No bathyarchaeotal species have as yet been successfully cultured in pure cultures, despite their widespread distribution in the marine, terrestrial and limnic environments (Kuboetal.2012), which hampers their direct physiological characterization. Combined with the aforementioned specific heterotrophic metabolic potentials of members within bathyarchaeotal subgroups and their occurrence in sediment layers of distinct biogeochemical properties (Lazaretal.2015), it was proposed that the acquisition of diverse physiological capacities by Bathyarchaeota is driven by adaptation to specific habitats rather than there being a common metabolic capacity. Lineage (full): cellular organisms; Archaea; TACK group. The archaeal phylum Bathyarchaeota, which is composed of a large number of diverse lineages, is widespread and abundant in marine sediments. Genomic inferences from the two reconstructed bathyarchaeotal genomic bins from the coal-bed methane wells suggest that some Bathyarchaeota are methylotrophic methanogens feeding on a wide variety of methylated compounds, possessing an additional ability to ferment peptides, glucose and fatty acids (Evansetal.2015). 2). The phylogenetic affiliation of sequences found in peat suggest that members of the thus-far-uncultivated group Candidatus Bathyarchaeota (representing a fourth phylum) may be involved in methane cycling, either anaerobic oxidation of methane and/or methanogenesis, as at least a few organisms within this group contain the essential To increase the permeability of the cell wall and obtain a good amplification signal, a 10-min 0.01 M HCl treatment may be employed (Kuboetal.2012). Several pre-/non-enriched sediment cultures afforded preliminary evidence for the trophic properties and metabolic capacities of Bathyarchaeota. The Subgroups-1, -6 and -15 genomes also encoded the methyl glyoxylate pathway, which is typically activated when slow-growing cells are exposed to an increased supply of sugar phosphates (Weber, Kayser and Rinas 2005). Together with evidence of few phylogenetic changes throughout the incubation, it was suggested that the microbial community detected by stable isotopic probing could serve well in reflecting the metabolically active components. Recent data point to the global occurrence of Bathyarchaeota and their potential impact on global carbon transformation, highlighting their important role as a group of global generalists participating in carbon cycling, similar to euryarchaeotal methanogens and Thaumarchaeota. Markers for individual pathway/function were scanned against genomes using the HMM and KEGG databases (Anantharamanetal.2016; Kanehisa, Sato and Morishima 2016; Spang, Caceres and Ettema 2017). Members of Bathyarchaeota are able to use CO2 and H2 from natural sources and fermentation products to fuel acetogenesis (Heetal.2016; Martinetal.2016). Metabolic pathways of the Peptidases targeting d-amino acids, which are highly enriched in the peptidoglycan of bacterial cell walls, are encoded as well, indicating that Bathyarchaeota may have acquired the capacity to degrade recalcitrant components of bacterial cell walls, i.e. Amend JP, McCollom TM, Hentscher M et al. On the other hand, the subgroups MCG-18 and MCG-19 were also named in Fillol et al.s research (Filloletal.2016). Four major heterotrophic pathways centralized on the acetyl-CoA generation are summarized below, reflecting the core metabolism of fermentation and acetogenesis (Fig. Evans PN, Parks DH, Chadwick GL et al. Results In the current study, nine It harbors methyl-coenzyme M reductase (MCR)-encoding genes, and many identified and unidentified methyltransferase-encoding genes for the utilization of various methylated compounds, but lacks most of the genes encoding the subunits of Na+-translocating methyl-H4MPT:coenzyme M methyltransferase, suggesting that the organism does not engage in hydrogenotrophic methanogenesis. Combined with the large amount of carbon deposited in the subseafloor (ca 15 1021 g) (Fryetal.2008), the high abundance of MCG archaea in marine sediments (10100% of total archaeal abundance) (Parkesetal.2005; Biddleetal.2006; Fryetal.2008; Kuboetal.2012; Lloydetal.2013) and their heterotrophic properties on detrital proteins, acetate, aromatic compounds and/or other organic substrates (Biddleetal.2006; Websteretal.2010; Websteretal.2011; Lloydetal.2013; Naetal.2015), naturally led to the proposal that this group of archaea may play an important role in global carbon biogeochemical cycling (Kuboetal.2012; Lloydetal.2013; Filloletal.2016; Heetal.2016). The