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Microbial Ecology of Methanotrophy in Streams Along a Gradient of CH4 Availability

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Microbial Ecology of Methanotrophy in Streams Along a Gradient of CH4 Availability fmicb-11-00771 May 12, 2020 Time: 19:58 # 1 ORIGINAL RESEARCH published: 14 May 2020 doi: 10.3389/fmicb.2020.00771 Microbial Ecology of Methanotrophy in Streams Along a Gradient of CH4 Availability Alexandre Bagnoud1†, Paraskevi Pramateftaki1, Matthew J. Bogard2†, Tom J. Battin1 and Hannes Peter1* 1 Stream Biofilm and Ecosystem Research Laboratory, School of Architecture, Civil and Environmental Engineering, École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland, 2 Groupe de recherche interuniversitaire en limnologie, Département des sciences biologiques, Université du Québec à Montréal, Montréal, QC, Canada Edited by: Jonathan P. Zehr, Despite the recognition of streams and rivers as sources of methane (CH4) to the University of California, Santa Cruz, atmosphere, the role of CH4 oxidation (MOX) in these ecosystems remains poorly United States understood to date. Here, we measured the kinetics of MOX in stream sediments of Reviewed by: 14 sites to resolve the ecophysiology of CH4 oxidizing bacteria (MOB) communities. Adrian Ho, Leibniz University Hannover, Germany The streams cover a gradient of land cover and associated physicochemical parameter Ronald Oremland, and differed in stream- and porewater CH4 concentrations. Michealis–Menten kinetic United States Geological Survey (USGS), United States parameter of MOX, maximum reaction velocity (Vmax), and CH4 concentration at half *Correspondence: Vmax (KS) increased with CH4 supply. KS values in the micromolar range matched the Hannes Peter CH4 concentrations measured in shallow stream sediments and indicate that MOX is hannes.peter@epfl.ch mostly driven by low-affinity MOB. 16S rRNA gene sequencing identified MOB classified † Present address: as Methylococcaceae and particularly Crenothrix. Their relative abundance correlated Alexandre Bagnoud, Haute École d’Ingénierie et de with pmoA gene counts and MOX rates, underscoring their pivotal role as CH4 oxidizers Gestion du Canton de Vaud, Institut in stream sediments. Building on the concept of enterotypes, we identify two distinct de Génie Thermique, Yverdon-les-Bains, Switzerland groups of co-occurring MOB. While there was no taxonomic difference among the Matthew J. Bogard, members of each cluster, one cluster contained abundant and common MOB, whereas Department of Biological Sciences, the other cluster contained rare operational taxonomic units (OTUs) specific to a University of Lethbridge, Lethbridge, AB, Canada subset of streams. These integrated analyses of changes in MOB community structure, gene abundance, and the corresponding ecosystem process contribute to a better Specialty section: understanding of the distal controls on MOX in streams. This article was submitted to Aquatic Microbiology, Keywords: Methylococcaceae, Crenothrix, Michaelis–Menten, enterotypes, pmoA a section of the journal Frontiers in Microbiology Received: 17 January 2020 INTRODUCTION Accepted: 31 March 2020 Published: 14 May 2020 In streambeds, microorganisms organized in surface-attached and matrix-enclosed biofilms drive Citation: biogeochemical processes that regulate the outgassing of carbon dioxide (CO2), methane (CH4), Bagnoud A, Pramateftaki P, and nitrous oxide (N2O) (Battin et al., 2016). While the study of stream CO2 biogeochemistry has Bogard MJ, Battin TJ and Peter H received considerable attention over the last decade, the biogeochemistry of CH4 (Crawford et al., (2020) Microbial Ecology of Methanotrophy in Streams Along 2014; Trimmer et al., 2015; Stanley et al., 2016) and specifically the microbial ecology of aerobic a Gradient of CH4 Availability. CH4 oxidizing bacteria (MOB) in streams remains poorly understood (Trimmer et al., 2015, 2010; Front. Microbiol. 11:771. Stanley et al., 2016). Streams outgas substantial amounts of CH4 originating from groundwater doi: 10.3389/fmicb.2020.00771 and methanogenesis within the streambed (Chaudhary et al., 2014; Stanley et al., 2016). Globally, Frontiers in Microbiology| www.frontiersin.org 1 May 2020| Volume 11| Article 771 fmicb-11-00771 May 12, 2020 Time: 19:58 # 2 Bagnoud et al. MOX in Stream Sediments streams and rivers may emit 26.8 Tg CH4 per year, which recent predictions that eutrophication will continue to increase is similar to the combined contribution of other natural during the 21st century (Sinha et al., 2017). CH4 sources such as wildfires, termites, CH4 hydrates, and permafrost (Stanley et al., 2016). The presence of wetlands and human activity in stream catchments are positively related MATERIALS AND METHODS to streamwater CH4 concentrations and CH4 evasion fluxes (Chaudhary et al., 2014; Stanley et al., 2016). Land cover is Study Sites and Sampling linked to