Methanotrophs
MicrobiologyFundamentals andBiotechnologicalApplications
Editor EunYeolLee DepartmentofChemicalEngineering
KyungHeeUniversity
Yongin-si,Gyeonggi-do RepublicofKorea
ISSN1862-5576ISSN1862-5584(electronic)
MicrobiologyMonographs
ISBN978-3-030-23260-3ISBN978-3-030-23261-0(eBook) https://doi.org/10.1007/978-3-030-23261-0
© SpringerNatureSwitzerlandAG2019
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Preface
Methanotrophsareaerobicproteobacteriathatcanutilizemethaneassolecarbonand energysource.Inecology,methanotrophshaveanessentialroleintheglobalcarbon cyclebyremovingmethanegeneratedgeothermallyandbymethanogens. Methanotrophshavebeenemployedasthebiocatalystformitigatingmethaneas greenhousegasandremediatinghalogenatedhydrocarbonsinsoilandunderground water.Recently,methaneisconsideredthenext-generationcarbonfeedstockfor industrialbiotechnologybecauseofitshighabundanceandlowprice.Methanotrophs canbeusedasthebiocatalystfortheproductionofchemicals,fuels,andbiomaterials frommethaneunderenvironmentallybenignproductionconditions.
Despitethegrowingimportanceofbasicandappliedresearcheson methanotrophs,therewasnocomprehensivetextbookforseniorundergraduateand postgraduatelevels. Methanotroph:MicrobiologicalFundamentalsandBiotechnologicalApplications waswritteninanattempttogivethereadersasystematicand comprehensiveoverviewforbothbasicandappliedaspectsofmethanotrophs.Thus, thisbookcanbeusedasareferenceformicrobiologistsandbiochemistsdealingwith ecology,environmentalfunctioning,andunderstandingofphysiologyandmetabolismofmethanotrophsaswellasthefundamentalsandapplicationsofmethane monooxygenases.Thisbookisalsovaluabletobiotechnologistsandbiochemical engineerswhoresearchontheomics-basedunderstandingofmethanemetabolism, metabolicengineeringforstrainimprovement,methanobactinbiosynthesis,and environmentalapplicationsofmethanotrophs.
Iwouldliketothankthepeoplewhowereinstrumentalinthewritingofthisbook. Firstofall,Iwouldliketothankthecontributorsfortakingtheirvaluabletimein writingthechapters.IappreciateverymuchProfessorAlexanderSteinbüchelofthe InstituteofMicrobiologyatMünsterUniversity,the MicrobiologyMonographs SeriesEditor,forgivingmetheopportunitytopublishthisbook.Iwouldliketo acknowledgeC1gasrefineryR&DprogramoftheNationalResearchFoundationof Koreaforsupportingtheresearchfundfordevelopmentofmethanotrophicplatform v
strainsforbioconversionofmethaneandmethanol.Lastly,Iamgratefultomy belovedfamilyandlabmembersfortheirconsiderationandsupportsothatIcould concentrateonwritingbookscomfortably.
Yongin-si,Gyeonggi-do,RepublicofKoreaEunYeolLee April2019
vi Preface
Contents
MethanotrophEcology,EnvironmentalDistributionandFunctioning ...1
PaulL.E.Bodelier,GermanPérez,AnneliesJ.Veraart, andSaschaM.B.Krause
EnrichmentandIsolationofAerobicandAnaerobicMethanotrophs ...39
Sung-KeunRhee,SamuelImisiAwala,andNgoc-LoiNguyen
TheBiochemistryofMethaneMonooxygenases ...................71
SunneyI.ChanandSeungJaeLee
Multi-omicsUnderstandingofMethanotrophs ....................121 YueZhengandLudmilaChistoserdova
Diversity,Physiology,andBiotechnologicalPotential ofHalo(alkali)philicMethane-ConsumingBacteria ................139 SnehalNariyaandMarinaG.Kalyuzhnaya
MetabolicEngineeringofMethanotrophsfortheProduction ofChemicalsandFuels ......................................163
OkKyungLee,DiepT.N.Nguyen,andEunYeolLee
Methanobactin:ANovelCopper-BindingCompoundProduced byMethanotrophs .........................................205
JeremyD.SemrauandAlanA.DiSpirito
EnvironmentalApplicationsofMethanotrophs ...................231 AdrianHo,MiyeKwon,MarcusA.Horn,andSukhwanYoon
Index ...................................................257
vii
ListofContributors
SamuelImisiAwala DepartmentofMicrobiology,ChungbukNationalUniversity, Cheongju,RepublicofKorea
PaulL.E.Bodelier DepartmentofMicrobialEcology,NetherlandsInstituteof Ecology(NIOO-KNAW),Wageningen,TheNetherlands
SunneyI.Chan NoyesLaboratory,CaliforniaInstituteofTechnology,Pasadena, CA,USA
InstituteofChemistry,AcademiaSinica,Nankang,Taipei,Taiwan
LudmilaChistoserdova DepartmentofChemicalEngineering,Universityof Washington,Seattle,WA,USA
AlanA.DiSpirito RoyJ.CarverDepartmentofBiochemistry,Biophysicsand MolecularBiology,IowaStateUniversity,Ames,IA,USA
AdrianHo InstituteforMicrobiology,LeibnizUniversitätHannover,Hannover, Germany
MarcusA.Horn InstituteforMicrobiology,LeibnizUniversitätHannover, Hannover,Germany
MarinaG.Kalyuzhnaya DepartmentofBiology,SanDiegoStateUniversity,San Diego,CA,USA
ViralInformationInstitute,SanDiegoStateUniversity,SanDiego,CA,USA
SaschaM.B.Krause JohannHeinrichvonThünenInstitute,FederalResearch InstituteforRuralAreas,ForestryandFisheries,Braunschweig,Germany
MiyeKwon DepartmentofCivilandEnvironmentalEngineering,KAIST, Daejeon,SouthKorea
EunYeolLee DepartmentofChemicalEngineering,KyungHeeUniversity, Yongin-si,SouthKorea
ix
OkKyungLee DepartmentofChemicalEngineering,KyungHeeUniversity, Yongin-si,SouthKorea
SeungJaeLee DepartmentofChemistryandInstituteforMolecularBiologyand Genetics,ChonbukNationalUniversity,Jeonju,RepublicofKorea
SnehalNariya DepartmentofBiology,SanDiegoStateUniversity,SanDiego, CA,USA
DiepT.N.Nguyen DepartmentofChemicalEngineering,KyungHeeUniversity, Yongin-si,SouthKorea
Ngoc-LoiNguyen DepartmentofMicrobiology,ChungbukNationalUniversity, Cheongju,RepublicofKorea
GermanPérez DepartmentofMicrobialEcology,NetherlandsInstituteofEcology (NIOO-KNAW),Wageningen,TheNetherlands
Sung-KeunRhee DepartmentofMicrobiology,ChungbukNationalUniversity, Cheongju,RepublicofKorea
JeremyD.Semrau DepartmentofCivilandEnvironmentalEngineering, UniversityofMichigan,AnnArbor,MI,USA
AnneliesJ.Veraart DepartmentofAquaticEcologyandEnvironmentalBiology, InstituteforWaterandWetlandResearch,RadboudUniversity,Nijmegen,The Netherlands
SukhwanYoon DepartmentofCivilandEnvironmentalEngineering,KAIST, Daejeon,SouthKorea
YueZheng DepartmentofChemicalEngineering,UniversityofWashington, Seattle,WA,USA
CASKeyLaboratoryofUrbanPollutantConversion,InstituteofUrban Environment,ChineseAcademyofSciences,Xiamen,China UniversityofChineseAcademyofSciences,Beijing,China
xListofContributors
MethanotrophEcology,Environmental DistributionandFunctioning
PaulL.E.Bodelier,GermanPérez,AnneliesJ.Veraart, andSaschaM.B.Krause
4.1ControllingAbioticFactors..
