Simultaneous Anaerobic and Aerobic Ammonia and Methane Oxidation under Oxygen Limitation Conditions

Methane and ammonia have to be removed from wastewater treatment effluent in order to discharge it to receiving water bodies. A potential solution for this is a combination of simultaneous ammonia and methane oxidation by anaerobic ammonia oxidation (anammox) bacteria and nitrite/nitrate-dependent anaerobic methane oxidation (N-damo) microorganisms. When applied, these microorganisms will be exposed to oxygen, but little is known about the effect of a low concentration of oxygen on a culture containing these microorganisms. In this study, a stable coculture containing anammox and N-damo microorganisms in a laboratory scale bioreactor was established under oxygen limitation. Membrane inlet mass spectrometry (MIMS) was used to directly measure the in situ simultaneous activity of N-damo, anammox, and aerobic ammonia-oxidizing microorganisms. In addition, batch tests revealed that the bioreactor also harbored aerobic methanotrophs and anaerobic methanogens. Together with fluorescence in situ hybridization (FISH) analysis and metagenomics, these results indicate that the combination of N-damo and anammox activity under the continuous supply of limiting oxygen concentrations is feasible and can be implemented for the removal of methane and ammonia from anaerobic digester effluents. IMPORTANCE Nitrogen in wastewater leads to eutrophication of the receiving water bodies, and methane is a potent greenhouse gas; it is therefore important that these are removed from wastewater. A potential solution for the simultaneous removal of nitrogenous compounds and methane is the application of a combination of nitrite/ nitrate-dependent methane oxidation (N-damo) and anaerobic ammonia oxidation (annamox). In order to do so, it is important to investigate the effect of oxygen on these two anaerobic processes. In this study, we investigate the effect of a continuous oxygen supply on the activity of an anaerobic methane- and ammonia-oxidizing coculture. The findings presented in this study are important for the potential application of these two microbial processes in wastewater treatment

Interactions of anaerobic ammonium oxidizers and sulfide-oxidizing bacteria in a substrate-limited model system mimicking the marine environment

In nature anaerobic ammonium oxidation (anammox) and denitrification processes convert fixed nitrogen to gaseous nitrogen compounds, which are then released to the atmosphere. While anammox bacteria produce N2 from ammonium and nitrite, in the denitrification process nitrate and nitrite are converted to N2 and the greenhouse gas nitrous oxide (N2O). Furthermore, nitrite needed by the anammox bacteria can be supplied by nitrate reduction to nitrite. Consequently, the interplay between nitrogen-transforming microorganisms control the amount of harmless N2 or the greenhouse gas N2O released to the atmosphere. Therefore, it is important to understand the interactions of these microorganisms in the natural environment, where dynamic conditions result in fluctuating substrate concentrations. Here, we studied the interactions between the sulfide-oxidizing denitrifier Sedimenticola selenatireducens and the anammox bacterium Scalindua brodae in a bioreactor mimicking the marine environment by creating sulfide, ammonium and nitrate limitation in distinct operational phases. Through a microbial interaction, Se. selenatireducens reduced nitrate to nitrite, which together with the supplied ammonium was converted to N2 by Sc. Brodae. Using comparative transcriptomics, we determined that Sc. Brodae and Se. selenatireducens had significant responses to ammonium and nitrate limitation, respectively, indicating that the activities of these microorganisms are regulated by different nitrogen compounds

FACIL: Fast and Accurate Genetic Code Inference and Logo

Motivation: The intensification of DNA sequencing will increasingly unveil uncharacterized species with potential alternative genetic codes. A total of 0.65% of the DNA sequences currently in Genbank encode their proteins with a variant genetic code, and these exceptions occur in many unrelated taxa. Results: We introduce FACIL (Fast and Accurate genetic Code Inference and Logo), a fast and reliable tool to evaluate nucleic acid sequences for their genetic code that detects alternative codes even in species distantly related to known organisms. To illustrate this, we apply FACIL to a set of mitochondrial genomic contigs of Globobulimina pseudospinescens. This foraminifer does not have any sequenced close relative in the databases, yet we infer its alternative genetic code with high confidence values. Results are intuitively visualized in a Genetic Code Logo

