Chemosynthetic symbiont with a drastically reduced genome serves as primary energy storage in the marine flatworm Paracatenula.

Hosts of chemoautotrophic bacteria typically have much higher biomass than their symbionts and consume symbiont cells for nutrition. In contrast to this, chemoautotrophic Candidatus Riegeria symbionts in mouthless Paracatenula flatworms comprise up to half of the biomass of the consortium. Each species of Paracatenula harbors a specific Ca Riegeria, and the endosymbionts have been vertically transmitted for at least 500 million years. Such prolonged strict vertical transmission leads to streamlining of symbiont genomes, and the retained physiological capacities reveal the functions the symbionts provide to their hosts. Here, we studied a species of Paracatenula from Sant'Andrea, Elba, Italy, using genomics, gene expression, imaging analyses, as well as targeted and untargeted MS. We show that its symbiont, Ca R. santandreae has a drastically smaller genome (1.34 Mb) than the symbiont´s free-living relatives (4.29-4.97 Mb) but retains a versatile and energy-efficient metabolism. It encodes and expresses a complete intermediary carbon metabolism and enhanced carbon fixation through anaplerosis and accumulates massive intracellular inclusions such as sulfur, polyhydroxyalkanoates, and carbohydrates. Compared with symbiotic and free-living chemoautotrophs, Ca R. santandreae's versatility in energy storage is unparalleled in chemoautotrophs with such compact genomes. Transmission EM as well as host and symbiont expression data suggest that Ca R. santandreae largely provisions its host via outer-membrane vesicle secretion. With its high share of biomass in the symbiosis and large standing stocks of carbon and energy reserves, it has a unique role for bacterial symbionts-serving as the primary energy storage for its animal host.

Molecular mechanism of sulfur chemolithotrophy in the betaproteobacterium Pusillimonas ginsengisoli SBSA.

Chemolithotrophic sulfur oxidation represents a significant part of the biogeochemical cycling of this element. Due to its long evolutionary history, this ancient metabolism is well known for its extensive mechanistic and phylogenetic diversification across a diverse taxonomic spectrum. Here we carried out whole-genome sequencing and analysis of a new betaproteobacterial isolate, Pusillimonas ginsengisoli SBSA, which is found to oxidize thiosulfate via the formation of tetrathionate as an intermediate. The 4.7 Mb SBSA genome was found to encompass a soxCDYZAXOB operon, plus single thiosulfate dehydrogenase (tsdA) and sulfite : acceptor oxidoreductase (sorAB) genes. Recombination-based knockout of tsdA revealed that the entire thiosulfate is first converted to tetrathionate by the activity of thiosulfate dehydrogenase (TsdA) and the Sox pathway is not functional in this bacterium despite the presence of all necessary sox genes. The ∆soxYZ and ∆soxXA knockout mutants exhibited a wild-type-like phenotype for thiosulfate/tetrathionate oxidation, whereas ∆soxB, ∆soxCD and soxO::KanR mutants only oxidized thiosulfate up to tetrathionate intermediate and had complete impairment in tetrathionate oxidation. The substrate-dependent O2 consumption rate of whole cells and the sulfur-oxidizing enzyme activities of cell-free extracts, measured in the presence/absence of thiol inhibitors/glutathione, indicated that glutathione plays a key role in SBSA tetrathionate oxidation. The present findings collectively indicate that the potential glutathione : tetrathionate coupling in P. ginsengisoli involves a novel enzymatic component, which is different from the dual-functional thiol dehydrotransferase (ThdT), while subsequent oxidation of the sulfur intermediates produced (e.g. glutathione : sulfodisulfane molecules) may proceed via the iterative action of soxBCD .

Nitrate reductases in Hydrogenobacter thermophilus with evolutionarily ancient features: distinctive localization and electron transfer.