Abstract Thedynamicsofmethaneconcentrationsintheatmosphereinrecent decadeshasdemonstratedmanyanomalieswhicharepoorlyunderstood.Theonly biologicalwayofdegradingthispotentgreenhousegasisbymicrobialoxidation. Aerobicmethanotrophicbacteria(MB)playanimportantroleinmanyecosystems worldwidedegradingmethanebeforeitcanescapetotheatmosphere.Thisgroupof bacteriahasintensivelybeenstudiedasamodelmicrobialfunctionalguildbecause thereisastronglinkbetweentheconsumptionofmethaneandthecompositionof MBcommunities,facilitatingthestudyofmicrobial “behavior” intheenvironment. ThesestudieshaverevealedastrongbiogeographyofMBwhichisdisplayedintheir phylogenynotonlyonthebasisofsinglefunctionalmarkergenesbutalsoon
P.L.E.Bodelier(*)·G.Pérez
DepartmentofMicrobialEcology,NetherlandsInstituteofEcology(NIOO-KNAW), Wageningen,TheNetherlands e-mail: P.Bodelier@nioo.knaw.nl
A.J.Veraart
DepartmentofAquaticEcologyandEnvironmentalBiology,InstituteforWaterandWetland Research,RadboudUniversity,Nijmegen,TheNetherlands
S.M.B.Krause
JohannHeinrichvonThünenInstitute,FederalResearchInstituteforRuralAreas,Forestryand Fisheries,Braunschweig,Germany
© SpringerNatureSwitzerlandAG2019
E.Y.Lee(ed.), Methanotrophs,MicrobiologyMonographs32, https://doi.org/10.1007/978-3-030-23261-0_1
Contents 1Introduction...................................................................................2 2TaxonomyandPhylogeny. ...................................................................3 3EnvironmentalDistribution ...................................................................7
.................................................................7
........................................................................10 4Ecology.......................................................................................21
3.1Biogeography..........
3.2SelectedHabitats
..........................................................21 4.2BioticInteractions......................................................................23 4.3LifeStrategies ...........................................................................25 5SynthesisandOutlook ........................................................................27 References .........................................................................................28
1
genomesequencebasis.Novelenvironmentalcontrollingfactorshavebeenrevealed (e.g.rareearthmetals)aswellasnovelorganismswithasyetunknowntraitsfor MB.Theresistanceandresilienceofmethaneconsumptionandmethaneconsuming communitieshavebeenshowntodependonspeci ficcommunitymembers.The currentknowledgeonenvironmentaldistributionandofMBhasledtoproposea life-historyscheme,classifyingMBcommunitiesontheircollectivetraitsratherthan singlyontheircapacitytheoxidisemethanealone.
1Introduction
Workingtowardsasustainablefutureforourplanetrequiresadrasticchangeinthe wayssocietyaddressesitsprimaryneeds:energy,food,resourcesandclimate.Finite stocksoffossilfuelsandcorrespondingrelatedclimateissuesleavesonlylow-carbon economieswithhighcontributionofrenewablesasalternatives,asrecentlyacknowledgedandratifiedinthe “ParisAgreement“ duringtheUnitedNationsFramework ConventiononClimateChange(http://unfccc.int/resource/docs/2015/cop21/eng/ l09r01.pdf).ToreachtheParistargetsdrasticreductionsinemissionsofgreenhouse gases(GHG)arenecessary(80%below1990levels),includingobtaining20%ofour energyfromrenewablesources.Inthiscontext,methane(CH4)playsakeyrole, beinga24timesmorepotentGHGthancarbondioxide,andcontributing40%to globalwarmingona20-yeartimescale(IPCC 2014).Hugeamountsofmethaneare releasedintotheatmosphereassociatedtogasandoilproductionandbybiological productioninnaturalandagriculturalwetlands,landfills,wastewatertreatmentand agriculturalproductionsystems,lakes,oceansandtermites(Deanetal. 2018). However,alargeportionofthismethanehasalreadybeenmitigatedbeforeescaping totheatmospherebytheactivityofmethane-oxidizingbacteria(MB)whichutilise methaneascarbonandenergysource(BodelierandSteenbergh 2014b).Thisreaction,turningmethaneintobiomassandmetabolitesisofhighvalueforproductionof bio-products(Fig. 1),isuniquetothesebacteria,andrequiresasetofenzymeswitha rangeofspecialfeatures(seeChaps. “Diversity,Physiology,andBiotechnological PotentialofHalo(alkali)philicMethane-ConsumingBacteria” and “MetabolicEngineeringofMethanotrophsfortheProductionofChemicalsandFuels”).Centralare themethanemonooxygenase(MMO)variants(particulate(pMMO)andsoluble methanemonooxygenase(sMMO))whichhaveaverybroadsubstraterange,making themhighlysuitableforapplicationsinsyntheticorganicchemistryandfordegrading environmentalcontaminantssuchastrichloroethylene(TCE)(Piejaetal. 2017). Theirimportanceforclimateandindustryhasspurredintensiveresearchintothe environmentaldistribution,diversity,ecology,physiologyandgenomicsofMB.The long-standingnotionthatthesemicrobesonlybelongtotheProteobacteria,andare exclusivelyobligatorymethanotrophicandaerobic,hasbeenprovenwronginthelast twodecades.NovelVerrucomicrobial(Dunfieldetal. 2007;vanTeeselingetal. 2014),anaerobic(Ettwigetal. 2010;Haroonetal. 2013;in ‘tZandtetal. 2018),
2P.L.E.Bodelieretal.
Fig.1 Methanecontainingbiogasoriginatingfrome.g.wasteorbeing flaredofduringenergy productioncanbeturnedintosustainablestorageofcarbonandenergy.Byusingnaturalconsortia ofmethaneconsumingmicrobesorengineeredstrains,methanecanbeturnedintoindustrially relevantchemicalswhichcanbestoredandtransportedeasilyandusedfortheproductionof valuablebio-basedchemicals
facultative(CrombieandMurrell 2014),and/ordenitrifyingMB(Dametal. 2013; Kitsetal. 2015)havebeendiscovered.Also,numerousenvironmentalstudies facilitatedbythedevelopmentofcultivationindependentassessmentmethodshave revealedavastasyetuntappeddiversityofMBwithdistinctbiogeography,indicatingalinkbetweenMBtraitsandhabitat(Knief 2015)ofinterestforfuturebiotechnologicalapplications.
Crucially,MBhavealsorecentlybeenshowntoperformbetterinnaturalaswellas syntheticmultispeciescommunities(Hoetal. 2014;Schnyderetal. 2018)depending oninteractionsandexchangesofnutrientswithothercommunitymembers(Zheng etal. 2014;Iguchietal. 2011)orbyexchangeofvolatileorganiccompounds(Veraart etal. 2018).However,thecultivationandindustrialhandlingofMBremaindifficult, resultinginonlyahandfulofstrainscurrentlyexploitedforindustrialpurposes.Next tothis,globalclimatemodelsstillgivepoorpredictionsoffutureatmospheric methaneconcentrations(Deanetal. 2018;Nisbetetal. 2014),whichdisplaya rangeofdistinctdynamicsandanomaliesinthepastdecades.Lackofadeeper understandingoftheecologyandfunctioningofMBinnaturalandman-made habitatsunderliesmanyoftheissueswehaveinusingMBinbio-industryandto explainatmosphericmethanedynamics.Thepresentchapterwillhighlightfeaturesof theecologyandenvironmentaldistributionofMBwithafocusontheaerobicMB.
