Euryarchaeal genomes are folded into SMC-dependent loops and domains, but lack transcription-mediated compartmentalization.

Hi-C has become a routine method for probing the 3D organization of genomes. However, when applied to prokaryotes and archaea, the current protocols are expensive and limited in their resolution. We develop a cost-effective Hi-C protocol to explore chromosome conformations of these two kingdoms at the gene or operon level. We first validate it on E. coli and V. cholera, generating sub-kilobase-resolution contact maps, and then apply it to the euryarchaeota H. volcanii, Hbt. salinarum, and T. kodakaraensis. With a resolution of up to 1 kb, we explore the diversity of chromosome folding in this phylum. In contrast to crenarchaeota, these euryarchaeota lack (active/inactive) compartment-like structures. Instead, their genomes are composed of self-interacting domains and chromatin loops. In H. volcanii, these structures are regulated by transcription and the archaeal structural maintenance of chromosomes (SMC) protein, further supporting the ubiquitous role of these processes in shaping the higher-order organization of genomes.

Teaching old dogs new tricks: genetic engineering methanogens.

Methanogenic archaea, which are integral to global carbon and nitrogen cycling, currently face challenges in genetic manipulation due to unique physiology and limited genetic tools. This review provides a survey of current and past developments in the genetic engineering of methanogens, including selection and counterselection markers, reporter systems, shuttle vectors, mutagenesis methods, markerless genetic exchange, and gene expression control. This review discusses genetic tools and emphasizes challenges tied to tool scarcity for specific methanogenic species. Mutagenesis techniques for methanogens, including physicochemical, transposon-mediated, liposome-mediated mutagenesis, and natural transformation, are outlined, along with achievements and challenges. Markerless genetic exchange strategies, such as homologous recombination and CRISPR/Cas-mediated genome editing, are also detailed. Finally, the review concludes by examining the control of gene expression in methanogens. The information presented underscores the urgent need for refined genetic tools in archaeal research. Despite historical challenges, recent advancements, notably CRISPR-based systems, hold promise for overcoming obstacles, with implications for global health, agriculture, climate change, and environmental engineering. This comprehensive review aims to bridge existing gaps in the literature, guiding future research in the expanding field of archaeal genetic engineering.

Evolutionary considerations of the oligosaccharyltransferase AglB and other aspects of N-glycosylation across Archaea.

Various biological markers in members of the TACK and Asgard archaeal super-phyla show Eukarya-like traits. These include the oligosaccharyltransferase, responsible for transferring glycans from the lipid carrier upon which they are assembled onto selected asparagine residues of target proteins during N-glycosylation. In Archaea, oligosaccharyltransferase activity is catalyzed by AglB. To gain deeper insight into AglB and N-glycosylation across archaeal phylogeny, bioinformatics approaches were employed to address variability in AglB sequence motifs involved in enzyme activity, construct a phylogenetic tree based on AglB sequences, search for archaeal homologues of non-catalytic subunits of the multimeric eukaryal oligosaccharyltransferase complex and predict the presence of aglB-based clusters of glycosylation-related genes in the Euryarchaeota and the DPANN, TACK and Asgard super-phyla. In addition, site-directed mutagenesis and mass spectrometry were employed to study the natural variability in the WWDXG motif central to oligosaccharyltransferase activity seen in archaeal AglB. The results clearly distinguish AglB from members of the DPANN super-phylum and the Euryarchaeota from the same enzyme in members of the TACK and Asgard super-phyla, which showed considerable similarity to its eukaryal homologue Stt3. The results thus support the evolutionary proximity of Eukarya and the TACK and Asgard archaea.

Unveiling the Biodiversity of Hyperthermophilic Archaea in Jharia Coal Mines: Potential Threat to Methanogenesis?

