Anja Poehlein

The genome of the ammonia-oxidizing Candidatus Nitrososphaera gargensis: insights into metabolic versatility and environmental adaptations.

The cohort of the ammonia-oxidizing archaea (AOA) of the phylum Thaumarchaeota is a diverse, widespread and functionally important group of microorganisms in many ecosystems. However, our understanding of their biology is still very rudimentary in part because all available genome sequences of this phylum are from members of the Nitrosopumilus cluster. Here we report on the complete genome sequence of Candidatus Nitrososphaera gargensis obtained from an enrichment culture, representing a different evolutionary lineage of AOA frequently found in high numbers in many terrestrial environments. With its 2.83 Mb the genome is much larger than that of other AOA. The presence of a high number of (active) IS elements/transposases, genomic islands, gene duplications and a complete CRISPR/Cas defence system testifies to its dynamic evolution consistent with low degree of synteny with other thaumarchaeal genomes. As expected, the repertoire of conserved enzymes proposed to be required for archaeal ammonia oxidation is encoded by N. gargensis, but it can also use urea and possibly cyanate as alternative ammonia sources. Furthermore, its carbon metabolism is more flexible at the central pyruvate switch point, encompasses the ability to take up small organic compounds and might even include an oxidative pentose phosphate pathway. Furthermore, we show that thaumarchaeota produce cofactor F420 as well as polyhydroxyalkanoates. Lateral gene transfer from bacteria and euryarchaeota has contributed to the metabolic versatility of N. gargensis. This organisms is well adapted to its niche in a heavy metal-containing thermal spring by encoding a multitude of heavy metal resistance genes, chaperones and mannosylglycerate as compatible solute and has the genetic ability to respond to environmental changes by signal transduction via a large number of two-component systems, by chemotaxis and flagella-mediated motility and possibly even by gas vacuole formation. These findings extend our understanding of thaumarchaeal evolution and physiology and offer many testable hypotheses for future experimental research on these nitrifiers.

An ancient pathway combining carbon dioxide fixation with the generation and utilization of a sodium ion gradient for ATP synthesis.

Synthesis of acetate from carbon dioxide and molecular hydrogen is considered to be the first carbon assimilation pathway on earth. It combines carbon dioxide fixation into acetyl-CoA with the production of ATP via an energized cell membrane. How the pathway is coupled with the net synthesis of ATP has been an enigma. The anaerobic, acetogenic bacterium Acetobacterium woodii uses an ancient version of this pathway without cytochromes and quinones. It generates a sodium ion potential across the cell membrane by the sodium-motive ferredoxin:NAD oxidoreductase (Rnf). The genome sequence of A. woodii solves the enigma: it uncovers Rnf as the only ion-motive enzyme coupled to the pathway and unravels a metabolism designed to produce reduced ferredoxin and overcome energetic barriers by virtue of electron-bifurcating, soluble enzymes.

New Mode of Energy Metabolism in the Seventh Order of Methanogens as Revealed by Comparative Genome Analysis of “Candidatus Methanoplasma termitum”

ABSTRACT The recently discovered seventh order of methanogens, the Methanomassiliicoccales (previously referred to as “Methanoplasmatales”), so far consists exclusively of obligately hydrogen-dependent methylotrophs. We sequenced the complete genome of “Candidatus Methanoplasma termitum” from a highly enriched culture obtained from the intestinal tract of termites and compared it with the previously published genomes of three other strains from the human gut, including the first isolate of the order. Like all other strains, “Ca. Methanoplasma termitum” lacks the entire pathway for CO2 reduction to methyl coenzyme M and produces methane by hydrogen-dependent reduction of methanol or methylamines, which is consistent with additional physiological data. However, the shared absence of cytochromes and an energy-converting hydrogenase for the reoxidation of the ferredoxin produced by the soluble heterodisulfide reductase indicates that Methanomassiliicoccales employ a new mode of energy metabolism, which differs from that proposed for the obligately methylotrophic Methanosphaera stadtmanae. Instead, all strains possess a novel complex that is related to the F420:methanophenazine oxidoreductase (Fpo) of Methanosarcinales but lacks an F420-oxidizing module, resembling the apparently ferredoxin-dependent Fpo-like homolog in Methanosaeta thermophila. Since all Methanomassiliicoccales also lack the subunit E of the membrane-bound heterodisulfide reductase (HdrDE), we propose that the Fpo-like complex interacts directly with subunit D, forming an energy-converting ferredoxin:heterodisulfide oxidoreductase. The dual function of heterodisulfide in Methanomassiliicoccales, which serves both in electron bifurcation and as terminal acceptor in a membrane-associated redox process, may be a unique characteristic of the novel order.