2TaxonomyandPhylogeny
Currentlydescribedspeciesandtheirphylogenyhaveextensivelybeendescribed recently(DedyshandKnief 2018)andwewillnotgointodetailedtaxonomyhere. However,insightsfromcomparativegenomicsandmanydiscoveriesinthelasttwo
MethanotrophEcology,EnvironmentalDistributionandFunctioning3
decadeshasresultedinamuchmoredifferentiatedviewonthismicrobialguild. Sincethe firstisolationofabacteriumcapableofgrowthonmethaneassoleenergy andcarbonsource(Söhngen 1906),obligateaerobicMBhavetraditionallybeen studiedandconceptuallybeentreatedasacoherentfunctionalmicrobialgroup.The firsttaxonomicframeworkforMBemergedaftertwodecadesofpioneeringworkin thelastcentury(Whittenburyetal. 1970;Bowmanetal. 1993)basicallydividing MBintwomaingroupsbasedonphysiological,morphological,ultrastructural,and chemotaxonomiccharacteristics.TypeIMBdistinguishedfromtypeIIbyhaving intracytoplasmicmembranes(ICM)structuredasvesiculardiscswhiletheseare arrangedalongtheperipheryofthecellaspairedmembraneswithtypeIIMB.Next tothis,typeIMB fixcarbonviatheribulosemonophosphatepathway(RuMP)while typeIIusetheserinepathway.Furthermore,bothgroupsclearlydifferedintheir fattyacidcompositionwithinthepolarlipidfractionofthecellmembranes(PLFA), withC16dominatedPLFApredominantlyintypeIandC18intypeII(Bowman etal. 1993;Bodelieretal. 2009)withevenPLFAwhichtodatehaveonlybeen detectedinMB(C16:1ω8candC18:1ω8c)enablingtheseparatedetectionofMB fromotherbacteriainenvironments(Bodelieretal. 2013).Besidestheability fix molecularnitrogen,whichatthattimeseparatedtypeIfromtypeII,theproduction ofrestingstages(cystsorspores)havealsobeenconsideredastaxonomicfeatureto distinguishmorebetweengeneraratherthanmaintypes.Theseparationintwomain groupsatthattimewasalsoreflectedinthephylogenybasedon16SrRNA,placing typeIwithinthe Gammaproteobacteria andtypeIIinthe Alphaproteobacteria (Bowmanetal. 1993).However,thesuccessfulcultivationofmanynewspecies (Knief 2015;DedyshandKnief 2018)generatedinformationthatrenderedthis dichotomousclassificationsystemnotrepresentativeforthephylogenomicand physiologicalviewwehavenow.Internalmembranesystemsdonotoccurinall MB(DunfieldandDedysh 2014;Vorobevetal. 2011);PLFAspeci ficfortypeIMB occuralsointypeII(Dedyshetal. 2007);nitrogen fixationalsooccursintypeIMB (Methylobactertundripaludum)andMBhavebeendiscoveredwhichhavepartsor evencompletegenesetsforbothtypesofcarbon fixationpathways(Nguyenetal. 2018),althoughithasnotbeendemonstratedthatbothpathwayscanbeoperational ineithertypeIortypeIIMB.
Despitetheinadequacyofthe “typeI/II” classificationsystemasanintegrated reflectionofphysiology,biochemistryandmorphology,thesystemisstillinuse sincecurrentphylogenybasedon16SandMMO(particulate(pMMO)andsoluble (sMMO)methanemonooxygenase)(Knief 2015;DedyshandKnief 2018)reflects thisclassification.CurrenttaxonomicradiationofaerobicMBcoverstaxonomically classifiedpurecultures,MBwithacandidatestatusduetothefactthatthesestrains areonlyavailableashighlyenrichedculturesanduncultivatedlineagesrepresented byenvironmental pmoA sequences(seeFig. 2).MostMBtypestrainsandclassified generafallwiththe Gammaproteobacteria inthefamilies Methylococcacea (type Ia/Ib)and Methylotermaceae (typeIc),whilethe Alphaproteobacteria harborthe familiesof Methylocystaceae (typeIIa)and Beijerinckiaceae (typeIIb)(Fig. 2). WiththediscoveryofMBinthephylumofthe Verrucomicrobia (Dunfieldetal. 2007;vanTeeselingetal. 2014;Poletal. 2007)belongingtothefamilyofthe
4P.L.E.Bodelieretal.
Fig.2 PhylogenyofdescribedaerobicMBbasedon16SrRNA( a )andtherelatednessofuncultivatedlineagesinrelationdescribedrepresentativesonthebasis ofthe pmoA gene( b ).FromDedyshandKnief( 2018 )withpermission
MethanotrophEcology,EnvironmentalDistributionandFunctioning5
Methylacidophyliceae atypeIIIclassifierwasnecessarytobeinlinewiththe “phylogeny-based” typing.WiththeadventofthisMBphylumanotherbiochemical variationwasintroduced,bythefactthattheseMBareautotrophic, fixingcarbon usingtheCalvincycle(Khademetal. 2011).
Thediscoveryoftheabilitytogrowonmethaneofanalreadydescribed filamentousbacterium Crenothrixpolyspora (Stoeckeretal. 2006)belongingtothe Gammaproteobacteria wasanothersurprisewithintheMBrealm,addinganother family(Crenotrichideae)totheMBtreewithadeviating pmoA,similarto amoA of ammoniaoxidizers.Lateranovelgenuswithinthisfamilywithwasdescribed Clonotrix (Vigliottaetal. 2007),whichsimilarto Crenotrix hasnotbeenbrought intopureculturerenderingtaxonomicallyacandidatusstatus.
Althoughstrictlyspeakingbeingananaerobe,microbesbelongingtothenovel NC10phylumandrepresentedbythe Candidatus Methylomirabilisoxyfera,canbe consideredaerobicMBsincetheygenerateoxygenfromthereductionofnitriteand thesubsequentdisproportionationofnitricoxide(NO)(Ettwigetal. 2010)anduse thatoxygentooxidizemethaneaerobically.Recently,novelspeciesbelongingtothe Methylomirabilis genushavebeenidentified(Grafetal. 2018;Versantvoortetal. 2018)butasyethasnotbroughtintopureculture.
NexttotheMBinpureculturesorenrichmentsarangeof pmoA-basedlineages havebeenidentifiedwhichmostlyclusterwithtaxonomicallydescribedorganisms butalsorepresentarangeofsequencesdistinctfromexisting pmoA sequences (Knief 2015;DedyshandKnief 2018)(Fig. 2).Foralmostalloftheseclustersit remainstobeelucidatedwhatorganismsarebehindthesesequences.However, recentlyforthesocalledUSCα (foruplandsoilcluster α)usingsinglecellsequencingofactivelyatmosphericmethaneoxidizingcells,agenomesequencewas obtaineddemonstratingthecloseassociationofrepresentativesofthisclusterwith thegenus Methylocapsa (Pratscheretal. 2018).
Recently,microbialtaxonomyhasbeenrevisedbytheinclusionofwholegenome sequencesoftaxonomicallydescribedmicrobesaswellasmetagenomeassembled genomes(MAG)fromenvironmentalsamples(Parksetal. 2018).Thisextensive effortcompared120singlecopyubiquitousproteincodinggenesofalmost95,000 genomes.Thisanalysesledtoasubstantialrevisionoftaxonomyremovingmanyof theinconsistenciesandpolyphyleticcasesofincongruencyofphenotypicandsingle genebasedphylogeny.AlsoforMB,changesweresuggested,ofwhichthepropositionofdividingtheorderof Methylococcales intothreefamiliesbeing Methylomonadaceae (Ia), Methylococcaceae (Ib)and Methylothermaceae (Ic)(Parksetal. 2018).Anindetailwholegenomeanalyses,comparingANI(average nucleotideidentity),dDDH(digitalDNA-DNAhybridization)andAAI(average aminoacididentity)ofexistingMethylococcalesgenomesalsoledtoreclassification ofpolyphyleticMBgenera(Orataetal. 2018)leadingforexampletothenewgenus Methylotuvimicrobium.ForacompleterevisionoftheMBtaxonomyandphylogeny, theamountofsequencedtypestrainsisstillnotcompleteenough.Itisobviousthough fromtheGTDB(http://gtdb.ecogenomic.org/)database,wherenon-MMOcarrying bacteriaareclassifiedintoMBorders(e.g. Cyclocystaceae intotheorderof Methylococcales)aswellasrecentevolutionarystudiesofcoppercontaining
6P.L.E.Bodelieretal.
monooxygenases(OsborneandHaritos 2018;Khadkaetal. 2018)thatmethane oxidationisatraitandpropertythathasbeenpassedonbyseverallateralgene transferevents.However,stillremnantsandrelatedfunctions,necessaryfor performingmethanotrophyarestillpresentinancestors.Thelattermayleadto morereshuf flingasgenomesbecomeavailableandleadmoreandmoretothe realizationthatwearenotlookingsimplyatafunctionalguildbutatgroupof microbeswithmanymoreevolutionaryselectedtraitsthanmethaneoxidationalone.