AIM: To examine the biodiversity of archaeal sulfate reducers and methanogens present in the underground coal mines of Jharia using metagenomics and pyrosequencing. OBJECTIVES: 1) Bioinformatical analysis of the metagenomic data related to a taxonomic analysis obtained from the coal to investigate complete archaeal taxonomic features of the coal bed methane (CBM) microbiome. 2) Bioinformatical analysis of the metagenomic data related to a functional analysis obtained from the coal to investigate functional features relating to taxonomic diversity of the CBM microbiome. 3) The functional attributes have been examined specifically for ORFs related to sulfite reduction and methanogenesis.The taxonomic and functional biodiversity related to euryarchaeota will help in a better understanding of the obstacles associated with methane production imposed by the sulfate reducers. BACKGROUND: The microbial methanogenesis in the coal microbiome is a resultant of substrate utilization by primarily fermentative bacteria and methanogens. The present work reveals the biodiversity of archaeal sulfate reducers and methanogens present in the underground coal mines of Jharia using metagenomics and pyrosequencing. METHODOLOGY: Bioinformatical analysis for structural and functional attributes was accomplished using MG-RAST. The structural analysis was accomplished using RefSeq database, whereas the functional analysis was done via CoG database with a cut off value, a sequence percent identity, and sequence alignment length cut off of 1e-5, 60% and 45, respectively. RESULTS: Attained communities revealed the dominance of hyperthermophilic archaea Pyrococcus furiosus along with Thermococcus kodakarensis in the coal metagenome.The obtained results also suggest the presence of dissimilatory sulfite reductase and formylmethanofuran dehydrogenase, formylmethanofuran: tetrahydromethanopterin formyltransferase involved in sulfite reduction and methanogenesis, respectively, in the microbiome. CONCLUSION: This report is the first attempt to showcase the existence of specific euryarchaeal diversity and their related functional attributes from Jharia coal mines through high throughput sequencing. The study helps in developing a better understanding of the presence of indigenous microbes (archaea) and their functions in the coal microbiome, which can be utilized further to resolve the energy crisis.

Elucidating functions of DP1 and DP2 subunits from the Thermococcus kodakarensis family D DNA polymerase.

DNA polymerase D (PolD), originally discovered in Pyrococcus furiosus, has no sequence homology with any other DNA polymerase family. Genes encoding PolD are found in most of archaea, except for those archaea in the Crenarchaeota phylum. PolD is composed of two proteins: DP1 and DP2. To date, the 3D structure of the PolD heteromeric complex is yet to be determined. In this study, we established a method that prepared highly purified PolD from Thermococcus kodakarensis, and purified DP1 and DP2 proteins formed a stable complex in solution. An intrinsically disordered region was identified in the N-terminal region of DP1, but the static light scattering analysis provided a reasonable molecular weight of DP1. In addition, PolD forms as a complex of DP1 and DP2 in a 1:1 ratio. Electron microscope single particle analysis supported this composition of PolD. Both proteins play an important role in DNA synthesis activity and in 3'-5' degradation activity. DP1 has extremely low affinity for DNA, while DP2 is mainly responsible for DNA binding. Our work will provide insight and the means to further understand PolD structure and the molecular mechanism of this archaea-specific DNA polymerase.

The Tubulin Superfamily in Archaea.

In comparison with bacteria and eukaryotes, the large and diverse group of microorganisms known as archaea possess a great diversity of cytoskeletal proteins, including members of the tubulin superfamily. Many species contain FtsZ, CetZ and even possible tubulins; however, some major taxonomic groups do not contain any member of the tubulin superfamily. Studies using the model archaeon, Halferax volcanii have recently been instrumental in defining the fundamental roles of FtsZ and CetZ in archaeal cell division and cell shape regulation. Structural studies of archaeal tubulin superfamily proteins provide a definitive contribution to the cytoskeletal field, showing which protein-types must have developed prior to the divergence of archaea and eukaryotes. Several regions of the globular core domain - the "signature" motifs - combine in the 3D structure of the common molecular fold to form the GTP-binding site. They are the most conserved sequence elements and provide the primary basis for identification of new superfamily members through homology searches. The currently well-characterised proteins also all share a common mechanism of GTP-dependent polymerisation, in which GTP molecules are sandwiched between successive subunits that are arranged in a head-to-tail manner. However, some poorly-characterised archaeal protein families retain only some of the signature motifs and are unlikely to be capable of dynamic polymerisation, since the promotion of depolymerisation by hydrolysis to GDP depends on contributions from both subunits that sandwich the nucleotide in the polymer.

