Hydrogenases from methanogenic archaea, nickel, a novel cofactor, and H2 storage.

Most methanogenic archaea reduce CO(2) with H(2) to CH(4). For the activation of H(2), they use different [NiFe]-hydrogenases, namely energy-converting [NiFe]-hydrogenases, heterodisulfide reductase-associated [NiFe]-hydrogenase or methanophenazine-reducing [NiFe]-hydrogenase, and F(420)-reducing [NiFe]-hydrogenase. The energy-converting [NiFe]-hydrogenases are phylogenetically related to complex I of the respiratory chain. Under conditions of nickel limitation, some methanogens synthesize a nickel-independent [Fe]-hydrogenase (instead of F(420)-reducing [NiFe]-hydrogenase) and by that reduce their nickel requirement. The [Fe]-hydrogenase harbors a unique iron-guanylylpyridinol cofactor (FeGP cofactor), in which a low-spin iron is ligated by two CO, one C(O)CH(2)-, one S-CH(2)-, and a sp(2)-hybridized pyridinol nitrogen. Ligation of the iron is thus similar to that of the low-spin iron in the binuclear active-site metal center of [NiFe]- and [FeFe]-hydrogenases. Putative genes for the synthesis of the FeGP cofactor have been identified. The formation of methane from 4 H(2) and CO(2) catalyzed by methanogenic archaea is being discussed as an efficient means to store H(2).

How clear is our current view on microbial dark matter? (Re-)assessing public MAG & SAG datasets with MDMcleaner.

As of today, the majority of environmental microorganisms remain uncultured and is therefore referred to as 'microbial dark matter' (MDM). Hence, genomic insights into these organisms are limited to cultivation-independent approaches such as single-cell- and metagenomics. However, without access to cultured representatives for verifying correct taxon-assignments, MDM genomes may cause potentially misleading conclusions based on misclassified or contaminant contigs, thereby obfuscating our view on the uncultured microbial majority. Moreover, gradual database contaminations by past genome submissions can cause error propagations which affect present as well as future comparative genome analyses. Consequently, strict contamination detection and filtering need to be applied, especially in the case of uncultured MDM genomes. Current genome reporting standards, however, emphasize completeness over purity and the de facto gold standard genome assessment tool, checkM, discriminates against uncultured taxa and fragmented genomes. To tackle these issues, we present a novel contig classification, screening, and filtering workflow and corresponding open-source python implementation called MDMcleaner, which was tested and compared to other tools on mock and real datasets. MDMcleaner revealed substantial contaminations overlooked by current screening approaches and sensitively detects misattributed contigs in both novel genomes and the underlying reference databases, thereby greatly improving our view on 'microbial dark matter'.

Single-cell transcriptomics and data analyses for prokaryotes-Past, present and future concepts.

Transcriptomics, or more specifically mRNA sequencing, is a powerful tool to study gene expression at the single-cell level (scRNA-seq) which enables new insights into a plethora of biological processes. While methods for single-cell RNA-seq in eukaryotes are well established, application to prokaryotes is still challenging. Reasons for that are rigid and diverse cell wall structures hampering lysis, the lack of polyadenylated transcripts impeding mRNA enrichment, and minute amounts of RNA requiring amplification steps before sequencing. Despite those obstacles, several promising scRNA-seq approaches for bacteria have been published recently, albeit difficulties in the experimental workflow and data processing and analysis remain. In particular, bias is often introduced by amplification which makes it difficult to distinguish between technical noise and biological variation. Future optimization of experimental procedures and data analysis algorithms are needed for the improvement of scRNA-seq but also to aid in the emergence of prokaryotic single-cell multi-omics. to help address 21st century challenges in the biotechnology and health sector.

A photobioreactor for production of algae biomass from gaseous emissions of an animal house.