3EnvironmentalDistribution
MBplayacrucialroleinmethanemitigationfromanumberofimportantmethane emittingsourceslikewetlands.Hence,inhabitatswhereyouhavehighmethaneand oxygeninclosevicinityMBareexpectedto flourish.However,therearemany habitatswhereMBwouldnotbeexpected.Inthissection,theenvironmental distributionofMBaswellastheirbiogeographywillbediscussed.Recent findings inafewkey-habitatswillbehighlighted.
3.1Biogeography
Principally,MBcanbefoundinallhabitatsweremethaneisavailable.Thiscanbe methaneproducinghabitatswithcloseproximityofanoxicandoxichabitats (e.g.wetlands,peat,lakesediments,marinesediments,landfills,etc.),butalsoin allotherhabitatsincontactwithanormalatmospherewhichcontainsapproximately 1.86ppmofmethane.ThedirectconnectionbetweenMBandclimatechangehas spurredthousandsofstudiesfocusingonenvironmentaldistributionofMB,their activityandthefactorscontrollingthelatter.Manyoftheseaspectshavebeen describedinarangeofexcellentreviewpapers(e.g.Knief 2015;Hansonand Hanson 1996;Conrad 1996, 2007;BodelierandLaanbroek 2004;Semrauetal. 2010;Kolb 2009;BodelierandSteenbergh 2014a, b;Hoetal. 2013a, b).Thewealth ofstudiesisalsoduetothefactthatthepresenceandactivityofMBcanbescreened verystraightforwardbydetectingthepresenceofthe pmoA/sMMO geneorby measuringthemethaneoxidationpotentialofincubatedsamples(e.g.Bodelier etal. 2013).Nexttothis,arangeofstudieshasshownthatcommunitycomposition ofMBislinkedtotheconsumptionofmethane(Schnyderetal. 2018;Bodelieretal. 2013;Nazariesetal. 2011;Levineetal. 2011)whichisenforcedbythepossibilityto trace 13C-labelledmethaneintaxonomicallyrelevantcompounds(e.g.PLFA,DNA, proteins)therebyassessingtheactivespeciesincomplexenvironments(Bodelier etal. 2013;Boschkeretal. 1998;Daebeleretal. 2014).Hence,thedirectlink betweenecosystemfunctionanddiversityoftheresponsiblespecieshasmadeMB amodelgroupwithinthe fieldofenvironmentalmicrobiology,beingmaybethebest describedenvironmentalmicrobialfunctionalgrouporguild.Using pmoA based
MethanotrophEcology,EnvironmentalDistributionandFunctioning7
analysisofallcultivatedanduncultivatedMBdiversity,(Knief 2015)KniefexecutedacomprehensivebiogeographyanalysesofallMBdiversityknownatthat moment.Itshowednotonlythatmorethanhalfofallenvironmentalsequenceswere relatedtoknowncultivatedspeciesbutalsothatMBdisplaydistinctbiogeography andhabitatpreference(Knief 2015).Typically,aquatichabitats(freshandmarine) tendtoharborlineagesof pmoA sequenceswhichdonotoccurelsewhere,while terrestrialenvironmentslikesoilcontainMBspecieswithamuchbroaderenvironmentaldistribution,maybereflectingthelargerenvironmentalheterogeneityand influencingbioticandabioticfactorsaffectingMBfunctioningandsurvivalin terrestrialecosystems.Typicalexceptions,arethe pmoA sequencelineages(typeId e.g.USCα,USCγ)associatedwiththeuptakeofatmosphericmethaneinsoilswhich werefoundalmostexclusivelyinuplandsoilsaswellastheVerrucomicrobialMB whichonlyoccurinextreme(lowpH,hightemperature)terrestrialsoils.Thestudyby Kniefwasbasedon371studiesbasedonthe pmoA gene.WeutilizedtheEarth MicrobiomeProject(EMP)(Thompsonetal. 2017)databasewhichisthemost comprehensivedatacollectiononmicrobial16SrRNAsequencingreadsandassociatedmetadataonvarioushabitats,tocharacterizepatternsofmethanotrophsacross differentbiomesandhabitatsonanevenbiggerscale(Table 1).
WhencomparingdistributionofMBgenerainterrestrialandaquaticbiomes (Fig. 4;Table 2)itisobviousthatterrestrialhabitatsarepopulatedbothbytypeI andIIMB.Themoststrikingdifferencebetweenterrestrialandaquaticbiomesisthe farlessevenrelativeabundancesofdetectedgeneraintheaquatichabitatswhichis duetoastrongrepresentationofthegenera Methylobacter, Methylocaldum ,and Methylomonas (Fig. 4).
Incontrast,proportionsofabsolutereadcountsweremoreevenlydistributed amongmajormethanotrophicgenerainterrestrialbiomes(Fig. 4;Table 2),possibly reflectingthedifferencesinselectivefactorsinterrestrialandaquatichabitats.An exceptiontothebroaddistributionofthegenerabelongingtothefamily Methylococcaceae isthegenus Methylosoma,whichexclusivelyisfoundinaquatic systems.Incontrasttothegenerallydisturbedrepresentativesofthegenera Methylobacter, Methylocaldum,and Methylomonas,membersofthefamily Methylocystaceae,moreinparticularofthegenus Methylocystis aremorestrongly associatedwithterrestrialthanwithaquaticbiomes(Fig. 4)althoughmanyisolates havebeenobtainedfromaquaticenvironments(Heyeretal. 2002).Membersofthe genus Methylosinus canbefoundpredominantlyinaquatichabitats.Withinthe familyof Beijerinckiaceae,thegenus Methylocella isonlyprevalentinterrestrial biomes.Representativesofthisgenusdonotcontainthe pmoA geneandthereforeare generallyunderrepresentedinstudiesofmethanotrophs(Rahmanetal. 2010). Methylocella isafacultativemethanotrophs,whichcanuseotherhydrocarbons suchasacetate,ethaneandpropane(Farhanetal. 2018),whichmayberelatedto itsenvironmentaldistribution.IthastobenotedthattheanalysesoftheEarth Microbiomedatasetarebasedonrelativelyshort16SrRNAsequencingreads (90–151bp),butevenwhenlookingonlyatsequencesrelatedcultivatedrepresentativestheanalysesofKnief(2015)isconfirmed,demonstratingclearbiogeographyof MBgenerawhichistheresultsofevolutionandselectionbasedonmanymoretraits thanthecapacitytooxidizemethane(Hoetal. 2013a, b, 2017a).Averyrecentstudy
8P.L.E.Bodelieretal.
Table1 Environmentsin researchstudiesoftheEarth MicrobiomeProject(EMP) (Thompsonetal. 2017)that includedmethanotrophic genera
Numberofstudies
Freshwater2757
Soil1581
Rhizosphere389
Biofilm387
Freshwatersediment328
Marinesediment321
Sand(biofilter)99
Organicmaterial(both)71
Mucus(humanandanimal)46
Feces(humanandanimal)35
Dustandair34
Animalnest28
Sebum(humanandanimal)22
Undergroundwater17
Seaandhypersalinewater15
Lakestromatolitemat13
Bodily fluid(animal)4
Brackishwater3
Saliva(humanandanimal)3
Animalexcreta3
Environmentalcategoriesweresimplifiedbutlargelyfollowedthe terminologyusedintheEMPmetadata.PleaserefertoFig. 3 for detailedmethodology
investigatingthebiogeographyofMBin697soilsamplesdistributedoverScotland providesstrongevidencethatevenonaregionalscaleclimo-edaphicfactors(Temperature,moisture,soilphysico-chemistry)influencesdistributionoftypeIIMBand USCα relatedsequencetypes(Nazariesetal. 2018).Land-use,moisture/rainfall, nutrientsandmetalswerethemostimportantfactorsinfluencingdistribution.The authorsofthisstudyusedtheclimo-edaphicrelationshipstomapMBdistribution overScotland,deliveringpotentialimportantinformationtoinformmodelspredicting greenhousegasemissionanduptakeinsoilunderfutureclimateorland-usechange scenarios(Nazariesetal. 2018).Similargeostatisticalapproacheswereusedonaplot scaletomapactivity,abundanceanddiversityofMBcommunitiesinawetlandsoil (Bodelieretal. 2013;Krauseetal. 2013;Wangetal. 2012)representingahydrologicalgradient.Onaplotscale(10m 10m),coveringthewhole floodingand subsequentmoisturegradient,methaneoxidationactivityanddistributionofthe activetypesofMBwereverywellexplainedbymoistureandthesubsequent availabilityofmethane(Bodelieretal. 2013;Wangetal. 2012).However,ona smallerscale(1m 1mand20cm 20cm)spatialdistributionwasaffectedby other,morelocalparametersnotdirectlyinfluencedbythelargerscalehydrological gradients(Krauseetal. 2013).Hence,MBdisplayclearbiogeographysubjectedto largescaledifferencesinabioticfactors,butonasmallscalelocalfactorsmayplaya moreprominentroleinnichedifferentiationandspeciationdependingontherespectiveenvironmentalproperties.
