Thaumarchaeota from deep-sea methane seeps provide novel insights into their evolutionary history and ecological implications

Abstract

Background: Ammonia-oxidizing archaea (AOA) of the phylum Thaumarchaeota mediate the rate-limiting step of nitrification and remove the ammonia that inhibits the aerobic metabolism of methanotrophs. However, the AOA that inhabit deep-sea methane-seep surface sediments (DMS) are rarely studied. Here, we used global DMS metagenomics and metagenome-assembled genomes (MAGs) to investigate the metabolic activity, evolutionary history, and ecological contributions of AOA. Expression of AOA-specific ammonia-oxidizing gene (amoA) was examined in the sediments collected from the South China Sea (SCS) to identify their active ammonia metabolism in the DMS.

Results: Our analysis indicated that AOA contribute > 75% to the composition of ammonia-utilization genes within the surface layers (above 30 cm) of global DMS. The AOA-specific ammonia-oxidizing gene was actively expressed in the DMS collected from the SCS. Phylogenomic analysis of medium-/high-quality MAGs from 18 DMS-AOA indicated that they evolved from ancestors in the barren deep-sea sediment and then expanded from the DMS to shallow water forming an amoA-NP-gamma clade-affiliated lineage. Molecular dating suggests that the DMS-AOA origination coincided with the Neoproterozoic oxidation event (NOE), which occurred ~ 800 million years ago (mya), and their expansion to shallow water coincided with the Sturtian glaciation (~ 713 mya). Comparative genomic analysis suggests that DMS-AOA exhibit higher requirement of carbon source for protein synthesis with enhanced genomic capability for osmotic regulation, motility, chemotaxis, and utilization of exogenous organic compounds, suggesting it could be more heterotrophic compared with other lineages.

Conclusion: Our findings provide new insights into the evolutionary history of AOA within the Thaumarchaeota, highlighting their critical roles in nitrogen cycling in the global DMS ecosystems. Video Abstract.

Keywords: Ammonia-oxidizing archaea; Deep-sea sediment; Genomic evolution; Methane seep.

Fig. 1

Geographical distribution of the sediment sampling sites. Maps to show the eight locations where surface sediment samples were collected and described previously (left) and the nine South China Sea stations where the deep-sea methane-seep sediment-surface samples were collected in this study (right). The locations of the various methane-seep sites are indicated by the site name and the abbreviation along with the water depth in meters below sea level (m)

Fig. 2

The isotopic signal and distribution of Thaumarchaeota in the DMS. A, B The isotopic signal of δ¹⁵N ‰ (nitrogen) and δ¹³C ‰ (carbon) in the short push cores (A) and the stable isotopic signal of δ¹⁵N ‰ (NH₃) and δ¹³C ‰ (CH₄) in the long push cores and non-seep push core (B) were collected from the SCS. C, D The relative abundance of Thaumarchaeota in gas hydrate seeps (C) and other types of methane seep collected globally (D). The relative abundance was calculated using the extracted 16S rRNA gene from each metagenomic dataset. E The relative abundance of genes involved in ammonia oxidation (both AOA and AOB affiliated) and anammox in each metagenomic dataset collected globally. mbsf, meters below the seafloor

Fig. 3

Phylogenomic tree of Thaumarchaeota (A) and phylogenetic tree of the amoA gene (B). The phylogenomic tree of Thaumarchaeota was constructed with 122 conservative genes. The Thaumarchaeota contain the terrestrial, deep-water, shallow-water (NP-epsilon), deep-sea sediment, methane-seep, and shallow-water (NP-gamma) groups, as well as genomes from the sediment of the Mariana Trench

Fig. 4

Evolutionary timeline of the Thaumarchaeota. The timeline was estimated using MCMCTree on top of a rooted ML concatenated tree consisting of Aigarchaeota, Crenarchaeota, and Euryarchaeota I, as well as various AOA and non-AOA Thaumarchaeota, with 62 conserved genes. Nodes with known time constraints are labeled with a transparent dark circle, and these are also listed in supplementary Table S4. The two blue vertical bars represent the timing of the Sturtian and Marinoan glacial periods, whereas the single green vertical bar represents the timing of the Neoproterozoic Oxygenation Event (NOE). The concentration of ancient atmospheric oxygen is represented as a percentage of the present atmospheric level; these data were retrieved from the TimeTree website (http://www.timetree.org/)

Fig. 5

The phylogenetic pattern of genes for environmental adaptation. The phylogenomic tree (left panel) was generated with GTDB-Tk using 122 conserved marker genes. In the heatmap (right panel), the intensity of red represents the copy number of the selected genes. These are as follows: speE, spermidine synthase; gcvH, glycine cleavage system proteins H; kat, putative 3-aminobutyric-CoA aminotransferase; proDH, proline dehydrogenase; metH, metH methionine synthase II (cobalamin independent); FDH, formate dehydrogenase; rocA, 1-pyrroline-5-carboxylate dehydrogenase; Kal, 3-aminobutyryl-CoA ammonia-lyase; LarA, lactate racemase; pgi, phosphoglucose isomerase; pcm, protein-L-isoaspartate carboxyl methyltransferase; alkA, DNA-3-methyladenine glycosylase; udg4, uracil DNA glycosylase family 4; mpg, methylpurine/alkyladenine-DNA glycosylase; ogg, 8-oxoguanine DNA glycosylase; radA, DNA repair protein RadA/Sms; polD, DNA polymerase; Amt, urease subunit gamma; nit2, nitrilase/omega-amidase; ipct/dipps bifunctional CTP, inositol-1-phosphate; CAP1, the monovalent cation proton antiporter-1 (CPA1) family; CPA2, the monovalent cation proton antiporter-2 (CPA2) family; CheA and CheB, chemotactic sensor histidine kinase CheA and methylesterase CheB; CheY, chemotaxis response regulator; cspC, cold-shock protein; cshA, cold-shock DEAD-box protein A; FlaJ, archaeal preflagellin peptidases; HAAT, the hydrophobic amino acid uptake transporter; APC, amino acid-polyamine-organocation transporter family; mpnS, methylphosphonate synthase; hcd, 4-hydroxybutyryl-CoA dehydratase

Fig. 6

The phylogenetic pattern of genes for transporters. The phylogenomic tree (left panel) was generated with GTDB-Tk using 122 conserved marker genes. In the heatmap (right panel), the intensity of blue represents the number of genes involved in the selected transporter families. The detailed information of these families is listed in Supplementary Table S9

Fig. 7

Basic metabolic modules for the carbon and energy metabolism pathways of the DMS-AOA. The pie chart indicates the percentage of DMS-AOA detected with the gene. A complete list of genes in the metabolic pathways can be found in Table S11. 1, acetyl-CoA carboxylase; 2, malonyl-CoA reductase; 3, malonate semialdehyde reductase; 4, 3-hydroxypropionyl-CoA synthetase; 5, 3-hydroxypropionyl-CoA dehydratase; 6, acryloyl-CoA reductase; 7, propionyl-CoA carboxylase; 8, methylmalonyl-CoA epimerase; 9, methylmalonyl-CoA mutase; 10, succinyl-CoA reductase; 11, succinate semialdehyde reductase; 12, 4-hydroxybutyryl-CoA synthetase; 13, 4-hydroxybutyryl-CoA dehydratase; 14, crotonyl-CoA hydratase; 15, 3-hydroxybutyryl-CoA dehydrogenase; 16, acetoacetyl-CoAβ-ketothiolase