Sustainable approaches to circular economy in animal agriculture are still poorly developed. Here, we report an approach to reduce gaseous emissions of CO2 and NH3 from animal housing while simultaneously using them to produce value-added biomass. To this end, a cone-shaped, helical photobioreactor was developed that can be integrated into animal housing by being freely suspended, thereby combining a small footprint with a physically robust design. The photobioreactor was coupled with the exhaust air of a chicken house to allow continuous cultivation of a mixed culture of Arthrospira spec. (Spirulina). Continuous quantification of CO2 and NH3 concentration showed that the coupled algae reactor effectively purifies the exhaust air from the chicken house while producing algal biomass. Typical production rates of greater than 0.3 g/l*day dry mass were obtained, and continuous operation was possible for several weeks. Morphological, biochemical, and genomic characterization of Spirulina cultures yielded insights into the dynamics and metabolic processes of the microbial community. We anticipate that further optimization of this approach will provide new opportunities for the generation of value-added products from gaseous CO2 and NH3 waste emissions, linking resource-efficient production of microalgae with simultaneous sequestration of animal emissions. KEY POINTS: • Coupling a bioreactor with exhaust gases of chicken coop for production of biomass. • Spirulina mixed culture removes CO2 and NH3 from chicken house emissions. • High growth rates and biodiversity adaptation for nitrogen metabolism. Towards a sustainable circular economy in livestock farming. The functional coupling of a helical tube photobioreactor with exhaust air from a chicken house enabled the efficient cultivation of Spirulina microalgae while simultaneously sequestering the animals' CO2 and NH3 emissions.

Back to Basics: A Simplified Improvement to Multiple Displacement Amplification for Microbial Single-Cell Genomics.

Microbial single-cell genomics (SCG) provides access to the genomes of rare and uncultured microorganisms and is a complementary method to metagenomics. Due to the femtogram-levels of DNA in a single microbial cell, sequencing the genome requires whole genome amplification (WGA) as a preliminary step. However, the most common WGA method, multiple displacement amplification (MDA), is known to be costly and biased against specific genomic regions, preventing high-throughput applications and resulting in uneven genome coverage. Thus, obtaining high-quality genomes from many taxa, especially minority members of microbial communities, becomes difficult. Here, we present a volume reduction approach that significantly reduces costs while improving genome coverage and uniformity of DNA amplification products in standard 384-well plates. Our results demonstrate that further volume reduction in specialized and complex setups (e.g., microfluidic chips) is likely unnecessary to obtain higher-quality microbial genomes. This volume reduction method makes SCG more feasible for future studies, thus helping to broaden our knowledge on the diversity and function of understudied and uncharacterized microorganisms in the environment.

IncP-type plasmids carrying genes for antibiotic resistance or for aromatic compound degradation are prevalent in sequenced Aromatoleum and Thauera strains.

Self-transferable plasmids of the incompatibility group P-1 (IncP-1) are considered important carriers of genes for antibiotic resistance and other adaptive functions. In the laboratory, these plasmids have a broad host range; however, little is known about their in situ host profile. In this study, we discovered that Thauera aromatica K172T , a facultative denitrifying microorganism capable of degrading various aromatic compounds, contains a plasmid highly similar to the IncP-1 ε archetype pKJK5. The plasmid harbours multiple antibiotic resistance genes and is maintained in strain K172T for at least 1000 generations without selection pressure from antibiotics. In a subsequent search, we found additional nine IncP-type plasmids in a total of 40 sequenced genomes of the closely related genera Aromatoleum and Thauera. Six of these plasmids form a novel IncP-1 subgroup designated θ, four of which carry genes for anaerobic or aerobic degradation of aromatic compounds. Pentanucleotide sequence analyses (k-mer profiling) indicated that Aromatoleum spp. and Thauera spp. are among the most suitable hosts for the θ plasmids. Our results highlight the importance of IncP-1 plasmids for the genetic adaptation of these common facultative denitrifying bacteria and provide novel insights into the in situ host profile of these plasmids.

A marine plasmid hitchhiking vast phylogenetic and geographic distances.

