Skip to main content
SpringerLink
Log in

Appropriate nonmycorrhizal controls in arbuscular mycorrhiza research: a microbiome perspective

  • Original Article
  • Published:
Mycorrhiza Aims and scope Submit manuscript

Abstract

Establishment of nonmycorrhizal controls is a “classic and recurrent theme” in mycorrhizal research. For decades, authors reported mycorrhizal plant growth/nutrition as compared to various nonmycorrhizal controls. In such studies, uncertainties remain about which nonmycorrhizal controls are most appropriate and, in particular, what effects the control inoculations have on substrate and root microbiomes. Here, different types of control and mycorrhizal inoculations were compared with respect to plant growth and nutrition, as well as the structure of root and substrate microbiomes, assessed by next-generation sequencing. We compared uninoculated (“absolute”) control to inoculation with blank pot culture lacking arbuscular mycorrhizal fungi, filtrate of that blank inoculum, and filtrate of complex pot-produced mycorrhizal inoculum. Those treatments were compared to a standard mycorrhizal treatment, where the previously sterilized substrate was inoculated with complex pot-produced inoculum containing Rhizophagus irregularis SYM5. Besides this, monoxenically produced inoculum of the same fungus was applied either alone or in combination with blank inoculum. The results indicate that the presence of mycorrhizal fungus always resulted in stimulation of Andropogon gerardii plant biomass as well as in elevated phosphorus content of the plants. The microbial (bacterial and fungal) communities developing in the differently inoculated treatments, however, differed substantially from each other and no control could be obtained comparable with the treatment inoculated with complex mycorrhizal inoculum. Soil microorganisms with significant biological competences that could potentially contribute to the effects of the various inoculants on the plants were detected in roots and in plant cultivation substrate in some of the treatments.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

References

  • Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ (1990) Basic local alignment search tool. Mol Biol 215:403–410

    Article  CAS  Google Scholar 

  • Apprill A, McNally S, Parsons R, Laura Weber L (2015) Minor revision to V4 region SSU rRNA 806R gene primer greatly increases detection of SAR11 bacterioplankton. Aquat Microb Ecol 75:129–137

    Article  Google Scholar 

  • Campos MAD, da Silva FSB, Yano-Melo AM, de Melo NF, Maia LC (2017) Application of arbuscular mycorrhizal fungi during the acclimatization of Alpinia purpurata to induce tolerance to Meloidogyne arenaria. Plant Pathol J 33:329–336

    Article  Google Scholar 

  • Cavagnaro TR, Jackson LE, Six J, Ferris H, Goyal S, Asami D, Scow KM (2006) Arbuscular mycorrhizas, microbial communities, nutrient availability, and soil aggregates in organic tomato production. Plant Soil 282:209–225

    Article  CAS  Google Scholar 

  • Christensen MJ, Leuchtmann A, Rowan DD, Tapper BA (1993) Taxonomy of Acremonium endophytes of tall fescue (Festuca arundinacea), meadow fescue (Festuca pratensis) and perennial ryegrass (Lolium perenne). Mycol Res 97:1083–1092

    Article  Google Scholar 

  • Chu-Chou M, Guo B, An ZQ, Hendrix JW, Ferriss RS, Siegel MR, Dougherty CT, Burrus PB (1992) Suppression of mycorrhizal fungi in fescue by the Acremonium coenophialum endophyte. Soil Biol Biochem 24:633–637

    Article  Google Scholar 

  • Cimmino A, Andolfi A, Berestetskiy A, Evidente A (2008) Production of phytotoxins by Phoma exigua var. exigua, a potential mycoherbicide against perennial thistles. J Agric Food Chem 56:6304–6309

    Article  CAS  Google Scholar 

  • de Andrade S, Malik S, Sawaya ACHF, Bottcher A, Mazzafera P (2013) Elicitation of tobacco alkaloid biosynthesis by disrupted spores and filtrate of germinating spores of the arbuscular mycorrhizal fungi Glomus etunicatum. J Plant Interact 8:162–169

    Article  Google Scholar 

  • Den Belder E, Jansen E (1994) Capture of plant-parasitic nematodes by an adhesive hyphae forming isolate of Arthrobotrys oligospora and some other nematode-trapping fungi. Nematologica 40:423–437

    Article  Google Scholar 

  • Edgar RC, Flyvbjerg H (2015) Error filtering, pair assembly and error correction for next-generation sequencing reads. Bioinformatics 31:3476–3482