Carbon monoxide as a metabolic energy source for extremely halophilic microbes: implications for microbial activity in Mars regolith.

Carbon monoxide occurs at relatively high concentrations (≥800 parts per million) in Mars' atmosphere, where it represents a potentially significant energy source that could fuel metabolism by a localized putative surface or near-surface microbiota. However, the plausibility of CO oxidation under conditions relevant for Mars in its past or at present has not been evaluated. Results from diverse terrestrial brines and saline soils provide the first documentation, to our knowledge, of active CO uptake at water potentials (-41 MPa to -117 MPa) that might occur in putative brines at recurrent slope lineae (RSL) on Mars. Results from two extremely halophilic isolates complement the field observations. Halorubrum str. BV1, isolated from the Bonneville Salt Flats, Utah (to our knowledge, the first documented extremely halophilic CO-oxidizing member of the Euryarchaeota), consumed CO in a salt-saturated medium with a water potential of -39.6 MPa; activity was reduced by only 28% relative to activity at its optimum water potential of -11 MPa. A proteobacterial isolate from hypersaline Mono Lake, California, Alkalilimnicola ehrlichii MLHE-1, also oxidized CO at low water potentials (-19 MPa), at temperatures within ranges reported for RSL, and under oxic, suboxic (0.2% oxygen), and anoxic conditions (oxygen-free with nitrate). MLHE-1 was unaffected by magnesium perchlorate or low atmospheric pressure (10 mbar). These results collectively establish the potential for microbial CO oxidation under conditions that might obtain at local scales (e.g., RSL) on contemporary Mars and at larger spatial scales earlier in Mars' history.

Comparative Analysis of Archaeal Lipid-linked Oligosaccharides That Serve as Oligosaccharide Donors for Asn Glycosylation.

The glycosylation of asparagine residues is the predominant protein modification in all three domains of life. An oligosaccharide chain is preassembled on a lipid-phospho carrier and transferred onto asparagine residues by the action of a membrane-bound enzyme, oligosaccharyltransferase. The oligosaccharide donor for the oligosaccharyl transfer reaction is dolichol-diphosphate-oligosaccharide in Eukaryota and polyprenol-diphosphate-oligosaccharide in Eubacteria. The donor in some archaeal species was reportedly dolichol-monophosphate-oligosaccharide. Thus, the difference in the number of phosphate groups aroused interest in whether the use of the dolichol-monophosphate type donors is widespread in the domain Archaea. Currently, all of the archaeal species with identified oligosaccharide donors have belonged to the phylum Euryarchaeota. Here, we analyzed the donor structures of two species belonging to the phylum Crenarchaeota, Pyrobaculum calidifontis and Sulfolobus solfataricus, in addition to two species from the Euryarchaeota, Pyrococcus furiosus and Archaeoglobus fulgidus The electrospray ionization tandem mass spectrometry analyses confirmed that the two euryarchaeal oligosaccharide donors were the dolichol-monophosphate type and newly revealed that the two crenarchaeal oligosaccharide donors were the dolichol-diphosphate type. This novel finding is consistent with the hypothesis that the ancestor of Eukaryota is rooted within the TACK (Thaum-, Aig-, Cren-, and Korarchaeota) superphylum, which includes Crenarchaea. Our comprehensive study also revealed that one archaeal species could contain two distinct oligosaccharide donors for the oligosaccharyl transfer reaction. The A. fulgidus cells contained two oligosaccharide donors bearing oligosaccharide moieties with different backbone structures, and the S. solfataricus cells contained two oligosaccharide donors bearing stereochemically different dolichol chains.

Cultivation of moonmilk-born non-extremophilic Thaum and Euryarchaeota in mixed culture.

