Biology and Fertility of Soils

, Volume 55, Issue 8, pp 777–788 | Cite as

Temporal and spacial dynamics of metabolically active bacteria associated with ectomycorrhizal roots of Betula pubescens

  • Hironari IzumiEmail author
Original Paper


Bacteria, mycorrhizal fungi and their host plants are ubiquitous terrestrial associations but their interactions are poorly understood. To investigate the effects of seasonal change of host plant phenology on the bacterial communities in ectomycorrhizal (ECM) roots, bacterial communities in birch root tips with different ECM fungal species, those of non-mycorrhizal roots and those in bulk soil, were examined over a 15-month period, including two autumn seasons of 2008 and 2009, in field settings. Morphotyping and sequencing to identify the fungal species and RNA-based DGGE, cloning and sequencing were used to characterise the active bacterial communities. The most frequently observed fungus was Leccinum scabrum while other ECM fungi including Hebeloma velutipes, Russula versicolor and unidentified Cortinarius and Tomentella species were commonly found. ECM roots with L. scabrum hosted declining numbers of bacterial sequence types, observed as distinct bands on the DGGE profile, from spring to autumn in 2009. The numbers of sequence types associated with L. scabrum and non-mycorrhizal roots were reduced in autumn 2009 than in autumn 2008, but the numbers associated with other ECM roots and bulk soil were constant over the same period. Roots with L. scabrum harboured unique bacterial communities which were distinct from bulk soils, non-mycorrhizal roots and roots with other ECM fungi in 2008 but the differences were less clear in 2009. The results indicate that the metabolically active bacterial communities in ECM are influenced by fungal species, seasonal host plant phenology and show year to year variation.


Ectomycorrhizal Betula Microbial community Lactobacillus RNA 



I acknowledge Katarina Ihrmark and Maria Jonsson for their laboratory assistance and Drs Anna Rosling, Petra M.A. Fransson and Roger D. Finlay for their helpful input to the project.

Funding information

This work was financially supported by a postdoc grant from the Department of Forest Mycology and Pathology, SLU.

Supplementary material

374_2019_1393_MOESM1_ESM.doc (285 kb)
ESM 1 (DOC 282 kb)


