Plant and Soil

, Volume 429, Issue 1–2, pp 437–450 | Cite as

Effects of Bacillus amyloliquefaciens and different phosphorus sources on Maize plants as revealed by NMR and GC-MS based metabolomics

  • Giovanni Vinci
  • Vincenza Cozzolino
  • Pierluigi Mazzei
  • Hiarhi Monda
  • Davide Savy
  • Marios Drosos
  • Alessandro Piccolo
Regular Article



Plant growth-promoting bacteria of the genus Bacillus are known to solubilize phosphates and enhance plant growth in many plant species. We explored the effects of the inoculation with a commercial isolate Bacillus amyloliquefaciens on the growth and metabolic processes of maize plants in pot soils treated with triple superphosphate, rock phosphate, and either cow- or horse-manure composts, as P-fertilizers.


The metabolic profiles of maize leaves in the different treatments were determined by both Gas Chromatography–Mass Spectrometry and Nuclear Magnetic Resonance spectroscopy. Principle Components Analysis (PCA) based on data matrix from both techniques revealed a relationship between treatments and specific plant metabolites.


Inoculated plants showed larger P and N contents and a more differentiated metabolome when treated with the two composts than with inorganic fertilizers. B. amyloliquefaciens in combination with composts significantly increased glucose, fructose, alanine and GABA metabolites in maize leaves, thus suggesting an improved photosynthetic activity due to enhanced P and N uptake. Both composts sustained plant growth and the phosphate solubilizing activity of B. amyloliquefaciens, while differences in P and N contents in plant leaves were attributed to the different content in compost of lignin residues and alkyl moieties, and consequent impact on microbial growth.


The combination of B. amyloliquefaciens inoculation with composted organic P-fertilizers rich in available metabolic carbon appears as an efficient alternative to mineral fertilizers to enhance nutrients uptake and foster growth mechanisms in maize plants.


Metabolomics Phosphate-solubilizing-bacteria Compost Rock phosphate Triple Superphosphate GC-MS 1H-NMR Thermochemolysis 



This work was conducted in partial fulfillment of first author PhD requirements, and received funding from the European Community’s Seventh Framework Programme (FP7/2007–2013) under grant agreement n° 312117 (BIOFECTOR).

Supplementary material

11104_2018_3701_MOESM1_ESM.docx (249 kb)
ESM 1 (DOCX 249 kb)