metabolic properties are also considerably diverse based on genomic analysis (Fig. with 12C-acetate added); this indicated that the acetate might participate in microbial biosynthesis rather than being used for energy production (Naetal.2015). Co-occurrence networks in the archaeal clone libraries indicated the role of Bathyarchaeota as keystone species, and suggested their function in maintaining the stability and adaptability of the archaeal community (Xiangetal.2017). The central product, acetyl-CoA, would either (i) be involved in substrate-level phosphorylation to generate acetate and ATP, catalyzed by an ADP-forming acetyl-CoA synthase as in other peptide-degrading archaea; (ii) be metabolized to generate acetate through the Pta-Ack pathway, similarly to bona fide bacterial homoacetogens; or (iii) be utilized for biosynthesis, e.g. Later on, members of Bathyarchaeota were also found to be abundant in deep marine subsurface sediments (Reedetal.2002; Inagakietal.2003), suggesting that this group of archaea is not restricted to terrestrial environments, and the name has been changed to MCG archaea (Inagakietal.2003). BA1 (Subgroup-3) genome contains many genes of the reductive acetyl-CoA (WoodLjungdahl) pathway and key genes of the methane metabolism pathway. Bathyarchaeota was the most abundant archaeal phylum in most samples, accounting for 13.8164.14% of archaeal sequences (Fig. Subgroup-15 was recently found to be enriched in 13C-labeled DNA after a 3-month incubation experiment using sulfate-reducing sediments from Aarhus Bay, but was not present in the corresponding total DNA library or in a control incubation sample (i.e. In summary, there are a total of 25 subgroups of Bathyarchaeota based on all available 16S rRNA gene sequences at this moment, and the former names for each subgroup are also labeled in the tree (Fig. The incorporation of 13C-bicarbonate into the archaeal lipids (potential bathyarchaeotal-specific biphytanes) was significantly observed only with lignin addition. Introduction. The percentages in every row stand for the proportions of subgroups in each environmental category. n. Bathyarchaeota Gender: neuter (iii) The relatively small 13C signature of the archaeal intact polar lipids in comparison with the archaeal biomass suggests that the C isotopic fractionation during lipid biosynthesis is different from that of typical methylotrophic methanogens (Summons, Franzmann and Nichols 1998). That approach revealed an order of magnitude increase in bathyarchaeotal abundance in both the control and experimental groups compared with time zero; however, no significant increase of bathyarchaeotal abundance was observed in experimental groups with substrate additives and various cultivation processing steps, compared with control groups with basal medium alone. bathys, meaning deep as it locates deep branching with Thaumarchaeota and Aigarchaeaota, and frequently detected in the deep subsurface sediments; N.L. Because of their high sequence coverage and bathyarchaeotal sequence specificity, MCG528 and MCG732 primers are recommended for the detection and quantification of Bathyarchaeota (Kuboetal.2012); nevertheless, this primer pair is not suitable for quantifying Bathyarchaeota in freshwater columns and sediments (Filloletal.2015). A segregated distribution of bathyarchaeotal subgroups was also observed in the water column and sediments in freshwater karstic lakes (Filloletal.2015). These results have not only demonstrated multiple and important ecological functions of this archaeal phylum, but also paved the way for a detailed understanding of the evolution and metabolism of archaea as such. WebHost. In addition, the catalyzed reporter deposition-fluorescent in situ hybridization (CARD-FISH) studies for the detection and quantification of bathyarchaeotal cells suggest that they are abundant in the center and marine invertebrate-inhabited layers in the Haakon Mosby Mud Volcano, and in the marine subsurface sediments in the Equatorial ODP site 1125 and Peru Basin ODP site 1231 (Kuboetal.2012). stands for formamide concentration in the hybridization buffer (%, vol/vol). These indicative subgroups are the dominant ones in the environment, as evaluated by relatively abundant fraction of Bathyarchaeota in corresponding archaeal communities (on average 44% among all studies). The current genomic and physiological information of these subgroups also suggests their potential ecological strategies and functions in specific habitats, further highlighting their important roles in global biogeochemical cycling (Xiangetal.2017). RNA slot blot hybridization can also be used for the quantification of functionally active bathyarchaeotal 16S rRNA. Hence, the primer pair MCG242dF and MCG678R was developed based on a collection of bathyarchaeotal sequences of freshwater origin (Filloletal.2016). However, after allowing for a single nucleotide mismatch, the coverage efficiency markedly increased, to around 8090%. However, according to the genomic information on most archaeal acetogens and bathyarchaeotal genomic bins obtained by Lazaretal. Given the diverse and complex phylogeny of the Bathyarchaeota (Kuboetal.2012; Filloletal.2016), the occurrence of commonly shared physiological and metabolic properties in different lineages seems unlikely, with the evolutionary diversification of bathyarchaeotal lineages largely driven by the adaptation to various environmental conditions and available carbon and energy sources, etc. Sequences longer than 940 bp were first used to construct the backbone of the tree, and additional sequences were then added without altering the general tree topology. A group called Peat MCG (pMCG) (Xiangetal.2017) was also listed on the tree; however, because there was only one represented sequence after dereplication at 90% similarity of all bathyarchaeotal 16S rRNA gene sequences, we did not list pMCG as a separate subgroup in this tree (Fig. (A) Phylogenetic tree of ribosomal proteins obtained from currently available bathyarchaeotal genomes (from GenBank, 29 November 2017 updated). The energy landscape of a local environment, i.e. To compare the coverage and specificity of analysis using the qPCR primer pairs MCG242dF/MCG678R and MCG528F/MCG732R for freshwater and marine sediment samples, amplicons obtained with these two primer pairs were analyzed and community structures compared (Filloletal.2015). This study represents the first report on the diversity and spatial distribution of microbial communities in methane-rich tropical shallow water ecosystems, highlighting the most abundant archaea detected, the Bathyarchaeia Class. Study sites and sampling Furthermore, a principal coordinate analysis also clearly separates the bathyarchaeotal community into freshwater and saline sediment groups. The total RNA is blotted onto nylon membranes and subsequently hybridized with 33P-labeled Bathyarchaeota-specific probes (Table 1). They also acquired some subunits of coenzyme F420 hydrogenase; this enzyme generates reduced ferredoxin, with hydrogen as the electron donor, as an alternative to MvhADG in many Methanomicrobiales (Thaueretal.2008; Lazaretal.2016; Sousaetal.2016). (2017) investigated the bathyarchaeotal community in two sediment cores from the South China Sea; the authors revealed a direct strong positive correlation between bathyarchaeotal 16S rRNA gene abundance and total organic carbon content along the core depth, suggesting an overall heterotrophic lifestyle of Bathyarchaeota in the South China Sea. This was confirmed by a permutational analysis of variance, with salinity as the best explanatory variable for the variance within the bathyarchaeotal community (R2 = 0.04, P < 0.001) (Filloletal.2016). (Kuboetal.2012), and the outgroup sequences of Crenarchaeota, YNPFFA group and Korarchaeota were added. Phylogenetic analysis of the Pta and Ack coding sequences in He et al.s study revealed that these genes form a monophyletic clade and are different from all other know sequences, indicating that they evolved independently of the currently known bacterial counterparts (Heetal.2016). Methanogens and acetogenic Clostridia are the most frequent basal-branching archaea and bacteria, respectively, in phylogenetic reconstructions reflecting the descendants of the last universal common ancestor; gene categories proposed for the last universal common ancestor also point to the acetogenic and methanogenic roots, reflecting its autotrophic lifestyle as H2-dependent and N2-fixing, utilizing the WoodLjungdahl pathway and originating from a hydrothermal environmental setting (Weissetal.2016). The metagenome Kubo et al. Multiple genomic and physiological traits of these microorganisms have been coming to light in recent decades with the advent of stable isotope labeling and metagenomic profiling methods. neut. Currently available bathyarchaeotal genomes (from GenBank, 29 November 2017 updated) with 16S rRNA gene sequences were labeled in the tree. WebArchaea are tiny, simple organisms. The assignment of bathyarchaeotal subgroups was made based on either having been formerly defined or being monophyletic, using both