  1. Agerer R (1987) Colour atlas of ectomycorrhizae. Einhorn-Verlag, MunichGoogle Scholar
  2. Artursson V, Finlay RD, Jansson JK (2005) Combined bromodeoxyuridine immunocapture and terminal-restriction fragment length polymorphism analysis highlights differences in the active soil bacterial metagenome due to Glomus mosseae inoculation or plant species. Environ Microbiol 7:1952–1966PubMedGoogle Scholar
  3. Baldrian P, Kolařík M, Stursová M, Kopecký J, Valášková V, Větrovský T, Zifčáková L, Snajdr J, Rídl J, Vlček C, Voříšková J (2012) Active and total microbial communities in forest soil are largely different and highly stratified during decomposition. ISME J:248–258PubMedPubMedCentralGoogle Scholar
  4. Blazewicz SJ, Barnard RL, Daly RA, Firestone MK (2013) Evaluating rRNA as an indicator of microbial activity in environmental communities: limitations and uses. ISME J 7:2061–2068PubMedPubMedCentralGoogle Scholar
  5. Bonfante P, Anca I-A (2009) Plants, mycorrhizal fungi, and bacteria: a network of interactions. Annu Rev Microbiol 63:363–383PubMedGoogle Scholar
  6. Burke DJ, Dunham SM, Kretzer AM (2008) Molecular analysis of bacterial communities associated with the roots of Douglas fir (Pseudotsuga menziesii) colonized by different ectomycorrhizal fungi. FEMS Micro Ecol 65:299–309Google Scholar
  7. Calvaruso C, Turpault MP, Leclerc E, Frey-Klett P (2006) Impact of ectomycorrhizosphere on the functional diversity of soil bacterial and fungal communities from a forest stand in relation to nutrient mobilization processes. Microb Ecol 54:567–577Google Scholar
  8. Daranas N, Rosello G, Cabrefiga J, Donati I, Frances J, Badosa E, Spinelli F, Montesinos E, Bonaterra A (2018) Biological control of bacterial plant diseases with Lactobacillus plantarum strains selected for their broad-spectrum activity. Ann Appl Biol 2018:1–14Google Scholar
  9. Den Bakker HC, Zuccarello GC, Kuyper TW, Noordeloos ME (2004) Evolution and host specificity in the ectomycorrhizal genus Leccinum. New Phytol 163:201–215Google Scholar
  10. Dennis PG, Miller AJ, Hirsch PR (2010) Are root exudates more important than other sources of rhizodeposits in structuring rhizosphere bacterial communities? FEMS Microbiol Ecol 72:313–327PubMedGoogle Scholar
  11. Deveau A, Palin B, Delaruelle C, Peter M, Kohler A, Pierrat JC, Sarniguet A, Garbaye J, Martin F, Frey-Klett P (2005) The mycorrhiza helper Pseudomonas fluorescens BBc6R8 has a specific priming effect on the growth, morphology and gene expression of the ectomycorrhizal fungus Laccaria bicolor S238N. New Phytol 175:743–755Google Scholar
  12. Felske A, Wolterink A, Van Lis R, De Vos WM, Akkermans ADL (2000) Response of a soil bacterial community to grassland succession as monitored by 16S rRNA levels of the predominant ribotypes. Appl Environ Microbiol 66:3998–4003PubMedPubMedCentralGoogle Scholar
  13. Frey-Klett P, Garbaye J, Tarkka M (2007) The mycorrhiza helper bacteria revisited. New Phytol 176:22–36PubMedGoogle Scholar
  14. Galperin MY, Cochrane GR (2009) Nucleic acids research annual database issue and the NAR online molecular biology database collection in 2009. Nucleic Acids Res 37:D1–D4PubMedGoogle Scholar
  15. Garbaye J (1994) Helper bacteria - a new dimension to the mycorrhizal symbiosis. New Phytol 128:197–210Google Scholar
  16. Gardes M, Bruns TD (1993) Its primers with enhanced specificity for basidiomycetes - application to the identification of mycorrhizae and rusts. Mol Ecol 2:113–118PubMedGoogle Scholar
  17. Grady EN, MacDonald J, Liu L, Richman A, Yuan Z-C (2016) Current knowledge and perspectives of Paenibacillus: a review. Microb Cell Factories 15:203. CrossRefGoogle Scholar
  18. Gremion F, Chatzinotas A, Harms H (2003) Comparative 16S rDNA and 16S rRNA sequence analysis indicates that Actinobacteria might be a dominant part of the metabolically active bacteria in heavy metal-contaminated bulk and rhizosphere soil. Environ Microbiol 5:896–907PubMedGoogle Scholar
  19. Hauben L, Vauterin L, Swings J, Moore ERB (1997) Comparison of 16S ribosomal DNA sequences of all Xanthomonas species. Int J Syst Bacteriol 47:328–335PubMedGoogle Scholar