PCR-DGGE, qPCR and sequencing highlighted a quite homogenous archaeal community prevailing in secondary calcite deposits, so-called moonmilk, within the cold alpine Hundalm cave in Tyrol (Austria). Furthermore, the depth profile of this moonmilk could prove that the Archaea are located in oxygen-rich near- and oxygen-depleted sub-surface layers. To gather these communities we therefore applied an aerobic and anaerobic cultivation approach in oligotrophic and methanotrophic media. The mixed moonmilk community was analyzed with a combination of molecular methods using qPCR, PCR-DGGE and sequencing. Anaerobic and aerobic cultures were additionally investigated with GC and HPLC analyses. It was possible to initially cultivate and enrich the supposed aerobic/microaerophilic and anaerobic archaeal fraction, representing the natural archaeal community. While the naturally less abundant near-surface Archaea are closely related to members of the Thaumarchaeota (Nitrosopumilus maritimus), the highly abundant anaerobic Archaea are more distantly related to members within the Euryarchaeota. It is possible that these cultivable moonmilk-born Archaea represent new ecotypes or are so far undescribed. Based on the sequencing results and the production of very low amounts of methane, a corresponding methanogenic community is thought to represent only a minor abundant archaeal fraction. On a physiological level the cultivated moonmilk community is cold-adapted and basically of oligotrophic and organotrophic character.

Draft genome sequence of Thermococcus waiotapuensis WT1T, a thermophilic sulfur-dependent archaeon from the order Thermococcales.

Thermococcus waiotapuensis WT1T is a thermophilic, peptide, and amino acid-fermenting archaeon from the order Thermococcales. It was isolated from Waiotapu, Aotearoa-New Zealand, and has a genome size of 1.80 Mbp. The genome contains 2,000 total genes, of which 1,913 encode proteins and 46 encode tRNA.

Temporal dynamics of volatile fatty acids profile, methane production, and prokaryotic community in an in vitro rumen fermentation system fed with maize silage.

Anaerobic in vitro fermentation is widely used to simulate rumen kinetics and study the microbiome and metabolite profiling in a controlled lab environment. However, a better understanding of the interplay between the temporal dynamics of fermentation kinetics, metabolic profiles, and microbial composition in in vitro rumen fermentation batch systems is required. To fill that knowledge gap, we conducted three in vitro rumen fermentations with maize silage as the substrate, monitoring total gas production (TGP), dry matter degradability (dDM), and methane (CH4) concentration at 6, 12, 24, 36, and 48 h in each fermentation. At each time point, we collected rumen fluid samples for microbiome analysis and volatile fatty acid (VFA) analysis. Amplicon sequencing of 16S rRNA genes (V4 region) was used to profile the prokaryotic community structure in the rumen during the fermentation process. As the fermentation time increased, dDM, TGP, VFA concentrations, CH4 concentration, and yield (mL CH4 per g DM at standard temperature and pressure (STP)) significantly increased. For the dependent variables, CH4 concentration and yield, as well as the independent variables TGP and dDM, polynomial equations were fitted. These equations explained over 85% of the data variability (R2 > 0.85) and suggest that TGP and dDM can be used as predictors to estimate CH4 production in rumen fermentation systems. Microbiome analysis revealed a dominance of Bacteroidota, Cyanobacteria, Desulfobacterota, Euryarchaeota, Fibrobacterota, Firmicutes, Patescibacteria, Proteobacteria, Spirochaetota, and Verrucomicrobiota. Significant temporal variations in Bacteroidota, Campylobacterota, Firmicutes, Proteobacteria, and Spirochaetota were detected. Estimates of alpha diversity based on species richness and the Shannon index showed no variation between fermentation time points. This study demonstrated that the in vitro fermentation characteristics of a given feed type (e.g., maize silage) can be predicted from a few parameters (CH4 concentration and yield, tVFA, acetic acid, and propionic acid) without running the actual in vitro trial if the rumen fluid is collected from similar donor cows. Although the dynamics of the rumen prokaryotes changed remarkably over time and in accordance with the fermentation kinetics, more time points between 0 and 24 h are required to provide more details about the microbial temporal dynamics at the onset of the fermentation.