distance and maximum-likelihood estimations (Kuboetal.2012). Institute for Advanced Study, Shenzhen University, Shenzhen 518060, People's Republic of China, Laboratory of Environmental Microbiology and Toxicology, School of Biological Sciences, The University of Hong Kong, Pokfulam Road, Hong Kong SAR, People's Republic of China. Because of the universal distribution and predominance of Bathyarchaeota, not only in the marine sediments but also in terrestrial sediments and various other eco-niches, and because of their versatile metabolism (including acetogenesis, methane metabolism, and dissimilatory nitrate and sulfate reduction) and potential interactions with ANME archaea, acetoclastic methanogens and heterotrophic bacteria, the ecological importance of this group of generalists has entered the limelight and needs further exploration. 1) (Heetal.2016; Lazaretal.2016). The potential AOM metabolic capacity of Bathyarchaeota could help to fully address the isotopic relationship between the archaeal biomass and the ambient environmental carbon pools, as follows. The concatenated ribosomal protein (RP) alignment contained 12 RPs, and those genomes with <25% RPs were excluded from tree construction. Regarding the functional properties, metabolic pathway analysis revealed that BA1 is a peptide and glucose fermenter, while BA2 is a fatty-acid oxidizer (Evansetal.2015). (2018) described a predominance of the phylum Bathyarchaeota (now class Bathyarchaeia from phylum Crenarchaeota) in mid-latitude estuaries, CARD-FISH can be utilized for the detection, identification and enumeration of microorganisms in various environments, independently of culturing (Kubota 2013). 2) based on currently available bathyarchaeotal 16S rRNA gene sequences from SILVA SSU 128 by adding the information from pervious publications (Kuboetal.2012; Lazaretal.2015; Filloletal.2016; Heetal.2016; Xiangetal.2017). Further, based on genomic inferences, Evansetal. They include Euryarchaeota, and members of the DPANN and Asgard archaea. adj. However, their life strategies have remained largely elusive. The major bathyarchaeotal community comprises Subgroups-1, -8, -12 and -15, and is relatively stable during the hypoxic/oxic change, thus being independent of the sedimentary chemistry change, such as manganese and iron redox cycling during different seasons (Devereuxetal.2015). the classification proposed by Zhou et al. Several sets of PCR primers and probes have been developed to detect and quantify Bathyarchaeota in natural community (Table 1). Bathyarchaeota are believed to have roles in the carbon cycle in marine systems. In addition, some regions of the bathyarchaeotal genome might have been acquired from bacteria because of the aberrant tetranucleotide frequency in the genomic fragments of Bathyarchaeota and bacterial phylogenetic origins of these genomic fragments (Lietal.2012). Bathyarchaeota, formerly known as the Miscellaneous Crenarchaeotal Group, is a phylum of global generalists that are widespread in anoxic sediments, which host relatively high abundance archaeal communities. Furthermore, the phylogeny of concatenated alignments constituting 12 ribosomal proteins obtained from currently available bathyarchaeotal genomes (from GenBank, 29 November 2017 updated) was also reconstructed, which showed a similar topology to those of 16S rRNA genes with a few exceptions in Subgroup-17 (Fig. It has been suggested that Bathyarchaeota is one of the cosmopolitan groups frequently detected in the freshwater and marine sediments (68% of all sediments analyzed), accounting for a large proportion of the sediment microbial communities (average 36 22%) (Filloletal.2016). The in silico tests revealed that primers MCG528, MCG493, MCG528 and MCG732 cover 87, 79, 44 and 27% of sequences of Subgroups-1 to -12 on average, respectively. 1) (for details see Kuboetal.2012). Until now, WebGiven the wide environmental and phylogenetic diversity of Bathyarchaeota, additional genomes are required to understand the metabolic capabilities of this understudied Among these are Subgroups-1 and -8 with high IndVal values in marine sediments, and Subgroups-5 and -11 with high IndVal values in fresh sediments (Filloletal.2016). Meanwhile, the ability to utilize a wide variety of substrates could have allowed Bathyarchaeota to avoid a direct competition with other substrate specialists, such as methanogens and sulfate reducers; in contrast, organic matter degradation to generate acetate might be more energetically favorable for Bathyarchaeota than for other bacterial acetogens, as