  20. Högberg NM, Briones MJI, Keel SG, Metcalfe DB, Campbell C, Midwood AJ, Thornton B, Hurry B, Linder S, Näsholm T, Högberg P (2010) Quantification of effects of season and nitrogen supply on tree below-ground carbon transfer to ectomycorrhizal fungi and other soil organisms in a boreal pine forest. New Phytol 187:485–493PubMedGoogle Scholar
  21. Huang X-F, Chaparro JM, Reardon KF, Zhang R, Shen Q, Vivanco JM (2014) Rhizosphere interactions: root exudates, microbes, and microbial communities. Botany 92:267–275Google Scholar
  22. Izumi H, Finlay RD (2011) Selective association of bacterial and ascomycete communities with ectomycorrhizas in Swedish subarctic birch forest. Environ Microbiol 13:819–830PubMedGoogle Scholar
  23. Izumi H, Anderson IC, Alexander IJ, Killham K, Moore ERB (2006) Endobacteria in some ectomycorrhiza of Scots pine (Pinus sylvestris). FEMS Microb Ecol 56:34–43Google Scholar
  24. Izumi H, Moore ERB, Killham K, Alexander IJ, Anderson IC (2007) Characterisation of endobacterial communities in ectomycorrhizas by DNA- and RNA-based molecular methods. Soil Biol Biochem 39:891–899Google Scholar
  25. Izumi H, Cairney JW, Killham K, Moore E, Alexander IJ, Anderson IC (2008) Bacteria associated with ectomycorrhizas of slash pine (Pinus elliottii) in south-eastern Queensland, Australia. FEMS Microbiol Lett 282:196–204PubMedGoogle Scholar
  26. Johansson T (1993) Seasonal changes in contents of root starch and soluble carbohydrates in 4-6 year old Betula pubescens and Populus tremula. Scand J For Res 8:94–106Google Scholar
  27. Klappenbach JA, Dunbar JM, Schmidt TM (2000) rRNA operon copy number reflects ecological strategies of bacteria. Appl Environ Microbiol 66:1328–1333PubMedPubMedCentralGoogle Scholar
  28. Kõljalg U, Larsson KH, Abarenkov K, Nilsson RH, Alexander IJ, Eberhardt U, Erland S, Høiland K, Kjøller R, Larsson E, Pennanen T, Sen R, Taylor AF, Tedersoo L, Vrålstad T, Ursing BM (2005) UNITE: a database providing web-based methods for the molecular identification of ectomycorrhizal fungi. New Phytol 166:1063–1068PubMedGoogle Scholar
  29. Koranda M, Schnecker J, Kaiser C, Fuchslueger L, Kitzler B, Stange CF, Sessitsch A, Zechmeister-Boltenstern S, Richter A (2011) Microbial process and community composition in the rhizosphere of European beech-the influence of plant C exudates. Soil Biol Biochem 43:551–558PubMedPubMedCentralGoogle Scholar
  30. Kozlowski TT, Pallardy SG (1997) Physiology of woody plants, 2nd edn. Academic Press, San Diego. USAGoogle Scholar
  31. Lee ZMP, Bussema C, Schmidt TM (2009) rrnDB: documenting the number of rRNA and tRNA genes in bacteria and archaea. Nucleic Acids Res 37:D489–D493PubMedGoogle Scholar
  32. Mahmood S, Paton GI, Prosser JI (2005) Cultivation-independent in situ molecular analysis of bacteria involved in degradation of pentachlorophenol in soil. Environ Microbiol 7:1349–1360PubMedGoogle Scholar
  33. Marupakula S, Mahmood S, Finlay RD (2015) Analysis of single root tip microbiomes suggests that distinctive bacterial communities are selectetd by Pinus sylvestris roots colonized by different ectomycorrhizal fungi. Environ Microbiol 18:1470–1483PubMedGoogle Scholar
  34. Marupakula S, Mahmood S, Jernberg J, Nallanchakravarthula S, Fahad ZA, Finlay RD (2017) Bacterial microbiomes of individual ectomycorrhizal Pinus sylvestris roots are shaped by soil horizon and differentially sensitive to nitrogen addition. Environ Microbiol 19:4736–4753PubMedGoogle Scholar
  35. McCune B, Grace JB (2002) Analysis of ecological communities. MjM Software design, Gleneden BeachGoogle Scholar
  36. Mocali S, Bertelli E, Di Cello F, Mengoni A, Sfalanga A, Viliani F, Caciottia A, Teglib S, Giuseppe S, Fan R (2003) Fluctuation of bacteria isolated from elm tissues during different seasons and from different plant organs. Res Microbiol 154:105–114PubMedGoogle Scholar
  37. Morrison E, Lagos L, Al-Agely A, Glaab H, Johnson W, Jorquera MA, Ogram A (2017) Mycorrhizal inoculation increases genes associated with nitrification and improved nutrient retention in soil. Biol Fertil Soils 53:275–279Google Scholar
  38. Muyzer G, Dewaal EC, Uitterlinden AG (1993) Profiling of complex microbial populations by denaturing gradient gel electrophoresis analysis of polymerase chain reaction amplified genes-coding for 16s ribosomal RNA. Appl Environ Microbiol 59:695–700PubMedPubMedCentralGoogle Scholar
