Diversity and community structure of ectomycorrhizal fungi in a wooded meadow Leho TEDERSOO a, * ,y , Triin SUVI a,y , Ellen LARSSON b , Urmas KO ˜ LJALG a a Institute of Botany and Ecology, University of Tartu, 40 Lai Str., 51005 Tartu, Estonia b Botanical Institute, Go ¨teborg University, PO Box 461, SE 40530 Go ¨teborg, Sweden article info Article history: Received 1 September 2005 Received in revised form 13 February 2006 Accepted 16 February 2006 Published online 12 June 2006 Corresponding Editor: John W. G. Cairney Keywords: Deciduous forest Ectomycorrhizal fungal community ITS sequencing Nature conservation Rarefaction Seminatural ecosystems Soil horizons Species richness extrapolation abstract Wooded meadows are seminatural plant communities that support high diversity of various taxa. Due to changes in land use, wooded meadows have severely declined during the last century. The dominant trees in wooded meadows acquire mineral nutrients via ectomycor- rhizal fungi. Using anatomotyping and sequencing of root tips, interpolation and extrapola- tion methods, we studied the diversity and community structure of ectomycorrhizal fungi in two soil horizons of both managed and forested parts of a wooded meadow in Estonia. Species of Thelephoraceae, Sebacinaceae and the genus Inocybe dominated the whole ectomy- corrhizal fungal community of 172 observed species. Forested and managed parts of the wooded meadow harboured different communities of ectomycorrhizal fungi, whereas soil horizon had a negligible effect on the fungal community composition. Diverse soil condi- tions and host trees likely support the high richness of ectomycorrhizal fungi in the wooded meadow ecosystem. Direct sequencing integrated with interpolation and extrapolation methods are promising to identify the fungi at the species level and to compare species rich- ness between communities of ectomycorrhizal fungi. ª 2006 The British Mycological Society. Published by Elsevier Ltd. All rights reserved. Introduction Wooded meadows are seminatural, sparsely wooded ecosys- tems that have developed due to hay-making or sheep grazing in low-productive areas in Europe. A few retained trees have created heterogeneous light conditions and soil nutrient gra- dients. Such patchiness, together with shallow, nutrient- poor soils, support high small-scale species richness of plants in wooded meadows (Kull & Zobel 1991). During the last century, industrialization and urbaniza- tion have strongly affected land use and reduced the importance of traditional farming methods, driving vast countryside areas to abandonment (Vitousek 1994; DeFries 2002). Seminatural meadows and wooded meadows have been most affected among seminatural ecosystems (Kukk & Kull 1997; van Dijk 2002). Wooded meadows covered im- mense areas especially at low-productive coastal and moun- tainous sites throughout Europe. At present more than 99 % of the wooded meadows have been abandoned, and have developed naturally into thickets, bushlands and marsh- lands (Kukk & Kull 1997). Of similar ecosystems, North American oak savannas have declined 5000-fold due to ces- sation of prescribed burning in the last few hundred years (Nuzzo 1986). * Corresponding author. E-mail address: [email protected]. y These authors contributed equally to this work. available at www.sciencedirect.com journal homepage: www.elsevier.com/locate/mycres mycological research 110 (2006) 734–748 0953-7562/$ – see front matter ª 2006 The British Mycological Society. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.mycres.2006.04.007
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m y c o l o g i c a l r e s e a r c h 1 1 0 ( 2 0 0 6 ) 7 3 4 – 7 4 8
Diversity and community structure of ectomycorrhizalfungi in a wooded meadow
Leho TEDERSOOa,*,y, Triin SUVIa,y, Ellen LARSSONb, Urmas KOLJALGa
aInstitute of Botany and Ecology, University of Tartu, 40 Lai Str., 51005 Tartu, EstoniabBotanical Institute, Goteborg University, PO Box 461, SE 40530 Goteborg, Sweden
a r t i c l e i n f o
Article history:
Received 1 September 2005
Received in revised form
13 February 2006
Accepted 16 February 2006
Published online 12 June 2006
Corresponding Editor:
John W. G. Cairney
Keywords:
Deciduous forest
Ectomycorrhizal fungal community
ITS sequencing
Nature conservation
Rarefaction
Seminatural ecosystems
Soil horizons
Species richness extrapolation
a b s t r a c t
Wooded meadows are seminatural plant communities that support high diversity of various
taxa. Due to changes in land use, wooded meadows have severely declined during the last
century. The dominant trees in wooded meadows acquire mineral nutrients via ectomycor-
rhizal fungi. Using anatomotyping and sequencing of root tips, interpolation and extrapola-
tion methods, we studied the diversity and community structure of ectomycorrhizal fungi in
two soil horizons of both managed and forested parts of a wooded meadow in Estonia.
