The hunt for the most-wanted chemolithoautotrophic spookmicrobes.

Microorganisms are the drivers of biogeochemical methane and nitrogen cycles. Essential roles of chemolithoautotrophic microorganisms in these cycles were predicted long before their identification. Dedicated enrichment procedures, metagenomics surveys and single-cell technologies have enabled the identification of several new groups of most-wanted spookmicrobes, including novel methoxydotrophic methanogens that produce methane from methylated coal compounds and acetoclastic 'Candidatus Methanothrix paradoxum', which is active in oxic soils. The resultant energy-rich methane can be oxidized via a suite of electron acceptors. Recently, 'Candidatus Methanoperedens nitroreducens' ANME-2d archaea and 'Candidatus Methylomirabilis oxyfera' bacteria were enriched on nitrate and nitrite under anoxic conditions with methane as an electron donor. Although 'Candidatus Methanoperedens nitroreducens' and other ANME archaea can use iron citrate as an electron acceptor in batch experiments, the quest for anaerobic methane oxidizers that grow via iron reduction continues. In recent years, the nitrogen cycle has been expanded by the discovery of various ammonium-oxidizing prokaryotes, including ammonium-oxidizing archaea, versatile anaerobic ammonium-oxidizing (anammox) bacteria and complete ammonium-oxidizing (comammox) Nitrospira bacteria. Several biogeochemical studies have indicated that ammonium conversion occurs under iron-reducing conditions, but thus far no microorganism has been identified. Ultimately, iron-reducing and sulfate-dependent ammonium-oxidizing microorganisms await discovery.

Current perspectives on the application of N-damo and anammox in wastewater treatment.

The efficient treatment of wastewater for the removal of nitrogen is of key importance to prevent eutrophication and deoxygenation of receiving water bodies. In addition, ineffective wastewater treatment can be a source of greenhouse gasses. The application of newly discovered microbial processes, such as nitrite/nitrate-dependent methane oxidation (N-damo), can make wastewater treatment systems more sustainable; especially when they are combined with anaerobic ammonium oxidation (anammox). A treatment system based on these microbial processes will need oxygen supply for the production of nitrite. This oxygen may inhibit N-damo and anammox and careful regulation of the oxygen supply is of key importance for the success of the application of N-damo in wastewater treatment.

Iron assimilation and utilization in anaerobic ammonium oxidizing bacteria.

The most abundant transition metal in biological systems is iron. It is incorporated into protein cofactors and serves either catalytic, redox or regulatory purposes. Anaerobic ammonium oxidizing (anammox) bacteria rely heavily on iron-containing proteins - especially cytochromes - for their energy conservation, which occurs within a unique organelle, the anammoxosome. Both their anaerobic lifestyle and the presence of an additional cellular compartment challenge our understanding of iron processing. Here, we combine existing concepts of iron uptake, utilization and metabolism, and cellular fate with genomic and still limited biochemical and physiological data on anammox bacteria to propose pathways these bacteria may employ.

Methylacidiphilum fumariolicum SolV, a thermoacidophilic 'Knallgas' methanotroph with both an oxygen-sensitive and -insensitive hydrogenase.

Methanotrophs play a key role in balancing the atmospheric methane concentration. Recently, the microbial methanotrophic diversity was extended by the discovery of thermoacidophilic methanotrophs belonging to the Verrucomicrobia phylum in geothermal areas. Here we show that a representative of this new group, Methylacidiphilum fumariolicum SolV, is able to grow as a real 'Knallgas' bacterium on hydrogen/carbon dioxide, without addition of methane. The full genome of strain SolV revealed the presence of two hydrogen uptake hydrogenases genes, encoding an oxygen-sensitive (hup-type) and an oxygen-insensitive enzyme (hhy-type). The hhy-type hydrogenase was constitutively expressed and active and supported growth on hydrogen alone up to a growth rate of 0.03 h-1, at O2 concentrations below 1.5%. The oxygen-sensitive hup-type hydrogenase was expressed when oxygen was reduced to below 0.2%. This resulted in an increase of the growth rate to a maximum of 0.047 h-1, that is 60% of the rate on methane. The results indicate that under natural conditions where both hydrogen and methane might be limiting strain SolV may operate primarily as a methanotrophic 'Knallgas' bacterium. These findings argue for a revision of the role of hydrogen in methanotrophic ecosystems, especially in soil and related to consumption of atmospheric methane.