Horizontal gene transfer (HGT) plays an important role in bacterial evolution and serves as a driving force for bacterial diversity and versatility. HGT events often involve mobile genetic elements like plasmids, which can promote their own dissemination by associating with adaptive traits in the gene pool of the so-called mobilome. Novel traits that evolve through HGT can therefore lead to the exploitation of new ecological niches, prompting an adaptive radiation of bacterial species. In this study, we present phylogenetic, biogeographic, and functional analyses of a previously unrecognized RepL-type plasmid found in diverse members of the marine Roseobacter group across the globe. Noteworthy, 100% identical plasmids were detected in phylogenetically and geographically distant bacteria, revealing a so-far overlooked, but environmentally highly relevant vector for HGT. The genomic and functional characterization of this plasmid showed a completely conserved backbone dedicated to replication, stability, and mobilization as well as an interchangeable gene cassette with highly diverse, but recurring motifs. The majority of the latter appear to be involved in mechanisms coping with toxins and/or pollutants in the marine environment. Furthermore, we provide experimental evidence that the plasmid has the potential to be transmitted across bacterial orders, thereby increasing our understanding of evolution and microbial niche adaptation in the environment.

Genome-inferred spatio-temporal resolution of an uncultivated Roizmanbacterium reveals its ecological preferences in groundwater.

Subsurface ecosystems like groundwater harbour diverse microbial communities, including small-sized, putatively symbiotic organisms of the Candidate Phyla Radiation, yet little is known about their ecological preferences and potential microbial partners. Here, we investigated a member of the superphylum Microgenomates (Cand. Roizmanbacterium ADI133) from oligotrophic groundwater using mini-metagenomics and monitored its spatio-temporal distribution using 16S rRNA gene analyses. A Roizmanbacteria-specific quantitative PCR assay allowed us to track its abundance over the course of 1 year within eight groundwater wells along a 5.4 km hillslope transect, where Roizmanbacteria reached maximum relative abundances of 2.3%. In-depth genomic analyses suggested that Cand. Roizmanbacterium ADI133 is a lactic acid fermenter, potentially able to utilize a range of complex carbon substrates, including cellulose. We hypothesize that it attaches to host cells using a trimeric autotransporter adhesin and inhibits their cell wall biosynthesis using a toxin-antitoxin system. Network analyses based on correlating Cand. Roizmanbacterium ADI133 abundances with amplicon sequencing-derived microbial community profiles suggested one potential host organism, classified as a member of the class Thermodesulfovibrionia (Nitrospirae). By providing lactate as an electron donor Cand. Roizmanbacterium ADI133 potentially mediates the transfer of carbon to other microorganisms and thereby is an important connector in the microbial community.

Nocardia macrotermitis sp. nov. and Nocardia aurantia sp. nov., isolated from the gut of the fungus-growing termite Macrotermes natalensis.

The taxonomic positions of two novel aerobic, Gram-stain-positive Actinobacteria, designated RB20T and RB56T, were determined using a polyphasic approach. Both were isolated from the fungus-farming termite Macrotermes natalensis. Results of 16S rRNA gene sequence analysis revealed that both strains are members of the genus Nocardia with the closest phylogenetic neighbours Nocardia miyunensis JCM12860T (98.9 %) and Nocardia nova DSM44481T (98.5 %) for RB20T and Nocardia takedensis DSM 44801T (98.3 %), Nocardia pseudobrasiliensis DSM 44290T (98.3 %) and Nocardia rayongensis JCM 19832T (98.2 %) for RB56T. Digital DNA-DNA hybridization (DDH) between RB20T and N. miyunensis JCM12860T and N. nova DSM 44481T resulted in similarity values of 33.9 and 22.0 %, respectively. DDH between RB56T and N. takedensis DSM44801T and N. pseudobrasiliensis DSM44290T showed similarity values of 20.7 and 22.3 %, respectively. In addition, wet-lab DDH between RB56T and N. rayongensis JCM19832T resulted in 10.2 % (14.5 %) similarity. Both strains showed morphological and chemotaxonomic features typical for the genus Nocardia, such as the presence of meso-diaminopimelic acid (A2pm) within the cell wall, arabinose and galactose as major sugar components within whole cell-wall hydrolysates, the presence of mycolic acids and major phospholipids (diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol), and the predominant menaquinone MK-8 (H4, ω-cyclo). The main fatty acids for both strains were hexadecanoic acid (C16 : 0), 10-methyloctadecanoic acid (10-methyl C18 : 0) and cis-9-octadecenoic acid (C18 : 1 ω9c). We propose two novel species within the genus Nocardia: Nocardia macrotermitis sp. nov. with the type strain RB20T (=VKM Ac-2841T=NRRL B65541T) and Nocardia aurantia sp. nov. with the type strain RB56T (=VKM Ac-2842T=NRRL B65542T).