Species of Thelephoraceae, Sebacinaceae and the genus Inocybe dominated the whole ectomy-
corrhizal fungal community of 172 observed species. Forested and managed parts of the
wooded meadow harboured different communities of ectomycorrhizal fungi, whereas soil
horizon had a negligible effect on the fungal community composition. Diverse soil condi-
tions and host trees likely support the high richness of ectomycorrhizal fungi in the wooded
meadow ecosystem. Direct sequencing integrated with interpolation and extrapolation
methods are promising to identify the fungi at the species level and to compare species rich-
ness between communities of ectomycorrhizal fungi.
ª 2006 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.
Introduction
Wooded meadows are seminatural, sparsely wooded ecosys-
tems that have developed due to hay-making or sheep grazing
in low-productive areas in Europe. A few retained trees have
created heterogeneous light conditions and soil nutrient gra-
dients. Such patchiness, together with shallow, nutrient-
poor soils, support high small-scale species richness of plants
in wooded meadows (Kull & Zobel 1991).
During the last century, industrialization and urbaniza-
tion have strongly affected land use and reduced the
importance of traditional farming methods, driving vast
countryside areas to abandonment (Vitousek 1994; DeFries
2002). Seminatural meadows and wooded meadows have
been most affected among seminatural ecosystems (Kukk
& Kull 1997; van Dijk 2002). Wooded meadows covered im-
mense areas especially at low-productive coastal and moun-
tainous sites throughout Europe. At present more than 99 %
of the wooded meadows have been abandoned, and have
developed naturally into thickets, bushlands and marsh-
lands (Kukk & Kull 1997). Of similar ecosystems, North
American oak savannas have declined 5000-fold due to ces-
sation of prescribed burning in the last few hundred years
y These authors contributed equally to this work.0953-7562/$ – see front matter ª 2006 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.doi:10.1016/j.mycres.2006.04.007
The genus Inocybe comprised 16 species on root tips. Inocybe
was among the most species rich genera in a Corsican decid-
uous forest (Richard et al. 2005), and in Oregon and Californian
coniferous forests [Cullings et al. 2001; Izzo et al. 2005 (se-
quences reanalysed by us using fasta3 queries and compared
with our unpublished fruit body sequences)]. We believe
that high variation in EcM anatomy and ITS sequences, low
alignment power of blastn algorithm, and the lack of public
sequences have generally hampered identification of Inocybe
spp. In this study, Inocybe spp. appeared more common in
the A-horizon, especially in forest pits. This may be related
to a high tolerance to elevated ground water and seasonal
flooding or preference for soil mineral particles. Inocybe spp.
most often fruit on organic-poor and damp soils (L. Tedersoo,
pers. obs.).
In contrast to the dominant genera, nearly all Russula and
Lactarius spp. were identified to species. Both R. velenovskyi
and R. sphagnophila were sequenced from root tips represent-
ing typical beige russuloid and dark brown thelephoroid ana-
tomotypes. This finding indicates either specific relationships
between fungi, unnoticed double colonization, or DNA han-
dling mistakes.
Boletus radicans, a locally red-listed species, both fruited
and formed EcM only in well-managed parts of the wooded
meadow. In addition, B. luridus occurred in abundance both
above and below ground exclusively in the wooded meadow.