Intensive nitrogen loss over the Omani Shelf due to anammox coupled with dissimilatory nitrite reduction to ammonium.

A combination of stable isotopes ((15)N) and molecular ecological approaches was used to investigate the vertical distribution and mechanisms of biological N(2) production along a transect from the Omani coast to the central-northeastern (NE) Arabian Sea. The Arabian Sea harbors the thickest oxygen minimum zone (OMZ) in the world's oceans, and is considered to be a major site of oceanic nitrogen (N) loss. Short (<48 h) anoxic incubations with (15)N-labeled substrates and functional gene expression analyses showed that the anammox process was highly active, whereas denitrification was hardly detectable in the OMZ over the Omani shelf at least at the time of our sampling. Anammox was coupled with dissimilatory nitrite reduction to ammonium (DNRA), resulting in the production of double-(15)N-labeled N(2) from (15)NO(2)(-), a signal often taken as the lone evidence for denitrification in the past. Although the central-NE Arabian Sea has conventionally been regarded as the primary N-loss region, low potential N-loss rates at sporadic depths were detected at best. N-loss activities in this region likely experience high spatiotemporal variabilities as linked to the availability of organic matter. Our finding of greater N-loss associated with the more productive Omani upwelling region is consistent with results from other major OMZs. The close reliance of anammox on DNRA also highlights the need to take into account the effects of coupling N-transformations on oceanic N-loss and subsequent N-balance estimates.

Pyrosequencing of 16S rRNA gene amplicons to study the microbiota in the gastrointestinal tract of carp (Cyprinus carpio L.).

The microbes in the gastrointestinal (GI) tract are of high importance for the health of the host. In this study, Roche 454 pyrosequencing was applied to a pooled set of different 16S rRNA gene amplicons obtained from GI content of common carp (Cyprinus carpio) to make an inventory of the diversity of the microbiota in the GI tract. Compared to other studies, our culture-independent investigation reveals an impressive diversity of the microbial flora of the carp GI tract. The major group of obtained sequences belonged to the phylum Fusobacteria. Bacteroidetes, Planctomycetes and Gammaproteobacteria were other well represented groups of micro-organisms. Verrucomicrobiae, Clostridia and Bacilli (the latter two belonging to the phylum Firmicutes) had fewer representatives among the analyzed sequences. Many of these bacteria might be of high physiological relevance for carp as these groups have been implicated in vitamin production, nitrogen cycling and (cellulose) fermentation.

A comparative genomics study of genetic products potentially encoding ladderane lipid biosynthesis.

BACKGROUND: The fatty acids of anaerobic ammonium oxidizing (anammox) bacteria contain linearly concatenated cyclobutane moieties, so far unique to biology. These moieties are under high ring strain and are synthesised by a presently unknown biosynthetic pathway. RESULTS: Gene clusters encoding enzymes of fatty acid biosynthesis in the anammox bacterium Kuenenia stuttgartiensis and 137 other organisms were analysed and compared in silico to gain further insight into the pathway of (ladderane) fatty acid biosynthesis. In K. stuttgartiensis four large gene clusters encode fatty acid biosynthesis. Next to the regular enzyme complex needed for fatty acid biosynthesis (FASII), the presence of four putative S-adenosyl-methionine (SAM) radical enzymes, two enzymes similar to phytoene desaturases and many divergent paralogues of beta-ketoacyl-ACP synthase (fabF) were unusual. Surprisingly, extensive synteny was observed with FASII gene clusters in the deltaproteobacterium Desulfotalea psychrophila. No ladderane lipids were detected in lipid extracts of this organism but we did find unusual polyunsaturated hydrocarbons (PUHC), not detected in K. stuttgartiensis. CONCLUSION: We suggest that the unusual gene clusters of K. stuttgartiensis and D. psychrophila encode a novel pathway for anaerobic PUFA biosynthesis and that K. stuttgartiensis further processes PUFA into ladderane lipids, in similar fashion to the previously proposed route of ladderane lipid biosynthesis. However, the presence of divergent paralogues of fabF with radically different active site topologies may suggest an alternative pathway where ladderane moieties are synthesised externally and are recruited into the pathway of fatty acid biosynthesis.