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Combination of Siderophore-Producing Bacteria and Piriformospora indica Provides an Efficient Approach to Improve Cadmium Tolerance in Alfalfa

  • Plant Microbe Interactions
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Abstract

Application of siderophore-producing microorganisms (SPMs), as an environmentally friendly approach, facilitates plant growth and survival under heavy metals toxicity. This study evaluated the effectiveness of SPMs, belonging to the bacterial genera Rhizobium and Pseudomonas and a root endophytic fungus (Piriformospora indica) to improve the fitness of alfalfa under cadmium (Cd) stress. A greenhouse experiment was performed as a randomized design with factorial arrangement of treatments. Treatments included microbial inoculations (Sinorhizobium meliloti, Pseudomonas fluorescence, and P. indica) and different Cd concentrations (0, 2, 5, 10 mg/kg) with three replications in potting media containing sand and sterile perlite (v/v, 2:1). The effect of Cd on plant growth and development, antioxidant enzymes activities, and accumulation of Cd and nutrients in alfalfa plant was investigated. Alfalfa inoculated with SPMs showed significantly higher biomass and nutrients uptake under both normal and Cd stress conditions than the controls. Under the highest Cd concentration (10 mg/kg), alfalfa plants inoculated with P. fluorescens and P. indica, either alone or in combination, showed the highest shoot dry weights. Cd-induced oxidative stress was mitigated by SPMs through enhanced antioxidant enzyme activities of catalase, ascorbate peroxidase, and guaiacol peroxidase. We showed that P. indica either alone or in combination with the siderophore producing bacteria (SPB) minimized the toxicity of Cd by enhanced growth rate and the lower Cd concentration in the shoots. In conclusion, metal-resistant SPMs could assist alfalfa to survive in Cd-contaminated soil by enhancing plant growth and development. Application of plant-associated microbes is an efficient, environmentally friendly approach to surmount the adverse effects of heavy metals toxicity on plants, animals, and humans.

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Abbreviations

APX:

Ascorbate peroxidase

CAT:

Catalase

CFU:

Colony forming units

CM:

Complex medium

DTT:

Dithiothreitol

EDTA:

Ethylenediaminetetraacetic acid

GPX:

Glutathione peroxidase

HM:

Heavy metal

PGPB:

Plant growth promoting bacteria

PGPF:

Plant growth promoting fungi

PGPMs:

Plant growth promoting microorganisms

PVP:

Polyvinylpyrrolidone

PBS:

Phosphate buffered saline

SNF:

Symbiotic nitrogen fixation

SPB:

Siderophore-producing bacteria

SPMs:

Siderophore-producing microorganisms

References

  1. Aliasgharzad N, Shirmohamadi E, Oustan S (2009) Siderophore production by mycorrhizal sorghum roots under micronutrient deficient condition. Soil Environ 28:119–123

    CAS  Google Scholar 

  2. Andrad SA, Abreu MF, Silveira AP (2004) Influence of lead additions on arbuscular mycorrhiza and Rhizobium symbioses under soybean plants. Appl Soil Ecol 26:123–131

    Google Scholar 

  3. Antunes PM, Rajcan I, Goss MJ (2006) Specific flavonoids as interconnecting signals in the tripartite symbiosis formed by arbuscular mycorrhizal fungi, Bradyrhizobium japonicum (Kirchner) Jordan and soybean (Glycine max L. Merrill.). Soil Biol Biochem 38:533–543

    CAS  Google Scholar 

  4. Bakkaus E, Collins RN, Morel JL, Gouget B (2006) Anion exchange liquid chromatography inductively coupled plasma- mass spectrometry of the Co2+, Cu2+, Fe3+ and Ni2+ complexes of mugineic and deoxymugineic acid. J Chromatogr 1129:208–215

    CAS  Google Scholar 

  5. Baltruschat H, Fodor J, Harrach BD, Niemczyk E, Barna B, Gullner G, Janeczko A, Kogel KH, Schaferm P, Schwarczinger I, Zuccaro A, Skoczowski A (2008) Salt tolerance of barley induced by the root endophyte Piriformospora indica is associated with a strong increase in antioxidants. New Phytol 180:501–510

    CAS  PubMed  Google Scholar 

  6. Barzanti R, Ozino F, Bazzicalupo M, Gabbrielli R, Galardi F, Gonnelli C, Mengoni A (2007) Isolation and characterization of endophytic bacteria from the nickel hyperaccumulator plant Alyssum bertolonii. Microb Ecol 53:306–316