Actinomadura rubteroloni sp. nov. and Actinomadura macrotermitis sp. nov., isolated from the gut of the fungus growing-termite Macrotermes natalensis.

The taxonomic positions of two novel aerobic, Gram-positive actinobacteria, designated strains RB29T and RB68T, were determined using a polyphasic approach. Based on 16S rRNA gene sequence analysis, the closest phylogenetic neighbours of RB29T were identified as Actinomadura rayongensis DSM 102126T (99.2 % similarity) and Actinomadura atramentaria DSM 43919T (98.7 %), and for strain RB68T was Actinomadura hibisca DSM 44148T (98.3 %). Digital DNA-DNA hybridization (dDDH) between RB29T and its closest phylogenetic neighbours, A. rayongensis DSM 102126T and A. atramentaria DSM 43919T, resulted in similarity values of 53.2 % (50.6-55.9 %) and 26.4 % (24.1-28.9 %), respectively. Additionally, the average nucleotide identity (ANI) was 93.2 % (94.0 %) for A. rayongensis DSM 102126T and 82.3 % (78.9 %) for A. atramentaria DSM 43919T. dDDH analysis between strain RB68T and A. hibisca DSM 44148T gave a similarity value of 24.5 % (22.2-27.0 %). Both strains, RB29T and RB68T, revealed morphological characteristics and chemotaxonomic features typical for the genus Actinomadura, such as the presence of meso-diaminopimelic acid in the cell wall, galactose and glucose as major sugar components within whole-cell hydrolysates and the absence of mycolic acids. The major phospholipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol and phosphatidylinositol mannoside. Predominant menaquinones were MK-9(H6) and MK-9(H8) for RB29T and MK-9(H4) and MK-9(H6) for RB68T. The main fatty acids were identified as 10-methyloctadecanoic acid (10-methyl C18:0), 14-methylpentadecanoic acid (iso-C16:0), hexadecanoic acid (C16:0) and cis-9-octadecanoic acid (C18 : 1 ω9c). Here, we propose two novel species of the genus Actinomadura: Actinomadura rubteroloni sp. nov. with the type strain RB29T (=CCUG 72668T=NRRL B-65537T) and Actinomadura macrotermitis sp. nov. with the type strain RB68T (=CCUG 72669T=NRRL B-65538T).

Streptomyces smaragdinus sp. nov., isolated from the gut of the fungus growing-termite Macrotermes natalensis.

The taxonomic position of a novel aerobic, Gram-positive actinobacteria, designated strain RB5T, was determined using a polyphasic approach. The strain, isolated from the gut of the fungus-farming termite Macrotermes natalensis, showed morphological, physiological and chemotaxonomic properties typical of the genus Streptomyces. Based on 16S rRNA gene sequence analysis, the closest phylogenetic neighbour of RB5T was Streptomyces polyrhachis DSM 42102T (98.87 %). DNA-DNA hybridization experiments between strain RB5T and S. polyrhachis DSM 42102T resulted in a value of 27.4 % (26.8 %). The cell wall of strain RB5T contained ll-diaminopimelic acid as the diagnostic amino acid. Mycolic acids and diagnostic sugars in whole-cell hydrolysates were not detected. The strain produced the following major phospholipids: diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol, phosphatidylinositol-mannoside and phosphatidylserine. The menaquinone profile showed hexa- and octahydrogenated menaquinones containing nine isoprene units [MK-9(H6) and MK-9(H8)]. The strain exhibited a fatty acid profile containing the following major fatty acids: 12-methyltridecanoic acid (iso-C14 : 0) 12-methyltetradecanoic acid (anteiso-C15 : 0), 13-methyltetradecanoic acid (iso-C15 : 0) and 14-methylpentadecanoic acid (iso-C16 : 0). Here, we propose a novel species of the genus Streptomyces - Streptomyces smaragdinus with the type strain RB5T (=VKM Ac-2839T=NRRL B65539T).