Dissimilatory nitrate reductase (NAR) and assimilatory nitrate reductase (NAS) serve as key enzymes for nitrogen catabolism and anabolism in many organisms. We purified NAR and NAS from H. thermophilus, a hydrogen-oxidizing chemolithoautotroph belonging to the phylogenetically deepest branch in the Bacteria domain. Physiological contribution of these enzymes to nitrate respiration and assimilation was clarified by transcriptomic analysis and gene disruption experiments. These enzymes showed several features unreported in bacteria, such as the periplasmic orientation of NAR anchored with a putative transmembrane subunit and the specific electron transfer from a [4Fe-4S]-type ferredoxin to NAS. While some of their enzymatic properties are shared with NARs from archaea and with NASs from phototrophs, phylogenetic analysis indicated that H. thermophilus NAR and NAS have deep evolutionary origins that cannot be explained by a recent horizontal gene transfer event from archaea and phototrophs. These findings revealed the diversity of NAR and NAS in nonphotosynthetic bacteria, and they also implied that the outward orientation of NAR and the ferredoxin-dependent electron transfer of NAS are evolutionarily ancient features preserved in H. thermophilus.

Spatially distinct, temporally stable microbial populations mediate biogeochemical cycling at and below the seafloor in hydrothermal vent fluids.

At deep-sea hydrothermal vents, microbial communities thrive across geochemical gradients above, at, and below the seafloor. In this study, we determined the gene content and transcription patterns of microbial communities and specific populations to understand the taxonomy and metabolism both spatially and temporally across geochemically different diffuse fluid hydrothermal vents. Vent fluids were examined via metagenomic, metatranscriptomic, genomic binning, and geochemical analyses from Axial Seamount, an active submarine volcano on the Juan de Fuca Ridge in the NE Pacific Ocean, from 2013 to 2015 at three different vents: Anemone, Marker 33, and Marker 113. Results showed that individual vent sites maintained microbial communities and specific populations over time, but with spatially distinct taxonomic, metabolic potential, and gene transcription profiles. The geochemistry and physical structure of each vent both played important roles in shaping the dominant organisms and metabolisms present at each site. Genomic binning identified key populations of SUP05, Aquificales and methanogenic archaea carrying out important transformations of carbon, sulfur, hydrogen, and nitrogen, with groups that appear unique to individual sites. This work highlights the connection between microbial metabolic processes, fluid chemistry, and microbial population dynamics at and below the seafloor and increases understanding of the role of hydrothermal vent microbial communities in deep ocean biogeochemical cycles.

Transcription of mcrA Gene Decreases Upon Prolonged Non-flooding Period in a Methanogenic Archaeal Community of a Paddy-Upland Rotational Field Soil.

Methanogenic archaea survive under aerated soil conditions in paddy fields, and their community is stable under these conditions. Changes in the abundance and composition of an active community of methanogenic archaea were assessed by analyzing mcrA gene (encoding α subunit of methyl-coenzyme M reductase) and transcripts during a prolonged drained period in a paddy-upland rotational field. Paddy rice (Oryza sativa L.) was planted in the flooded field and rotated with soybean (Glycine max [L.] Merr.) under upland soil conditions. Soil samples were collected from the rotational plot in the first year, with paddy rice, and in the two successive years, with soybean, at six time points, before seeding, during cultivation, and after harvest as well as from a consecutive paddy (control) plot. By the time that soybean was grown in the second year, the methanogenic archaeal community in the rotational plot maintained high mcrA transcript levels, comparable with those of the control plot community, but the levels drastically decreased by over three orders of magnitude after 2 years of upland conversion. The composition of active methanogenic archaeal communities that survived upland conversion in the rotational plot was similar to that of the active community in the control plot. These results revealed that mcrA gene transcription of methanogenic archaeal community in the rotational field was affected by a prolonged non-flooding period, longer than 1 year, indicating that unknown mechanisms maintain the stability of methanogenic archaeal community in paddy fields last up to 1 year after the onset of drainage.

Specific detection of form IA RubisCO genes in chemoautotrophic bacteria.