Metagenome Survey of a Multispecies and Alga-Associated Biofilm Revealed Key Elements of Bacterial-Algal Interactions in Photobioreactors

ABSTRACT Photobioreactors (PBRs) are very attractive for sunlight-driven production of biofuels and capturing of anthropogenic CO2. One major problem associated with PBRs however, is that the bacteria usually associated with microalgae in nonaxenic cultures can lead to biofouling and thereby affect algal productivity. Here, we report on a phylogenetic, metagenome, and functional analysis of a mixed-species bacterial biofilm associated with the microalgae Chlorella vulgaris and Scenedesmus obliquus in a PBR. The biofilm diversity and population dynamics were examined through 16S rRNA phylogeny. Overall, the diversity was rather limited, with approximately 30 bacterial species associated with the algae. The majority of the observed microorganisms were affiliated with Alphaproteobacteria, Betaproteobacteria, and Bacteroidetes. A combined approach of sequencing via GS FLX Titanium from Roche and HiSeq 2000 from Illumina resulted in the overall production of 350 Mbp of sequenced DNA, 165 Mbp of which was assembled in larger contigs with a maximum size of 0.2 Mbp. A KEGG pathway analysis suggested high metabolic diversity with respect to the use of polymers and aromatic and nonaromatic compounds. Genes associated with the biosynthesis of essential B vitamins were highly redundant and functional. Moreover, a relatively high number of predicted and functional lipase and esterase genes indicated that the alga-associated bacteria are possibly a major sink for lipids and fatty acids produced by the microalgae. This is the first metagenome study of microalga- and PBR-associated biofilm bacteria, and it gives new clues for improved biofuel production in PBRs.

The Janthinobacterium sp. HH01 Genome Encodes a Homologue of the V. cholerae CqsA and L. pneumophila LqsA Autoinducer Synthases

Janthinobacteria commonly form biofilms on eukaryotic hosts and are known to synthesize antibacterial and antifungal compounds. Janthinobacterium sp. HH01 was recently isolated from an aquatic environment and its genome sequence was established. The genome consists of a single chromosome and reveals a size of 7.10 Mb, being the largest janthinobacterial genome so far known. Approximately 80% of the 5,980 coding sequences (CDSs) present in the HH01 genome could be assigned putative functions. The genome encodes a wealth of secretory functions and several large clusters for polyketide biosynthesis. HH01 also encodes a remarkable number of proteins involved in resistance to drugs or heavy metals. Interestingly, the genome of HH01 apparently lacks the N-acylhomoserine lactone (AHL)-dependent signaling system and the AI-2-dependent quorum sensing regulatory circuit. Instead it encodes a homologue of the Legionella- and Vibrio-like autoinducer (lqsA/cqsA) synthase gene which we designated jqsA. The jqsA gene is linked to a cognate sensor kinase (jqsS) which is flanked by the response regulator jqsR. Here we show that a jqsA deletion has strong impact on the violacein biosynthesis in Janthinobacterium sp. HH01 and that a jqsA deletion mutant can be functionally complemented with the V. cholerae cqsA and the L. pneumophila lqsA genes.

First Insights into the Genome of the Gram-Negative, Endospore-Forming Organism Sporomusa ovata Strain H1 DSM 2662

ABSTRACT The genome of Sporomusa ovata strain H1 DSM 2662, an anaerobic, Gram-negative endospore-forming bacterium, was sequenced. S. ovata uses N-methyl compounds, primary alcohols, fatty acids, and H2 and CO2 as energy and carbon sources to produce acetate. The genome harbors one chromosome, which encodes proteins typical for sporulation.

Genome-guided analysis of physiological and morphological traits of the fermentative acetate oxidizer Thermacetogenium phaeum

BackgroundThermacetogenium phaeum is a thermophilic strictly anaerobic bacterium oxidizing acetate to CO2 in syntrophic association with a methanogenic partner. It can also grow in pure culture, e.g., by fermentation of methanol to acetate. The key enzymes of homoacetate fermentation (Wood-Ljungdahl pathway) are used both in acetate oxidation and acetate formation. The obvious reversibility of this pathway in this organism is of specific interest since syntrophic acetate oxidation operates close to the energetic limitations of microbial life.ResultsThe genome of Th. phaeum is organized on a single circular chromosome and has a total size of 2,939,057 bp. It comprises 3.215 open reading frames of which 75% could be assigned to a gene function. The G+C content is 53.88 mol%. Many CRISPR sequences were found, indicating heavy phage attack in the past. A complete gene set for a phage was found in the genome, and indications of phage action could also be observed in culture. The genome contained all genes required for CO2 reduction through the Wood-Ljungdahl pathway, including two formyl tetrahydrofolate ligases, three carbon monoxide dehydrogenases, one formate hydrogenlyase complex, three further formate dehydrogenases, and three further hydrogenases. The bacterium contains a menaquinone MQ-7. No indications of cytochromes or Rnf complexes could be found in the genome.ConclusionsThe information obtained from the genome sequence indicates that Th. phaeum differs basically from the three homoacetogenic bacteria sequenced so far, i.e., the sodium ion-dependent Acetobacterium woodii, the ethanol-producing Clostridium ljungdahlii, and the cytochrome-containing Moorella thermoacetica. The specific enzyme outfit of Th. phaeum obviously allows ATP formation both in acetate formation and acetate oxidation.

