We report eight genomes from representatives of the phylum Acidobacteria subdivisions 1 and 3, isolated from soils. The genome sizes range from 4.9 to 6.7 Mb. Genomic analysis reveals putative genes for low- and high-affinity respiratory oxygen reductases, high-affinity hydrogenases, and the capacity to use a diverse collection of carbohydrates

Clum, Alicia

Daum, Chris

Eichorst, Stephanie A

Goodwin, Lynne A

Huntemann, Marcel

Ivanova, Natalia

Kyrpides, Nikos

Mikhailova, Natalia

Palaniappan, Krishnaveni

Pillay, Manoj

Reddy, TBK

Shapiro, Nicole

Stamatis, Dimitrios

Trojan, Daniela

Varghese, Neha

Woebken, Dagmar

Woyke, Tanja

English

eScholarship - University of California

Lawrence Berkeley National LaboratoryLBL PublicationsTitleOne Complete and Seven Draft Genome Sequences of Subdivision 1 and 3 Acidobacteria Isolated from Soil.Permalinkhttps://escholarship.org/uc/item/8kv096skJournalMicrobiology resource announcements, 9(5)ISSN2576-098XAuthorsEichorst, Stephanie ATrojan, DanielaHuntemann, Marcelet al.Publication Date2020DOI10.1128/mra.01087-19 Peer reviewedeScholarship.org Powered by the California Digital LibraryUniversity of CaliforniaOne Complete and Seven Draft Genome Sequences ofSubdivision 1 and 3 Acidobacteria Isolated from SoilStephanie A. Eichorst,a Daniela Trojan,a Marcel Huntemann,b Alicia Clum,b Manoj Pillay,b Krishnaveni Palaniappan,bNeha Varghese,b Natalia Mikhailova,b Dimitrios Stamatis,b T. B. K. Reddy,b Chris Daum,b Lynne A. Goodwin,c Nicole Shapiro,bNatalia Ivanova,b Nikos Kyrpides,b Tanja Woyke,b Dagmar WoebkenaaDivision of Microbial Ecology, Department of Microbiology and Ecosystem Science, Centre for Microbiology and Environmental Systems Science, University of Vienna,Vienna, AustriabDepartment of Energy, Joint Genome Institute, Berkeley, California, USAcMaterials Science and Technology, Engineered Materials, Los Alamos National Laboratory, Los Alamos, New Mexico, USAABSTRACT We report eight genomes from representatives of the phylum Acidobac-teria subdivisions 1 and 3, isolated from soils. The genome sizes range from 4.9 to6.7 Mb. Genomic analysis reveals putative genes for low- and high-affinity respiratoryoxygen reductases, high-affinity hydrogenases, and the capacity to use a diverse col-lection of carbohydrates.The Acidobacteria constitute a large, diverse, and phylogenetically distinct phylum withcurrently 26 defined subdivisions based on the 16S rRNA gene phylogeny (1). Typically,members of subdivisions 1, 2, 3, 4, 5, and 6 populate terrestrial environments at a highrelative sequence abundance based on rRNA gene libraries (as high as 20 to 40% of thetotal bacterial community) (2, 3). Phylogenetically representative acidobacterium strainswere chosen for genome sequencing, which were isolated from soils (meadow grassland,agricultural, or peat bog) (4–7). More specifically, six strains in subdivision 1 (Terriglobusroseus KBS 63, Acidobacteriaceae bacterium strain KBS 83, Acidobacteriaceae bacteriumstrain KBS 89, Acidobacteriaceae bacterium strain KBS 146, Terriglobus sp. strain TAA 43, andAcidobacteriaceae bacterium strain TAA 166) and two strains in subdivision 3 (Acidobacteriabacterium strain KBS 96 and Bryobacter aggregatus MPL3) were sequenced to