    Article  CAS  Google Scholar 

  • Fahrbach M, Kuever J, Meinke R, Kämpfer P, Hollender J (2006) Denitratisoma oestradiolicum gen. nov., sp. nov., a 17beta-oestradiol-degrading, denitrifying betaproteobacterium. Int J Syst Evol Microbiol 56:1547–1552

    Article  CAS  Google Scholar 

  • Fisher JB, Jayachandran K (2002) Arbuscular mycorrhizal fungi enhance seedling growth in two endangered plant species from south Florida. Int J Plant Sci 163:559–566

    Article  Google Scholar 

  • Fortin JA, Becard G, Declerck S, Dalpe Y, St-Arnaud M, Coughlan AP, Piche Y (2002) Arbuscular mycorrhiza on root-organ cultures. Can J Bot 80:1–20

    Article  CAS  Google Scholar 

  • Gryndler M, Černá L, Bukovská P, Hršelová H, Jansa J, (2014) Tuber aestivum association with non-host roots. Mycorrhiza 24 (8):603-610

    Article  Google Scholar 

  • Ihrmark K, Bödeker ITM, Cruz-Martinez K, Friberg H, Kubartova A, Schenck J, Strid Y, Stenlid J, Brandström-Durling M, Clemmensen KE, Lindahl BD (2012) New primers to amplify the fungal ITS2 region—evaluation by 454-sequencing of artificial and natural communities. FEMS Microbiol Ecol 82:666–677

    Article  CAS  Google Scholar 

  • Kahiluoto H, Vestberg M (2000) Creation of a non-mycorrhizal control for a bioassay of AM effectiveness. 2. Benomyl application and soil sampling time. Mycorrhiza 9:259–270

    Article  Google Scholar 

  • Kahiluoto H, Ketoja E, Vestberg M (2000) Creation of a non-mycorrhizal control for a bioassay of AM effectiveness. 1. Comparison of methods. Mycorrhiza 9:241–258

    Article  Google Scholar 

  • Kiers ET, Duhamel M, Beesetty Y, Mensah JA, Franken O, Verbruggen E, Fellbaum CR, Kowalchuk GA, Hart MM, Bago A, Palmer TM, West SA, Vandenkoornhuyse P, Jansa J, Bücking H (2011) Reciprocal rewards stabilize cooperation in the mycorrhizal symbiosis. Science 333:880–882

    Article  CAS  Google Scholar 

  • Koide R, Li M (1989) Appropriate controls for vesicular-arbuscular mycorrhiza research. New Phytol 111:35–44

    Article  Google Scholar 

  • Kojima T, Saito K, Oba H, Yoshida Y, Terasawa J, Umehara Y, Suganuma N, Kawaguchi M, Ohtomo R (2014) Isolation and phenotypic characterization of Lotus japonicus mutants specifically defective in arbuscular mycorrhizal formation. Plant Cell Physiol 55:928–941

    Article  CAS  Google Scholar 

  • Kreutzer WA (1960) Soil treatment. In: Horsfall JG, Dimond AE (eds) Plant pathology III—the diseased population epidemics and control. Academic Press, New York, pp 431–476

    Google Scholar 

  • Leigh J, Fitter AH, Hodge A (2011) Growth and symbiotic effectiveness of an arbuscular mycorrhizal fungus in organic matter in competition with soil bacteria. FEMS Microbiol Ecol 76:428–438

    Article  CAS  Google Scholar 

  • Manian S, Edathil TT, Udayian K (1995) Vesicular-arbuscular mycorrhizal colonization and growth of tomato (Lycopersicon esculentum) in autoclaved soil. Pertanika J Trop Agric Sci 18:95–101

    Google Scholar 

  • Marschner P, Crowley DE, Lieberei R (2001) Arbuscular mycorrhizal infection changes the bacterial 16S rDNA community composition in the rhizosphere of maize. Mycorrhiza 11:297–302

    Article  CAS  Google Scholar 

  • Marsh JF, Schultze M (2001) Analysis of arbuscular mycorrhizas using symbiosis-defective plant mutants. New Phytol 150:525–532

    Article  Google Scholar 

  • Merryweather J, Fitter A (1996) Phosphorus nutrition of an obligately mycorrhizal plant treated with the fungicide benomyl in the field. New Phytol 132:307–311

    Article  CAS  Google Scholar 

  • Mortimer PE, Perez-Fernandez MA, Valentine AJ (2008) The role of arbuscular mycorrhizal colonization in the carbon and nutrient economy of the tripartite symbiosis with nodulated Phaseolus vulgaris. Soil Biol Biochem 40:1019–1027