the former do not need to invest in ATP to activate formate; subsequently, Bathyarchaeota plays the role of active carbon transformers, especially in the subsurface sediments, to fuel the heterotrophy and acetoclastic methanogenesis processes and facilitate coupled carbon cycling (Fig. However, due to the great diversity of them, there is limited genomic information that accurately encompasses the metabolic potential of the entire archaeal phylum. JCYJ20170818091727570). The reconstructed bathyarchaeotal genomes (except for Subgroup-15) also encode proteins with the ability to import extracellular carbohydrates. Furthermore, evidence of fatty acid and aromatic compound utilization by Bathyarchaeota has been presented (Mengetal.2014; Evansetal.2015; Heetal.2016); these transformations would be supported by the beta-oxidation pathway and a potential anaerobic aromatic compound degradation pathway. Archaea are abundant in lake sediments [14].Particularly, members of the phylum Bathyarchaeota and the class Thermoplasmata are widespread and considered as core generalists in sediment habitats [], where they have been recognized as key players in the carbon cycle [69].Archaea are also common Subgroup-5 thrives in the euxinic bottom water layer, characterized as anoxic and sulfide-rich, with accumulated inorganic and organic reduced compounds; Subgroup-6 is a group of generalists that are adapted to both planktonic and sediment habitats with a wide range of sulfidic conditions. ( 2012) conducted a comprehensive analysis of the biogeographical distribution of Bathyarchaeota and found that it was the dominant archaeal population in anoxic, low-activity subsurface sediments. Third, only limited reports on the distribution patterns of bathyarchaeotal subgroups and the associated environmental factors are available. Background Bathyarchaeota, a newly proposed archaeal phylum, is considered as an important driver of the global carbon cycle. The Bathyarchaeota formerly known as the Miscellaneous Crenarchaeotal Group is an evolutionarily diverse group of microorganisms found in a wide range of Physiological incubation experiments with stable isotopic probing demonstrated that members of Bathyarchaeota are able to assimilate a wide variety of the tested 13C-organic compounds, including acetate, glycine, urea, simple biopolymers (extracted algal lipids) and complex biopolymers (ISOGRO) (Websteretal.2010; Seyler, McGuinness and Kerkhof 2014). Based on the genomic evidence, the authors concluded that some lineages of Bathyarchaeota are similar to bona fide bacterial homoacetogens, with pathways for acetogenesis and fermentative utilization of a variety of organic substrates (Heetal.2016). However, the global methane cycle should be reconsidered since the previously unrecognized methane metabolic capacity appears to be present within such a widespread and abundant phylum. The product, acetate, would then be used by acetate-consuming SRB to benefit the thermodynamic efficiency of AOM. The ability to use a wide range of substrates for energy conservation and biosynthesis, rather than a single reductive acetyl-CoA pathway, enhances the survival of Bathyarchaeota in energy-limited environments (Lazaretal.2016). Subgroup-5 is divided into Subgroups-5a and -5b, each with intragroup similarity >90% according to a maximum-likelihood estimation. Thaumarchaeota MG-I was present in the 12C-DNA library in the corresponding zone but was not detected in the 13C-DNA library, suggesting that these microbes are not able to use 13C-acetate (Websteretal.2010). Recently, two more bathyarchaeotal fosmid clones were screened from estuarine mangrove sediments (Mengetal.2014). Moreover, the carbonyl branch of the WoodLjungdahl pathway might reduce CO2 into acetyl-CoA. During the enriching process with lignin addition, the Subgroup-8 abundance climbed over 10 times compared with the initial stage and became the most dominant archaeal species. Genomic characterization and metabolic potentials of Bathyarchaeota. The possibility of the replacement of the AOM function of ANME by Bathyarchaeota was also suggested by a microbial community composition in a study of the microbial colonization within an artificial micro-niche, basaltic glass imposed by hydrothermal conditions (Callacetal.2013). In this process, methane is not assimilated by Bathyarchaeota but serves as an energy source. 2. (2016) reconstructed six nearly complete bathyarchaeotal genomes (Subgroups-13, -15, -16, -18 and -19) from the Guaymas Basin subsurface sediment. 