    CAS  PubMed  Google Scholar 

  7. Berendsen RL, Pieterse C, Bakker P (2012) The rhizosphere microbiome and plant health. Trends Plant Sci 17(8):428–486

    Google Scholar 

  8. Bhargava A, Carmona FC, Bhargava M, Srivastava S (2012) Review: approaches for enhanced phytoextraction of heavy metals. J Environ Manag 105:103–120

    CAS  Google Scholar 

  9. Bisht R, Chaturvedi S, Srivastava R, Sharma AK, Johri BN (2009) Effect of arbuscular mycorrhizal fungi, Pseudomonas fluorescens and Rhizobium leguminosarum on the growth and nutrient status of Dalbergia sissoo Roxb. Trop Ecol 50(2):231–242

    CAS  Google Scholar 

  10. Bremmer J, Mulvaney C (eds) (1982) Methods of soil analysis. Nitrogen-Total. Part 2. Chemical and Microbiollogical Properties. 2rd edn. American Society of Agronomy, Inc., Madison, pp 295–624

    Google Scholar 

  11. Burd GI, Dixon DG, Glick BR (2000) Plant growth-promoting bacteria that decrease heavy metal toxicity in plants. Can J Microbiol 46(3):237–245

    CAS  PubMed  Google Scholar 

  12. Bürkert B, Robson A (1994) Zn uptake in subterranean clover (Trifolium subterraneum L.) by three vesicular-arbuscula r mycorrhizal fungi in a root-free sandy soil. Soil Biol Biochem 26:1117–1124

    Google Scholar 

  13. Burleigh SH, Kristensen BK, Bechmann IE (2003) A plasma membrane zinc transporter from Medicago truncatula is up regulated in roots by Zn fertilization, yet down-regulated by arbuscular mycorrhizal colonization. Plant Mol Biol 52(5):1077–1088

    CAS  PubMed  Google Scholar 

  14. Chen D, Qian P, Wang W (2008) Biokinetics of cadmium and zinc in a marine bacterium: Influences of metal interaction and pre-exposure. Environ Toxicol Chem 27:1794–1808

    CAS  PubMed  Google Scholar 

  15. Das K, Roychoudhury A (2014) Reactive oxygen species (ROS) and response of antioxidants as ROS-scavengers during environmental stress in plants. Front Environ Sci 2:1–13

    CAS  Google Scholar 

  16. Dell’Amico E, Cavalca L, Andreoni V (2008) Improvement of Brassica napus growth under cadmium stress by cadmium resistant rhizobacteria. Soil Biol Biochem 40:74–84

    Google Scholar 

  17. Deshmukh SD, Kogel KH (2007) Piriformospora indica protects barley from root rot caused by Fusarium graminearum. J Plant Dis Prot 14(6):263–268

    Google Scholar 

  18. Dhiman SS, Zhao X, Li J, Kim D, Kalia VC, Lee J (2017) Correction: metal accumulation by sunflower (Helianthus annuus L.) and the efficacy of its biomass in enzymatic saccharification. PLoS One 12(6):e0179746. https://doi.org/10.1371/journal.pone.0175845

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Dimkpa C (2016) Microbial siderophores: production, detection and application in agriculture and environment. Endocytobiosis Cell Res 27(2):7–16

    Google Scholar 

  20. Dodd IC, Zinovkina NY, Safronova VI, Belimov AA (2010) Rhizobacterial mediation of plant hormone status. Ann Appl Biol 157:361–379

    CAS  Google Scholar 

  21. Doornbos RF, Van Loon LC, Bakker PAHM (2012) Impact of root exudates and plant defense signaling on bacterial communities in the rhizosphere. Agron Sustain Dev 32:227–243

    Google Scholar 

  22. Du YX, Pan GX, Li LQ, Hu ZL, Wang XZ (2011) Leaf N/P ratio and nutrient reuse between dominant species and stands: predicting phosphorus deficiencies in karst ecosystems, southwestern China. Environ Earth Sci 64:299–309

    CAS  Google Scholar 

  23. Gill SS, Gill R, Trivedi DK, Anjum NA, Sharma KK, Ansari MW, Ansari AA, Johri AK, Prasad R, Pereira E, Varma A, Tuteja N (2016) Piriformospora indica: potential and significance in plant stress tolerance. Front Microbiol 7:1–20

    Google Scholar 

  24. Gómez-Sagasti M, Marino D (2015) PGPRs and nitrogen-fixing legumes: a perfect team for efficient Cd phytoremediation? Front Plant Sci 6:81

