Genomic Insights into the Ecological Role and Evolution of a Novel Thermoplasmata Order, " Candidatus Sysuiplasmatales"

Abstract

Recent omics studies have provided invaluable insights into the metabolic potential, adaptation, and evolution of novel archaeal lineages from a variety of extreme environments. We utilized a genome-resolved metagenomic approach to recover eight medium- to high-quality metagenome-assembled genomes (MAGs) that likely represent a new order ("Candidatus Sysuiplasmatales") in the class Thermoplasmata from mine tailings and acid mine drainage (AMD) sediments sampled from two copper mines in South China. 16S rRNA gene-based analyses revealed a narrow habitat range for these uncultured archaea limited to AMD and hot spring-related environments. Metabolic reconstruction indicated a facultatively anaerobic heterotrophic lifestyle. This may allow the archaea to adapt to oxygen fluctuations and is thus in marked contrast to the majority of lineages in the domain Archaea, which typically show obligately anaerobic metabolisms. Notably, "Ca. Sysuiplasmatales" could conserve energy through degradation of fatty acids, amino acid metabolism, and oxidation of reduced inorganic sulfur compounds (RISCs), suggesting that they may contribute to acid generation in the extreme mine environments. Unlike the closely related orders Methanomassiliicoccales and "Candidatus Gimiplasmatales," "Ca. Sysuiplasmatales" lacks the capacity to perform methanogenesis and carbon fixation. Ancestral state reconstruction indicated that "Ca. Sysuiplasmatales," the closely related orders Methanomassiliicoccales and "Ca. Gimiplasmatales," and the orders SG8-5 and RBG-16-68-12 originated from a facultatively anaerobic ancestor capable of carbon fixation via the bacterial-type H₄F Wood-Ljungdahl pathway (WLP). Their metabolic divergence might be attributed to different evolutionary paths. IMPORTANCE A wide array of archaea populate Earth's extreme environments; therefore, they may play important roles in mediating biogeochemical processes such as iron and sulfur cycling. However, our knowledge of archaeal biology and evolution is still limited, since the majority of the archaeal diversity is uncultured. For instance, most order-level lineages except Thermoplasmatales, Aciduliprofundales, and Methanomassiliicoccales within Thermoplasmata do not have cultured representatives. Here, we report the discovery and genomic characterization of a novel order, "Ca. Sysuiplasmatales," within Thermoplasmata in extremely acidic mine environments. "Ca. Sysuiplasmatales" are inferred to be facultatively anaerobic heterotrophs and likely contribute to acid generation through the oxidation of RISCs. The physiological divergence between "Ca. Sysuiplasmatales" and closely related Thermoplasmata lineages may be attributed to different evolutionary paths. These results expand our knowledge of archaea in the extreme mine ecosystem.

Keywords: acid mine drainage; ancestral state reconstruction; genome-resolved metagenomics; “Ca. Sysuiplasmatales; ” facultatively anaerobic heterotrophic lifestyle.

FIG 1

Phylogenomic tree of the novel order based on 122 archaeon-specific conserved marker genes, OrthoANI values, and 16S rRNA gene similarity values. White spaces indicate cases where no 16S rRNA gene was identified or the 16S rRNA gene was too short to be used for 16S rRNA sequence similarity calculation. The tree was constructed based on the concatenated alignment using IQ-TREE with 1,000 ultrafast bootstrapping iterations. The concatenated alignment was generated by GTDB-Tk. Support values greater than 75% are shown with black solid circles. Crenarchaeota was selected as the outgroup.

FIG 2

Metabolic potentials of the novel order. Genes identified in “Ca. Sysuiplasma acidicola,” “Ca. Sysuiplasma superficiale,” and “Ca. Sysuiplasma jiujiangense” are represented by light green, orange, and dark green circles, respectively. The copy numbers of each gene in each genome are listed in Data Set S5.

FIG 3

Occurrence of key proteins of interest in the MAGs of the novel order, Methanomassiliicoccales, “Ca. Gimiplasmatales,” RBG-16-68-12, and SG8-5. The MAGs were grouped according to their phylogenetic relationships, and the gene names were grouped by functional categories. The copy numbers of each gene in each genome are provided in Data Set S6. The genes composed of multiple subunits are marked as present if half or more than half of the subunits were identified.

FIG 4

Ancestral genome content reconstruction of Thermoplasmata, including the novel order. The numbers of gene gain and loss events are marked on the nodes and tips of the phylogenomic tree. The COG category information of the gained and lost genes for the key nodes is shown using pie charts. Some key gene gain and lost events are also marked. The complete topology of the phylogenomic tree and ancestral state reconstruction results are shown in Fig. S5. A list of gained and lost genes for the nodes and tips is presented in Data Set S7.