    Article  CAS  Google Scholar 

  • Naumann M, Schüßler A, Bonfante P (2010) The obligate endobacteria of arbuscular mycorrhizal fungi are ancient heritable components related to the Mollicutes. ISME J 4:862–871

    Article  Google Scholar 

  • Nazeri NK, Lambers H, Tibbett M, Ryan MH (2013) Do arbuscular mycorrhizas or heterotrophic soil microbes contribute toward plant acquisition of a pulse of mineral phosphate? Plant Soil 373:699–710

    Article  CAS  Google Scholar 

  • Ohno T, Zibilske LM (1991) Determination of low concentrations of phosphorus in soil extracts using malachite green. Soil Sci Soc Am J 55:892–895

    Article  CAS  Google Scholar 

  • Pizano C, Mangan SA, Graham JH, Kitajima K (2017) Host-specific effects of soil microbial filtrates prevail over those of arbuscular mycorrhizae in a fragmented landscape. Ecol Appl 27:1946–1957

    Article  Google Scholar 

  • Püschel D, Janoušková M, Voříšková A, Gryndlerová H, Vosátka M, Jansa J (2017) Arbuscular mycorrhiza stimulates biological nitrogen fixation in two Medicago spp. through improved phosphorus acquisition. Front Plant Sci 8:390

    Article  Google Scholar 

  • Řezáčová V, Slavíková R, Sochorová L, Konvalinková T, Procházková V, Šťovíček V, Hršelová H, Beskid O, Hujslová M, Gryndlerová H, Gryndler M, Püschel D, Jansa J (2018) Mycorrhizal symbiosis induces plant carbon re-allocation differently in C3 and C4 Panicum grasses. Plant Soil 425:441–456. https://doi.org/10.1007/s11104-018-3606-9

    Article  CAS  Google Scholar 

  • Rivero J, Gamir J, Aroca R, Pozo MJ, Flors V (2015) Metabolic transition in mycorrhizal tomato roots. Front Microbiol 6:598

    Article  Google Scholar 

  • Rosikiewicz P, Bonvin J, Sanders IR (2017) Cost-efficient production of in vitro Rhizophagus irregularis. Mycorrhiza 27:477–486

    Article  Google Scholar 

  • Schliemann W, Ammer C, Strack D (2008) Metabolite profiling of mycorrhizal roots of Medicago truncatula. Phytochemistry 69:112–146

    Article  CAS  Google Scholar 

  • Schouteden N, De Waele D, Panis B, Vos CM (2015) Arbuscular mycorrhizal fungi for the biocontrol of plant-parasitic nematodes: a review of the mechanisms involved. Front Microbiol 6:1280

    Article  Google Scholar 

  • Shaw LJ, Beaton Y, Glover LA, Killham K, Meharg AA (1999) Re-inoculation of autoclaved soil as a non-sterile treatment for xenobiotic sorption and biodegradation studies. Appl Soil Ecol 11:217–226

    Article  Google Scholar 

  • Slavíková R, Püschel D, Janoušková M, Hujslová M, Konvalinková T, Gryndlerová H, Gryndler M, Weiser M, Jansa J (2017) Monitoring CO2 emissions to gain a dynamic view of carbon allocation to arbuscular mycorrhizal fungi. Mycorrhiza 27:35–51

    Article  Google Scholar 

  • Smith SE, Smith FA, Jakobsen I (2003) Mycorrhizal fungi can dominate phosphate supply to plants irrespective of growth responses. Plant Physiol 133:16–20

    Article  CAS  Google Scholar 

  • Souza RC, Hungria M, Cantao ME, Vasconcelos ATR, Nogueira MA, Vicente VA (2015) Metagenomic analysis reveals microbial functional redundancies and specificities in a soil under different tillage and crop-management regimes. Appl Soil Ecol 86:106–112

    Article  Google Scholar 

  • Ter Braak CJF, Šmilauer P (2012) Canoco reference manual and user’s guide: software for ordination (version 5). Microcomputer Power, Ithaca 496 pp

    Google Scholar 

  • Thonar C, Erb A, Jansa J (2012) Real-time PCR to quantify composition of arbuscular mycorrhizal fungal communities—marker design, verification, calibration and field validation. Mol Ecol Resour 12:219–232

