Arbuscular mycorrhizal fungi (AMF) and dark septate endophytes (DSE) form symbiotic relationships with plants influencing their productivity, diversity and ecosystem functions. Only a few studies on these fungi, however, have been conducted in extreme elevations and none over 5500 m a.s.l., although vascular plants occur up to 6150 m a.s.l. in the Himalayas. We quantified AMF and DSE in roots of 62 plant species from contrasting habitats along an elevational gradient (3400-6150 m) in the Himalayas using a combination of optical microscopy and next generation sequencing. We linked AMF and DSE communities with host plant evolutionary history, ecological preferences (elevation and habitat type) and functional traits. We detected AMF in elevations up to 5800 m, indicating it is more constrained by extreme conditions than the host plants, which ascend up to 6150 m. In contrast, DSE were found across the entire gradient up to 6150 m. AMF diversity was unimodally related to elevation and positively related to the intensity of AMF colonization. Mid-elevation steppe and alpine plants hosted more diverse AMF communities than plants from deserts and the subnival zone. Our results bring novel insights to the abiotic and biotic filters structuring AMF and DSE communities in the Himalayas.

Department of Forest Ecology Faculty of Forestry and Wood Sciences Czech University of Life Sciences Prague Kamýcká 129 Prague 6 Suchdol CZ 165 21 Czech Republic

Faculty of Science University of South Bohemia Branišovská 1760 370 05 České Budějovice Czech Republic

Institute of Botany The Czech Academy of Sciences Zámek 1 252 43 Průhonice Czech Republic

Institute of Low Temperature Science Hokkaido University Kita 19 Nishi 8 Kita ku Sapporo 060 0819 Japan

Zobrazit více v PubMed

Spatafora JW, et al. A phylum-level phylogenetic classification of zygomycete fungi based on genome-scale data. Mycologia. 2016;108:1028–1046. doi: 10.3852/16-042. PubMed DOI PMC

Smith, S. E. & Read, D. J. Mycorrhizal Symbiosis. Soil Science Society of America Journal137 (2008).

Rillig MC. Arbuscular mycorrhizae, glomalin, and soil aggregation. Can. J. soil Sci. 2004;84:355–363. doi: 10.4141/S04-003. DOI

van der Heijden MGa, Boller T, Wiemken A, Sanders IR. Different arbuscular mycorrhizal fungal species are potential determinants of plant community structure. Ecology. 1998;79:2082–2091. doi: 10.1890/0012-9658(1998)079[2082:DAMFSA]2.0.CO;2. DOI

Hiiesalu I, et al. Species richness of arbuscular mycorrhizal fungi: Associations with grassland plant richness and biomass. New Phytol. 2014;203:233–244. doi: 10.1111/nph.12765. PubMed DOI

Davison J, et al. Global assessment of arbuscular mycorrhizal fungus diversity reveals very low endemism. Science (80-.). 2015;349:970–973. doi: 10.1126/science.aab1161. PubMed DOI

Doležal J, et al. Vegetation dynamics at the upper elevational limit of vascular plants in Himalaya. Sci. Rep. 2016;6:24881. doi: 10.1038/srep24881. PubMed DOI PMC

Oehl F, Körner C. Multiple mycorrhization at the coldest place known for Angiosperm plant life. Alp. Bot. 2014;124:193–198. doi: 10.1007/s00035-014-0138-7. DOI

Ruotsalainen AL, Väre H, Vestberg M. Seasonality of root fungal colonization in low-alpine herbs. Mycorrhiza. 2002;12:29–36. doi: 10.1007/s00572-001-0145-6. PubMed DOI

Schmidt SK, Sobieniak-Wiseman LC, Kageyama Sa, Halloy SRP, Schadt CW. Mycorrhizal and Dark-Septate Fungi in Plant Roots Above 4270 Meters Elevation in the Andes and Rocky Mountains. Arctic, Antarct. Alp. Res. 2008;40:576–583. doi: 10.1657/1523-0430(07-068)[SCHMIDT]2.0.CO;2. DOI