Polyhalogenation of Isoflavonoids by the Termite-Associated Actinomadura sp. RB99.

Based on high-resolution tandem mass spectrometry (HR-MS2) and global natural products social molecular networking (GNPS), we found that plant-derived daidzein and genistein derivatives are polyhalogenated by termite-associated Actinomadura species RB99. MS-guided purification from extracts of bacteria grown under optimized conditions led to the isolation of eight polychlorinated isoflavones, including six unreported derivatives, and seven novel polybrominated derivatives, two of which showed antimicrobial activity.

Microbial single-cell omics: the crux of the matter.

Single-cell genomics and transcriptomics can provide reliable context for assembled genome fragments and gene expression activity on the level of individual prokaryotic genomes. These methods are rapidly emerging as an essential complement to cultivation-based, metagenomics, metatranscriptomics, and microbial community-focused research approaches by allowing direct access to information from individual microorganisms, even from deep-branching phylogenetic groups that currently lack cultured representatives. Their integration and binning with environmental 'omics data already provides unprecedented insights into microbial diversity and metabolic potential, enabling us to provide information on individual organisms and the structure and dynamics of natural microbial populations in complex environments. This review highlights the pitfalls and recent advances in the field of single-cell omics and its importance in microbiological and biotechnological studies. KEY POINTS: • Single-cell omics expands the tree of life through the discovery of novel organisms, genes, and metabolic pathways. • Disadvantages of metagenome-assembled genomes are overcome by single-cell omics. • Functional analysis of single cells explores the heterogeneity of gene expression. • Technical challenges still limit this field, thus prompting new method developments.

Additions to the genus Gimesia: description of Gimesia alba sp. nov., Gimesia algae sp. nov., Gimesia aquarii sp. nov., Gimesia aquatilis sp. nov., Gimesia fumaroli sp. nov. and Gimesia panareensis sp. nov., isolated from aquatic habitats of the Northern Hemisphere.

Thirteen novel planctomycetal strains were isolated from five different aquatic sampling locations. These comprise the hydrothermal vent system close to Panarea Island (Italy), a biofilm on the surface of kelp at Monterey Bay (CA, USA), sediment and algae on Mallorca Island (Spain) and Helgoland Island (Germany), as well as a seawater aquarium in Braunschweig, Germany. All strains were shown to belong to the genus Gimesia. Their genomes cover a size range from 7.22 to 8.29 Mb and have a G+C content between 45.1 and 53.7%. All strains are mesophilic (Topt 26-33 °C) with generation times between 12 and 32 h. Analysis of fatty acids yielded palmitic acid (16:0) and a fatty acid with the equivalent chain length of 15.817 as major compounds. While five of the novel strains belong to the already described species Gimesia maris and Gimesia chilikensis, the other strains belong to novel species, for which we propose the names Gimesia alba (type strain Pan241wT = DSM 100744T = LMG 31345T = CECT 9841T = VKM B-3430T), Gimesia algae (type strain Pan161T = CECT 30192T = STH00943T = LMG 29130T), Gimesia aquarii (type strain V144T = DSM 101710T = VKM B-3433T), Gimesia fumaroli (type strain Enr17T = DSM 100710T = VKM B-3429T) and Gimesia panareensis (type strain Enr10T = DSM 100416T = LMG 29082T). STH numbers refer to the Jena Microbial Resource Collection (JMRC).