    PubMed  PubMed Central  Google Scholar 

  25. Hao SX, Taghavi S, Xie P, Orbach MJ, Alwathnani HA, Rensing C, Wei G (2014) Phyotoremediation of heavy and transition metals aides by legume-rhizobia symbiosis. Int J Phytoremediation 16:179–202

    CAS  PubMed  Google Scholar 

  26. Haselwandter K (2008) Structure and function of siderophores produced by mycorrhizal fungi. Mineral Mag 72:61–64

    CAS  Google Scholar 

  27. Hassan MJ, Shao G, Zhang G (2005) Influence of cadmium toxicity on antioxidant enzymes activity in rice cultivars with different grain Cd accumulation. J Plant Nutr 28:1259–1270

    CAS  Google Scholar 

  28. Husaini AM, Abdin MZ, Khan S, Xu YW, Aquil S, Anis M (2012) Modifying strawberry for better adaptability to adverse impact of climate change. Curr Sci 102:1660–1673

    CAS  Google Scholar 

  29. Joner E, Leyval C (2008) Uptake of 109 cd by roots and hyphae of a Glomus Mosseae/Trifolium Subterraneum Mycorrhiza from soil amended with high and low concentrations of cadmium. New Phytol 135(2):353–360

    Google Scholar 

  30. Jouili H, Ferjani EL (2004) Effect of copper excess on superoxide dismutase, catalase, and peroxidase activities in sunflower seedlings (Helianthus annuus L.). Acta Physiol Plant 26(1):29–35

    CAS  Google Scholar 

  31. Kirk JL, Klironomos JN, Lee H, Trevors JT (2005) The effects of perennial ryegrass and alfalfa on microbial abundance and diversity in petroleum contaminated soil. Environ Pollut 133:455–465

    CAS  PubMed  Google Scholar 

  32. Lane EA, Canty MJ, More SJ (2015) Cadmium exposure and consequence for the health and productivity of farmed ruminants. Res Vet Sci 101:132–139

    CAS  PubMed  Google Scholar 

  33. Lee YJ, George E (2005) Contributions of mycorrhizal hyphae to the uptake of metal cations by cucumber plants at two levels of phosphorus supply. Plant Soil 278:361–370

    CAS  Google Scholar 

  34. Ma Y, Rajkumar M, Freitas H (2009) Improvement of plant growth and nickel uptake by nickel resistant-plant-growth promoting bacteria. J Hazard Mater 166:1154–1161

    CAS  PubMed  Google Scholar 

  35. Ma Y, Prasad MNV, Rajkumar M, Freitas H (2011) Plant growth promoting rhizobacteria and endophytes accelerate phytoremediation of metalliferous soils. Biotechnol Adv 29:248–258

    CAS  PubMed  Google Scholar 

  36. Malla R, Prasad R, Giang PH, Pokharel U, Oelmüller R, Varma A (2004) Characteristic features of symbiotic fungus Piriformospora indica. Endocytobiosis. Cell Res 5:579–600

    Google Scholar 

  37. Mansotra P, Sharma P, Sharma S (2015) Bioaugmentation of Mesorhizobium cicer, Pseudomonas spp. and Piriformospora indica for sustainable chickpea production. Physiol Mol Biol Plants 21(3):385–393

    PubMed  PubMed Central  Google Scholar 

  38. Meda AR, Scheuermann EB, Prechsl UE, Erenoglu B, Schaaf G, Hayen H, Weber G, von Wirén N (2007) Iron acquisition by Phytosiderophores contributes to cadmium tolerance. Plant Physiol 143(4):1761–1773

    CAS  PubMed  PubMed Central  Google Scholar 

  39. Mehta S, Nautiyal CS (2001) An efficient method for qualitative screening of phosphate solubilizing bacteria. Curr Microbiol 43:51–58

    CAS  PubMed  Google Scholar 

  40. Milagres AMF, Machuca A, Napoleao D (1999) Detection of siderophore production from several fungi and bacteria by a modification of chrome azurol S (CAS) agar plate assay. J Microbiol Methods 37:1–6

    CAS  PubMed  Google Scholar 

  41. Mishra J, Singh R, Arora NK (2017) Alleviation of heavy metal stress in plants and remediation of soil by Rhizosphere microorganisms. Front Microbiol 8:1706