Gai JP, et al. Occurrence and distribution of arbuscular mycorrhizal fungal species in three types of grassland community of the Tibetan Plateau. Ecol. Res. 2009;24:1345–1350. doi: 10.1007/s11284-009-0618-1. DOI

Pan J, et al. Arbuscular mycorrhizal and dark septate endophytic fungi at 5,500 m on a glacier forefront in the Qinghai-Tibet Plateau, China. Symbiosis. 2013;60:101–105. doi: 10.1007/s13199-013-0245-z. DOI

Upson R, Newsham KK, Read D. Root-fungal associations of Colobanthus quitensis and Deschampsia antarctica in the maritime and subantarctic. Arctic, Antarct. Alp. Res. 2008;40:592–599. doi: 10.1657/1523-0430(07-057)[UPSON]2.0.CO;2. DOI

Casanova-Katny MA, Torres-Mellado GA, Palfner G. & Cavieres, L. a. The best for the guest: High Andean nurse cushions of Azorella madreporica enhance arbuscular mycorrhizal status in associated plant species. Mycorrhiza. 2011;21:613–622. doi: 10.1007/s00572-011-0367-1. PubMed DOI

Urcelay C, Acho J, Joffre R. Fungal root symbionts and their relationship with fine root proportion in native plants from the Bolivian Andean highlands above 3,700 m elevation. Mycorrhiza. 2011;21:323–330. doi: 10.1007/s00572-010-0339-x. PubMed DOI

Ruotsalainen, A. A. L., Väre, H., Oksanen, J. & Tuomi, J. Root Fungus Colonization along an Altitudinal Gradient in North Norway Root Fungus Colonization along an Altitudinal Gradient in North Norway. 36, 239–243 (2004).

Haselwandter K, Read DJ. The significance of a root fungus association in 2 Carex species of high alpine plant communities. Oecologia. 1982;53:352–354. doi: 10.1007/BF00389012. PubMed DOI

Wu L, Guo S. Interaction between an isolate of dark-septate fungi and its host plant Saussurea involucrata. Mycorrhiza. 2008;18:79–85. doi: 10.1007/s00572-007-0159-9. PubMed DOI

Newsham KK. A meta-analysis of plant responses to dark septate root endophytes. New Phytol. 2011;190:783–793. doi: 10.1111/j.1469-8137.2010.03611.x. PubMed DOI

Jumpponen A. Dark septate endophytes - Are they mycorrhizal? Mycorrhiza. 2001;11:207–211. doi: 10.1007/s005720100112. DOI

Öpik M, et al. The online database MaarjAM reveals global and ecosystemic distribution patterns in arbuscular mycorrhizal fungi (Glomeromycota) New Phytol. 2010;188:223–241. doi: 10.1111/j.1469-8137.2010.03334.x. PubMed DOI

Kessler M, Güdel R, Salazar L, Homeier J, Kluge J. Impact of mycorrhization on the abundance, growth and leaf nutrient status of ferns along a tropical elevational gradient. Oecologia. 2014;175:887–900. doi: 10.1007/s00442-014-2941-7. PubMed DOI

Gai JP, et al. Arbuscular mycorrhizal fungal diversity along a Tibetan elevation gradient. Pedobiologia (Jena). 2012;55:145–151. doi: 10.1016/j.pedobi.2011.12.004. DOI

Angel R, et al. The root-associated microbial community of the world’s highest growing vascular plants. Microb. Ecol. 2016;72:394–406. doi: 10.1007/s00248-016-0779-8. PubMed DOI PMC

Dvorský M, et al. Vascular plants at extreme elevations in eastern Ladakh, northwest Himalayas. Plant Ecol. Divers. 2015;8:571–584. doi: 10.1080/17550874.2015.1018980. DOI

Dvorský M, et al. Gardening in the zone of death: an experimental assessment of the absolute elevation limit of vascular plants. Sci. Rep. 2016;6:24440. doi: 10.1038/srep24440. PubMed DOI PMC