These results corroborate with several observations that Bole-
tus spp. fruit predominantly in the managed wooded
meadows (Kalamees 2004). Calciphilous Boletus spp. may pre-
fer wooded meadows because of more alkaline soils or more
abundant sunlight as Estonia is the northernmost habitat for
several Boletus spp. We anticipate that the fungi we found
only in the wooded meadow are also likely to inhabit the for-
est soil, but below our detection limits.
Conclusions and future considerations
Integrating anatomotyping and sequencing enabled us to dis-
tinguish EcM fungi from saprobes and endophytic fungi and
provided species level identification for many EcM fungi on
root tips. The results show that traditionally managed areas
differ most strongly from forested areas, with no apparent de-
pendence on soil variables or soil horizon. However, repli-
cated wooded meadows are needed to reveal the
characteristic EcM fungi and to address the effects of manage-
ment per se on changes in community structure and diversity.
Rarefaction and extrapolation offer promising alternatives to
species richness comparisons between sites and treatments,
but high sampling effort is unavoidable. To compare the fun-
gal diversity and community composition between studies,
considering the presence of cryptic species, molecular species
criteria need to be established together with appropriate soft-
ware (Schloss & Handelsman 2005). Due to differences in the
rate of evolution in the ITS region, molecular taxonomists
should develop molecular species criteria separately for each
genus.
Acknowledgements
We thank John W. G. Cairney and referees for comments. We
thank Saaremaa Keskkonnateenistus for permission to sam-
ple in Tagamoisa wooded meadow; Teele Jairus, Sergei Polme,
Marko Peterson, Eele Ounapuu, Erki Laaneoks and Helena
Faust for assistance during root sampling; Tiina Ojala and
Mari Reitalu for providing information on history of the study
site; Martin Ryberg and R. Henrik Nilsson for commenting on
an earlier draft of the manuscript. This study was funded by
ESF grants nos 5232 and 6606 and WFS.
r e f e r e n c e s
Agerer R, 1991. Characterization of ectomycorrhiza. In: Norris JR,Read DJ, Varma AK (eds), Techniques for the Study of Mycorrhiza.Academic Press, London, pp. 25–73.
Agerer R, Gottlein A, 2003. Correlations between projection areaof ectomycorrhizae and H2O extractable nutrients in organicsoil layers. Mycological Progress 2: 45–52.
AOAC, 1990. Official Methods of Analysis, 15th edn. AOAC,Kensington.
Avis PG, McLaughlin DJ, Dentinger BC, Reich PB, 2003. Long-termincrease in nitrogen supply alters above- and below-groundectomycorrhizal communities and increases the dominanceof Russula spp. in a temperate oak savanna. New Phytologist160: 239–253.
Bruns TD, 1995. Thoughts on the processes that maintain localspecies diversity of ectomycorrhizal fungi. Plant and Soil 170:63–73.
Burnham KP, Overton WS, 1979. Robust estimation of populationsize when capture probabilities vary among animals. Ecology60: 927–936.
Butler MJ, Day AW, 1998. Fungal melanins: a review. CanadianJournal of Microbiology 44: 1115–1136.
Chao A, 1987. Estimating the population size for capture–recap-ture data with unequal catchability. Biometrics 43: 783–791.
Cline ET, Ammirati JF, Edmonds RL, 2005. Does proximity tomature trees influence ectomycorrhizal fungus communitiesof Douglas-fir seedlings? New Phytologist 166: 993–1009.
Colwell RK, 2004. EstimateS: statistical estimate of species richnessand shared species from samples. Version 7. Persistent URL:purl.oclc.org/estimates
Colwell RK, Coddington JA, 1994. Estimating terrestrial biodiver-sity through extrapolation. Philosophical Transactions of theRoyal Society of London, Series B 345: 101–118.
Colwell RK, Mao CX, Chang J, 2004. Interpolating, extrapolating,and comparing incidence-based species accumulation curves.Ecology 85: 2717–2727.