MethanotrophEcology,EnvironmentalDistributionandFunctioning9
Fig.3 EnvironmentsinresearchstudiesoftheEarthMicrobiomeProject(EMP)(Thompsonetal. 2017)thatincludedmethanotrophicgenera.Environmentalcharacterizationwassimplifiedand basedonavailablemetadataoftheEMPproject.Fordatageneration,we firstusedtheoverview table “ChooseYourOwnEMPBIOMTable” fromtheEMPFTPsitetoselectaclosedreference OTUbasedquality filteredbiomedataset(filename:emp_cr_silva_16S_123.qc_filtered.biome). Subsequently,acustomizedR(RDevelopmentCoreTeam 2008)andQIIMEscriptswasusedto extractallmethanotrophicgenera,readcounts,andmetadata.Scriptsandrawdatacanbeprovided uponrequestandother filesareprovidedat ftp://ftp.microbio.me/emp/release1/ .Sampleprocessing, sequencing,andcoreamplicondataanalysiswereperformedbytheEarthMicrobiomeProject (www.earthmicrobiome.org ),andallampliconsequencedataandmetadatahavebeenmadepublic throughtheEMPdataportal(qiita.microbio.me/emp)
3.2SelectedHabitats
Consideringthewidedistributiondescribedabove,itwouldbeachallengeto describeMBinallhabitatsweretheyaredetected.Hence,inthischapterwewill focusonafewclimaterelevanthabitatswithanemphasisonnovelrecent developments.
3.2.1MethaneSinkHabitats
MBaretheonlybiologicalwayofmitigatingmethaneproducedinanoxichabitats butalsototakeupandoxidizemethanefromtheatmosphere.Uptakeofmethanein oxicuplandsoils(forest,grassland,deserts)represents6%ofthesinkpartintotal
10P.L.E.Bodelieretal.
Fig.4 Distributionofmethanotrophicgenerainterrestrial(a)andaquaticbiomes(b)accordingto metadataoftheEMPproject(Thompsonetal. 2017). TheseOTUswereonlyclassifiedtofamily levelintheEMPproject(Thompsonetal. 2017).Differentmethanotrophicgeneraweregroupedby theirphylogeneticgroupsofthefamilies Methylocystaceae, Beijerinckiaceae , Methylococcaceae , Methylothermaceae, Methylacidiphilaceae ,andCandidatedivisionNC10.Circleswerearranged usingtheRfunctioncircleProgressiveLayoutintheRpackagepackcirclesversion0.3.3 (RDevelopmentCoreTeam 2008).Thesizeofeachcircleisproportionaltotheabsoluteread counts.Circlesweredetachedforbettervisibility.Dataisbasedon4.067,99316SrRNAgene sequencingreads(Table 2)from6156studiespublishedintheEarthMicrobiomeProject(Thompson etal. 2017).TheoriginaldataisshowninTable 2.PleaserefertolegendofFig. 3 fordetailed methodology
globalmethanebalance(Deanetal. 2018)andisaprocessproposedlycarriedoutby asyetuncultivatedMBbelongingtotypeIdandIIbsequenceclusters(Dedyshand Knief 2018),betterknownasUSCγ andUSCα andrelatedclusters.Theprocess carriedoutbythesesocalled “highaffinity” MB(degradingmethanetoconcentrationsbelow1.8ppm)isaffectedbyarangeofenvironmentalparameters(CH4,O2, moisture,ammonium,agriculturalpractices,land-usechange,plants,organicacids
MethanotrophEcology,EnvironmentalDistributionandFunctioning11
Table2 Absolutenumberofsequencingreadsofmethanotrophicgenerainaquaticandterrestrial biomes
TaxonomyAquatic_biomeTerrestrial_biome
Methylocystaceaea 182651272
Methylocystis 682830242
Methylosinus 106567165801
Methylocapsa 15241638
Methylocella 756061966
Methyloferula 14251457
Methylococcaceaea 173025272
Methylobacter 209128699226
Methyloglobulus 3470220175
Methylomarinum 161136
Methylomicrobium 2658918095
Methylomonas 382196158153
Methylosarcina 15815
Methylosoma 32430564
Methylosphaera 26785
Methylovulum 460198630
Methylocaldum 60483728607
Methylococcus 1460212032
Methylogaea 6022148
Methylothermus 60
Candidatus.Methylomirabilis 5485514
Methyloceanibacter 195858
aTheseOTU’swereclassifiedtothefamilylevel.Differentmethanotrophicgeneraweregroupedby theirphylogeneticgroupsofthefamilies Methylocystaceae, Beijerinckiaceae , Methylococcaceae , Methylothermaceae, Methylacidiphilaceae ,andCandidatedivisionNC10.PleaserefertoFig. 4 for detailedmethodology
andmonoterpenesetc.)ashasbeendescribedinarangeofstudies(Kolb 2009; Nazariesetal. 2011;Dunfield 2007;KolbandHorn 2012;Maureretal. 2008; Menyailoetal. 2010;Wieczoreketal. 2011).Recoveryafterdisturbancecantake decadesandmaydependonthediversityofMBpresent(Levineetal. 2011).Dueto thefactthatatmosphericmethanealoneisnotsufficienttogrow,allattemptstogrow andisolateatmosphericmethaneMBhassofarfailedandhence,novelinformation onthedistributionandfunctioningofthesemicrobeshasbeenverysparse.Novel habitatspotentiallycontributingtoatmosphericmethanedegradation,havebeen reportedrecently.Glacierfore fieldsoils(Naueretal. 2012),higharticcryosols (Lauetal. 2015),thawingpermafrost(Singletonetal. 2018),limestone(Waring etal. 2017),Lava(Pratscheretal. 2018)andkarstcaves(Zhaoetal. 2018).Genomes relatedtoUSCγ (Edwardsetal. 2017)andUSCα (Pratscheretal. 2018;Singleton etal. 2018)havebeenrecoveredfromarticsoilandfromforestsoils.Thesegenomes haveconfirmedtherelatednessofUSCα withgenus Methylocapsa andforUSCγ with Methylocaldum .ThenovelUSCα genomesrecoveredfromamireinNorthern
12P.L.E.Bodelieretal.
Swedenrevealedthepotentialforutilizationofacetate,confirmingwhatwasalready detectedbystableisotopeprobingearlier(Pratscheretal. 2011)aswellasthe presenceofgenesnecessaryforCOconsumption(Singletonetal. 2018).Pratscher etal.(2018)wasabletovisualizethe firstUSCα cellfromforestsoilsusinga combinationof fluorescencecellsortingusingacombinationofasuicidesubstrate and fluorescentinsituhybridization.Inthiswaysufficientcells,activelyoxidizing atmosphericmethanecouldbecapturedtoperformagenomeanalyseswhich revealedthatthesemicrobesonlyhaveXoxF(lanthanidedependent)methanol dehydrogenaseonly,genesinvolvedinexopolymerproductionandexcretionas wellasthecapacitytoformtrehalose.Thelattermaypointtowardsadaptationtodry orcoldconditionsandgrowthinbiofilms.Thelatter fittedwiththehighabundance ofthesemicrobesinsubterraneancavewallbiofilmsincavesglobally(Pratscher etal. 2018),asurveywhichwaspossiblewith16SrRNAprimersdevelopedusing thegenomeinfo.Allinall,thesenewdevelopmentspointintothedirectionthat atmosphericMBthriveinoligotrophichabitats,wheretheymakepartofpioneering microflora.