Updates to the recently introduced family Lacipirellulaceae in the phylum Planctomycetes: isolation of strains belonging to the novel genera Aeoliella, Botrimarina, Pirellulimonas and Pseudobythopirellula and the novel species Bythopirellula polymerisocia and Posidoniimonas corsicana.

Eight novel strains of the phylum Planctomycetes were isolated from different aquatic habitats. Among these habitats were the hydrothermal vent system close to Panarea Island, a public beach at Mallorca Island, the shore of Costa Brava (Spain), and three sites with brackish water in the Baltic Sea. The genome sizes of the novel strains range from 4.33 to 6.29 Mb with DNA G+C contents between 52.8 and 66.7%. All strains are mesophilic (Topt 24-30 °C) and display generation times between 17 and 94 h. All eight isolates constitute novel species of either already described or novel genera within the family Lacipirellulaceae. Two of the novel species, Posidoniimonas polymericola (type strain Pla123aT = DSM 103020T = LMG 29466T) and Bythopirellula polymerisocia (type strain Pla144T = DSM 104841T = VKM B-3442T), belong to established genera, while the other strains represent the novel genera Aeoliella gen. nov., Botrimarina gen. nov., Pirellulimonas gen. nov. and Pseudobythopirellula gen. nov. Based on our polyphasic analysis, we propose the species Aeoliella mucimassa sp. nov. (type strain Pan181T = DSM 29370T = LMG 31346T = CECT 9840T = VKM B-3426T), Botrimarina colliarenosi sp. nov. (type strain Pla108T = DSM 103355T = LMG 29803T), Botrimarina hoheduenensis sp. nov. (type strain Pla111T = DSM 103485T = STH00945T, Jena Microbial Resource Collection JMRC), Botrimarina mediterranea sp. nov. (type strain Spa11T = DSM 100745T = LMG 31350T = CECT 9852T = VKM B-3431T), Pirellulimonas nuda sp. nov. (type strain Pla175T = DSM 109594T = CECT 9871T = VKM B-3448T) and Pseudobythopirellula maris sp. nov. (type strain Mal64T = DSM 100832T = LMG 29020T).

Communal metabolism by Methylococcaceae and Methylophilaceae is driving rapid aerobic methane oxidation in sediments of a shallow seep near Elba, Italy.

The release of abiotic methane from marine seeps into the atmosphere is a major source of this potent greenhouse gas. Methanotrophic microorganisms in methane seeps use methane as carbon and energy source, thus significantly mitigating global methane emissions. Here, we investigated microbial methane oxidation at the sediment-water interface of a shallow marine methane seep. Metagenomics and metaproteomics, combined with 13 C-methane stable isotope probing, demonstrated that various members of the gammaproteobacterial family Methylococcaceae were the key players for methane oxidation, catalysing the first reaction step to methanol. We observed a transfer of carbon to methanol-oxidizing methylotrophs of the betaproteobacterial family Methylophilaceae, suggesting an interaction between methanotrophic and methylotrophic microorganisms that allowed for rapid methane oxidation. From our microcosms, we estimated methane oxidation rates of up to 871 nmol of methane per gram sediment per day. This implies that more than 50% of methane at the seep is removed by microbial oxidation at the sediment-water interface, based on previously reported in situ methane fluxes. The organic carbon produced was further assimilated by different heterotrophic microbes, demonstrating that the methane-oxidizing community supported a complex trophic network. Our results provide valuable eco-physiological insights into this specialized microbial community performing an ecosystem function of global relevance.

Unravelling the Identity, Metabolic Potential and Global Biogeography of the Atmospheric Methane-Oxidizing Upland Soil Cluster α.