better populatethe phylogenetic branches in the phylum Acidobacteria with genomic representation.Strains were grown on either a modified minimal medium or R2 medium as describedpreviously (5–7). Genomic DNA (gDNA) was isolated using a modified cetyltrimethylam-monium bromide (CTAB) DNA extraction protocol as recommended by the Department ofEnergy (DOE) Joint Genome Institute (JGI). Details on the isolation and growth conditionsalong with the sequencing can be found in Table 1. The genomes across all strains rangedin size from 4.9 to 6.7 Mb (Table 1), which is similar to the size range of the previouslysequenced acidobacterial strains (8, 9). The GC content ranged from 57 to 60 mol%(Table 1). Although many of these genomes are considered to be permanent drafts(Table 1), CheckM analysis indicated that they are ca. 95% complete (10).The genomes of KBS 63 and KBS 96 harbored the catalytic subunit for the high-affinitycbb3-type cytochrome c oxidase (heme-copper-oxygen reductase [HCO] type C, EC 1.9.3.1,KEGG orthology [KO] number K00404), while the genomes of KBS 146 and TAA 166harbored the high-affinity cytochrome bd-type quinol oxidase (EC 7.1.1, KO numberK00425) (10). All strains harbored at least one homologue of the low-affinity terminaloxidase HCO type A (EC 1.9.3.1, KO number K02274) (10). Numerous glycoside hydrolase(GH) families were detected across the genomes, with GH109, GH74, and GH13 being someof the most prevalent (10). The genomes of KBS 83 and KBS 96 harbored a high-affinitygroup 1h hydrogenase (EC 1.12.99.6, KO number K06281), presumably giving them thepotential to scavenge atmospheric concentrations of hydrogen during periods of starvationCitation Eichorst SA, Trojan D, Huntemann M,Clum A, Pillay M, Palaniappan K, Varghese N,Mikhailova N, Stamatis D, Reddy TBK, Daum C,Goodwin LA, Shapiro N, Ivanova N, Kyrpides N,Woyke T, Woebken D. 2020. One complete andseven draft genome sequences of subdivision1 and 3 Acidobacteria isolated from soil.Microbiol Resour Announc 9:e01087-19.https://doi.org/10.1128/MRA.01087-19.Editor Irene L. G. Newton, Indiana University,BloomingtonCopyright © 2020 Eichorst et al. This is anopen-access article distributed under the termsof the Creative Commons Attribution 4.0International license.Address correspondence to Stephanie A.Eichorst, stephanie.eichorst@univie.ac.at.This article is Los Alamos unclassified reportLA-UR-18-24028.Received 1 October 2019Accepted 17 December 2019Published 30 January 2020GENOME SEQUENCEScrossmVolume 9 Issue 5 e01087-19 mra.asm.org 1TABLE1Isolationconditions,librarypreparationinformation,genomesequencingstatistics,andaccessionnumbersforthedescribedacidobacterialgenomesStrainIsolationsource;yrIsolationandgrowthconditions(reference)LibraryprepnNo.ofreads(technology)Avgreadlength(bp)Assembler(reference)No.ofcontigs(no.ofscaffolds)N/L50of:Avggenomecoverage()Genomesize(Mb)GCcontent(mol%)AssemblylevelGenBankaccessionno.SRAaccessionno.ScaffoldsContigsKBS63KelloggBiologicalStation,HickoryCorners,MI,USA,grasslandsoil(treatment8);2003VSB-7mixorganicCsubstratesinCO2-enrichedair(5)454paired-endlibrary;aIlluminastd.shotgunlibraryb69,481,460(IlluminaSolexa);514,324(454pairedend)76(IlluminaSolexa);151(454pairedend)Newblerv2.3-Prerelease6/30/2009,Velvetv1.0.13(13)33(2)1/5.2Mb1/5.2Mb1083.1(IlluminaSolexa);8.5(454pairedend)5.2360.3CompleteCP003379SRS1568612KBS83KelloggBiologicalStation,HickoryCorners,MI,USA,agriculturalsoil(treatment1);2006VL55mixofplantpolymericCinair(7)Illuminastd.shotgunlibraryb12,985,884(IlluminaHiSeq2000)2150Velvetv1.1.04(13),wgsime ,Allpaths-LGvr41043(14)25(25)5/504.7kb5/504.7kb3906.2559.2PermanentdraftARMD00000000SRS844143KBS89KelloggBiologicalStation,HickoryCorners,MI,USA,grasslandsoil(treatment8);2002VSB-6.8mixorganicCsubstrates,acylhomoserinelactonesinCO2-enrichedair(4,5)Illuminastd.shotgunlibraryb12,475,762(IlluminaHiSeq2000)2150Velvetv1.1.04(13),wgsime ,Allpaths-LGvr41043(14)14(14)3/943.6kb3/943.6kb122.66.0157.6PermanentdraftARME00000000SRS844144KBS146KelloggBiologicalStation,HickoryCorners,MI,USA,grasslandsoil(treatment8);2006VL55mixoforganicCsubstrates,CO2-enrichedhypoxia(11)PacBioSMRTbelllibraryd185,131(PacBioRSplatform)3,4932,894HGAPv2.0.0(15)2(2)1/5.0Mb1/5.0Mb64.15.0056.7PermanentdraftJHVA00000000SRS1534005TAA43HindgutofReticulitermesflavipes(Kollar)(Phinotermitidae),Dansville,MI,USA;2002VSB-7yeastextractandpeptoneinCO2-enrichedair(4,5)Illuminastd.shotgunlibraryb13,649,630(Illuminastd.pairedend,IlluminaHiSeq2000)2151Velvetv1.2.07(13),wgsime ,Allpaths-LGvr46652(14)7(7)1/3.5Mb1/3.5Mb302.04.9556.7PermanentdraftJUGR00000000SRS1366045TAA166HindgutofR.flavipes(Kollar)(Phinotermitidae),Dansville,MI,USA;2002VSB-7yeastextractandpeptoneinCO2-enrichedair(4,5)Illuminastd.shotgunlibrarybandlong-insertmatepairlibraryc ;PacBioSMRTbelllibraryd40,920,398(IlluminaCLIPpairedend,IlluminaHiSeq2000);15,055,388(Illuminastd.pairedend,IlluminaHiSeq2000);233,258(PacBioRSplatform)290;2150;12259AllpathsLGvr42328(14)3(3)1/4.7Mb1/4.7Mb973.9(Illumina);85.4(PacBio)6.1458.8PermanentdraftATWD00000000SRS438888KBS96KelloggBiologicalStation,HickoryCorners,MI,USA,agriculturalsoil(treatment1);2006VL55mixofplantpolymericCinair(7)Illuminastd.shotgunlibrarybandlong-insertmatepairlibraryc14,485,381(Illuminastd.pairedend,HiSeq2000);58,296,769(IlluminaCLIPpairedend,IlluminaHiSeq2000)2150;289Allpaths-LGvr41043(14)13(2)1/6.7Mb2/1.6Mb1,098.66.6957.2PermanentdraftARMF00000000SRS438892(Continuedonnextpage)Eichorst et al.Volume 9 Issue 5 e01087-19 mra.asm.org 2TABLE1(Continued)StrainIsolationsource;yrIsolationandgrowthconditions(reference)LibraryprepnNo.ofreads(technology)Avgreadlength(bp)Assembler(reference)No.ofcontigs(no.ofscaffolds)N/L50of:Avggenomecoverage()Genomesize(Mb)GCcontent(mol%)AssemblylevelGenBankaccessionno.SRAaccessionno.ScaffoldsContigsMPL3AcidicSphagnumpeatbog,Bakchar,Tomskregion,WestSiberia;2004Biofilm-mediatedenrichmentapproach(6,12)PacBioSMRTbelllibraryd208,346(PacBioRSplatform)3,0872,325HGAPv2.0.0(15)4(4)1/4.3Mb1/4.3Mb176.85.7557.0PermanentdraftJNIF00000000SRS1520682a454Titanium,pairedends,8kb;15ggenomicDNAareshearedbytheHydroshearto~8-kbsizefragments.Theshearedsamplesarethengelselectedforthe8-kbbands,purified,andligatedtothe42-bploxPlinkersoneitherend.TheseloxPlinkersarelabeledbybiotin.TheloxPlinker-ligatedfragmentsarethencircularizedbytheCrerecombinase.Asaresult,theendsof20-kbfragmentsarebroughttogetherandbridgedbyasingleloxPlinker.ThesecircularDNAsarefurthershearedto500-bpfragments,andthefragmentscarryingtheloxPlinkersarerecoveredbytheStreptavidin-coatedmagneticbeads.Consequently,theloxPlinker-containingfragmentsareligatedtothe454TitaniumadaptersAandBinthesamewaythattheshotgunlibrariesarecreated.The454libraryfragmentsarethenclonallyamplifiedinbulkbycapturingthemthroughhybridizationonmicroparticlebeadsandsubjectingthemtoemulsion-basedPCR.ThisresultsinbeadsthatarecoveredwithmillionsofcopiesofasingleDNAfragment(range,400--800bp),whereeachbeadcontainsadifferentclonallyamplifiedlibraryfragment.Afteramplification,thebeadsarerecoveredfromtheemulsionsandareloadedintothewellsofaPicoTiterPlate(PTP)devicesuchthatwellscontainsingleDNAbeads.ThePTPdeviceistheninsertedintothe454genomesequencerFLX-TitaniuminstrumentforsequencingwheresequencingreagentsaresequentiallyflowedoverthePTPwells.Eachincorporationofanucleotidecomplementarytothetemplatestrandresultsinachemiluminescentlightsignalthatisrecordedbyacamera,andthesequenceoftheDNAfragmentsisdetermined.Thissequencing-by-synthesismethodisknownaspyrosequencing.bIlluminaregularfragment,300bp;100ngofDNAwasshearedto300bpusingtheCovarisLE220instrumentandsizeselectedusingsolid-phasereversibleimmobilization(SPRI)beads(BeckmanCoulter).Thefragmentsweretreatedwithendrepair,A-tailing,andligationofIllumina-compatibleadapters(IDT,Inc.)usingtheKAPA-Illuminalibrarycreationkit(KapaBiosystems).ThepreparedlibrarieswerequantifiedusingKapaBiosystem’snext-generationsequencinglibraryquantitativePCR(qPCR)kitandrunonaRocheLightCycler480real-timePCRinstrument.Thequantifiedlibrarieswerethenmultiplexedwithotherlibraries,andthepooloflibrarieswasthenpreparedforsequencingontheIlluminaHiSeqortheIlluminaGAIIx(KBS63)sequencingplatformutilizingaTruSeqpaired-endclusterkitv3orapaired-endclustergenerationkit,v4(KBS63)andIllumina’scBotinstrumenttogenerateaclusteredflowcellforsequencing.SequencingoftheflowcellwasperformedontheIlluminaHiSeq2000sequencerusingaTruSeqSBSsequencingkitv3followinga2150indexedrunrecipeandontheIlluminaGAIIxsequencerusingSBSsequencingkits,v4,followinga276runrecipe(KBS63).cIlluminaregularlongmatepair,8kb,Cre-Loxrandomshearmatepairsequences;5gofDNAwasshearedusingCovarisg-TUBEsandwasgelsizeselectedfor8kb.TheshearedDNAwastreatedwithendrepairandligatedwithbiotinylatedadapterscontainingloxP.Theadapter-ligatedDNAfragmentswerecircularizedviarecombinationwithaCreexcisionreaction(NEB).ThecircularizedDNAtemplateswerethenrandomlyshearedusingtheCovarisLE220instrument.TheshearedfragmentsweretreatedwithendrepairandA-tailingusingtheKAPA-Illuminalibrarycreationkit(KapaBiosystems)followedbyimmobilizationofmatepairfragmentsonstreptavidinbeads(Invitrogen).Illumina-compatibleadapters(IDT,Inc.)wereligatedtothematepairfragments,and14cyclesofPCRwereusedtoenrichforthefinallibrary(KapaBiosystems).ThepreparedlibrarieswerequantifiedusingKapaBiosystem’snext-generationsequencinglibraryqPCRkitandrunonaRocheLightCycler480real-timePCRinstrument.Thequantifiedlibrarieswerethenmultiplexedwithotherlibraries,andthepooloflibrarieswasthenpreparedforsequencingontheIlluminaHiSeqsequencingplatformutilizingaTruSeqpaired-endclusterkitv3andIllumina’scBotinstrumenttogenerateaclusteredflowcellforsequencing.SequencingoftheflowcellwasperformedontheIlluminaHiSeq2000sequencerusingaTruSeqSBSsequencingkitv3followinga2100indexedrunrecipe.dPacBio10-kblibrarieswithAMPurebeadsizeselection;unamplifiedlibrariesweregeneratedusingthePacificBiosciencesstandardtemplatepreparationprotocolforcreating10-kblibraries.Atotalof5gofgenomicDNAwasusedtogenerateeachlibrary,andtheDNAwasshearedusingCovarisg-TUBEstogenerateshearedfragments10kbinlength.TheshearedDNAfragmentswerethenpreparedusingPacificBiosciencesSMRTbelltemplatepreparationkitv1.0,wherethefragmentsweretreatedwithDNAdamagerepair,hadtheirendsrepairedsothattheywerebluntended,andwere5=phosphorylated.PacificBioscienceshairpinadapterswerethenligatedtothefragmentstocreatetheSMRTbelltemplateforsequencing.TheSMRTbelltemplateswerethenpurifiedusingexonucleasetreatmentsandsizeselectedusingAMPurePBbeads.SequencingprimerwasthenannealedtotheSMRTbelltemplates,andversionXLsequencingpolymerasewasboundtothem.ThepreparedSMRTbelltemplatelibrarieswerethensequencedonaPacificBiosciencesRSIIsequencerusingversionC2chemistryand2-hsequencingmovieruntimes.ehttps://github.com/lh3/wgsim.Microbiology Resource AnnouncementVolume 9 Issue 5 e01087-19 mra.asm.org 3(10). The addition of these genomes in the public databases will help to further ourunderstanding of this prevalent and diverse phylum.Data availability. The genome sequences are available in GenBank; accessionnumbers can be found in Table 1.ACKNOWLEDGMENTSWe thank Svetlana Dedysh for donating her strain Bryobacter aggregatus MPL3T and theDivision of Computational Systems Biology for providing and maintaining the Life ScienceCompute Cluster (LiSC) at the University of Vienna.This work was supported by an Austrian Science Fund FWF project grant (P26392-B20to D.W. and S.A.E.) and a Joint Genome Institute Community Sequencing Program, “Pop-ulating the Branches of the Phylum Acidobacteria” (number 386 to S.A.E., Cheryl R. Kuske,and Jonathan Eisen). The work conducted by the U.S. Department of Energy Joint GenomeInstitute, a DOE Office of Science User Facility, is supported by the Office of Science of theU.S. Department of Energy under contract number DE-AC02-05CH11231.REFERENCES1. Barns SM, Cain EC, Sommerville L, Kuske CR. 2007. Acidobacteria phylumsequences in uranium-contaminated subsurface sediments greatly ex-pand the known diversity within the phylum. Appl Environ Microbiol73:3113–3116. https://doi.org/10.1128/AEM.02012-06.2. Lipson DA, Schmidt SK. 2004. Seasonal changes in an alpine soil bacterialcommunity in the Colorado Rocky Mountains. Appl Environ Microbiol70:2867–2879. https://doi.org/10.1128/aem.70.5.2867-2879.2004.3. Janssen PH. 2006. Identifying the dominant soil bacterial taxa in librariesof 16S rRNA and 16S rRNA genes. Appl Environ Microbiol 72:1719 –1728.https://doi.org/10.1128/AEM.72.3.1719-1728.2006.4. Stevenson BS, Eichorst SA, Wertz JT, Schmidt TM, Breznak JA. 2004. Newstrategies for cultivation and detection of previously uncultured mi-crobes. Appl Environ Microbiol 70:4748 – 4755. https://doi.org/10.1128/AEM.70.8.4748-4755.2004.5. Eichorst SA, Breznak JA, Schmidt TM. 2007. Isolation and characterizationof soil bacteria that define Terriglobus gen. nov., in the phylum Acido-bacteria. Appl Environ Microbiol 73:2708 –2717. https://doi.org/10.1128/AEM.02140-06.6. Kulichevskaya IS, Suzina NE, Liesack W, Dedysh SN. 2010. Bryobacteraggregatus gen. nov., sp. nov., a