Biology of a widespread uncultivated archaeon that contributes to carbon fixation in the subsurface

Subsurface microbial life contributes significantly to biogeochemical cycling, yet it remains largely uncharacterized, especially its archaeal members. This 'microbial dark matter' has been explored by recent studies that were, however, mostly based on DNA sequence information only. Here, we use diverse techniques including ultrastuctural analyses to link genomics to biology for the SM1 Euryarchaeon lineage, an uncultivated group of subsurface archaea. Phylogenomic analyses reveal this lineage to belong to a widespread group of archaea that we propose to classify as a new euryarchaeal order ('Candidatus Altiarchaeales'). The representative, double-membraned species 'Candidatus Altiarchaeum hamiconexum' has an autotrophic metabolism that uses a not-yet-reported Factor(420)-free reductive acetyl-CoA pathway, confirmed by stable carbon isotopic measurements of archaeal lipids. Our results indicate that this lineage has evolved specific metabolic and structural features like nano-grappling hooks empowering this widely distributed archaeon to predominate anaerobic groundwater, where it may represent an important carbon dioxide sink

The responses of an anaerobic microorganism, Yersinia intermedia MASE-LG-1 to individual and combined simulated Martian stresses

The limits of life of aerobic microorganisms are well understood, but the responses of anaerobic microorganisms to individual and combined extreme stressors are less well known. Motivated by an interest in understanding the survivability of anaerobic microorganisms under Martian conditions, we investigated the responses of a new isolate, Yersinia intermedia MASE-LG-1 to individual and combined stresses associated with the Martian surface. This organism belongs to an adaptable and persistent genus of anaerobic microorganisms found in many environments worldwide. The effects of desiccation, low pressure, ionizing radiation, varying temperature, osmotic pressure, and oxidizing chemical compounds were investigated. The strain showed a high tolerance to desiccation, with a decline of survivability by four orders of magnitude during a storage time of 85 days. Exposure to X-rays resulted in dose-dependent inactivation for exposure up to 600 Gy while applied doses above 750 Gy led to complete inactivation. The effects of the combination of desiccation and irradiation were additive and the survivability was influenced by the order in which they were imposed. Ionizing irradiation and subsequent desiccation was more deleterious than vice versa. By contrast, the presence of perchlorates was not found to significantly affect the survival of the Yersinia strain after ionizing radiation. These data show that the organism has the capacity to survive and grow in physical and chemical stresses, imposed individually or in combination that are associated with Martian environment. Eventually it lost its viability showing that many of the most adaptable anaerobic organisms on Earth would be killed on Mars today

First Insights into the Diverse Human Archaeome: Specific Detection of Archaea in the Gastrointestinal Tract, Lung, and Nose and on Skin

Human-associated archaea remain understudied in the field of microbiome research, although in particular methanogenic archaea were found to be regular commensals of the human gut, where they represent keystone species in metabolic processes. Knowledge on the abundance and diversity of human-associated archaea is extremely limited, and little is known about their function(s), their overall role in human health, or their association with parts of the human body other than the gastrointestinal tract and oral cavity. Currently, methodological issues impede the full assessment of the human archaeome, as bacteria-targeting protocols are unsuitable for characterization of the full spectrum of Archaea. The goal of this study was to establish conservative protocols based on specifically archaea-targeting, PCR-based methods to retrieve first insights into the archaeomes of the human gastrointestinal tract, lung, nose, and skin. Detection of Archaea was highly dependent on primer selection and the sequence processing pipeline used. Our results enabled us to retrieve a novel picture of the human archaeome, as we found for the first time Methanobacterium and Woesearchaeota (DPANN superphylum) to be associated with the human gastrointestinal tract and the human lung, respectively. Similar to bacteria, human-associated archaeal communities were found to group biogeographically, forming (i) the thaumarchaeal skin landscape, (ii) the (methano)euryarchaeal gastrointestinal tract, (iii) a mixed skin-gastrointestinal tract landscape for the nose, and (iv) a woesearchaeal lung landscape. On the basis of the protocols we used, we were able to detect unexpectedly high diversity of archaea associated with different body parts

S-layers at second glance? Altiarchaeal grappling hooks (hami) resemble archaeal S-layer proteins in structure and sequence.