The Complete Genome Sequence of Clostridium aceticum: a Missing Link between Rnf- and Cytochrome-Containing Autotrophic Acetogens

ABSTRACT Clostridium aceticum was the first isolated autotrophic acetogen, converting CO2 plus H2 or syngas to acetate. Its genome has now been completely sequenced and consists of a 4.2-Mbp chromosome and a small circular plasmid of 5.7 kbp. Sequence analysis revealed major differences from other autotrophic acetogens. C. aceticum contains an Rnf complex for energy conservation (via pumping protons or sodium ions). Such systems have also been found in C. ljungdahlii and Acetobacterium woodii. However, C. aceticum also contains a cytochrome, as does Moorella thermoacetica, which has been proposed to be involved in the generation of a proton gradient. Thus, C. aceticum seems to represent a link between Rnf- and cytochrome-containing autotrophic acetogens. In C. aceticum, however, the cytochrome is probably not involved in an electron transport chain that leads to proton translocation, as no genes for quinone biosynthesis are present in the genome. IMPORTANCE Autotrophic acetogenic bacteria are receiving more and more industrial focus, as CO2 plus H2 as well as syngas are interesting new substrates for biotechnological processes. They are both cheap and abundant, and their use, if it results in sustainable products, also leads to reduction of greenhouse gases. Clostridium aceticum can use both gas mixtures, is phylogenetically not closely related to the commonly used species, and may thus become an even more attractive workhorse. In addition, its energy metabolism, which is characterized here, and the ability to synthesize cytochromes might offer new targets for improving the ATP yield by metabolic engineering and thus allow use of C. aceticum for production of compounds by pathways that currently present challenges for energy-limited acetogens. Autotrophic acetogenic bacteria are receiving more and more industrial focus, as CO2 plus H2 as well as syngas are interesting new substrates for biotechnological processes. They are both cheap and abundant, and their use, if it results in sustainable products, also leads to reduction of greenhouse gases. Clostridium aceticum can use both gas mixtures, is phylogenetically not closely related to the commonly used species, and may thus become an even more attractive workhorse. In addition, its energy metabolism, which is characterized here, and the ability to synthesize cytochromes might offer new targets for improving the ATP yield by metabolic engineering and thus allow use of C. aceticum for production of compounds by pathways that currently present challenges for energy-limited acetogens.

Complete Genome Sequence of the Type Strain Cupriavidus necator N-1

Here we announce the complete genome sequence of the copper-resistant bacterium Cupriavidus necator N-1, the type strain of the genus Cupriavidus. The genome consists of two chromosomes and two circular plasmids. Based on genome comparison, the chromosomes of C. necator N-1 share a high degree of similarity with the two chromosomal replicons of the bioplastic-producing hydrogen bacterium Ralstonia eutropha H16. The two strains differ in their plasmids and the presence of hydrogenase genes, which are absent in strain N-1.

Analysis and comparative genomics of ICEMh1, a novel integrative and conjugative element (ICE) of Mannheimia haemolytica

OBJECTIVES The aim of this study was to identify and analyse the first integrative and conjugative element (ICE) from Mannheimia haemolytica, the major bacterial component of the bovine respiratory disease (BRD) complex. METHODS The novel ICEMh1 was discovered in the whole-genome sequence of M. haemolytica 42548 by sequence analysis and comparative genomics. Transfer of ICEMh1 was confirmed by conjugation into Pasteurella multocida recipient cells. RESULTS ICEMh1 has a size of 92,345 bp and harbours 107 genes. It integrates into a chromosomal tRNA(Leu) copy. Within two resistance gene regions of ∼ 7.4 and 3.3 kb, ICEMh1 harbours five genes, which confer resistance to streptomycin (strA and strB), kanamycin/neomycin (aphA1), tetracycline [tetR-tet(H)] and sulphonamides (sul2). ICEMh1 is related to the recently described ICEPmu1 and both ICEs seem to have evolved from a common ancestor. A region of ICEMh1 that is absent in ICEPmu1 was found in putative ICE regions of other M. haemolytica genomes, suggesting a recombination event between two ICEs. ICEMh1 transfers to P. multocida by conjugation, in which it also uses a tRNA(Leu) as the integration site. PCR assays and susceptibility testing confirmed the presence and activity of the ICEMh1-associated resistance genes in the P. multocida recipient. CONCLUSIONS These findings showed that ICEs, with structurally variable resistance gene regions, are present in BRD-associated Pasteurellaceae, can easily spread across genus borders and enable the acquisition of multidrug resistance via a single horizontal gene transfer event. This poses a threat to efficient antimicrobial chemotherapy of BRD-associated bacterial pathogens.