Methane oxidation in anoxic lake water stimulated by nitrate and sulfate addition.

Methanotrophic bacteria play a key role in limiting methane emissions from lakes. It is generally assumed that methanotrophic bacteria are mostly active at the oxic-anoxic transition zone in stratified lakes, where they use oxygen to oxidize methane. Here, we describe a methanotroph of the genera Methylobacter that is performing high-rate (up to 72 µM day-1 ) methane oxidation in the anoxic hypolimnion of the temperate Lacamas Lake (Washington, USA), stimulated by both nitrate and sulfate addition. Oxic and anoxic incubations both showed active methane oxidation by a Methylobacter species, with anoxic rates being threefold higher. In anoxic incubations, Methylobacter cell numbers increased almost two orders of magnitude within 3 days, suggesting that this specific Methylobacter species is a facultative anaerobe with a rapid response capability. Genomic analysis revealed adaptations to oxygen-limitation as well as pathways for mixed-acid fermentation and H2 production. The denitrification pathway was incomplete, lacking the genes narG/napA and nosZ, allowing only for methane oxidation coupled to nitrite-reduction. Our data suggest that Methylobacter can be an important driver of the conversion of methane in oxygen-limited lake systems and potentially use alternative electron acceptors or fermentation to remain active under oxygen-depleted conditions.

Hydrogen-independent CO2 reduction dominates methanogenesis in five temperate lakes that differ in trophic states.

Emissions of microbially produced methane (CH4) from lake sediments are a major source of this potent greenhouse gas to the atmosphere. The rates of CH4 production and emission are believed to be influenced by electron acceptor distributions and organic carbon contents, which in turn are affected by anthropogenic inputs of nutrients leading to eutrophication. Here, we investigate how eutrophication influences the abundance and community structure of CH4 producing Archaea and methanogenesis pathways across time-resolved sedimentary records of five Swiss lakes with well-characterized trophic histories. Despite higher CH4 concentrations which suggest higher methanogenic activity in sediments of eutrophic lakes, abundances of methanogens were highest in oligotrophic lake sediments. Moreover, while the methanogenic community composition differed significantly at the lowest taxonomic levels (OTU), depending on whether sediment layers had been deposited under oligotrophic or eutrophic conditions, it showed no clear trend in relation to in situ distributions of electron acceptors. Remarkably, even though methanogenesis from CO2-reduction was the dominant pathway in all sediments based on carbon isotope fractionation values, taxonomic identities, and genomes of resident methanogens, CO2-reduction with hydrogen (H2) was thermodynamically unfavorable based on measured reactant and product concentrations. Instead, strong correlations between genomic abundances of CO2-reducing methanogens and anaerobic bacteria with potential for extracellular electron transfer suggest that methanogenic CO2-reduction in lake sediments is largely powered by direct electron transfer from syntrophic bacteria without involvement of H2 as an electron shuttle.

Assessing the Effect of Humic Substances and Fe(III) as Potential Electron Acceptors for Anaerobic Methane Oxidation in a Marine Anoxic System.

Marine anaerobic methane oxidation (AOM) is generally assumed to be coupled to sulfate reduction, via a consortium of anaerobic methane-oxidizing archaea (ANME) and sulfate-reducing bacteria (SRB). ANME-1 are, however, often found as single cells, or only loosely aggregated with SRB, suggesting they perform a form of AOM independent of sulfate reduction. Oxidized metals and humic substances have been suggested as potential electron acceptors for ANME, but up to now, AOM linked to reduction of these compounds has only been shown for the ANME-2 and ANME-3 clades. Here, the effect of the electron acceptors anthraquinone-disulfonate (AQDS), a humic acids analog, and Fe3+ on anaerobic methane oxidation were assessed by incubation experiments with anoxic Black Sea water containing ANME-1b. Incubation experiments with 13C-methane and AQDS showed a stimulating effect of AQDS on methane oxidation. Fe3+ enhanced the ANME-1b abundance but did not substantially increase methane oxidation. Sodium molybdate, which was added as an inhibitor of sulfate reduction, surprisingly enhanced methane oxidation, possibly related to the dominant abundance of Sulfurospirillum in those incubations. The presented data suggest the potential involvement of ANME-1b in AQDS-enhanced anaerobic methane oxidation, possibly via electron shuttling to AQDS or via interaction with other members of the microbial community.

Impact of Electron Acceptor Availability on Methane-Influenced Microorganisms in an Enrichment Culture Obtained From a Stratified Lake.

Methanotrophs are of major importance in limiting methane emissions from lakes. They are known to preferably inhabit the oxycline of stratified water columns, often assumed due to an intolerance to atmospheric oxygen concentrations, but little is known on the response of methanotrophs to different oxygen concentrations as well as their preference for different electron acceptors. In this study, we enriched a methanotroph of the Methylobacter genus from the oxycline and the anoxic water column of a stratified lake, which was also present in the oxic water column in the winter. We tested the response of this Methylobacter-dominated enrichment culture to different electron acceptors, i.e., oxygen, nitrate, sulfate, and humic substances, and found that, in contrast to earlier results with water column incubations, oxygen was the preferred electron acceptor, leading to methane oxidation rates of 45-72 pmol cell-1 day-1. Despite the general assumption of methanotrophs preferring microaerobic conditions, methane oxidation was most efficient under high oxygen concentrations (>600 µM). Low (<30 µM) oxygen concentrations still supported methane oxidation, but no methane oxidation was observed with trace oxygen concentrations (<9 µM) or under anoxic conditions. Remarkably, the presence of nitrate stimulated methane oxidation rates under oxic conditions, raising the methane oxidation rates by 50% when compared to oxic incubations with ammonium. Under anoxic conditions, no net methane consumption was observed; however, methanotroph abundances were two to three times higher in incubations with nitrate and sulfate compared to anoxic incubations with ammonium as the nitrogen source. Metagenomic sequencing revealed the absence of a complete denitrification pathway in the dominant methanotroph Methylobacter, but the most abundant methylotroph Methylotenera seemed capable of denitrification, which can possibly play a role in the enhanced methane oxidation rates under nitrate-rich conditions.