The uncultivated "Candidatus Altiarchaeum hamiconexum" (formerly known as SM1 Euryarchaeon) carries highly specialized nano-grappling hooks ("hami") on its cell surface. Until now little is known about the major protein forming these structured fibrous cell surface appendages, the genes involved or membrane anchoring of these filaments. These aspects were analyzed in depth in this study using environmental transcriptomics combined with imaging methods. Since a laboratory culture of this archaeon is not yet available, natural biofilm samples with high Ca. A. hamiconexum abundance were used for the entire analyses. The filamentous surface appendages spanned both membranes of the cell, which are composed of glycosyl-archaeol. The hami consisted of multiple copies of the same protein, the corresponding gene of which was identified via metagenome-mapped transcriptome analysis. The hamus subunit proteins, which are likely to self-assemble due to their predicted beta sheet topology, revealed no similiarity to known microbial flagella-, archaella-, fimbriae- or pili-proteins, but a high similarity to known S-layer proteins of the archaeal domain at their N-terminal region (44-47% identity). Our results provide new insights into the structure of the unique hami and their major protein and indicate their divergent evolution with S-layer proteins

Coupling Genetic and Chemical Microbiome Profiling Reveals Heterogeneity of Archaeome and Bacteriome in Subsurface Biofilms That Are Dominated by the Same Archaeal Species

Earth harbors an enormous portion of subsurface microbial life, whose microbiome flux across geographical locations remains mainly unexplored due to difficult access to samples. Here, we investigated the microbiome relatedness of subsurface biofilms of two sulfidic springs in southeast Germany that have similar physical and chemical parameters and are fed by one deep groundwater current. Due to their unique hydrogeological setting these springs provide accessible windows to subsurface biofilms dominated by the same uncultivated archaeal species, called SM1 Euryarchaeon. Comparative analysis of infrared imaging spectra emonstrated great variations in archaeal membrane composition between biofilms of the two springs, suggesting different SM1 euryarchaeal strains of the same species at both aquifer outlets. This strain variation was supported by ultrastructural and metagenomic analyses of the archaeal biofilms, which included intergenic spacer region sequencing of the rRNA gene operon. At 16S rRNA gene level, PhyloChip G3 DNA microarray detected similar biofilm communities for archaea, but site-specific communities for bacteria. Both biofilms showed an enrichment of different deltaproteobacterial operational taxonomic units, whose families were, however, congruent as were their lipid spectra. Consequently, the function of the major proportion of the bacteriome appeared to be conserved across the geographic locations studied, which was confirmed by dsrB-directed quantitative PCR. Consequently, microbiome differences of these subsurface biofilms exist at subtle nuances for archaea (strain level variation) and at higher taxonomic levels for predominant bacteria without a substantial perturbation in bacteriome function. The results of this communication provide deep insight into the dynamics of subsurface microbial life and warrant its future investigation with regard to metabolic and genomic analyses

Lack of correlation of desiccation and radiation tolerance in microorganisms from diverse extreme environments tested under anoxic conditions

Four facultative anaerobic and two obligate anaerobic bacteria were isolated from extreme environments (deep subsurface halite mine, sulfidic anoxic spring, mineral-rich river) in the frame MASE (Mars Analogues for Space Exploration) project. The isolates were investigated under anoxic conditions for their survivability after desiccation up to 6 months and their tolerance to ionizing radiation up to 3000 Gy. The results indicated that tolerances to both stresses are strain-specific features. Yersinia intermedia MASE-LG-1 showed a high desiccation tolerance but its radiation tolerance was very low. The most radiation-tolerant strains were Buttiauxella sp. MASE-IM-9 and Halanaerobium sp. MASE-BB-1. In both cases, cultivable cells were detectable after an exposure to 3 kGy of ionizing radiation, but cells only survived desiccation for 90 and 30 days, respectively. Although a correlation between desiccation and ionizing radiation resistance has been hypothesized for some aerobic microorganisms, our data showed that there was no correlation between tolerance to desiccation and ionizing radiation, suggesting that the physiological basis of both forms of tolerances is not necessarily linked. In addition, these results indicated that facultative and obligate anaerobic organisms living in extreme environments possess varied species-specific tolerances to extremes