The analysis of RubisCO genes is a highly useful instrument to explore the diversity of chemoautotrophic bacteria using the Calvin-Benson-Bassham cycle for CO2 fixation. However, because of the wide taxonomic distribution of phylogenetically related RubisCO forms, environmental studies targeting chemoautotrophs are hampered in habitats dominated by phototrophs. Here, we report the development of a gene marker that specifically detects form IA RubisCO genes in bacteria, excluding photoautotrophic representatives. The high specificity of the PCR assay was confirmed by sequence analysis of DNA obtained from the photic zone of six lakes, were chemoautotrophs are outnumbered by Cyanobacteria also using form IA RubisCO for CO2 assimilation.

In-situ incubation of iron-sulfur mineral reveals a diverse chemolithoautotrophic community and a new biogeochemical role for Thiomicrospira.

Sulfide mineral precipitation occurs at mid-ocean ridge (MOR) spreading centers, both in the form of plume particles and seafloor massive sulfide structures. A common constituent of MOR is the iron-bearing sulfide mineral pyrrhotite, which was chosen as a substrate for in-situ incubation studies in shallow waters of Catalina Island, CA to investigate the colonization of iron-oxidizing bacteria. Microbial community datasets were obtained from in-situ incubated pyrrhotite, allowing for direct comparison to microbial communities of iron-sulfides from active and inactive chimneys in deep-sea environments. Unclassified Gammaproteobacteria and Alphaproteobacteria (Magnetovibrio) largely dominated the bacterial community on pyrrhotite samples incubated in the water column while samples incubated at the surface sediment showed more even dominance by Deltaproteobacteria (Desulfobulbus), Gammaproteobacteria (Piscirickettsiaceae), Alphaproteobacteria (Rhodobacteraceae), and Bacteroidetes (Flavobacteriia). Cultivations that originated from pyrrhotite samples resulted in the enrichment of both, sheath-forming and stalk-forming Zetaproteobacteria. Additionally, a putative novel species of Thiomicrospira was isolated and shown to grow autotrophically with iron, indicating a new biogeochemical role for this ubiquitous microorganism.

Microbiological oxidation of antimony(III) with oxygen or nitrate by bacteria isolated from contaminated mine sediments.

Bacterial oxidation of arsenite [As(III)] is a well-studied and important biogeochemical pathway that directly influences the mobility and toxicity of arsenic in the environment. In contrast, little is known about microbiological oxidation of the chemically similar anion antimonite [Sb(III)]. In this study, two bacterial strains, designated IDSBO-1 and IDSBO-4, which grow on tartrate compounds and oxidize Sb(III) using either oxygen or nitrate, respectively, as a terminal electron acceptor, were isolated from contaminated mine sediments. Both isolates belonged to the Comamonadaceae family and were 99% similar to previously described species. We identify these novel strains as Hydrogenophaga taeniospiralis strain IDSBO-1 and Variovorax paradoxus strain IDSBO-4. Both strains possess a gene with homology to the aioA gene, which encodes an As(III)-oxidase, and both oxidize As(III) aerobically, but only IDSBO-4 oxidized Sb(III) in the presence of air, while strain IDSBO-1 could achieve this via nitrate respiration. Our results suggest that expression of aioA is not induced by Sb(III) but may be involved in Sb(III) oxidation along with an Sb(III)-specific pathway. Phylogenetic analysis of proteins encoded by the aioA genes revealed a close sequence similarity (90%) among the two isolates and other known As(III)-oxidizing bacteria, particularly Acidovorax sp. strain NO1. Both isolates were capable of chemolithoautotrophic growth using As(III) as a primary electron donor, and strain IDSBO-4 exhibited incorporation of radiolabeled [(14)C]bicarbonate while oxidizing Sb(III) from Sb(III)-tartrate, suggesting possible Sb(III)-dependent autotrophy. Enrichment cultures produced the Sb(V) oxide mineral mopungite and lesser amounts of Sb(III)-bearing senarmontite as precipitates.

RubisCO-based CO2 fixation and C1 metabolism in the actinobacterium Pseudonocardia dioxanivorans†CB1190.