Liu Y, et al. Diverse communities of arbuscular mycorrhizal fungi inhabit sites with very high altitude in Tibet Plateau. FEMS Microbiol. Ecol. 2011;78:355–365. doi: 10.1111/j.1574-6941.2011.01163.x. PubMed DOI

Sundqvist MK, Liu Z, Giesler R, Wardle DA. Plant and microbial responses to nitrogen and phosphorus addition across an elevational gradient in subarctic tundra. Ecology. 2014;95:1819–1835. doi: 10.1890/13-0869.1. PubMed DOI

Mandyam K, Jumpponen A. Seeking the elusive function of the root-colonising dark septate endophytic fungi. Stud. Mycol. 2005;53:173–189. doi: 10.3114/sim.53.1.173. DOI

Newsham KK, Upson R, Read DJ. Mycorrhizas and dark septate root endophytes in polar regions. Fungal Ecol. 2009;2:10–20. doi: 10.1016/j.funeco.2008.10.005. PubMed DOI

Crush JR. Signifi cance of endomycorrhizas in tussock grassland in Otago, New Zealand. New Zeal. J. Bot. 1973;11:645–660. doi: 10.1080/0028825X.1973.10430306. DOI

Bledsoe C, Klein P, Bliss L. A survey of mycorrhizal plants on Truelove lowland, Devon island, NWT, Canada. Can. J. Bot. 1990;68:1848–1856. doi: 10.1139/b90-242. DOI

Pfeffer P, Douds D, Becard G, Shachar-Hill Y. Carbon Uptake and the Metabolism and Transport of Lipids in an Arbuscular Mycorrhiza. Plant Physiol. 1999;120:587–598. doi: 10.1104/pp.120.2.587. PubMed DOI PMC

Bücking H, Shachar-Hill Y. Phosphate uptake, transport and transfer by the arbuscular mycorrhizal fungus Glomus intraradices is stimulated by increased carbohydrate availability. New Phytol. 2005;165:899–912. doi: 10.1111/j.1469-8137.2004.01274.x. PubMed DOI

Kiers ET, et al. Reciprocal Rewards Stabilize Cooperation in the Mycorrhizal Symbiosis. Science (80-.). 2011;333:880–882. doi: 10.1126/science.1208473. PubMed DOI

Ackerly DDTS. Correlated Evolution, and Independent Contrasts. Evolution (N. Y). 2000;54:1480–1492. PubMed

Desdevises Y, Legendre P, Azouzi L, Morand S. Quantifying phylogenetically structured environmental variation. Evolution (N. Y). 2003;57:2647–2652. PubMed

Zobel M, Öpik M. Plant and arbuscular mycorrhizal fungal (AMF) communities - which driveswhich? J. Veg. Sci. 2014;25:1133–1140. doi: 10.1111/jvs.12191. DOI

Dvorský M, Doležal J, De Bello F, Klimešová J, Klimeš L. Vegetation types of East Ladakh: Species and growth form composition along main environmental gradients. Appl. Veg. Sci. 2011;14:132–147. doi: 10.1111/j.1654-109X.2010.01103.x. DOI

Chaurasia B, Pandey A, Palni LMS. Distribution, colonization and diversity of arbuscular mycorrhizal fungi associated with central Himalayan rhododendrons. For. Ecol. Manage. 2005;207:315–324. doi: 10.1016/j.foreco.2004.10.014. DOI

Mundra S, et al. Arctic fungal communities associated with roots of Bistorta vivipara do not respond to the same fine-scale edaphic gradients as the aboveground vegetation. New Phytol. 2015;205:1587–1597. doi: 10.1111/nph.13216. PubMed DOI

Körner, C. Alpine Plant Life - Functional Plant Ecology of High Mountain Ecosystems. (Springer, 2003).