However,atmosphericmethaneoxidationisnotrestrictedtotheseexotic,enigmaticmicrobesbutpotentiallycanbecarriedoutbysomewell-knownlowaffinity (i.e.thriveathighmethaneconcentrations)MBwhichhavebeendemonstratedto oxidizeatmosphericmethaneinpurecultureincubations(KniefandDunfield 2005) althoughtheseMBcannotgrownorsurviveonatmosphericmethanealone. Recently,itwasdemonstratedthatagriculturalsoilscanbeturnedintoastrong sinkforatmosphericmethaneuponadditionoforganicamendments(e.g.compost, sewagesludge;aquaticplantmaterial)(Hoetal. 2015, 2017b).Although,the mechanismsresponsibleforthisphenomenonarenotyetclear,theuseofstable isotopeprobingincombinationwithPLFA(phospholipidfattyacid)demonstrated thatconventionalMB(i.e. Methylocystis; Methylosinus; Methyloferula )were responsiblefortheuptakeofmethaneintheseagriculturalsoils(Hoetal. 2019). Sincecellspecificactivitieswerelowerthanwhatisneededformaintenanceofcells, leadingtotheconclusionthattheseMBneedalternativesourcesofenergy.ConventionalMBinrice fieldsoilwereabletoturnthesoilintoapersistentandstrong sinkforatmosphericmethaneafterbeingspikedwithhighconcentrationsofmethane(Caietal. 2016).
Consumptionofmoremethane(i.e.repeatedincubationwith10,000ppmv)ledto asoilcommunitythatwasabletotakeupmethaneformorethana100days, demonstratingthatevenawetlandsoilcanturnintoasinkformethanewithout involvementofhighaffinityMB.UtilizationofstoragecompoundslikePHB(poly hydroxylbutyricacid)thatcanaccumulateduringperiodsofhighmethaneavailabilitywerehypothesizedtobethesourceofenergytocontinue pmoA activityunder atmosphericmethanesupply.Thismaypartiallyalsoexplaintheeffectofadditionof organicamendmentsinagriculturalsoil,whichmaypromoteinternalmethane productioninsoilaggregatesfuelingconventionalMBwithenergybutpotentially alsowithnutrientsandotheressentialmineralslikemetals(Fig. 5).Hence,with respecttosoilmethanesinksrecentnoveldevelopmentsoffernewviewsonfunctioningofhighaffinityMBbutalsopointtowardsthepotentialofasyetnon-methane
MethanotrophEcology,EnvironmentalDistributionandFunctioning13
Fig.5 Proposedmechanismsofresidue-stimulateduptakeofatmosphericCH4 byagricultural soils.Additionoforganiccarboncanleadtoanaerobicconditionsmediatedbytheaggregate structureinthesoilwhichleadstointernalCH4 productionstimulatinggrowthofobligatoryand facultativeMBormethylotrophscarryingsMMOgenes.Thisstimulationmaybeenhancedby nutrients(N,P,metals,traceelements)presentintheresidues.Thesestimulatedcellswillhavea highercapacitytoco-oxidizeCH4 fromtheatmosphere
sinkhabitatstobeexploredfortheirroleinclimatechangemitigationscenariosby assessingthefunctioningofconventionalMBandtheirroleinatmosphericmethane uptake.Thelatterwillbenecessarysinceclimatechangephenomena(lowerrainfall, hightemperature)mayincreaseatmosphericmethaneconsumption(Festetal. 2017).
3.2.2Wetlands
Duetotheirtight-aquaticterrestrialcouplingandlargeorganicmatteraccumulation, wetlandsarehotspotsofbiogeochemicalprocessing.Thehighmethanogenicactivity intheiranoxic,carbon-richsoilshasmadethemlargestsourceofglobalatmospheric methane,emitting142-284TgCH4 peryear(Kirschkeetal. 2013).Thisprovidesan idealhabitatforanaerobicandaerobicmethanotrophs,whichbothareabundantin wetlands,andcanreduceCH4 emissionsby10–90%(Segers 1998).Thefunctioning ofaerobicmethanotrophsinwetlandshasbeenreviewedextensively(Conrad 2007; BodelierandSteenbergh 2014a, b;Dedysh 2009;Bridghametal. 2013;Bodelier 2011)mainlyalsoduetotherelevanceofthesemicrobesinmitigationofmethane emissionfromricepaddies(e.g.Bodelieretal. 2000;Krugeretal. 2002;Chowdhury andDick 2013).MBabundanceandcommunitycompositioninwetlandslargely dependonlocalenvironmentalconditions,inpartdeterminedbytheecosystemtype. Wecandistinguishseveraldifferenttypesofwetlands,includingpeatlands(bogsand fens,arcticpermafrost),saltmarshesandestuaries,tropical(natural)wetlands
Internal CH4 hypothesis
Residue Soil Anareobicity Obligatory MOB Facultative MOB Methylotrophs Internal CH4 production Metals/trace elements Nutrients: NPK Carbon Atmospheric CH4
Smart residue mixes
14P.L.E.Bodelieretal.
includingriver floodplains,agriculturalwetlandssuchasricepaddies,andgeothermalwetlands.Methanotrophcommunitieshavebeenbestdescribedfortemperate and(ant)arcticecosystems,althoughsomeresearchhasbeendoneinthetropics, whichalsoharbourlargepeatlandareas.Inpeatlands,aerobicproteobacterial methanotrophsdominate,withmembersofthealphaproteobacteriadominatingin oligotrophic,acidic(pH < 5)bogs,whereasingeneralgammaproteobacteria dominateinminerotrophic,nutrient-richandonlymildlyacidic(pH > 5)fens (reviewedbyVerbekeetal. 2018).Wetlandsoccuroveracontinuumofhydrologicalandgeologicalconditions,andthereforeoftenliesomewhereinbetweenthese twoconditions,thusharbouringamixtureofalphaandgammaproteobacteria. Withinthealphaproteobacteriaoccurringinbogs, Methylocystis strainshavebeen foundtobemostactive,while Methylocella and Methylocapsa speciesarealso abundant(Verbekeetal. 2018).Withinthegammaproteobacteria, Methylobacter, Methylomonas and Methylomicrobium speciesarethemostabundant(Verbeke etal. 2018).Geothermalwetlandsarecharacterizedbyhightemperaturesand tendtobeveryacidic(pH < 4)duetosulfuroxidation.Methanotrophsisolated fromthesewetlandshavebeenfoundtobelongtothe Verrucomicrobia,andare currentlydescribedasthecandidatecluster “Methylacidiphilum” andthecandidate genus “Methylacidimicrobium” (Sharpetal. 2014).
Typically,aerobicMBarepresentatorabovetheoxic-anoxicinterfaceof wetlandsoils(Reimetal. 2012;Vaksmaaetal. 2017b).However,inrecentyears MBhavealsobeendetectedinassociationwith Sphagnum mosses(Larmolaetal. 2010;Kipetal. 2010),eitheroccurringinair-filledporeswithin Sphagnum tissues, orassociatedtotheirroots.ItwassuggestedthatMBliveinsymbiosiswithpeat mosses(Raghoebarsingetal. 2005)providingthemosswithCO2 whilereceiving photosyntheticoxygenfromthemoss.Otherstudiesreportedonmethanedependent nitrogen fixationbymossessuggestinganimportantroleforaerobicMBinpeat growth(Larmolaetal. 2014),whichwascontradictedbyotherstudiesfailingto observeaneffectofmethaneadditiononN2 fixationbymosses(Koxetal. 2018).As yetconclusiveevidenceformechanismofsymbiosisbetweenMBandpeatmosses inmissing(HoandBodelier 2015)asaretheresponsiblemicrobes(Koxetal. 2018). Asimilar,potentiallyfarreachingsymbiosisisclaimedforMBandriceroots,where diazotrophicMBintherhizosphere fixnitrogenusingmethaneasenergysourceand transferringthisnitrogentothericeplants(Ikedaetal. 2014).Agenesimilartothe oneinvolvedinplant-fungalsymbiosiswasproposedtobeinvolvedintheactivation ofrice-MBnitrogentransfer(Baoetal. 2014).However,besidesthelocalizationof typeIIMBproteinsinriceroottissue(Baoetal. 2014)nodirectproofforthis symbiosisisavailable,whichcanbeofgreatinfluenceforthesustainablecultivation ofriceemittinglessmethanewhilelessnitrogenfertilizerisrequired.Withrespectto thelatter,therecentlydiscoveredabilityofsomeMBtodenitrify(Dametal. 2013; Kitsetal. 2015)shedsnewlightoninteractionsbetweenmethaneandnitrogen cyclinginwetlandsystemsandbeyond.Therecentlydescribedgenomesequenceof thedenitrifyingmethanotroph Methylomonasdenitrificans FGJ1revealsgenes involvedinoxygenscavengingunderhypoxicconditions,hypothesizedtomaintain oxygenprovisionforthe pmoA underlowoxygenconditions(Orataetal. 2018).