Influence of aphotic haloclines and euxinia on organic biomarkers and microbial communities in a thalassohaline and alkaline volcanic crater lake.

Studies on microbial communities, and their associated organic biomarkers, that are found thriving in the aphotic euxinic waters in modern stratified ecosystems are scarce compared to those undertaken in euxinic photic zones. The Dziani Dzaha (Mayotte, Indian Ocean) is a tropical, saline, alkaline crater lake that has recently been presented as a modern analog of Proterozoic Oceans due to its thalassohaline classification (having water of marine origin) and specific biogeochemical characteristics. Continuous intense photosynthetic production and microbial mineralization keep most of the water column permanently aphotic and anoxic preventing the development of a euxinic (sulfidic and anoxic) photic zone despite a high sulfide/sulfate ratio and the presence of permanent or seasonal haloclines. In this study, the molecular composition of the organic matter in Lake Dziani Dzaha was investigated and compared to the microbial diversity evaluated through 16S rRNA gene amplicon sequencing, over two contrasting seasons (rainy vs. dry) that influence water column stratification. Depth profiles of organic biomarker concentrations (chlorophyll-a and lipid biomarkers) and bacterial and archaeal OTU abundances appeared to be strongly dependent on the presence of aphotic haloclines and euxinia. OTU abundances revealed the importance of specific haloalkaliphilic bacterial and archaeal assemblages in phytoplanktonic biomass recycling and the biogeochemical functioning of the lake, suggesting new haloalkaline non-phototrophic anaerobic microbial precursors for some of the lipid biomarkers. Uncultured Firmicutes from the family Syntrophomonadaceae (Clostridiales), and Bacteroidetes from the ML635J-40 aquatic group, emerged as abundant chemotrophic bacterial members in the anoxic or euxinic waters and were probably responsible for the production of short-chain n-alkenes, wax esters, diplopterol, and tetrahymanol. Halocline-dependent euxinia also had a strong impact on the archaeal community which was dominated by Woesearchaeota in the sulfide-free waters. In the euxinic waters, methanogenic Euryarchaeota from the Methanomicrobia, Thermoplasmata, and WSA2 classes dominated and were likely at the origin of common hydrocarbon biomarkers of methanogens (phytane, pentamethyl-eicosenes, and partially hydrogenated squalene).

ADP-dependent glucose/glucosamine kinase from Thermococcus kodakarensis: cloning and characterization.

The genome sequence of Thermococcus kodakarensis contains an open reading frame, TK1110, annotated as ADP-dependent glucokinase. The encoding gene was expressed in Escherichia coli and the gene product, TK-GLK, was produced in soluble and active form. The recombinant enzyme was extremely thermostable. Thermostability was increased significantly in the presence of ammonium sulfate. ADP was the preferred co-factor for TK-GLK, which could be replaced with CDP but with a 60% activity. TK-GLK was a metal ion-dependent enzyme which exhibited glucokinase, glucosamine kinase and glucose 6-phosphatase activities. It catalyzed the phosphorylation of both glucose and glucosamine with nearly the same rate and affinity. The apparent Km values for glucose and glucosamine were 0.48 ± 0.03 and 0.47 ± 0.09 mM, respectively. The catalytic efficiency (kcat/Km) values against these two substrates were 6.2 × 105 ± 0.25 and 5.8 × 105 ± 0.75 M-1 s-1. The apparent Km value for dephosphorylation of glucose 6-phosphate was ~14-fold higher than that of glucose phosphorylation. Similarly, catalytic efficiency (kcat/Km) for phosphatase reaction was ~19-fold lower than that for the kinase reaction. To the best of our knowledge, this is the first report that describes the reversible nature of a euryarchaeal ADP-dependent glucokinase.

Influence of Environmental Factors on the Variability of Archaeal Communities in a Karst Wetland.