Cázares E, Trappe JM, Jumpponen A. Mycorrhiza-plant colonization patterns on a subalpine glacier forefront as a model system of primary succession. Mycorrhiza. 2005;15:405–416. doi: 10.1007/s00572-004-0342-1. PubMed DOI

Oehl F, Schneider D, Sieverding E, Burga Ca. Succession of arbuscular mycorrhizal communities in the foreland of the retreating Morteratsch glacier in the Central Alps. Pedobiologia (Jena). 2011;54:321–331. doi: 10.1016/j.pedobi.2011.07.006. DOI

Liu Y, et al. Direct and indirect influences of 8 yr of nitrogen and phosphorus fertilization on Glomeromycota in an alpine meadow ecosystem. New Phytol. 2012;194:523–535. doi: 10.1111/j.1469-8137.2012.04050.x. PubMed DOI

Schüßler A, Schwarzott D, Walker C. A new fungal phylum, the Glomeromycota: phylogeny and evolution. Mycol. Res. 2001;105:1413–1421. doi: 10.1017/S0953756201005196. DOI

Li X, et al. Contribution of arbuscular mycorrhizal fungi of sedges to soil aggregation along an altitudinal alpine grassland gradient on the Tibetan Plateau. Environ. Microbiol. 2015;17:2841–2857. doi: 10.1111/1462-2920.12792. PubMed DOI

Jumpponen A. Spatial distribution of discrete RAPD phenotypes of a root endophytic fungus Phialocephala fortinii, at a primary successional site on a glacier forefront. New Phytol. 1999;141:333–344. doi: 10.1046/j.1469-8137.1999.00344.x. PubMed DOI

Gardes M, Bruns TD. ITS primers with enhanced specificity for basidiomycetes, application to the identification of mycorrihiza and rusts. Mol. Ecol. 1993;2:113–118. doi: 10.1111/j.1365-294X.1993.tb00005.x. PubMed DOI

Dvorský, M., Macek, M., Kopecký, M., Wild, J. & Doležal, J. Niche asymmetry of vascular plants increases with elevation. J. Biogeogr. doi:10.1111/jbi.13001 (2017).

Karger, D. N. et al. Climatologies at high resolution for the earth’s land surface areas (2016). PubMed PMC

Janatková K, et al. Community structure of soil phototrophs along environmental gradients in arid Himalaya. Environ. Microbiol. 2013;15:2505–2516. doi: 10.1111/1462-2920.12132. PubMed DOI

Řeháková K, Chlumská Z, Doležal J. Soil cyanobacterial and microalgal diversity in dry mountains of Ladakh, NW Himalaya, as related to site, altitude, and vegetation. Microb. Ecol. 2011;62:337–346. doi: 10.1007/s00248-011-9878-8. PubMed DOI

Vierheilig H, Schweiger P, Brundrett M. An overview of methods for the detection and observation of arbuscular mycorrhizal fungi in roots. Physiol. Plant. 2005;125:393–404.

Trouvelot, A., Kough, J. & Gianinazzi-Pearson, V. In Physiological and Genetical Aspects of Mycorrhizae (eds Gianinazzi-Pearson, V. & Gianinazzi, S.) 217–221 (INRA Press, 1986).

Jumpponen A, Trappe JM. Dark septate endophytes: a review of facultative biotrophic root-colonizing fungi. New Phytol. 1998;140:295–310. doi: 10.1046/j.1469-8137.1998.00265.x. PubMed DOI

Doyle JJ, Doyle JL. A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochem. Bull. 1987;19:11–15.