Pseudonocardia is an actinobacterial genus of interest due to its potential biotechnological, medical and environmental remediation applications, as well as for the ecologically relevant symbiotic relationships it forms with attine ants. Some Pseudonocardia spp. can grow autotrophically, but the genetic basis of this capability has not previously been reported. In this study, we examined autotrophy in Pseudonocardia dioxanivorans CB1190, which can grow using H2 and CO2, as well as heterotrophically. Genomic and transcriptomic analysis of CB1190 cells grown with H2/bicarbonate implicated the Calvin-Benson-Bassham (CBB) cycle in growth-supporting CO2 fixation, as well as a [NiFe] hydrogenase-encoding gene cluster in H2 oxidation. The CBB cycle genes are evolutionarily most related to actinobacterial homologues, although synteny has not been maintained. Ribulose-1,5-bisphosphate carboxylase activity was confirmed in H2/bicarbonate-grown CB1190 cells and was detected in cells grown with the C1 compounds formate, methanol and carbon monoxide. We also demonstrated the upregulation of CBB cycle genes upon exposure of CB1190 to these C1 substrates, and identified genes putatively involved in generating CO2 from the C1 substrates by using RT-qPCR. Finally, the potential for autotrophic growth of other Pseudonocardia spp. was explored, and the ecological implications of autotrophy in attine ant- and plant root-associated Pseudonocardia discussed.

Draft genome sequence of an ammonia-oxidizing archaeon, "Candidatus Nitrosopumilus sediminis" AR2, from Svalbard in the Arctic Circle.

Ammonia-oxidizing archaea (AOA) typically predominate over ammonia-oxidizing bacteria in marine sediments. We herein present the draft genome sequence of an ammonia-oxidizing archaeon, "Candidatus Nitrosopumilus sediminis" AR2, which was enriched in culture from a marine sediment obtained off Svalbard, within the Arctic Circle. The typical genes involved in archaeal ammonia oxidation and carbon fixation necessary for chemolithoautotrophic growth were observed. Interestingly, the AR2 genome sequence was revealed to possess, uniquely among cultivated AOA from marine environments, a capability for urea utilization.

Complete genome and comparative analysis of the chemolithoautotrophic bacterium Oligotropha carboxidovorans OM5.

BACKGROUND: Oligotropha carboxidovorans OM5 T. (DSM 1227, ATCC 49405) is a chemolithoautotrophic bacterium capable of utilizing CO (carbon monoxide) and fixing CO2 (carbon dioxide). We previously published the draft genome of this organism and recently submitted the complete genome sequence to GenBank. RESULTS: The genome sequence of the chemolithoautotrophic bacterium Oligotropha carboxidovorans OM5 consists of a 3.74-Mb chromosome and a 133-kb megaplasmid that contains the genes responsible for utilization of carbon monoxide, carbon dioxide, and hydrogen. To our knowledge, this strain is the first one to be sequenced in the genus Oligotropha, the closest fully sequenced relatives being Bradyrhizobium sp. BTAi and USDA110 and Nitrobacter hamburgiensis X14. Analysis of the O. carboxidovorans genome reveals potential links between plasmid-encoded chemolithoautotrophy and chromosomally-encoded lipid metabolism. Comparative analysis of O. carboxidovorans with closely related species revealed differences in metabolic pathways, particularly in carbohydrate and lipid metabolism, as well as transport pathways. CONCLUSION: Oligotropha, Bradyrhizobium sp and Nitrobacter hamburgiensis X14 are phylogenetically proximal. Although there is significant conservation of genome organization between the species, there are major differences in many metabolic pathways that reflect the adaptive strategies unique to each species.

The thermal response of soil microbial methanogenesis decreases in magnitude with changing temperature.