    Article  CAS  Google Scholar 

  • Veresoglou SD (2012) Arbuscular mycorrhiza prevents suppression of actual nitrification rates in the (myco-)rhizosphere of Plantago lanceolata. Pedosphere 22:225–229

    Article  CAS  Google Scholar 

  • Větrovský T, Baldrian P (2013) Analysis of soil fungal communities by amplicon pyrosequencing: current approaches to data analysis and the introduction of the pipeline SEED. Biol Fertil Soils 49:1027–1037

    Article  Google Scholar 

  • Viollet A, Corberand T, Mougel C, Robin A, Lemanceau P, Mazurier S (2011) Fluorescent pseudomonads harboring type III secretion genes are enriched in the mycorrhizosphere of Medicago truncatula. FEMS Microbiol Ecol 75:457–467

    Article  CAS  Google Scholar 

  • Wagg C, Bender SF, Widmer F, van der Heijden MGA (2014) Soil biodiversity and soil community composition determine ecosystem multifunctionality. Proc Natl Acad Sci U S A 111:5266–5270

    Article  CAS  Google Scholar 

  • Wang X, Chen M, Xiao J, Hao L, Crowley DE, Zhang Z, Yu J, Huang N, Huo M, Wu J (2015) Genome sequence analysis of the naphthenic acid degrading and metal resistant bacterium Cupriavidus gilardii CR3. PLoS One 10:e0132881

    Article  Google Scholar 

  • Wehner J, Antunes PM, Powell JR, Mazukatow J, Rillig MC (2010) Plant pathogen protection by arbuscular mycorrhizas: a role for fungal diversity? Pedobiologia 53:197–201

    Article  Google Scholar 

  • White T, Bruns T, Lee S, Taylor J (1990) Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Innis MA, Gelfand DH, Sninsky JJ, White TJ (eds) PCR protocols: a guide to methods and applications. Academic Press, San Diego, pp 315–322

    Google Scholar 

  • Willmann M, Gerlach N, Buer B, Polatajko A, Nagy R, Koebke E, Jansa J, Flisch R, Bucher M (2013) Mycorrhizal phosphate uptake pathway in maize: vital for growth and cob development on nutrient poor agricultural and greenhouse soils. Front Plant Sci 4:533

    Article  Google Scholar 

  • Wu X, Wang W, Liu J, Pan D, Tu X, Lv P, Wang Y, Cao H, Wang Y, Rimao Hua R (2017) Rapid biodegradation of the herbicide 2,4-dichlorophenoxyacetic acid by Cupriavidus gilardii T-1. J Agric Food Chem 65:3711–3720

    Article  CAS  Google Scholar 

Download references

Funding

The research was financially supported by the Czech Scientific Foundation (project P504-12-1665), Czech Ministry of Education, Youth and Sports (project LK11224) and long-term development project of the Institute of Microbiology ASCR, Prague (project RVO61388971). PS was further supported by the Center of Excellence PLADIAS, Czech Scientific Foundation project 14-36079G.

Author information

Authors and Affiliations

  1. Institute of Microbiology, v.v.i., Academy of Sciences of the Czech Republic, Vídeňská 1083, 142 20, Prague 4, Czech Republic

    Milan Gryndler, David Püschel, Petra Bukovská, Hana Hršelová, Martina Hujslová, Hana Gryndlerová, Olena Beskid, Tereza Konvalinková & Jan Jansa

  2. Faculty of Science, Jan Evangelista Purkyně University in Ústí nad Labem, České mládeže 8, 400 96, Ústí nad Labem, Czech Republic

    Milan Gryndler

  3. Faculty of Science, University of South Bohemia, České Budějovice, Czech Republic

    Petr Šmilauer

Authors
  1. Milan Gryndler
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Petr Šmilauer
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. David Püschel
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Petra Bukovská
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Hana Hršelová
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Martina Hujslová
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Hana Gryndlerová
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Olena Beskid
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Tereza Konvalinková
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Jan Jansa
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Milan Gryndler.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Electronic supplementary material

ESM 1

(XLSX 5239 kb)

ESM 2

(DOCX 18 kb)

ESM 3

(PDF 318 kb)

ESM 4

(GIF 629 kb)

High resolution image (TIF 226 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Gryndler, M., Šmilauer, P., Püschel, D. et al. Appropriate nonmycorrhizal controls in arbuscular mycorrhiza research: a microbiome perspective. Mycorrhiza 28, 435–450 (2018). https://doi.org/10.1007/s00572-018-0844-x

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00572-018-0844-x

Keywords

Search

Navigation