  1. Amiour N, Imbaud S, Clément G, Agier N, Zivy M, Valot B, Tercet-Laforgue T (2012) The use of metabolomics integrated with transcriptomic and proteomic studies for identifying key steps involved in the control of nitrogen metabolism in crops such as maize. J Exp Bot 63:5017–5033CrossRefPubMedGoogle Scholar
  2. Babicki S, Arndt D, Marcu A, Liang Y, Grant JR, Maciejewski A, Wishart DS (2016) Heatmapper: web-enabled heat mapping for all. Nucleic Acids Res 44:147–153CrossRefGoogle Scholar
  3. Brady NC, Weil RR (2008) The nature and properties of soils (fourteenth ed.), Prentice Hall, Upper Saddle River, NJ, USAGoogle Scholar
  4. Bundy JG, Davey MP, Viant MR (2009) Environmental metabolomics: a critical review and future perspectives. Metabolomics 5:3–21CrossRefGoogle Scholar
  5. Chapman S, Barreto H (1997) Using a chlorophyll meter to estimate specific leaf nitrogen of tropical maize during vegetative growth. Agron J 89:557–562CrossRefGoogle Scholar
  6. Chien SH, Menon RG (1995) Factors affecting the agronomic effectiveness of phosphate rock for direct application. Fertil Res 41:227–234CrossRefGoogle Scholar
  7. Cordell D, Drangert JO, White S (2009) The story of phosphorus: global food security and food for thought. Glob Environ Chang 19:292–305CrossRefGoogle Scholar
  8. Cozzolino V, Di Meo V, Piccolo A (2013) Impact of arbuscular mycorrhizal fungi applications on maize production and soil phosphorus availability. J Geochem Explor 129:40–44Google Scholar
  9. Cozzolino V, Di Meo V, Monda H, Spaccini R, Piccolo, A (2016) The molecular characteristics of compost affect plant growth, arbuscular mycorrhizal fungi, and soil microbial community composition. Biol Fert Soils 52:15–29Google Scholar
  10. Fait A, Fromm H, Walter D, Galili G, Fernie AR (2008) Highway or byway: the metabolic role of the GABA shunt in plants. Trends Plant Sci 13:14–19CrossRefPubMedGoogle Scholar
  11. Foyer C, Spencer C (1986) The relationship between phosphate status and photosynthesis in leaves. Planta 167:369–375CrossRefPubMedGoogle Scholar
  12. Ganie AH, Ahmad A, Pandey R, Aref IM, Yousuf PY, Ahmad S, Iqbal M (2015) Metabolite profiling of low-P tolerant and low-P sensitive maize genotypes under phosphorus starvation and restoration conditions. PLoS One 10:e0129520CrossRefPubMedPubMedCentralGoogle Scholar
  13. Gericke S, Kurmies B (1952) Die kolorimetrische Phosphorsaurebestimmung mit Ammonium-Vanadat-Molybdat und ihre Anwendung in der Pflanzenanalyse. Z Pflanzenernähr Bodenkd 59:32–35Google Scholar
  14. Gyaneshwar P, Naresh KG, Parekh LJ, Poole PS (2002) Role of soil microorganisms in improving P nutrition of plants. Plant Soil 245:83–93CrossRefGoogle Scholar
  15. Hargreaves JC, Adl MS, Warman PR (2008) A review of the use of composted municipal solid waste in agriculture. Agric Ecosyst Environ 123:1–14CrossRefGoogle Scholar
  16. Hernández G, Ramírez M, Valdés-López O et al (2007) Phosphorus stress in common bean: root transcript and metabolic responses. Plant Physiol 144:752–767CrossRefPubMedPubMedCentralGoogle Scholar
  17. Hirel B, Andrieu B, Valadier MH, Renard S, Quilleré I, Chelle M, Drouet JL (2005) Physiology of maize II: identification of physiological markers representative of the nitrogen status of maize (Zea mays) leaves during grain filling. Physiol Plant 124:178–188CrossRefGoogle Scholar
  18. Hofstra G, Nelson CD (1969) The translocation of photosynthetically assimilated in 14C in corn. Can J Bot 47:1435–1442CrossRefGoogle Scholar
  19. Huang CY, Roessner U, Eickmeier I, Genc Y, Callahan DL, Shirley N, Bacic A (2008) Metabolite profiling reveals distinct changes in carbon and nitrogen metabolism in phosphate-deficient barley plants (Hordeum vulgare L.). Plant Cell Physiol 49:691–703CrossRefPubMedGoogle Scholar
  20. Iordachescu M, Imai R (2008) Trehalose biosynthesis in response to abiotic stresses. J Integr Plant Biol 50(10):1223–1229CrossRefPubMedGoogle Scholar