Microbial methanogenesis in anaerobic soils contributes greatly to global methane (CH4) release, and understanding its response to temperature is fundamental to predicting the feedback between this potent greenhouse gas and climate change. A compensatory thermal response in microbial activity over time can reduce the response of respiratory carbon (C) release to temperature change, as shown for carbon dioxide (CO2) in aerobic soils. However, whether microbial methanogenesis also shows a compensatory response to temperature change remains unknown. Here, we used anaerobic wetland soils from the Greater Khingan Range and the Tibetan Plateau to investigate how 160 days of experimental warming (+4°C) and cooling (-4°C) affect the thermal response of microbial CH4 respiration and whether these responses correspond to changes in microbial community dynamics. The mass-specific CH4 respiration rates of methanogens decreased with warming and increased with cooling, suggesting that microbial methanogenesis exhibited compensatory responses to temperature changes. Furthermore, changes in the species composition of methanogenic community under warming and cooling largely explained the compensatory response in the soils. The stimulatory effect of climate warming on soil microbe-driven CH4 emissions may thus be smaller than that currently predicted, with important consequences for atmospheric CH4 concentrations.

Autotrophic carbon fixation pathways along the redox gradient in oxygen-depleted oceanic waters.

Anoxic marine zones (AMZs), also known as 'oxygen-deficient zones', contribute to the loss of fixed nitrogen from the ocean by anaerobic microbial processes. While these microbial processes associated with the nitrogen cycle have been extensively studied, those linked to the carbon cycle in AMZs have received much less attention, particularly the autotrophic carbon fixation - a crucial component of the carbon cycle. Using metagenomic and metatranscriptomic data from major AMZs, we report an explicit partitioning of the marker genes associated with different autotrophic carbon fixation pathways along the redox gradient (from oxic to anoxic conditions) present in the water column of AMZs. Sequences related to the Calvin-Benson-Bassham cycle were found along the entire gradient, while those related to the reductive Acetyl-CoA pathway were restricted to suboxic and anoxic waters. Sequences putatively associated with the 3-hydroxypropionate/4-hydroxybutyrate cycle dominated in the upper and lower oxyclines. Genes related to the reductive tricarboxylic acid cycle were represented from dysoxic to anoxic waters. The taxonomic affiliation of the sequences is consistent with the presence of microorganisms involved in crucial steps of biogeochemical cycles in AMZs, such as the gamma-proteobacteria sulfur oxidisers, the anammox bacteria Candidatus Scalindua and the thaumarcheota ammonia oxidisers of the Marine Group I.

Expression of genes for sulfur oxidation in the intracellular chemoautotrophic symbiont of the deep-sea bivalve Calyptogena okutanii.

To understand sulfur oxidation in thioauto-trophic deep-sea clam symbionts, we analyzed the recently reported genomes of two chemoautotrophic symbionts of Calyptogena okutanii (Candidatus Vesicomyosocius okutanii strain HA: Vok) and C. magnifica (Candidatus Ruthia magnifica strain Cm: Rma), and examined the sulfur oxidation gene expressions in the Vok by RT-PCR. Both symbionts have genes for sulfide-quinone oxidoreductase (sqr), dissimilatory sulfite reductase (dsr), reversible dissimilatory sulfite reductase (rdsr), sulfur-oxidizing multienzyme system (sox)(soxXYZA and soxB but lacking soxCD), adenosine phosphosulfate reductase (apr), and ATP sulfurylase (sat). While these genomes share 29 orthologous genes for sulfur oxidation implying that both symbionts possess the same sulfur oxidation pathway, Rma has a rhodanese-related sulfurtransferase putative gene (Rmag0316) that has no corresponding ortholog in Vok, and Vok has one unique dsrR (COSY0782). We propose that Calyptogena symbionts oxidize sulfide and thiosulfate, and that sulfur oxidation proceeds as follows. Sulfide is oxidized to sulfite by rdsr. Sulfite is oxidized to sulfate by apr and sat. Thiosulfate is oxidized to zero-valence sulfur by sox, which is then reduced to sulfide by dsr. In addition, thiosulfate may also be oxidized into sulfate by another component of sox. The result of the RT-PCR showed that genes (dsrA, dsrB, dsrC, aprA, aprB, sat, soxB, and sqr) encoding key enzymes catalyzing sulfur oxidation were all equally expressed in the Vok under three different environmental conditions (aerobic, semioxic, and aerobic under high pressure at 9 MPa), indicating that all sulfur oxidation pathways function simultaneously to support intracellular symbiotic life.

The canonical network of autotrophic intermediary metabolism: minimal metabolome of a reductive chemoautotroph.