streamwater CH4 concentration, as the excess delivery We sampled 14 sites (located between 378 and 1,189 m above of organic matter and nutrients leads to oxygen depletion and sea level) of nine streams in Switzerland between March and thus stimulates anaerobic methanogenesis. July 2016. Using QGIS, we derived the percent land cover for MOB constitute the main microbial processors of CH4 at each of the catchments using European Environment Agency’s oxic/anoxic interfaces in stream sediments (Trimmer et al., 2010, 2018 Corine Land Cover (CLC, Version 20) database. Dominant 2015; Buriánková et al., 2012; Shelley et al., 2014, 2015) where land cover types included non-irrigated arable land, mixed forest, MOB can account for up to 30% of prokaryotic cell numbers coniferous forest, natural grasslands, bare rock, discontinuous (Buriánková et al., 2012). CH4 oxidation (MOX) can be an urban fabric, and land principally occupied by agriculture with important ecosystem process, equivalent to up to 46% of net significant areas of natural vegetation and transitional woodland- photosynthetic production in chalk streams (Trimmer et al., shrub (Supplementary Table S1). 2010; Shelley et al., 2014) and reducing CH4 evasion fluxes At each site, we collected triplicate benthic sediment samples by 70% (Zaiss et al., 1981; de Angelis and Scranton, 1993). (down to 5 cm depth) from which we retained the sandy fraction Among MOB, low-affinity CH4 oxidizers with substrate affinity (<2 mm). Sediment subsamples for DNA extraction were constants KS [i.e., the substrate concentration for which the immediately transferred to LifeGuard Soil Preservation Solution ◦ MOX rate is half the maximum reaction velocity (Vmax)] in the (Mobio) and stored at −80 C until processing. Subsamples micromolar range are adapted to elevated CH4 concentrations for flow cytometric cell counting were immediately fixed typical for hot spots of methanogenesis. In contrast, high-affinity with 2.5% formaldehyde (final concentration) and kept cool ◦ oxidizers with KS values in the nanomolar range are adapted (4 C). Sediments were transported cooled to the laboratory for to atmospheric CH4 concentrations (Bender and Conrad, 1992). kinetic analyses (see below). Porewater was sampled using a Independent of their kinetic affinity, known MOB are distributed 1.5-cm diameter piezometer connected to a syringe. For the across Methylocystaceae, Beijerinckiaceae (both a-Proteobacteria, determination of dissolved CH4 and CO2, we collected 30 ml or type II MOB), Methylococcaceae, Methylothermaceae (both stream- and porewater into 60 ml syringes flushed with N2 and g-Proteobacteria, or type I MOB), and Methylacidiphilaceae containing 0.05 g of NaN3 to stop biological activity. Stream- and (Verrucomicrobia) (Knief, 2015). Proteobacterial MOB dominate porewater dissolved oxygen (DO) was directly measured with an methanotrophic communities in most environments and were LDO probe (Hach) equipped with a flow-through cell connected previously associated with two life strategies (Ho et al., 2013): to the piezometer. Another set of stream- and porewater samples g-proteobacterial MOB are generally responsive to changes in were filtered [0.2 mm polytetrafluoroethylene (PTFE) filters] and CH4 concentrations by rapidly inducing pmoA gene expression treated with 1 M HCl for the measurement of Fe(II) or 5% (w/v) (coding for a subunit of the particulate CH4 monooxygenase). zinc acetate for S(-II) measurements. Stream- and porewater Although numerically not abundant in many environments, samples for the determination of dissolved organic carbon (DOC) ◦ they functionally dominate MOX when CH4 concentrations are concentration were filtered through pre-combusted (450 C for high. Recent studies revealed the importance of Crenothrix, a 4 h) GF/F filters (Whatman). filamentous type I MOB in lakes (Oswald et al., 2017) and caves (Karwautz et al., 2018). In contrast, a-proteobacterial MOB Geochemical Analyses typically feature more stable populations with less responsiveness Anions and cations were measured by ion chromatography to fluctuating CH4 concentrations (Ho et al., 2013). (DX-3000, Dionex) with an IonPac AS11-HC column (anions) The aim of this study was to link MOX kinetics to MOB and an IonPac CS16 Cation-Exchange column (cations). S(- community structure and genetic potential of stream sedimentary II) and Fe(II) were measured photometrically (Cline, 1969; communities. Experimenting with stream sediments sampled Stookey, 1970). DOC concentration was measured using a Sievers across a large environmental gradient (including alpine, natural, M5310C TOC Analyzer. CO2 and CH4 concentrations in the urban, and agriculture-dominated catchments), we hypothesized streamwater and porewater were measured with the headspace that the kinetics of MOX depend on CH4 supply, which we equilibration method and using cavity ring-down spectroscopy
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