Courty P-E, Pritsch K, Schloter M, Hartmann A, Garbaye J, 2005.Activity profiling of ectomycorrhiza communities in two forestsoils using multiple enzymatic tests. New Phytologist 167: 309–319.
Cullings KW, Vogler DR, Parker TV, Makhija S, 2001. Defoliationeffects on the ectomycorrhizal community of a mixed Pinuscontorta/Picea engelmannii stand in Yellowstone Park. Oecologia127: 533–539.
Dahlberg A, Jonsson L, Nylund J-E, 1997. Species diversity anddistribution of biomass above and below ground amongectomycorrhizal fungi in an old-growth Norway spruce forestin South Sweden. Canadian Journal of Botany 75: 1323–1335.
Dahlgren RA, Singer MJ, Huang X, 1997. Oak tree and grazingimpacts on soil properties and nutrients in a California oakwoodland. Biogeochemistry 39: 45–64.
DeFries R, 2002. Past and future sensitivity of primary productionto human modification of the landscape. Geophysics ResearchLetters 29: 36–39.
Dickie IA, Xu B, Koide RT, 2002. Vertical niche differentiation ofectomycorrhizal hyphae in soil as shown by T-RFLP analysis.New Phytologist 156: 527–535.
Edwards IP, Turco RF, 2005. Inter- and intraspecific resolution ofnrDNA TRFLP assessed by computer-simulated restrictionanalysis of a diverse collection of ectomycorrhizal fungi.Mycological Research 109: 213–226.
FAO, ISRIC, ISSS, 1998. World Reference Base for Soil Resources. FAO,Rome.
Gardes M, Bruns TD, 1993. ITS primers with enhancedspecificity for basidiomycetesdapplication to the identifi-cation of mycorrhizas and rusts. Molecular Ecology 2: 113–118.
Gotelli NJ, Colwell RK, 2001. Quantifying biodiversity: proceduresand pitfalls in the measurement and comparison of speciesrichness. Ecology Letters 4: 379–391.
Hibbett DS, 2001. Shiitake mushrooms and molecular clocks:historical biogeography of Lentinula. Journal of Biogeography 28:231–241.
Horton TR, Bruns TD, 2001. The molecular evolution in ectomy-corrhizal ecology: peeking into the black box. Molecular Ecology10: 1855–1871.
Izzo A, Agbowo J, Bruns TD, 2005. Detection of plot level changesin ectomycorrhizal communities across years in an old-growth mixed-conifer forest. New Phytologist 166: 619–629.
Kalamees K, 1979. The fungal cover and its seasonal dynamics ofthe Estonian meadows. Yearbook of The Estonian NaturalistsSociety 67: 39–54.
Kalamees K, 2004. Seenestik. In: Kukk T (ed), Parandkooslused.Parandkoosluste Kaitse Uhing, Tartu, pp. 136–142.
Kaldorf M, Renker C, Fladung M, Buscot F, 2004. Characterizationand spatial distribution of ectomycorrhizas colonizing aspenclones released in an experimental field. Mycorrhiza 14: 295–306.
Karen O, Hogberg N, Dahlberg A, Jonsson L, Nylund J-E, 1997.Inter- and intraspecific variation of the ITS region of rDNA ofectomycorrhizal fungi in Fennoscandia as detected by endo-nuclease analysis. New Phytologist 136: 313–325.
Keating KA, Quinn JF, Ivie MA, Ivie LL, 1998. Estimating the ef-fectiveness of further sampling in species inventories. Ecolog-ical Applications 8: 1239–1249.
Koljalg U, Dahlberg A, Taylor AFS, Larsson E, Hallenberg N,Stenlid J, Larsson K-H, Fransson PM, Karen O, Jonsson L, 2000.Diversity and abundance of resupinate thelephoroid fungi asectomycorrhizal symbionts in Swedish boreal forests. Molec-ular Ecology 9: 1985–1996.