    PubMed  PubMed Central  Google Scholar 

  42. Mittal M, Flora SJ (2007) Vitamin E supplementation protects oxidative stress during arsenic and fluoride antagonism in male mice. Drug Chem Toxicol 30:263–281

    CAS  PubMed  Google Scholar 

  43. Nadeem SM, Ahmad M, Zahir ZA, Javaid A, Ashraf M (2014) The role of mycorrhizae and plant growth promoting rhizobacteria (PGPR) in improving crop productivity under stressful environments. Biotechnol Adv 32:429–444

    PubMed  Google Scholar 

  44. Nakano Y, Asada K (1981) Hydrogen peroxide is scavenged by ascorbate specific peroxidase in spinach chloroplasts. Plant Cell Physiol 22:867–880

    CAS  Google Scholar 

  45. Pajuelo E, Rodríguez-Llorente ID, Lafuente A, Caviedes MA (2011) Legume–Rhizobium symbioses as a tool for bioremediation of heavy metal polluted soils. In: Khan MS, Zaidi A, Goel R, Musarrat J (eds) Biomanagement of metal-contaminated soils. Springer, Netherlands, pp 95–123

  46. Patreze CM, Cordeiro L (2004) Nitrogen-fixing and vesicular arbuscular mycorrhizal symbioses in some tropical legume trees of tribe Mimosease. For Ecol Manag 196:275–285

    Google Scholar 

  47. Penrose DM, Glick BR (2003) Methods for isolating and characterizing ACC deaminase-containing plant growth-promoting rhizobacteria. Physiol Plant 18:10–15

    Google Scholar 

  48. Pooja P, Kumari N, Sharma V (2013) Structural and functional alterations in photosynthetic apparatus of plants under cadmium stress. Bot Stud 54:45

    Google Scholar 

  49. Rabie GH (2005) Contribution of arbuscular mycorrhizal fungus to red kidney and wheat plants tolerance grown in heavy metal-polluted soil. Afr J Biotechnol 4(4):332–345

    CAS  Google Scholar 

  50. Rai M, Acharya D, Singh A, Varma A (2001) Positive growth responses of the medicinal plants Spilanthes calva and Withania somnifera to inoculation by Piriformospora indica in a field trial. Mycorrhiza 11:123–128

    PubMed  Google Scholar 

  51. Rajkumar M, Ae N, Prasad MN, Freitas H (2010a) Potential of siderophore-producing bacteria for improving heavy metal phytoextraction. Trends Biotechnol 28(3):142–149

    CAS  PubMed  Google Scholar 

  52. Rajkumar M, Sandhya S, Prasad MNV, Freitas H (2010b) Perspectives of plant-associated microbes in heavy metal phytoremediation. Biotechnol Adv 30:1562–1574

    Google Scholar 

  53. Ren X-M, Guo S-J, Tian W, Chen Y, Han H, Chen E, Li B-L, Li Y-Y, Chen Z-J (2019) Effects of plant growth-promoting bacteria (PGPB) inoculation on the growth, antioxidant activity, cu uptake, and bacterial community structure of rape (Brassica napus L.) grown in Cu-contaminated agricultural soil. Front Microbiol 10:1455

    PubMed  PubMed Central  Google Scholar 

  54. Sarwar M, Arshad M, Martens DA, Frankenberger WT (1992) Tryptophan dependent biosynthesis of auxins in soil. Plant Soil 147:207–215

    CAS  Google Scholar 

  55. Sarwar N, Saifullah S, Malhi S, Zia MH, Naeem A, Bibia S, Farida G (2010) Role of mineral nutrition in minimizing cadmium accumulation by plants. J Sci Food Agric 90:925–937

    CAS  PubMed  Google Scholar 

  56. Sayyed RZ, Patel PR (2011) Biocontrol potential of siderophore producing heavy metal resistant Alcaligenes sp and Acinetobacter sp. vis-A-vis organophosphorus fungicide. Indian J Microbiol 51(3):266–272

    CAS  PubMed  PubMed Central  Google Scholar 

  57. Schlesier B, Bréton F, Mock HP (2003) A hydroponic culture system for growing Arabidopsis thaliana plantlets under sterile conditions. Plant Mol Biol Report 21:449–456

    CAS  Google Scholar 

  58. Schreiner PR (2007) Effect of native and non-native arbuscular mycorrhizal fungi on growth and nutrient uptake of ‘Pinot noir’ (Vitis vinifera L.) in two soils with contrasting levels of phosphorus. Appl Soil. Ecol 36:205–215