Simon L, Lalonde M, Bruns TD. Specific amplification of 18s fungal ribosomal genes from vesicular–arbuscular endomycorrhizal fungi colonizing roots. Appl. Environ. Microbiol. 1992;58:291–295. PubMed PMC

Helgason T, Daniell TJ, Husband R, Fitter AH, Young JPW. Ploughing up the wood-wide web? Nature. 1998;394:431. doi: 10.1038/28764. PubMed DOI

Dumbrell AJ, et al. Distinct seasonal assemblages of arbuscular mycorrhizal fungi revealed by massively parallel pyrosequencing. New Phytol. 2011;190:794–804. doi: 10.1111/j.1469-8137.2010.03636.x. PubMed DOI

Větrovský T, Baldrian P. Analysis of soil fungal communities by amplicon pyrosequencing: Current approaches to data analysis and the introduction of the pipeline SEED. Biol. Fertil. Soils. 2013;49:1027–1037. doi: 10.1007/s00374-013-0801-y. DOI

Schloss PD. Introducing mothur: Open-source, platform-independent, community-supported software for describing and comparing microbial communities. Appl. Environ. Microbiol. 2009;75:7537–7541. doi: 10.1128/AEM.01541-09. PubMed DOI PMC

Edgar RC, Haas BJ, Clemente JC, Quince C, Knight R. UCHIME improves sensitivity and speed of chimera detection. Bioinformatics. 2011;27:2194–2200. doi: 10.1093/bioinformatics/btr381. PubMed DOI PMC

Edgar RC. Search and clustering orders of magnitude faster than BLAST. Bioinformatics. 2010;26:2460–2461. doi: 10.1093/bioinformatics/btq461. PubMed DOI

Katoh K, Toh H. Recent developments in the MAFFT multiple sequence alignment program. Brief. Bioinform. 2008;9:286–298. doi: 10.1093/bib/bbn013. PubMed DOI

Yang Z. A space-time process model for the evolution of DNA sequences. Genetics. 1995;139:993–1005. PubMed PMC

Tanabe AS. Kakusan4 and Aminosan: two programs for comparing nonpartitioned, proportional and separate models for combined molecular phylogenetic analyses of multilocus sequence data. Mol. Ecol. Resour. 2011;11:914–921. doi: 10.1111/j.1755-0998.2011.03021.x. PubMed DOI

Huelsenbeck JP, Ronquist F. MRBAYES: Bayesian inference of phylogenetic trees. Bioinformatics. 2001;17:754–755. doi: 10.1093/bioinformatics/17.8.754. PubMed DOI

Klimeš, L. & Dickoré, W. B. Flora of Ladakh (Jammu & Kashmir, India). A preliminary checklist published at http://www.butbn.cas.cz/klimes. Accessed 4 October 2010 (2006).

Farquhar GD, Ball MC, Voncaemmerer S, Roksandic Z. Effect of salinity and humidity on delta-C-13 value of halophytes - evidence for diffusional isotope fractionation determined by the ratio of inter-cellular atmospheric partial pressure of CO2 uncer different environmental conditions. Oecologia. 1982;52:121–124. doi: 10.1007/BF00349020. PubMed DOI

Chlumska Z, Janecek S, Dolezal J. How to Preserve Plant Samples for Carbohydrate Analysis? Test of Suitable Methods Applicable in Remote Areas. Folia Geobot. 2013;49:1–15. doi: 10.1007/s12224-013-9153-5. DOI

Legendre, P. & Legendre, L. Numerical Ecology (Elsevier Science, 1998).

Diniz-Filho JAF, de Sant Ana CER, Bini LM. An eigenvector method for estimating phylogenetic inertia. Evolution (N. Y). 1998;52:1247–1262. PubMed

ter Braak, C. J. F. & Šmilauer, P. Canoco reference manual and users’s guide: sofware for ordination (version 5.0). (Microcomputer Power, 2012).

R Core Team. R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. https://www.R-project.org/ (2015).

Paradis E, Claude J, Strimmer K. APE: analyses of phylogenetics and evolution in R language. Bioinformatics. 2004;20:289–290. doi: 10.1093/bioinformatics/btg412. PubMed DOI

De Caceres, M. & Jansen, F. INDICSPECIES: Relationship Between Species and Groups of Sites. R package version 1.7.5. https://cran.r-project.org/web/packages/indicspecies/indicspecies.pdf (2015).

Venables, W. N. & Ripley, B. D. Modern Applied Statistics with S. (Springer-Verlag, 2002).