine reductase (hcp) were the most abundant nitrogen metabolism genes in the mat. The recovered hao genes encode hydroxylamine dehydrogenase EC 1.7.2.6 (HAO) proteins lacking the tyrosine residue present in aerobic ammonia oxidizing bacteria (AOB). Phylogenetic analysis confirmed that those proteins were more closely related to ɛHao protein present in Campylobacterota lineages (previously known as Epsilonproteobacteria) rather than oxidative HAO of AOB. The presence of hao sequences related with ɛHao protein, as well as numerous hcp genes encoding a prismane protein, suggest the presence of a nitrogen cycling pathway previously described in Nautilia profundicola as ancestral to the most commonly studied present day nitrogen cycling pathways.Subject terms: Microbial ecology, Population dynamics, Population genetics, Metagenomics     

Carboxylase genes of Sulfolobus metallicus

Carbon dioxide limitation of Sulfolobus metallicus resulted in increased cellular concentrations of polypeptides that were predicted to be biotin carboxylase and biotin carboxyl-carrier-protein components of a protein complex. These polypeptides were coeluted from a native polyacrylamide gel and were estimated at 19 and 59 kDa after separation by denaturing gel electrophoresis. Their encoding genes were identified, sequenced and shown to code for polypeptides of 18,580 and 58,235 Da with similarities to biotin carboxyl carrier proteins and biotin carboxylases, respectively. The genes overlapped at the second of two stop codons that terminated the carboxylase gene. A third gene occurred on the opposite strand, 293 bp upstream of the biotin carboxylase gene. Its deduced amino acid sequence was similar to those of carboxyl transferase subunits of carboxylase enzymes, in particular to those of the propionyl-CoA carboxylases. It is proposed that the three described genes could encode the key enzyme complex responsible for carbon dioxide fixation during autotrophic growth of the thermoacidophilic archaea. Received: 24 February 1999 / Accepted: 30 July 1999     

河口生态系统氨氧化菌生态学研究进展

由amoA基因编码的氨单加氧酶(AMO)所调控的氨氧化作用,是硝化作用的限速步骤和中心环节,而含有amoA基因的氨氧化细菌(AOB)和氨氧化古菌(AOA)多样性与环境因子关系密切,对缓解河口生态系统因人类活动造成的富营养化等环境问题具有特别重要的意义。水、陆和海交汇形成高度变异的具环境因子梯度的河口生态系统,是研究AOA和AOB生态学的天然实验室。河口AOA与AOB的群落组成、丰富度特征和生物有效性,与河口主要环境因子盐度、富营养化程度、植被、温度、碳、氮、硫、铁等,尤其是对盐度和富营养化有着较为强烈的响应。AOA和AOB多样性变化规律及其与河口特有的环境因子之间的相关性,应当是今后我国河口氨氧化菌研究的方向和重点。包括:(1)建立有效的氨氧化菌活性评价方法;(2)研究AOA的同化作用方式;(3)依据氨氧化菌分类和组成对河口环境变化的适应进化机制,建议可作为指示河口环境质量变化的生物标记;(4)将传统的分离培养方法与现代分子生物学研究方法相结合,筛选我国河口高效的氨氧化菌,并将其应用于生产。     

The characteristics and comparative analysis of methanotrophs reveal genomic insights into Methylomicrobium sp. enriched from marine sediments

Methanotrophic bacteria are widespread and use methane as a sole carbon and energy source. They also play a crucial role in marine ecosystems by preventing the escape of methane into the atmosphere from diverse methane sources, such as methane seeps and hydrothermal vents. Despite their importance for methane carbon cycling, relatively few marine methanotrophic bacteria have been isolated and studied at the genomic level. Herein, we report the genome of a marine methanotrophic member of the genus Methylomicrobium, metagenome-assembled genome (MAG) wino1, which was obtained through enrichment using methane as the sole carbon source. Phylogenetic analysis based on 16S rRNA sequences and comparison of pmoA genes supported the close relationship of MAG-wino1 to the genus Methylomicrobium and it possessed a genome of 5.06 Mb encoding many specialized methanotrophic genes. A comparison of MAG-wino1 with the genomes of other strains (Methylomicrobium alcaliphilum 20Z^T and Methylomicrobium buryatense 5G) showed that genes (e.g. ectABC, ask, and mscLS) involved in the accumulation of compatible solutes required for survival in marine environments might be conserved. Methane utilization genes, including methanol dehydrogenase, and key enzymes related to ribulose monophosphate (RuMP) metabolism were identified. The wino1 genome harbored nitrogen fixation, urease, urea and nitrate transporter genes involved in the exploitation of nitrogen sources. Poly-β-hydroxybutyrate degradation and glycogen synthesis-related genes may facilitate survival under nutrient-limiting conditions. Additionally, genome analysis revealed three dominant taxa in the enrichment culture, methanotroph Methylomicrobium sp., methylotroph Methyloceanibacter sp., and non-methylotroph Labrenzia sp., which provided insights into microbial associations in marine sediments.     

Urease gene‐containing Archaea dominate autotrophic ammonia oxidation in two acid soils

The metabolic traits of ammonia‐oxidizing archaea (AOA) and bacteria (AOB) interacting with their environment determine the nitrogen cycle at the global scale. Ureolytic metabolism has long been proposed as a mechanism for AOB to cope with substrate paucity in acid soil, but it remains unclear whether urea hydrolysis could afford AOA greater ecological advantages. By combining DNA‐based stable isotope probing (SIP) and high‐throughput pyrosequencing, here we show that autotrophic ammonia oxidation in two acid soils was predominately driven by AOA that contain ureC genes encoding the alpha subunit of a putative archaeal urease. In urea‐amended SIP microcosms of forest soil (pH 5.40) and tea orchard soil (pH 3.75), nitrification activity was stimulated significantly by urea fertilization when compared with water‐amended soils in which nitrification resulted solely from the oxidation of ammonia generated through mineralization of soil organic nitrogen. The stimulated activity was paralleled by changes in abundance and composition of archaeal amoA genes. Time‐course incubations indicated that archaeal amoA genes were increasingly labelled by ¹³CO₂ in both microcosms amended with water and urea. Pyrosequencing revealed that archaeal populations were labelled to a much greater extent in soils amended with urea than water. Furthermore, archaeal ureC genes were successfully amplified in the ¹³C‐DNA, and acetylene inhibition suggests that autotrophic growth of urease‐containing AOA depended on energy generation through ammonia oxidation. The sequences of AOB were not detected, and active AOA were affiliated with the marine Group 1.1a‐associated lineage. The results suggest that ureolytic N metabolism could afford AOA greater advantages for autotrophic ammonia oxidation in acid soil, but the mechanism of how urea activates AOA cells remains unclear.