  21. Koliaei AA, Akbari Gh A, Armandpisheh O, Labbafi MR, Zarghami R (2011) Effects of phosphate chemical fertilizers and biologic fertilizers in various moisture regimes on some morphological characteristics and seeds performance in maize S.C.704. Asian Journal of Agriculture and Food Sciences 3:223–234Google Scholar
  22. Kowaljow E, Mazzarino MJ (2007) Soil restoration in semiarid Patagonia: Chemical and biological response to different compost quality. Soil Biol Biochem 39:1580–1588CrossRefGoogle Scholar
  23. Lazcano C, Gómez-Brandón M, Revilla P, Domínguez J (2013) Short-term effects of organic and inorganic fertilizers on soil microbial community structure and function. Biol Fertil Soils 49:723–733CrossRefGoogle Scholar
  24. Lekfeldt JDS, Rex M, Mercl F, Kulhánek M, Tlustoš P, Magid J, de Neergaard A (2016) Effect of bioeffectors and recycled P-fertiliser products on the growth of spring wheat. Chem Biol Technol Agric 3:22Google Scholar
  25. Lemaire G, Gastal F (1997) N uptake and distribution in plant canopies. In: Lemaire G (ed) Diagnosis of the nitrogen status in crops. Springer-Verlag, Berlin, pp 3–43CrossRefGoogle Scholar
  26. Li M, Welti R, Wang X (2006) Quantitative profiling of Arabidopsis polar glycerolipids in response to phosphorus starvation. Roles of phospholipases Dζ1 and Dζ2 in phosphatidylcholine hydrolysis and digalactosyldiacylglycerol accumulation in phosphorus-starved plants. Plant Physiol 142:750–761CrossRefPubMedPubMedCentralGoogle Scholar
  27. Li M, Cozzolino V, Mazzei P, Drosos M, Monda H, Hu Z, Piccolo A (2017) Effects of microbial bioeffectors and P amendements on P forms in a maize cropped soil as evaluated by 31 P–NMR spectroscopy. Plant Soil:1–18Google Scholar
  28. Lichtenthaler HK (1987) Chlorophylls and carotenoids, the pigments of photosynthetic biomembranes. In: Douce R, Packer L (eds) Methods Enzymol. 148, 350–382. Academic Press Inc., New YorkGoogle Scholar
  29. Lynch JP (2011) Root phenes for enhanced soil exploration and phosphorus acquisition: tools for future crops. Plant Physiol 156:1041–1049CrossRefPubMedPubMedCentralGoogle Scholar
  30. Malik MA, Marschner P, Khan KS (2012) Addition of organic and inorganic P sources to soil–effects on P pools and microorganisms. Soil Biol Biochem 49:106–113CrossRefGoogle Scholar
  31. Michaeli S, Fromm H (2015) Closing the loop on the GABA shunt in plants: are GABA metabolism and signaling entwined? Front Plant Sci 6:419CrossRefPubMedPubMedCentralGoogle Scholar
  32. Murphy J, Riley JP (1962) A modifed single solution method for the determination of phosphate in natural waters. Anal Chim Acta 27:31–36CrossRefGoogle Scholar
  33. Pane C, Piccolo A, Spaccini R, Celano G, Villecco D, Zaccardelli M (2013) Agricultural waste-based composts exhibiting suppressivity to diseases caused by the phytopathogenic soil-borne fungi Rhizoctonia solani and Sclerotinia minor. Appl Soil Ecol 65:43–51CrossRefGoogle Scholar
  34. Qiao J-Q, Wu H-J, Huo R, Gao X-W, Borriss R (2014) Stimulation of plant growth and biocontrol by Bacillus amyloliquefaciens subsp. plantarum FZB42 engineered for improved action. Chem Biol Technol Agric 1:12. CrossRefGoogle Scholar
  35. Reddy MS, Kumar S, Khosla B (2002) Biosolubilization of poorly soluble rock phosphates by Aspergillus tubingensis and Aspergillus niger. Bioresour Technol 84:187–189CrossRefPubMedGoogle Scholar
  36. Rodriguez H, Fraga R (1999) Phosphate solubilizing bacteria and their role in plant growth promotion. Biotechnol Adv 17:319–339CrossRefPubMedGoogle Scholar
  37. Rodziewicz P, Swarcewicz B, Chmielewska K, Wojakowska A, Stobiecki M (2014) Influence of abiotic stresses on plant proteome and metabolome changes. Acta Physiol Plant 36:1–19CrossRefGoogle Scholar
  38. Schlüter U, Colmsee C, Scholz U, Bräutigam A, Weber APM, Zellerhoff N, Bucher M, Fahnenstich H, Sonnewald U (2013) Adaptation of maize source leaf metabolism to stress related disturbances in carbon, nitrogen and phosphorus balance. BMC Genomics 14:442CrossRefPubMedPubMedCentralGoogle Scholar
