Denitrifying bacteria anaerobically oxidize methane in the absence of Archaea

Abstract

Recently, a microbial consortium was shown to couple the anaerobic oxidation of methane to denitrification, predominantly in the form of nitrite reduction to dinitrogen gas. This consortium was dominated by bacteria of an as yet uncharacterized division and archaea of the order Methanosarcinales. The present manuscript reports on the upscaling of the enrichment culture, and addresses the role of the archaea in methane oxidation. The key gene of methanotrophic and methanogenic archaea, mcrA, was sequenced. The associated cofactor F(430) was shown to have a mass of 905 Da, the same as for methanogens and different from the heavier form (951 Da) found in methanotrophic archaea. After prolonged enrichment (> 1 year), no inhibition of anaerobic methane oxidation was observed in the presence of 20 mM bromoethane sulfonate, a specific inhibitor of MCR. Optimization of the cultivation conditions led to higher rates of methane oxidation and to the decline of the archaeal population, as shown by fluorescence in situ hybridization and quantitative MALDI-TOF analysis of F(430). Mass balancing showed that methane oxidation was still coupled to nitrite reduction in the total absence of oxygen. Together, our results show that bacteria can couple the anaerobic oxidation of methane to denitrification without the involvement of Archaea.

Potential roles of anaerobic ammonium and methane oxidation in the nitrogen cycle of wetland ecosystems

“…The wide range in growth temperature of anammox bacteria may promote their existence in wetlands. So far, the NC10 bacteria have only been detected in moderate fresh water sediments with temperatures between 10°C and 30°C (Ettwig et al 2008(Ettwig et al , 2009Hu et al 2009). …”

“…This discovery and potential contribution of anammox bacteria is important because it may necessitate a re-evaluation of the major fluxes in the global nitrogen budget and regulation of the processes leading to losses of fixed nitrogen. The addition of nitritedependent anaerobic methane oxidation (ANME coupled to denitrification) to the nitrogen cycle is even more recent (Raghoebarsing et al 2006;Ettwig et al 2008). Anoxic sediment from a fresh water canal in The Netherlands was used as an inoculum and supplied with methane, nitrate, and nitrite in a bioreactor with efficient biomass retention.…”

Archaea catalyze iron-dependent anaerobic oxidation of methane

“…Ammonium was measured colorimetrically at 623 nm after a 30-min reaction of 100-μL samples with alkaline phenol catalyzed by sodium nitroprusside at 37°C (36). For the miniaturized setup (Exetainer incubations), nitrate was quantified with HPLC using a conductivity detector (26). Fe 2+ was measured using the ferrozine method (37).…”

Anaerobic oxidation of methane: an underappreciated aspect of methane cycling in peatland ecosystems?

“…Additions of 13 C-labelled CH 4 and analysis of lipid biomarkers indicated that CH 4 -derived C was incorporated into the biomass of both the bacterium and the archaeon, but to a lesser degree in the archaeon. It is not clear what is driving this pattern, but it is important to point out that the enrichment culture in this study utilized nitrite (NO Ettwig et al (2010Ettwig et al ( , 2008 showed that NO − 2 -driven AOM can occur in the absence of an Archaeal partner and is carried out by an anaerobic denitrifying bacterium that oxidizes CH 4 aerobically by reducing NO − 2 to N 2 and O 2 . This could explain why less of the labeled CH 4 -C was recovered from the archaeon.…”

“…Panganiban et al, 1979;Smith et al, 1993;Eller et al, 2005), two landfills (Bjerg et al, 1995;Grossman et al, 2002), anoxic waste slurries (Malek and Weismann, 1988), a contaminated aquifer (Smith et al, 1991), and flooded-rice paddies (Miura et al, 1992;Murase and Kimura, 1994b), but much of the evidence is anecdotal in nature and strong evidence for AOM in freshwater systems has been limited. Work by Islas-Lima et al (2004) and Raghoebarsing et al (2006) has demonstrated that AOM in some freshwater systems is linked to denitrification and dentrifying bacteria, which provides an energetically favorable alternative to marine AOM linked to SR. Further work has suggested that AOM can be carried out by denitrifying bacteria in the absence of an archeal consortium (Ettwig et al, 2008) and that this process might be linked to nitrite (NO − 2 ) reduction and the production of oxygen (O 2 ) as an electron acceptor ; hence, implying aerobic metabolism under anoxic conditions.…”