Chemoautorophs that fix carbon by the reductive tricarboxylic acid cycle represent one of the dominant bacterial life forms that make a major contribution to biomass production. From the viewpoint of biogenesis, construction of a canonical chart of intermediary metabolism for this class of organisms may help us to understand early cellular evolution and point us to the last universal common ancestor. Data-mining the KEGG Pathways database enabled us to integrate required biosynthetic pathways and derive a chart that represents the complete anabolic network of a reductive chemoautotroph. Compounds of this metabolic network together constitute a representative minimal metabolome that comprises 287 metabolites. These compounds have been classified into different groups including those compounds that form nodes in the network. It can be seen that a relatively sparse set of organic chemical reactions dominate the anabolic synthesis in the assembly of the minimal autotrophic metabolome. Empirical generalizations that result from analyzing this metabolic network may aid in elucidating selection rules that govern its emergence and further evolution and may also help in delineating attributes that impart the observed robustness to these metabolites.

Assessment of environmental gene tags linked with carbohydrate metabolism and chemolithotrophy associated microbial community in River Ganga.

The microbial community mediated biogeochemical cycles play important role in global C-cycle and display a sensitive response to environmental changes. Limited information is available on microbial composition and functional diversity controlling biogeochemical cycles in the riverine environment. The Ganga River water and sediment samples were studied for environmental gene tags with reference to carbohydrate metabolism, photoheterotrophy and chemolithotrophy using high throughput shotgun metagenomic sequencing and functional annotation. The diversity of environmental gene tags specific microbial community was annotated against reference sequence database using Kaiju taxonomic classifier. The metagenomic analyses revealed that the river harbored a broad range of carbohydrate and energy metabolism genes. The in-depth investigation of metagenomic data revealed that the enzymes associated with reverse TCA cycle, Calvin-Benson cycle enzyme RuBisCO, starch and sucrose metabolism genes were highly abundant. The enzymes associated with sulfur metabolism such as EC:2.7.7.4 (sulfate to ammonium per sulfate), EC:1.8.1.2, EC:1.8.7.1 (sulfite to H2S) were prevalent in both the class of samples. The principal component analysis of the functional profiles revealed that the water and sediment samples were clustered distinctly suggesting that both the sites had variable abundance of functional genes and associated microbiota. The taxonomic classification showed abundance of Proteobacteria, Actinobacteria and Bacteroidetes phyla. Also, the metagenomic study showed the presence of purple sulfur bacteria viz. Thiodictyon, Nitrosococcus and purple non-sulfur bacteria viz. Bradyrhozobium and Rhodobacter. The study demonstrates that the Ganga River microbiome has prevalence of functional genes involved in carbohydrate anabolism and catabolism, and CO2 fixation with great prospects in cellulose and sulfide degrading enzyme production and characterization.

Culture of Methanogenic Archaea from Human Colostrum and Milk.

Archaeal sequences have been detected in human colostrum and milk, but no studies have determined whether living archaea are present in either of these fluids. Methanogenic archaea are neglected since they are not detected by usual molecular and culture methods. By using improved DNA detection protocols and microbial culture techniques associated with antioxidants previously developed in our center, we investigated the presence of methanogenic archaea using culture and specific Methanobrevibacter smithii and Methanobrevibacter oralis real-time PCR in human colostrum and milk. M. smithii was isolated from 3 colostrum and 5 milk (day 10) samples. M. oralis was isolated from 1 milk sample. For 2 strains, the genome was sequenced, and the rhizome was similar to that of strains previously isolated from the human mouth and gut. M. smithii was detected in the colostrum or milk of 5/13 (38%) and 37/127 (29%) mothers by culture and qPCR, respectively. The different distribution of maternal body mass index according to the detection of M. smithii suggested an association with maternal metabolic phenotype. M. oralis was not detected by molecular methods. Our results suggest that breastfeeding may contribute to the vertical transmission of these microorganisms and may be essential to seed the infant's microbiota with these neglected critical commensals from the first hour of life.