Archaea are ubiquitous and play an important role in elemental cycles in Earth's biosphere; but little is known about their diversity, distribution, abundance, and impact in karst environments. The present study investigated the effect of environmental factors on the variability of archaeal communities in the sediment of the Huixian karst wetland, the largest karst wetland in South China. Sediment cores were obtained from four sampling sites with different water depths and macrophyte inhabitants in both the winter of 2016 and the summer of 2018. The community analysis was based on PacBio sequencing and quantitative PCR of the archaeal 16S rRNA gene. The results showed that Euryarchaeota (57.4%) and Bathyarchaeota (38.7%) were dominant in all the samples. Methanogenic Methanosarcinales (25.1%) and Methanomicrobiales (13.7%), and methanotrophic archaea ANME-2d (9.0%) were the dominant Euryarchaeota; MCG-11 (16.5%), MCG-6 (9.1%), and MCG-5b (5.5%) were the dominant Bathyarchaeota. The community composition remained stable between summer and winter, and the vertical distributions of the archaeal phyla conformed to two patterns among the four sampling sites. In the winter samples, the archaeal 16S rRNA gene abundance was approximately 1.0E+10 copies/g of wet sediment and the Shannon index was 7.3±5, which were significantly higher than in the summer samples and in other karst environments. A correlation analysis showed that the moisture content and pH were the factors that mostly affected the archaeal communities. The prevalence of nitrate in the summer may be a key factor causing a significant decrease in archaeal abundance and diversity. Two features specific to karst environments, calcium-richness and weak alkalescence of the water supplies, may benefit the prevalence of bathyarchaeotal subgroups MCG-11, MCG-5b, and MCG-6. These results suggest that in karst wetlands, most of the archaea belong to clades that have significant roles in carbon turnover; their composition remains stable, but their abundance and diversity vary significantly from season to season.

Crop rotation reduces the frequency of anaerobic soil bacteria in Red Latosol of Brazil.

Crop diversity affects the processes of soil physical structuring and most likely provokes changes in the frequencies of soil microbial communities. The study was conducted for soil prokaryotic diversity sequencing 16S rDNA genes from a 25-year no-tillage experiment comprised of two crop systems: crop succession (Triticum aestivum-Glycine max) and rotation (Vicia sativa-Zea mays-Avena sativa-Glycine max-Triticum aestivum-Glycine max). The hypothesis was that a crop system with higher crop diversification (rotation) would affect the frequencies of prokaryotic taxa against a less diverse crop system (succession) altering the major soil functions guided by bacterial diversity. Soils in both crop systems were dominated by Proteobacteria (31%), Acidobacteria (23%), Actinobacteria (10%), and Gemmatimonadetes (7.2%), among other common copiotrophic soil bacteria. Crop systems did not affect the richness and diversity indexes of soil bacteria and soil archaea. However, the crop rotation system reduced only the frequencies of anaerobic metabolism bacteria Chloroacidobacteria, Holophagae, Spirochaetes, Euryarchaeota, and Crenarchaeota. It can be concluded that crop succession, a system that is poorer in root diversity over time, may have conditioned the soil to lower oxygen diffusion and built up ecological niches that suitable for anaerobic bacteria tolerating lower levels of oxygen. On the other hand, it appeared that crop rotation has restructured the soil over the years while enabling copiotrophic aerobic bacteria to dominate the soil ecosystem. The changes prompted by crop succession have implications for efficient soil organic matter decomposition, reduced greenhouse gas emissions, higher root activity, and overall soil productivity, which compromise to agriculture sustainability.

Corrigendum: High Representation of Archaea Across All Depths in Oxic and Low-pH Sediment Layers Underlying an Acidic Stream.

[This corrects the article DOI: 10.3389/fmicb.2020.576520.].

Biochemical characterization of a highly active ADP-dependent phosphofructokinase from Thermococcus kodakarensis.