  39. Schoch G, Goepfert S, Morant M, Hehn A, Meyer D, Ullmann P, Werck-Reichhart D (2001) CYP98A3 from Arabidopsis thaliana is a 30-hydroxylase of phenolic esters, a missing link in the phenylpropanoid pathway. J Biol Chem 276:36566–36574CrossRefPubMedGoogle Scholar
  40. Schröder JJ, Smit a L, Cordell D, Rosemarin A (2011) Improved phosphorus use efficiency in agriculture: a key requirement for its sustainable use. Chemosphere 84:822–831CrossRefPubMedGoogle Scholar
  41. Sengupta S, Mukherjee S, Basak P, Majumder AL (2015) Significance of galactinol and raffinose family oligosaccharide synthesis in plants. Front Plant Sci. 26 6:656Google Scholar
  42. Spaccini R, Piccolo A (2009) Molecular characteristics of humic acids extracted from compost at increasing maturity stages. Soil Biol Biochem 41:1164–1172CrossRefGoogle Scholar
  43. Spohn M, Kuzyakov Y (2013) Phosphorus mineralization can be driven by microbial need for carbon. Soil Biol Biochem 61:69–75CrossRefGoogle Scholar
  44. Sugiharto B, Miyata K, Nakamoto H, Sasakawa H, Sugiyama T (1990) Regulation of expression of carbon-assimilating enzymes by nitrogen in maize leaf. Plant Physiol 92:963CrossRefPubMedPubMedCentralGoogle Scholar
  45. Sun CX, Gao XX, Li MQ, Fu JQ, Zhang YL (2015) Metabolic response of maize (Zea mays L.) plants to combined drought and salt stress. Plant Soil 388:99–117CrossRefGoogle Scholar
  46. Takahama U (1998) Ascorbic acid-dependant regulation of redox levels of chlorogenic acid and its isomers in the apoplast of leaves of Nicotiana tabacum L. Plant Cell Physiol 39:681–689CrossRefGoogle Scholar
  47. Thomas H, Stoddart JL (1980) Leaf senescence. Annu Rev Plant Physiol 31:83–111CrossRefGoogle Scholar
  48. Thonar C, Lekfeldt JDS, Cozzolino V, Kundel D, Kulhánek M, Mosimann C, Neumann G, Piccolo A, Rex M, Symanczik S, Walder F, Weinmann M, de Neergaard A, Mäder P (2017) Potential of three microbial bio-effectors to promote maize growth and nutrient acquisition from alternative phosphorous fertilizers in contrasting soils. Chem Biol Technol Agric 4:7. CrossRefGoogle Scholar
  49. Vane CH, Martin SC, Snape CE, Abbott GD (2001) Degradation of lignin in wheat straw during growth of the oyster mushroom (Pleurotus ostreatus) using off-line thermochemolysis with tetramethylammonium hydroxyde and solid state 13C NMR. J Agric Food Chem 49:2709–2716CrossRefPubMedGoogle Scholar
  50. Vogt T (2010) Phenylpropanoid Biosynthesis. Mol Plant 3:2–20CrossRefPubMedGoogle Scholar
  51. Zapata F, Zaharah AR (2002) Phosphorus availability from phosphate rock and sewage sludge as influence by the addition of water soluble phosphate fertilizer. Nutr Cycl Agroecosyst 63:43–48CrossRefGoogle Scholar
  52. Zhang L, Ding X, Chen S, He X, Zhang F, Feng G (2014) Reducing carbon: phosphorus ratio can enhance microbial phytin mineralization and lessen competition with maize for phosphorus. J Plant Interact 9:850–856CrossRefGoogle Scholar
  53. Zhang L, Xu M, Liu Y, Zhang F, Hodge A, Feng G (2016) Carbon and phosphorus exchange may enable cooperation between an arbuscular mycorrhizal fungus and a phosphate solubilizing bacterium. New Phytol 210:1022–1032CrossRefPubMedGoogle Scholar
  54. Zhao L, Hu J, Huang Y, Wang H, Adeleye A, Ortiz C, Keller A (2016) 1H NMR and GC MS based metabolomics reveal nano-Cu altered cucumber (Cucumis sativus) fruit nutritional supply. Plant Physiol Biochem.

Copyright information

© Springer International Publishing AG, part of Springer Nature 2018

Authors and Affiliations

  • Giovanni Vinci
    • 1
  • Vincenza Cozzolino
    • 1
  • Pierluigi Mazzei
    • 1
  • Hiarhi Monda
    • 1
  • Davide Savy
    • 1
  • Marios Drosos
    • 1
  • Alessandro Piccolo
    • 1
  1. 1.Centro Interdipartimentale di Ricerca sulla Risonanza Magnetica Nucleare per l’Ambiente, l’Agroalimentare ed i Nuovi Materiali (CERMANU)Università di Napoli Federico IIPorticiItaly

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