Detailed community profiling using PhyloChip G3 and SR-FTIR.

<p>A: Ordination analysis of PhyloChip G3 data based on weighted UniFrac measure of eOTU abundances followed by non-metric multidimensional scaling (NMDS). Stress for NMDS of archaeal eOTUs (#37): 0.0088. Stress for NMDS of bacterial eOTUs (#1300): 0.0223. B: Heatmap displaying significantly different families found between the two biofilm types, MSI-BF and SM-BF by PhyloChip G3 assay. Significance is based on aggregated HybScores of eOTUs on family level followed by a Welch-test. For false discovery detection please see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0099801#pone.0099801.s006" target="_blank">Fig. S6</a>. C: Ordination analysis of SR-FTIR data based on a linear discriminant analysis and principal component analysis (PCA-LDA) in the spectral region of 2800–3100 cm⁻¹ on the archaea spectra extracted from the maps from the three different locations. On the right there is the plot of PCA-LDA loadings. PCA-LDA1 explains for the 93.4% of the variance, PCA-LDA2 for 5.3% and PCA-LDA3 for 0.9%. Arrows point to the infrared signals used to explain the difference between the samples: 2975 cm⁻¹, 2965 cm⁻¹, 2924 cm⁻¹ and 2850 cm⁻¹. D: PCA-LDA in the spectral regions of 900–1280 cm⁻¹ and 2800–3100 cm⁻¹ on SR-FTIR spectra of the bacteria “pixels” from the chemical maps of the samples at the three different locations. On the right there is a plot of PCA-LDA loadings in the two spectral region of interest. PCA-LDA1 explains for the 54.5% of the variance, PCA-LDA2 for 28.6% and PCA-LDA3 for 7.3%. Arrows point to the main infrared signals used to explain the difference between the samples: 2958 cm⁻¹, 2925 cm⁻¹, 2870 cm⁻¹ and 2850 cm⁻¹, in the second panel 1250 cm⁻¹, 1110 cm⁻¹, 1080 cm⁻¹ and 1045 cm⁻¹.</p

Scanning and transmission electron micrographs of biofilms, cells and hami.

<p>Left panels: MSI, right panel: SM. A: Scanning electron micrograph of MSI biofilm, showing SM1 euryarchaeal cells with defined distances and cell-cell connections. Bar: 1 µm. B: Scanning electron micrograph of SM biofilm, showing SM1 euryarchaeal cells with defined distances and fine-structured cell-cell connections. In-between: Bacterial filamentous and rod-shaped cells. Bar: 1 µm. C: Scanning electron micrograph of dividing SM1 euryarchaeal cell (MSI) with cell surface appendages. Bar: 200 nm. D: Scanning electron micrograph of dividing SM1 euryarchaeal cell (SM) with cell surface appendages. Bar: 200 nm. E: Transmission electron micrograph of cell surface appendages (hami) of SM1 euryarchaeal cells from the MSI biofilm. The hami carry the nano-grappling hooks, but besides that appear bare (square), without prickles (Moissl et al 2005). Bar: 100 nm. F: Transmission electron micrograph of cell surface appendages and matrix of SM1 euryarchaeal cells from the SM biofilm. The hami reveal the typical ultrastructure, with nano-grappling hooks and barbwire-like prickle region (square, Moissl et al 2005). Bar: 100 nm.</p