Koljalg U, Larsson K-H, Abarenkov K, Nilsson RH, Alexander IJ,Eberhardt U, Erland S, Høiland K, Kjøller R, Larsson E,Pennanen T, Sen R, Taylor AFS, Tedersoo L, Vralstad T,Ursing BM, 2005. UNITE: a database providing web-basedmethods for the molecular identification of ectomycorrhizalfungi. New Phytologist 166: 1063–1068. http://unite.zbi.ee/.
Kukk T, Kull K, 1997. Puisniidud. Estonia Maritima 2: 1–249.Kull K, Zobel M, 1991. High species richness in an Estonian
wooded meadow. Journal of Vegetation Science 2: 711–714.Larsson K-H, Larsson E, Koljalg U, 2004. High phylogenetic
diversity among corticioid homobasidiomycetes. MycologicalResearch 108: 983–1002.
Lilleskov EA, Bruns TD, Horton TR, Taylor DL, Grogan P, 2004.Detection of forest stand-level spatial structure in ectomy-corrhizal fungal communities. FEMS Microbiology Ecology 49:319–332.
Lilleskov EA, Fahey TJ, Horton TR, Lovett GM, 2002. Belowgroundectomycorrhizal community change over a nitrogen deposi-tion gradient in Alaska. Ecology 83: 104–115.
Mao CX, Colwell RK, 2005. Estimation of species richness: mixturemodels, the role of rare species, and inferential challenges.Ecology 86: 1143–1153.
Mason PA, Last FT, Pelham J, Ingleby K, 1982. Ecology of somefungi associated with an ageing stand of birches (Betula pen-dula and B. pubescens). Forest Ecology and Management 4:19–39.
Melo AS, Froelich CG, 2001. Evaluation of methods for estimatingmacroinvertebrate species richness using individual stones intropical streams. Freshwater Biology 46: 711–721.
Murat C, Vizzini A, Bonfante P, Mello A, 2005. Morphological andmolecular typing of the below-ground fungal community innatural Tuber magnatum truffle-ground. FEMS MicrobiologyLetters 245: 307–313.
Nuzzo V, 1986. Extent and status of Midwest oak savanna: pre-settlement and 1985. Natural Areas Journal 6: 6–36.
Page AL, Miller RH, Keeney DR, 1982. Methods of Soil Analysis. II.Chemical and Microbiological Properties, 2nd edn. AmericanSociety of Agronomy, Madison.
Reich PB, Peterson DW, Wedin DA, Wrage K, 2001. Fire and veg-etation effects on productivity and nitrogen cycling acrossa forest–grassland continuum. Ecology 82: 1703–1719.
Richard F, Millot S, Gardes M, Selosse M-A, 2005. Diversity andspecificity of ectomycorrhizal fungi retrieved from an old-growth Mediterranean forest dominated by Quercus ilex. NewPhytologist 166: 1011–1023.
Rosling A, Landeweert R, Lindahl BD, Larsson K-H, Kuyper TW,Taylor AFS, Finlay RD, 2003. Vertical distribution of ectomy-corrhizal fungi in a podzol profile. New Phytologist 159: 775–783.
Schloss PD, Handelsman J, 2005. Introducing DOTUR, a computerprogram for defining operational taxonomic units and esti-mating species richness. Applied and Environmental Microbiology71: 1501–1506.
Selosse M-A, Weiß M, Jany J-L, Tillier A, 2002. Communities andpopulations of sebacinoid basidiomycetes associated with theachlorophyllous orchid Neottia nidus-avis and neighbouringtree ectomycorrhizae. Molecular Ecology 11: 1831–1844.
Stampfli A, Zeiter M, 1999. Plant species decline due to aban-donment of meadows cannot easily be reversed by mowing. Acase study from the southern Alps. Journal of Vegetation Science10: 151–164.
Swofford DL, 2002. PAUP*. Phylogenetic Analysis Using Parsimony(*and Other Methods). Version 4. Sinauer Associates, Sunderland.
Taylor AFS, 2002. Fungal diversity in ectomycorrhizal communi-ties: sampling effort and species detection. Plant and Soil 244:19–28.
Tedersoo L, Koljalg U, Hallenberg N, Larsson K-H, 2003. Fine scaledistribution of ectomycorrhizal fungi and roots across sub-strate layers including coarse woody debris in a mixed forest.New Phytologist 159: 153–165.
Tedersoo L, Suvi T, Larsson E, Koljalg U, 2006. Molecular andmorphological diversity of pezizalean ectomycorrhiza. NewPhytologist, 170: 581–596.
van der Heijden EW, Kuyper TW, 2003. Ecological strategies ofectomycorrhizal fungi of Salix repens: root manipulationversus root replacement. Oikos 103: 668–680.
van Dijk, G, 2002. Biodiversity indicators in agriculture: a combi-nation of species and habitat approaches. Proceedings of theOECD Expert Meeting on Agri-Biodiversity Indicators. OECD,Zurich
Vitousek PM, 1994. Beyond global warming: ecology and globalchange. Ecology 75: 1861–1876.
Waldrop MP, Firestone MK, 2004. Microbial community utilizationof recalcitrant and simple carbon compounds: impact of oak-woodland plant communities. Oecologia 138: 275–284.
Walker JF, Miller OK, Horton JL, 2005. Hyperdiversity of ectomy-corrhizal fungus assemblages on oak seedlings in mixed for-ests in the Southern Appalachian Mountains. Molecular Ecology14: 829–838.
Watling R, 2005. Fungal conservation: some impressionsdapersonal view. Exotic species and fungi: interactions withfungal, plant, and animal communities. In: Dighton J, White JF,Oudemans P (eds), The Fungal Community. Its Organization
and Role in the Ecosystem. CRC Press, Boca Ranton, pp.881–896.
Weiß M, Selosse M-A, Rexer K-H, Urban A, Oberwinkler F, 2004.Sebacinales: a hitherto overlooked cosm of heterobasidiomy-cetes with a broad mycorrhizal potential. Mycological Research108: 1003–1010.
Wilson SD, 1993. Belowground competition in forest and prairie.Oikos 68: 146–150.
Unpublished supplementary information (lost during the revising process)
Table 1. The effect of wooded meadow management and soil horizon on diversity and frequency ofectomycorrhizal fungi. Values represent means ± SE. Different letters indicate significant differences (based onBenjamini and Hochberg´s sharpening modification to Bonferroni correction; Benjamini & Hochberg, 2000 asimplemented in a spreadsheet program of Verhoeven et al. (2005)) for both factors separately based on three-way mixed ANOVAs. Neither block effects nor interactions were significant. Species with melanized cell walls(see table 2) were distinguished microscopically; species with hypogeous fruit bodies were distinguished basedon sequence similarity and phylogenetic analyses (table 2).
Wooded meadow management Soil horizon
Diversity variables / taxa Managed Unmanaged O-horizon A-horizonThe number of observedspecies per plot 14.3 ± 0.9 16.4 ± 0.9 16.9 ± 1.0x 13.8 ± 0.7y
The number of observedspecies per sample 4.63 ± 0.36 5.30 ± 0.28 5.65 ± 0.32x 4.27 ± 0.23y
The number of observedspecies per root fragment 1.50 ± 0.07a 1.83 ± 0.07b 1.82 ± 0.07x 1.50 ± 0.07y
Table 2. The effect of wooded meadow management and soil horizon on soil variables. Values represent means± SE. Different letters indicate significant differences (based on Benjamini and Hochberg´s sharpeningmodification to Bonferroni correction; Benjamini & Hochberg, 2000 as implemented in a spreadsheet programof Verhoeven et al. (2005)) for both factors separately based on three-way mixed ANOVAs. Neither blockeffects nor interactions were significant.
Benjamini Y, Hochberg Y. 2000. On the adaptive control of the false discovery rate in multiple testing withindependent statistics. Journal of Educational and Behavioral Statistics 25: 60-83.