The genome sequence of Thermococcus kodakarensis contains an open reading frame, TK0376, annotated as ADP-dependent phosphofructokinase belonging to pfkC family. The encoding gene was expressed in Escherichia coli and the gene product was characterized. The recombinant protein was produced in soluble and active form. Phosphofructokinase activity of TK0376 was metal-ion dependent and the highest activity (5090 µmol min-1 mg-1) was found in the presence of Co2+ followed by Mg2+ (3280 µmol min-1 mg-1) at 90°C and pH 7.5. TK0376 preferred ADP as phosphoryl donor, however, it could be replaced by ATP but with a 5-fold lower activity. It catalyzed the phosphorylation of fructose 6-phosphate and dephosphorylation of fructose 1,6-bisphosphate. In addition, it was able to phosphorylate glucose and nucleosides but with a much lower rate compared to that of fructose 6-phosphate. The apparent kcat and Km values against fructose 6-phosphate were 4238 s-1 and 0.74 mM, respectively. The rate of dephosphorylation of fructose 1,6-bisphosphate was 3-times lower at 50°C than the phosphorylation of fructose 6-phosphate. Similarly, the rate of phosphorylation of glucose was 450-fold lower than that of fructose 6-phosphate. Phosphofructokinase activity was not allosterically regulated, but it was slightly enhanced by phosphoenol pyruvate, and inhibited by ATP and AMP in a competitive manner.

Simulation of Enhanced Growth of Marine Group II Euryarchaeota From the Deep Chlorophyll Maximum of the Western Pacific Ocean: Implication for Upwelling Impact on Microbial Functions in the Photic Zone.

Mesoscale eddies can have a strong impact on regional biogeochemistry and primary productivity. To investigate the effect of the upwelling of seawater by western Pacific eddies on the composition of the active planktonic marine archaeal community composition of the deep chlorophyll maximum (DCM) layer, mesoscale cold-core eddies were simulated in situ by mixing western Pacific DCM layer water with mesopelagic layer (400 m) water. Illumina sequencing of the 16S rRNA gene and 16S rRNA transcripts indicated that the specific heterotrophic Marine Group IIb (MGIIb) taxonomic group of the DCM layer was rapidly stimulated after receiving fresh substrate from 400 m water, which was dominated by uncultured autotrophic Marine Group I (MGI) archaea. Furthermore, niche differentiation of autotrophic ammonia-oxidizing archaea (MGI) was demonstrated by deep sequencing of 16S rRNA, amoA, and accA genes, respectively. Similar distribution patterns of active Marine Group III (MGIII) were observed in the DCM layer with or without vertical mixing, indicating that they are inclined to utilize the substrates already present in the DCM layer. These findings underscore the importance of mesoscale cyclonic eddies in stimulating microbial processes involved in the regional carbon cycle.

Marine Group II Euryarchaeota Contribute to the Archaeal Lipid Pool in Northwestern Pacific Ocean Surface Waters.

Planktonic archaea include predominantly Marine Group I Thaumarchaeota (MG I) and Marine Group II Euryarchaeota (MG II), which play important roles in the oceanic carbon cycle. MG I produce specific lipids called isoprenoid glycerol dibiphytanyl glycerol tetraethers (GDGTs), which are being used in the sea surface temperature proxy named TEX86. Although MG II may be the most abundant planktonic archaeal group in surface water, their lipid composition remains poorly characterized because of the lack of cultured representatives. Circumstantial evidence from previous studies of marine suspended particulate matter suggests that MG II may produce both GDGTs and archaeol-based lipids. In this study, integration of the 16S rRNA gene quantification and sequencing and lipid analysis demonstrated that MG II contributed significantly to the pool of archaeal tetraether lipids in samples collected from MG II-dominated surface waters of the Northwestern Pacific Ocean (NWPO). The archaeal lipid composition in MG II-dominated NWPO waters differed significantly from that of known MG I cultures, containing relatively more 2G-OH-, 2G- and 1G- GDGTs, especially in their acyclic form. Lipid composition in NWPO waters was also markedly different from MG I-dominated surface water samples collected in the East China Sea. GDGTs from MG II-dominated samples seemed to respond to temperature similarly to GDGTs from the MG I-dominated samples, which calls for further study using pure cultures to determine the exact impact of MG II on GDGT-based proxies.