The thermoacidophilic methanotroph Methylacidiphilum fumariolicum SolV oxidizes subatmospheric H₂ with a high-affinity, membrane-associated [NiFe] hydrogenase

The trace amounts (0.53 ppmv) of atmospheric hydrogen gas (H2) can be utilized by microorganisms to persist during dormancy. This process is catalyzed by certain Actinobacteria, Acidobacteria, and Chloroflexi, and is estimated to convert 75 × 1012 g H2 annually, which is half of the total atmospheric H2. This rapid atmospheric H2 turnover is hypothesized to be catalyzed by high-affinity [NiFe] hydrogenases. However, apparent high-affinity H2 oxidation has only been shown in whole cells, rather than for the purified enzyme. Here, we show that the membrane-associated hydrogenase from the thermoacidophilic methanotroph Methylacidiphilum fumariolicum SolV possesses a high apparent affinity (Km(app) = 140 nM) for H2 and that methanotrophs can oxidize subatmospheric H2. Our findings add to the evidence that the group 1h [NiFe] hydrogenase is accountable for atmospheric H2 oxidation and that it therefore could be a strong controlling factor in the global H2 cycle. We show that the isolated enzyme possesses a lower affinity (Km = 300 nM) for H2 than the membrane-associated enzyme. Hence, the membrane association seems essential for a high affinity for H2. The enzyme is extremely thermostable and remains folded up to 95 °C. Strain SolV is the only known organism in which the group 1h [NiFe] hydrogenase is responsible for rapid growth on H2 as sole energy source as well as oxidation of subatmospheric H2. The ability to conserve energy from H2 could increase fitness of verrucomicrobial methanotrophs in geothermal ecosystems with varying CH4 fluxes. We propose that H2 oxidation can enhance growth of methanotrophs in aerated methane-driven ecosystems. Group 1h [NiFe] hydrogenases could therefore contribute to mitigation of global warming, since CH4 is an important and extremely potent greenhouse gas.</p

A novel mesocosm set-up reveals strong methane emission reduction in submerged peat moss Sphagnum cuspidatum by tightly associated methanotrophs

Wetlands present the largest natural sources of methane (CH_4) and their potential CH_4 emissions greatly vary due to the activity of CH_4-oxidizing bacteria associated with wetland plant species. In this study, the association of CH_4-oxidizing bacteria with submerged Sphagnum peat mosses was studied, followed by the development of a novel mesocosm set-up. This set-up enabled the precise control of CH_4 input and allowed for monitoring the dissolved CH_4in a Sphagnum moss layer while mimicking natural conditions. Two mesocosm set-ups were used in parallel: one containing a Sphagnum moss layer in peat water, and a control only containing peat water. Moss-associated CH_4 oxidizers in the field could reduce net CH_4 emission up to 93%, and in the mesocosm set-up up to 31%. Furthermore, CH_4 oxidation was only associated with Sphagnum, and did not occur in peat water. Especially methanotrophs containing a soluble methane monooxygenase enzyme were significantly enriched during the 32 day mesocosm incubations. Together these findings showed the new mesocosm setup is very suited to study CH_4 cycling in submerged Sphagnum moss community under controlled conditions. Furthermore, the tight associated between Sphagnum peat mosses and methanotrophs can significantly reduce CH_4 emissions in submerged peatlands