Understanding of global methane sources and sinks is a prerequisite for the design of strategies to counteract global warming. Microbial methane oxidation in soils represents the largest biological sink for atmospheric methane. However, still very little is known about the identity, metabolic properties and distribution of the microbial group proposed to be responsible for most of this uptake, the uncultivated upland soil cluster α (USCα). Here, we reconstructed a draft genome of USCα from a combination of targeted cell sorting and metagenomes from forest soil, providing the first insights into its metabolic potential and environmental adaptation strategies. The 16S rRNA gene sequence recovered was distinctive and suggests this crucial group as a new genus within the Beijerinckiaceae, close to Methylocapsa. Application of a fluorescently labelled suicide substrate for the particulate methane monooxygenase enzyme (pMMO) coupled to 16S rRNA fluorescence in situ hybridisation (FISH) allowed for the first time a direct link of the high-affinity activity of methane oxidation to USCα cells in situ. Analysis of the global biogeography of this group further revealed its presence in previously unrecognized habitats, such as subterranean and volcanic biofilm environments, indicating a potential role of these environments in the biological sink for atmospheric methane.

Isolation, Biosynthesis and Chemical Modifications of Rubterolones A-F: Rare Tropolone Alkaloids from Actinomadura sp. 5-2.

The discovery of six new, highly substituted tropolone alkaloids, rubterolones A-F, from Actinomadura sp. 5-2, isolated from the gut of the fungus-growing termite Macrotermes natalensis is reported. Rubterolones were identified by using fungus-bacteria challenge assays and a HRMS-based dereplication strategy, and characterised by NMR and HRMS analyses and by X-ray crystallography. Feeding experiments and subsequent chemical derivatisation led to a first library of rubterolone derivatives (A-L). Genome sequencing and comparative analyses revealed their putative biosynthetic pathway, which was supported by feeding experiments. This study highlights how gut microbes can present a prolific source of secondary metabolites.

Coupling of ferredoxin and heterodisulfide reduction via electron bifurcation in hydrogenotrophic methanogenic archaea.

In methanogenic archaea growing on H(2) and CO(2) the first step in methanogenesis is the ferredoxin-dependent endergonic reduction of CO(2) with H(2) to formylmethanofuran and the last step is the exergonic reduction of the heterodisulfide CoM-S-S-CoB with H(2) to coenzyme M (CoM-SH) and coenzyme B (CoB-SH). We recently proposed that in hydrogenotrophic methanogens the two reactions are energetically coupled via the cytoplasmic MvhADG/HdrABC complex. It is reported here that the purified complex from Methanothermobacter marburgensis catalyzes the CoM-S-S-CoB-dependent reduction of ferredoxin with H(2). Per mole CoM-S-S-CoB added, 1 mol of ferredoxin (Fd) was reduced, indicating an electron bifurcation coupling mechanism: 2H(2) + Fd(OX) + CoM-S-S-CoB-->Fd(red)(2-) + CoM-SH + CoB-SH + 2H(+). This stoichiometry of coupling is consistent with an ATP gain per mole methane from 4 H(2) and CO(2) of near 0.5 deduced from an H(2)-threshold concentration of 8 Pa and a growth yield of up to 3 g/mol methane.

Pseudomonas stutzeri KC carries the pdt genes for carbon tetrachloride degradation on an integrative and conjugative element.

Pseudomonas stutzeri KC can rapidly degrade carbon tetrachloride (CCl4) to CO2 by a fortuitous reaction with pyridine-2,6-bis(thiocarboxylic acid), a metal chelator encoded by pdt genes. These genes were first identified after a spontaneous mutant, strain CTN1, lost the ability to degrade CCl4. Here we report the complete genome of strain KC and show that these pdt genes are located on an integrative and conjugative element (ICE), designated ICEPsstKC. Comparative genome analyses revealed homologues of pdt genes in genomes of members of other gammaproteobacterial orders. Discrepancies between the tree topologies of the deduced pdt gene products and the host phylogeny based on 16S rRNA provided evidence for horizontal gene transfer (HGT) in several sequenced strains of these orders. In addition to ICEPsstKC, HGT may be have been facilitated by other mobile genetic elements, as indicated by the location of the pdt gene cluster adjacent to fragments of other ICEs and prophages in several genome assemblies. We could here show that the majority of cells from the culture collection DSMZ had lost the ICE. The presence of the pdt gene cluster on mobile genetic elements has important implications for the bioremediation of CCl4 for bioremediation of CCl4 and needs consideration when selecting suitable strains.