MethanotrophEcology,EnvironmentalDistributionandFunctioning15
Hence,consumptionofmethanemaybeprovidingwetlandplantswithnitrogenby diazotrophicMBbutontheotherhandmayalsoleadtoNlossbydenitrifying MB.MBcommunitycompositionwillplayacrucialroleinthisdelicatebalance. Whenitcomestowetlands,inmanycases Methylobacter speciesarethedominant activeMBpresent(Bodelieretal. 2013;Smithetal. 2018).Recentlyithasbeen shownthat Methylobacter genomesretrievedfromfreshwaterwetlandsharbour genesfordissimilatorydenitrificationaswellasgenesforperformingunderhypoxic conditions,suggestingthatthephenotypediscoveredinthedescribedstrain Methylomonasdenitrificans isfunctionalinnatureandismorewidespreadamong typeIMB(Smithetal. 2018).Thepreferenceoflowoxygenconcentrationsfor growthandactivityhasbeendescribedasanichedifferentiatingfactorforMBin otherwetland(Reimetal. 2012)andlakesediments(Hernandezetal. 2015)with always MethylobacterspeciesbeingdominantandactiveMBatlowoxygen(Oshkin etal. 2015). Methylobacter wasalsothemostactiveandabundantMBspeciesin articpeat(Tveitetal. 2013, 2014)aswellasinthehighmethanefensedimentlayers alonganpermafrostthawgradient(Singletonetal. 2018).Thelatterstudyrepresents themostextensivemetagenomeandmetatranscriptomeanalysesofMBcommunitiestodate.Permafostwetlandsreleasehighamountsofmethaneuponthawing whichmakesmitigationbyMBofhighenvironmentalrelevance.Theinvestigated thawinggradientinamiresysteminNorthernSwedenrevealedaclearniche differentiationoftypeIIaandb(USCα)MBinthepermafrostpalsaandinthe acidicbog,whiletypeIMBdominatedthehighmethanedeeperfensedimentlayers (Singletonetal. 2018).TheretrievedMBgenomesrevealedmanyfeaturesof relevanceofMBsurvivalandfunctioninginthesehabitatsandalsoyieldevidence for Hypomicrobia tobelongtothetypeIIMBfamilies.Theauthorsconcludedthat methaneconcentrationwasastrongselectivefactorshapingtheobservedniche differentiation.Fairlyrecent,anoveldescribedMB, CandidatusMethylospira mobilis (typeIb)fromanacidicpeatbog,showeddistinctchemotacticbehaviour towardshighmethaneandlowoxygen,demonstratingthepotentialforMBto activelypositionthemselveswithintheenvironmentalgradientsinherenttowetland habitats(Danilovaetal. 2016).
Thelatterwillbequiteimportantgiventhemostrecentviewonmethane oxidationinwetlandsystemswhereaerobicMBdonothaveexclusiveaccessto methaneconsideringthepresenceofanaerobicmethaneoxidisers(AOM)whichcan mediatedanaerobicmethaneconsumptionbyvariouselectronacceptors,including iron,sulphate,andmanganese(Deanetal. 2018;Vaksmaaetal. 2017b;Welteetal. 2016).Anaerobicmethaneoxidationcanbecoupledtonitriteornitratereduction. Thishasbeenobservedinthecandidatespecies Methylomirabilisoxyfera(nitriteAOM), firstenrichedfromnitrite-richsediments(Ettwigetal. 2010)andtheANME2Darchaeallineage “Candidatus Methanoperedenaceae”,whichhasbeenrecently enrichedfromrice-paddysoils(Haroonetal. 2013;Vaksmaaetal. 2017a).Inrice paddies(Vaksmaaetal. 2017b)aswellasincoastalwetlands(Heetal. 2019), anaerobicmethaneoxidationratesaswellasnumberscanbesubstantialandindicate thatastrongnichedifferentiationofAOMandMBinwetlandsystemtogether formingeffective filtersforthemethaneproducedbyoxidizingfromsourceto
16P.L.E.Bodelieretal.
atmosphere.However,theeffectivenesswillstronglydependontheenvironmental gradientsbutalsoonthetraitsandcompetitiveabilitiesoftherespectiveAOMand MBpresent.
Sometimes,importantcontrollingfactorsareoverlookedintheseinteractions.In arangeofwetlandditchesastrongcorrelationwithmethaneaerobicmethane oxidationandphosphorusavailabilitywasfoundinthesurfacecmofthesediments whichweredominatedbytypeIMB(Veraartetal. 2015).Scanningtheliteratureon these findingsrevealedstrongpositiveeffectsofPonmethaneoxidationinarctic permafrostwetlands,alsodominatedbytypeIMB(Grayetal. 2014).BrowsingMB genomesontraitsrelatingtoPuptakeandutilizationrevealedaclearseparation betweentypeIandIIMB,pointingtowardsastrongerrelianceonPintypeIMB, makingPanimportantnichedifferentiatingfactorinwetlandsystems(Veraartetal. 2015).
3.2.3AquaticSystems
Aquaticsystemsarewaterbodiesclassifiedasfreshwater,transitional,coastalor marine,coastalandmarine.Themarineecosystemisthebiggestcoveringalmost 70%oftheEarthsurfacebutCH4 releasedfromthemtotheatmosphereisasmall sourcewithameanvalueof12TgCH4 year 1 incomparisontoothernatural environments(Saunoisetal. 2016).Onthecontrary,freshwaterecosystems(~4% Earthsurface)cancontributeupto100TgCH4 year 1 (Bastvikenetal. 2011). EstimationsorfuturetrendsinCH4 emissionslackrobustnessduetotherestricted numberofmeasurements,differentmethodologyusedandalsothelittleknowledge onthedynamicsofCH4 emissionsinsomesystems(Reayetal. 2018).Thereleaseof CH4 fromaquaticsystemstotheatmosphereinvolvesprocessessuchasdiffusion, ebullition(Walteretal. 2007),seepage,resuspension(Bussmann 2005)orbioturbation(OliveiraJunioretal. 2019)ofthesediments.Typically,methaneproduction occursinsedimentsoranoxicbottomwaterlayers.However,fairlyrecentitwas discoveredthatoxicwatersaresupersaturatedwithCH4 (Grossartetal. 2011;Karl etal. 2008).Thisphenomenonofthesocalled “methane-paradox” hasspurred aquaticmethanecyclingresearchtopinpointthepossibleabiotic(DelSontroetal. 2017;McGinnisetal. 2017)orbiotic(Grossartetal. 2011;Tangetal. 2016;Yan etal. 2019)factorsunderlyingthisphenomenon(Yanetal. 2019).Thelargeroleof freshwatersystemsinglobalmethaneemission,combinedwiththeunexpectedhigh methaneconcentrationsinoxicwatercolumnshasspurredattentionforMBin aquaticsystems.