  39. Nguyen NH, Bruns TD (2015) The microbiome of Pinus muricata ectomycorrhizae: community assemblages, fungal species effects and Burkholderia as important bacteria in multipartnered symbiosis. Microb Ecol 69:914–921PubMedGoogle Scholar
  40. Nogales B, Moore ERB, Abraham WR, Timmis KN (1999) Identification of the metabolically active members of a bacterial community in a polychlorinated biphenyl polluted moorland soil. Environ Microbiol 1:199–212PubMedGoogle Scholar
  41. Offre P, Pivato B, Siblot S, Gamalero E, Corberand T, Lemanceau P, Mougel C (2007) Identification of bacterial groups preferentially associated with mycorrhizal roots of Medicago truncatula. Appl Environ Microbiol 73:913–921PubMedGoogle Scholar
  42. Palomino MR, Garcia JAL, Ramos B, Manero FJG, Probanza A (2005) Seasonal diversity changes in alder (Alnus glutinosa) culturable rhizobacterial communities throughout a phenological cycle. Appl Soil Ecol 29:215–224Google Scholar
  43. Pohjanen J, Koskimaki JJ, Sutela S, Ardanov P, Suorsa M, Niemi K, Sarjala T, Haggman H, Prittla AM (2014) Interaction with ectomycorrhizal fungi and endophytic Methylobacterium affects nutrient uptake and growth of pine seedlings in vitro. Tree Physiol 34:993–1008PubMedGoogle Scholar
  44. Romanowicz KJ, Freedman ZB, Upchurch RA, Argiroff WA, Zak DR (2016) Active microorganisms in forest soils differ from the total community yet are shaped by the same environmental factors: the influence of pH and soil moisture. FEMS Microbiol Ecol 92.
  45. Salvioli A, Chiapello M, Fontaine J, Hadj-Sahraoui AL, Grandmougin-Ferjani A, Lanfranco L, Bonfante P (2010) Endobacteria affect the metabolic profile of their host Gigaspora margarita, an arbuscular mycorrhizal fungus. Environ Microbiol 12:2083–2095PubMedGoogle Scholar
  46. Scholer A, Jacquiod S, Vestergaad G, Schulz S, Schloter M (2017) Analysis of soil microbial communities based on amplicon sequencing of marker genes. Biol Fertil Soils 53:485–489Google Scholar
  47. Schrey SD, Schellhammer M, Ecke M, Hampp R, Tarkka MT (2005) Mycorrhiza helper bacterium Streptomyces AcH 505 induces differential gene expression in the ectomycorrhizal fungus Amanita muscaria. New Phytol 168:205–216PubMedGoogle Scholar
  48. Timonen S, Hurek T (2006) Characterization of culturable bacterial populations associating with Pinus sylvestris-Suillus bovinus mycorrhizospheres. Can J Microbiol 52:769–778PubMedGoogle Scholar
  49. Timonen S, Jorgensen KS, Haahtela K, Sen R (1998) Bacterial community structure at defined locations of Pinus sylvestris Suillus bovinus and Pinus sylvestris Paxillus involutus mycorrhizospheres in dry pine forest humus and nursery peat. Can J Microbiol 44:499–513Google Scholar
  50. Vandenkoornhuyse P, Mahe S, Ineson P, Staddon P, Ostle N, Cliquet JB, Francez AJ, Fitter AH, Young JPW (2007) Active root-inhabiting microbes identified by rapid incorporation of plant-derived carbon into RNA. Proc Natl Acad Sci U S A 104:16970–16975PubMedPubMedCentralGoogle Scholar
  51. Vestergaad G, Schulz S, Scholer A, Schloter M (2017) Making big data smart-how to use metagenomics to understand soil quality. Biol Fertil Soils 53:479–484Google Scholar
  52. Vik U, Logares R, Blaalid R, Halvorsen R, Carlsen T, Bakke I, Kolsto A-B, Okstad OA, Kauserud H (2013) Different bacterial communities in ectomycorrhizae and surrounding soil. Sci Rep 3:3471. CrossRefPubMedPubMedCentralGoogle Scholar
  53. Warmink JA, Nazir R, van Elsas JD (2009) Universal and species-specific bacterial ‘fungiphiles’ in the mycospheres of different basidiomycetous fungi. Environ Microbiol 11:300–312PubMedGoogle Scholar
  54. Zadworny M, McCormack ML, Rawlik K, Jagodzinski AM (2015) Seasonal variation in chemistry, but not morphology, in roots of Quercus robur growing in different soil types. Tree Physiol 35:644–652PubMedGoogle Scholar
  55. Žifčáková L, Větrovský T, Howe A, Baldrian P (2016) Microbial activity in forest soil reflects the changes in ecosystem properties between summer and winter. Environ Microbiol 18:288–301PubMedGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Uppsala Biocenter, Department of Mycology and Plant PathologySwedish University of Agricultural SciencesUppsalaSweden
  2. 2.Microbial Ecology UnitThe Jurinji Buddhist templeKyotoJapan

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