Insights into microbial community structure and function from a shallow, simulated CO2 -leakage aquifer demonstrate microbial selection and adaptation.

Geological carbon storage is likely to be a part of a comprehensive strategy to minimize the atmospheric release of carbon dioxide (CO2 ), raising concerns that injected CO2 will leak into overlying freshwater aquifers. CO2(aq) leakage may impact the dominant microbial community responsible for important ecosystem functions such as nutrient cycling, metal cycling and carbon conversion. Here, we examined the impact of an experimental in situ CO2 -leakage on a freshwater aquifer microbial community. High-throughput 16S rRNA gene sequencing demonstrated lower microbial diversity in freshwater wells with CO2 concentrations above 1.15 g l-1 . Metagenomic sequencing and population genome binning were used to evaluate the metabolic potential of microbial populations across four CO2 exposed samples and one control sample. Population genome binning resulted in the recovery and annotation of three metagenome assembled genomes (MAGs). Two of the MAGs, most closely related to Curvibacter and Sulfuricurvum, had the functional capacity for CO2 utilization via carbon fixation coupled to sulfur and iron oxidation. The third draft genome was an Archaea, most closely related to Methanoregula, characterized by the metabolic potential for methanogenesis. Together, these findings show that CO2 leakage in a freshwater aquifer poses a strong selection, driving both microbial community structure and metabolic function.

Genomic inference of the metabolism and evolution of the archaeal phylum Aigarchaeota.

Microbes of the phylum Aigarchaeota are widely distributed in geothermal environments, but their physiological and ecological roles are poorly understood. Here we analyze six Aigarchaeota metagenomic bins from two circumneutral hot springs in Tengchong, China, to reveal that they are either strict or facultative anaerobes, and most are chemolithotrophs that can perform sulfide oxidation. Applying comparative genomics to the Thaumarchaeota and Aigarchaeota, we find that they both originated from thermal habitats, sharing 1154 genes with their common ancestor. Horizontal gene transfer played a crucial role in shaping genetic diversity of Aigarchaeota and led to functional partitioning and ecological divergence among sympatric microbes, as several key functional innovations were endowed by Bacteria, including dissimilatory sulfite reduction and possibly carbon monoxide oxidation. Our study expands our knowledge of the possible ecological roles of the Aigarchaeota and clarifies their evolutionary relationship to their sister lineage Thaumarchaeota.

A novel soxO gene, encoding a glutathione disulfide reductase, is essential for tetrathionate oxidation in Advenella kashmirensis.

Molecular mechanisms of chemolithotrophic tetrathionate oxidation are not clearly understood. Here we used transposon(Tn5-mob)-insertion mutagenesis to search for novel tetrathionate oxidation genes in the facultatively chemolithoautotrophic betaproteobacterium Advenella kashmirensis that not only oxidizes tetrathionate, but also produces the same as an intermediate during thiosulfate oxidation. Genome-wide random insertion of Tn5-mob occurred at a frequency of one per 104 donor E. coli cells. A library of 8000 transconjugants yielded five tetrathionate-oxidation-impaired mutants, of which, the one named Ak_Tn_16 was studied here in detail. When grown chemolithoautotrophically on thiosulfate, Ak_Tn_16 converted the total thiosulfate supplied to equivalent amount of tetrathionate, exactly in the same way as the wild type. It could not, however, oxidize the intermediary tetrathionate to sulfate; Ak_Tn_16 could not also oxidize tetrathionate when it was supplied as the starting chemolithotrophic substrate. In the Ak_Tn_16 genome, Tn5-mob was found to have transposed in a novel soxO gene, located just-upstream of soxB, within the sox gene cluster. SoxO was predicted, via iterative threading assembly simulation, to be a glutathione-disulfide (GSSG) reductase. When Ak_Tn_16 was grown in tetrathionate-based chemolithoautotrophic medium supplemented with reduced glutathione (GSH) its tetrathionate-oxidation deficiency, remarkably, was ameliorated. Implications for a key role of GSH in tetrathionate oxidation are discussed in the light of other molecular evidences available for A. kashmirensis.