Amsterdam urban canals contain novel niches for methane-cycling microorganisms

Urbanised environments have been identified as hotspots of anthropogenic methane emissions. Especially urban aquatic ecosystems are increasingly recognised as important sources of methane. However, the microbiology behind these emissions remains unexplored. Here, we applied microcosm incubations and molecular analyses to investigate the methane‐cycling community of the Amsterdam canal system in the Netherlands. The sediment methanogenic communities were dominated by Methanoregulaceae and Methanosaetaceae, with co‐occurring methanotrophic Methanoperedenaceae and Methylomirabilaceae indicating the potential for anaerobic methane oxidation. Methane was readily produced after substrate amendment, suggesting an active but substrate‐limited methanogenic community. Bacterial 16S rRNA gene amplicon sequencing of the sediment revealed a high relative abundance of Thermodesulfovibrionia. Canal wall biofilms showed the highest initial methanotrophic potential under oxic conditions compared to the sediment. During prolonged incubations the maximum methanotrophic rate increased to 8.08 mmol g(DW) (−1) d(−1) that was concomitant with an enrichment of Methylomonadaceae bacteria. Metagenomic analysis of the canal wall biofilm lead to the recovery of a single methanotroph metagenome‐assembled genome. Taxonomic analysis showed that this methanotroph belongs to the genus Methyloglobulus. Our results underline the importance of previously unidentified and specialised environmental niches at the nexus of the natural and human‐impacted carbon cycle

Complete Genome Sequence of the Aerobic Facultative Methanotroph Methylocella tundrae Strain T4

Methylocella tundrae T4T is a facultative aerobic methanotroph which was isolated from an acidic tundra wetland and possesses only a soluble methane monooxygenase. The complete genome, which includes two megaplasmids, was sequenced using a combination of Illumina and Nanopore technologies. One of the megaplasmids carries a propane monooxygenase gene cluster

Draft genome of a novel methanotrophic Methylobacter sp. from the volcanic soils of Pantelleria Island

The genus Methylobacter is considered an important and often dominant group of aerobic methane-oxidizing bacteria in many oxic ecosystems, where members of this genus contribute to the reduction of CH4 emissions. Metagenomic studies of the upper oxic layers of geothermal soils of the Favara Grande, Pantelleria, Italy, revealed the presence of various methane-oxidizing bacteria, and resulted in a near complete metagenome assembled genome (MAG) of an aerobic methanotroph, which was classified as a Methylobacter species. In this study, the Methylobacter sp. B2 MAG was used to investigate its metabolic potential and phylogenetic affiliation. The MAG has a size of 4,086,539&nbsp;bp, consists of 134 contigs and 3955 genes were found, of which 3902 were protein coding genes. All genes for CH4 oxidation to CO2 were detected, including pmoCAB encoding particulate methane monooxygenase (pMMO) and xoxF encoding a methanol dehydrogenase. No gene encoding a formaldehyde dehydrogenase was present and the formaldehyde to formate conversion follows the tetrahydromethanopterin (H4MPT) pathway. “Ca. Methylobacter favarea” B2 uses the Ribulose-Mono-Phosphate (RuMP) pathway for carbon fixation. Analysis of the MAG indicates that Na+/H+ antiporters and the urease system might be important in the maintenance of pH homeostasis of this strain to cope with acidic conditions. So far, thermoacidophilic Methylobacter species have not been isolated, however this study indicates that members of the genus Methylobacter can be found in distinct ecosystems and their presence is not restricted to freshwater or marine sediments

A novel mesocosm set-up reveals strong methane emission reduction in submerged peat moss Sphagnum cuspidatum by tightly associated methanotrophs

Wetlands present the largest natural sources of methane (CH_4) and their potential CH_4 emissions greatly vary due to the activity of CH_4-oxidizing bacteria associated with wetland plant species. In this study, the association of CH_4-oxidizing bacteria with submerged Sphagnum peat mosses was studied, followed by the development of a novel mesocosm set-up. This set-up enabled the precise control of CH_4 input and allowed for monitoring the dissolved CH_4in a Sphagnum moss layer while mimicking natural conditions. Two mesocosm set-ups were used in parallel: one containing a Sphagnum moss layer in peat water, and a control only containing peat water. Moss-associated CH_4 oxidizers in the field could reduce net CH_4 emission up to 93%, and in the mesocosm set-up up to 31%. Furthermore, CH_4 oxidation was only associated with Sphagnum, and did not occur in peat water. Especially methanotrophs containing a soluble methane monooxygenase enzyme were significantly enriched during the 32 day mesocosm incubations. Together these findings showed the new mesocosm setup is very suited to study CH_4 cycling in submerged Sphagnum moss community under controlled conditions. Furthermore, the tight associated between Sphagnum peat mosses and methanotrophs can significantly reduce CH_4 emissions in submerged peatlands