peat-inhabiting, aerobic chemo-organotroph from subdivision 3 of the Acidobacteria. Int J Syst EvolMicrobiol 60:301–306. https://doi.org/10.1099/ijs.0.013250-0.7. Eichorst SA, Kuske CR, Schmidt TM. 2011. Influence of plant polymerson the distribution and cultivation of bacteria in the phylum Acido-bacteria. Appl Environ Microbiol 77:586 –596. https://doi.org/10.1128/AEM.01080-10.8. Ward NL, Challacombe JF, Janssen PH, Henrissat B, Coutinho PM, Wu M,Xie G, Haft DH, Sait M, Badger J, Barabote RD, Bradley B, Brettin TS,Brinkac LM, Bruce D, Creasy T, Daugherty SC, Davidsen TM, DeBoy RT,Detter JC, Dodson RJ, Durkin AS, Ganapathy A, Gwinn-Giglio M, Han CS,Khouri H, Kiss H, Kothari SP, Madupu R, Nelson KE, Nelson WC, PaulsenI, Penn K, Ren Q, Rosovitz MJ, Selengut JD, Shrivastava S, Sullivan SA,Tapia R, Thompson LS, Watkins KL, Yang Q, Yu C, Zafar N, Zhou L, KuskeCR. 2009. Three genomes in the phylum Acidobacteria provide insightinto their lifestyles in soils. Appl Environ Microbiol 74:2046 –2056.https://doi.org/10.1128/AEM.02294-08.9. Rawat SR, Männistö MK, Bromberg Y, Häggblom MM. 2012. Comparativegenomic and physiological analysis provides insights into the role of Aci-dobacteria in organic carbon utilization in Arctic tundra soils. FEMS Micro-biol Ecol 82:341–355. https://doi.org/10.1111/j.1574-6941.2012.01381.x.10. Eichorst SA, Trojan D, Roux S, Herbold C, Rattei T, Woebken D. 2018.Genomic insights into the Acidobacteria reveal strategies for their success interrestrial environments. Environ Microbiol 20:1041–1063. https://doi.org/10.1111/1462-2920.14043.11. Eichorst SA. 2007. Isolation and characterization of members of thephylum Acidobacteria from soils. PhD thesis. Michigan State University,East Lansing, MI. https://doi.org/10.25335/M5804XZ94.12. Dedysh SN, Pankratov TA, Belova SE, Kulichevskaya IS, Liesack W.2006. Phylogenetic analysis and in situ identification of Bacteria com-munity composition in an acidic Sphagnum peat bog. Appl EnvironMicrobiol 72:2110 –2117. https://doi.org/10.1128/AEM.72.3.2110-2117.2006.13. Zerbino DR, Birney E. 2008. Velvet: algorithms for de novo short readassembly using de Bruijn graphs. Genome Res 18:821– 829. https://doi.org/10.1101/gr.074492.107.14. Gnerre S, MacCallum I, Przybylski D, Ribeiro FJ, Burton JN, Walker BJ,Sharpe T, Hall G, Shea TP, Sykes S, Berlin AM, Aird D, Costello M, Daza R,Williams L, Nicol R, Gnirke A, Nusbaum C, Lander ES, Jaffe DB. 2011.High-quality draft assemblies of mammalian genomes from massivelyparallel sequence data. Proc Natl Acad Sci U S A 108:1513–1518. https://doi.org/10.1073/pnas.1017351108.15. Chin CS, Alexander DH, Marks P, Klammer AA, Drake J, Heiner C, Clum A,Copeland A, Huddleston J, Eichler EE, Turner SW, Korlach J. 2013. Non-hybrid, finished microbial genome assemblies from long-read SMRTsequencing data. Nat Methods 10:563–569. https://doi.org/10.1038/nmeth.2474.Eichorst et al.Volume 9 Issue 5 e01087-19 mra.asm.org 4

One Complete and Seven Draft Genome Sequences of Subdivision 1 and 3 Acidobacteria Isolated from Soil.

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One Complete and Seven Draft Genome Sequences of Subdivision 1 and 3 Acidobacteria Isolated from Soil.

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