MarineCH4 oxidationcanbeperformedbyanaerobicandaerobicCH4 oxidizers (Ruffetal. 2019).Anaerobicoxidationisdrivenbysyntrophicinteractionbetween archaeaandSO4 2-reducing δ-Proteobacteria(Krukenbergetal. 2018).TheseconsortiaofANME-archaea(ANME1,2and3)andsulfate-reducingbacteria(SRB) dominatemethaneoxidationindeep-seasediments(Ruffetal. 2013;McGlynnetal. 2018;Durisch-Kaiseretal. 2005).Thisinteractiondependsoncontrolledgene expressiontoenhanceelectrontransferamongtheconsortiummembers(Krukenberg
MethanotrophEcology,EnvironmentalDistributionandFunctioning17
etal. 2018;McGlynnetal. 2018).AerobicMBinmarineenvironmentsarecomprised by fivesocalleddeep-seaclustersallassociatedwithtypeIlineages(reviewedin Knief 2015).Asyet,thereisnoclearevidenceofthefactorsthatdriveniche differentiationofMBbelongingtotheseclusters(Ruffetal. 2013).Despitethelow amountofO2 indeepregionsofmarinesystems,aerobicmethanotrophsfromthe orderMethylococcalesarepresent(Ruffetal. 2013;Tavorminaetal. 2008).Ithas beenrecentlydescribedthatANME-archaeaandTypeIMBforma “methanotrophic microbiome” whereTypeIMBarethe firsttocolonizethedeep-seasediments(Ruff etal. 2019).RegardingTypeI,somelineagesrelatedtoterrestrialenvironmentsare presentinadeep-seafanbecauseoftheinputsofsoilderivedorganicmatter(Bessette etal. 2017).The pmoA phylotypesfoundinthewater-columndifferfromtheones inhabitantthesedimentsintermsofdiversityandabundance.Forinstance,the planktonicMBmembersaredistributedinthreecladesbelongingtotheorderof Methylococcaleswhereasthebenthiconesarelessdiverse(Tavorminaetal. 2008). Similartowetlands,MBcanbefoundinOxygenMinimumZones(OMZ),regionsin oceansandseaswhereO2 dropsdowntonanoorpicomolarlevels.Theseregions showanactiveCH4 oxidationwhereTypeIMBandNC10(Chronopoulouetal. 2017)seemedtoplayarelevantrole.
Freshwaterecosystemsharborenvironmentssuchaslakes,rivers,pondsand reservoirsplayingarelevantroleintheCcycleaswellasinthecyclingofother nutrients.Theyalsoprovidemanyservicesaswater,electricalenergyandfood supplyorcanserveasrecreationalareas(Vollmeretal. 2016).Still,theyarevery vulnerabletoclimatechangeespeciallyregardingtheincreaseoftemperaturewhich altersthethermalstructureandwaterchemistryofthesesystems(Yvon-Durocher etal. 2011).Amongthefreshwatersystems,lakesarethemoststudiedinrelationto CH4 dynamics.Lakescoverasmallarea(0.9%)oftheEarth’ssurfacebuthavea largecontribution(6–16%)tothetotalglobalCH4 budget(Tangetal. 2016; Bastvikenetal. 2004).Incomparisontootherenvironments,freshwatersystems hold “endemic” MBwhichwemaycalltypicalfreshwaterMB.Theselineages comprisesixsequenceclusterswithintypeIaandIbMB(Knief 2015;Dedyshand Knief 2018).MostoftheseclustersareubiquitousinFWsystems,exceptfortheso calledcluster4awhichisonlydetectedsofarinsedimentswhilecluster2ismore frequentinthewatercolumn(Knief 2015).FirstreportsonfreshwaterMBcommunitiesfocusedonthesediment(NaguibandOverbeck 1970;ReeburghandHeggie 1977;Cappenberg 1974).Thesestudies,demonstratedclearlythatsedimentsharboredmuchhighernumbersofMBthanthepelagicpartofthelake(Zhouetal. 2018).Inmostpelagiczonesoflakes,typeIMBbelongingtothegenus Methylobacter arethedominatingMBgroup(Bleesetal. 2014;Biderre-Petitetal. 2011;Kojimaetal. 2009),Recentlyanovelgenuswithinthefamilyofthe Methylococcaceae, CandidatusMethylomidiphilusalinensis,wasdemonstratedto bethemostabundantMBinahumiclakeinFinland(Rissanenetal. 2018).Nextto this,MBrelatedthe filamentousbacterium Crenothrixpolyspora werereportedtobe themainCH4 sinkintwostratifiedlakes(Oswaldetal. 2017),demonstratingthe lackofknowledgeonMBinfreshwaterhabitats.Incontrast,otherreportsfoundthat TypeIIrepresentativesmayplayarelevantroleinthelacustrineCH4 cycle.For
18P.L.E.Bodelieretal.
instance,inanalkalinelaketherewasanalmostequalabundanceoftypeIIMB, represented2%ofthewholemicrobialcommunity,whiletypeIaccountedbetween 3%and5%(Carinietal. 2005).Intheoxicwatersofanutrientdepriveddeeplake (Zigahetal. 2015)typeIIwasmoreabundantandactiveinnutrientthantypeI (Parksetal. 2018).Similarresultswerefoundinsub-arcticlakes(Heetal. 2012). Surprisingly,highabundanceoftypeIMBhavebeendetectedinanoxicpartsof thewatercolumnoflakes(Schubertetal. 2011)suggestingthattheyareeither inactiveorperforminganaerobicmethaneoxidation.However,thisisnotthecaseas theywereactivelytranscribing pmoA andnoarchaealAOMweredetectable(Blees etal. 2014;Rissanenetal. 2018;Miluckaetal. 2015).Thiswouldsuggestthatthey areinteractingwithothermembersofthemicrobialplanktoniccommunityorusing otherelectronacceptorsratherthanO2.IncubationswithlightincreasedtheCH4 oxidationinanoxicwatersamplesdominatedbyTypeIfromboreal(Rissanenetal. 2018)andtemperatelakes(Oswaldetal. 2015).Thisimpliesaninteractionbetween MBandphototrophs(oxygenicphotosynthesis)asdemonstratedrecentlyina numberoflakes(Miluckaetal. 2015;Oswaldetal. 2016).Hence,thelowamount oflightpenetratingintothedeeperparsoflakesissufficientforphototrophsto produceoxygenusedbymethanotrophstooxidizemethaneaerobically.Recently,it wasalsoreportedthatmembersoftheNC10phylum,i.e. Candidatus Methylomirabilisoxyfera canplayanimportantroleinmethaneoxidation.This bacteriumlinkstheCandNcyclesbycouplingCH4 oxidationwithNO2 /NO3 reduction(nitrate/nitrite-dependentanaerobicmethaneoxidation,n-damo).Ina freshwaterreservoir,clonelibrarieswerecomprisedoftypeI,IIMBwithadominanceofmembersoftheNC10phylum.Thelatterdominatedindeepwatersofthis system(Kojimaetal. 2014).Morerecently,abloomofNC10 “Ca. Methylomirabilis limnetica” reaching27%ofthewholebacterialcommunitydominatedthedeepest waterlayersofastrati fiedlake(Grafetal. 2018).Asimilarstronglinkbetween nitrogencyclingandmethaneoxidationwasfoundin15Indiandam-reservoirs. Using 15N-incubationsitwasshownthatCH4 amendmentboostedNlossfromthis systemswithmorethan10timesduetodenitri ficationbyaerobicMB(Naqvietal. 2018).
AnaerobicCH4 oxidation(AMO)alsotakesplaceinfreshwatersystemsand playsasignificantroleinfreshwaterCH4 dynamics.Thisprocesscanhappen simultaneouslywiththeaerobicoxidation(Arpetal. 2018)orseasonally(Roland etal. 2017).CH4 oxidizersdifferentfromtheProteobacterialwere firstdescribedin anoxicwaterlayersofLakePluβsee(Elleretal. 2005).Thisstudyshowedthe existenceofanaerobicmethane-oxidizingarchaea(ANME1and2)andtheroleof theanaerobicCH4 oxidationinafreshwatersystem.Differenttotheirmarine counterparts,thesearchaealmethanotrophsseemednottodependonsulfatereducingpartners,astherewasnoevidenceofphysicalattachmenttothem.Ina meromictic,temperateandeutrophiclake,CH4 anaerobicoxidationwascoupled withSO4 2 reductiononceNO3 isscarceorbothacceptorsmaybeusedconcomitantly(Rolandetal. 2017).Inatropicallake,anewclusterofANME(differentfrom themarinemembers)wasdescribedandassociatedtoSO4 2 (Zigahetal. 2015).
MethanotrophEcology,EnvironmentalDistributionandFunctioning19