3A). Ancestral state reconstruction was used to estimate the diversification of bathyarchaeotal lineages previously subjected to the saline/freshwater transition. Materials and methods 2.1. (2012) demonstrated that the developed primers and probes result in poor coverage of Subgroups-13 to -17. The syntrophic relationship between Bathyarchaeota and SRB would be similar to the anaerobic methane-oxidizing archaea (ANME)/SRB consortium, and acetate would be maintained at a low level as a transient intermediate (Boetiusetal.2000; Hinrichs and Boetius 2002). (Fig. Subgroups were assigned from the corresponding 16S rRNA gene phylogenic tree (Fig. WebBathyarchaeota dominated the archaeal interaction network with 82% nodes, 96% edges, and 71% keystone species. Four genomes (Subgroups-1, -6, -7 and -15) were recovered from the sediment metagenome. Kuboetal. However, it has lost the majority of genes involved in the methyl branch of the WoodLjungdahl pathway and also lost energy-conserving complexes, similar to BA1. A meta-analysis of the distribution of sediment archaeal communities towards environmental eco-factors (7098 archaeal operational taxonomic units from 207 sediment sites worldwide) was performed and a multivariate regression tree was constructed to depict the relationship between archaeal lineages and the environmental origin matrix (Filloletal.2016). Yuetal. The discovery of BchG of archaeal origin in the genomic content of Bathyarchaeota also suggests that an archaeon-based photosynthetic pathway might exist in nature, and that photosynthesis might have evolved before the divergence of bacteria and archaea (Mengetal.2009). Subgroup-6 persists in such suboxic, sulfide-depleted shallow sediment layers, while Subgroups-1, -5 and -8 preferentially occur in deeper, more reducing subsurface layers (Lazaretal.2015). OTUs classified within Bathyarchaeota and Chloroflexi (Dehalococcoidia) showed positive correlation with methane concentrations, sediment depth and oxidation-reduction potential. Furthermore, both FISH labeling and intact polar lipid quantification suggest the presence of highly abundant and active bathyarchaeotal cells in the Peru offshore subsurface sediments collected during the Ocean Drilling Program Leg 201 (Biddleetal.2006; Lippetal.2008). Fillol M, Snchez-Melsi A, Gich F et al. Based on the lineage distribution pattern analysis of the archaeal community of seven major eco-niches, it is also evident that the different evolutionary lineages are habitat-specific, and salinity rather than temperature is the primary driving force of the variation of global archaea distribution, with a similar pattern also evident for the global bacterial distribution (Lozupone and Knight 2007; Auguet, Barberan and Casamayor 2010). A recent study found that the refractory aromatic polymer lignin stimulated the growth of Bathyarchaeota (Subgroup-8) and they incorporated CO2 as a carbon source autotrophically and utilized lignin as an energy source (Yuetal.2018). Hallam SJ, Putnam N, Preston CM et al. Bathyarchaeota is characterized by high intragroup diversity, with most subgroups showing within-sequence similarity <92% (Kuboetal.2012; Filloletal.2016). Lomstein BA, Langerhuus AT, DHondt S et al. Abstract. 2012 ). While Subgroups-18 and -19 were named to be consistent with subgroups MCG-18 and MCG-19 as proposed in two previous reports (Lazaretal.2015; Filloletal.2016), Subgroup-20 was renamed to replace the subgroup MCG-19 in Fillol et al.s tree (Filloletal.2016). To cover all bathyarchaeotal subgroups that are characterized by high intragroup diversity while retaining bathyarchaeotal sequence specificity is necessary but challenging. WebBathyarchaeota are abundant in sediments, and they may involve in sedimentary organic matter degradation, acetogenesis, and, potentially, methane metabolism, based on genomics. Following the four treatments, the viable bathyarchaeotal communities mainly comprised Subgroups-4 and -8, thus indicating that these two subgroups could tolerate the initial aerobic conditions (Gagenetal.2013). lipid and amino acid synthesis (Fig. In experiments towards cultivating Bathyarchaeota from the White Oak River estuary sediments, the abundance of Bathyarchaeota in control groups (basal medium) and in experimental groups containing various substrate additives and submitted to various culture processing steps were compared (Gagenetal.2013). Devereux R, Mosher JJ, Vishnivetskaya TA et al. Due to their prevalence in the microbial community, we also performed phylogenetic analysis to understand the closeness of our Bathyarchaeota OTUs with