    Google Scholar 

  59. Schwyn B, Neilands JB (1987) Universal chemical assay for the detection and determination of siderophores. Anal Biochem 160:47–56

    CAS  PubMed  Google Scholar 

  60. Seneviratne M, Seneviratne G, Madawala H, Vithanage M (2017) In: Singh JS, Seneviratne G (eds) Role of rhizospheric microbes in heavy metal uptake by plants, in agro-environmental sustainability: managing environmental pollution, vol 2. Springer International Publishing, Cham, pp 147–163

    Google Scholar 

  61. Shao G, Chen M, Wang W, Mou R, Zhang G (2007) Iron nutrition affects cadmium accumulation and toxicity in rice plants. Plant Growth Regul 53:33–42

    CAS  Google Scholar 

  62. Sharma P, Jha AB, Dubey RS, Pessarakli M (2012) Reactive oxygen species, oxidative damage, and antioxidative defense mechanism in plants under stressful conditions. J Bot 2012:1–26

    Google Scholar 

  63. Sun CA, Johnson J, Cai DG, Sherameti I, Oelmüller R, Lou BG (2010) Piriformospora indica confers drought tolerance in Chinese cabbage leaves by stimulating antioxidant enzymes, the expression of drought-related genes and the plastid-localized CAS protein. J Plant Physiol 167:1009–1017

    CAS  PubMed  Google Scholar 

  64. Talebi M, Bahar M, Saeidi G, Mengoni A, Marco Bazzicalupo M (2008) Diversity of Sinorhizobium strains nodulating Medicago sativa from different Iranian regions. FEMS Microbiol Lett 288:40–46

    CAS  PubMed  Google Scholar 

  65. Tangahu BV, Abdullah SRS, Basri H, Idris M, Anuar N, Mukhlisin M (2011) A review on heavy metals (As, Pb, and Hg) uptake by plants through phytoremediation. Int J Chem Eng 2011:1–31. https://doi.org/10.1155/2011/939161

    Article  Google Scholar 

  66. Teresa Cubo M, Ana MB, John EB, Jose ER (1988) Melanin production by Rhizobium strains. Appl Environ Microbiol 54(7):1812–1817

    PubMed Central  Google Scholar 

  67. Thirkell T, Cameron D, Hodge A (2019) Contrasting nitrogen fertilisation rates alter mycorrhizal contribution to barley nutrition in a field trial. Front Plant Sci 10:1312. https://doi.org/10.3389/fpls.2019.01312

    Article  PubMed  PubMed Central  Google Scholar 

  68. Unnikumar KR, Sowjanya SK, Varma A (2013) Piriformospora indica: a versatile root endophytic symbiont. Symbiosis 60:107–113

    Google Scholar 

  69. Upadhyaya A, Sankhla D, Davis TD, Sankhla N, Smith BN (1985) Effect of paclobutrazol on the activities of some enzymes of activated oxygen metabolism and lipid peroxidation in senescing soybean leaves. J Plant Physiol 121:453–461

    CAS  Google Scholar 

  70. Varma A, Chincholkar SB (eds) (2007) Microbial siderophores. New York Springer, New York, pp 43–61

  71. Verma S, Varma A, Rexer KH, Hassel A, Kost G, Sarbhoy A, Bisen P, Bütehorn B, Franken P (1998) Piriformospora indica, gen. et sp. nov., a new root-colonizing fungus. Mycologia 90:898–905

    Google Scholar 

  72. Vivas A, Biro B, Ruiz-Lozano JM, Barea JM, Azcon R (2006) Two bacterial strains isolated from a Zn-polluted soil enhance plant growth and mycorrhizal efficiency under Zn toxicity. Chemosphere 52:1523–1533

    Google Scholar 

  73. Walle F, Achatz B, Baltruschat H, Fodor J, Becker K, Fischer M, Heier T, Hückelhoven R, Neumann C, von Wettstein D, Franken P, Kogel KH (2005) The endophytic fungus Piriformospora indica reprograms barley to salt-stress tolerance, disease resistance, and higher yield. Proc Natl Acad Sci U S A 102:13386–13391

    Google Scholar 

  74. Wang Q, Xiong D, Zhao P, Yu X, Tu B, Wang G (2011) Effect of applying an arsenic-resistant and plant growth-promoting rhizobacterium to enhance soil arsenic phytoremediation by Populus eltoides LH05–17. J Appl Microbiol 111:1065–1074

    CAS  PubMed  Google Scholar 

  75. Wani PA, Khan MS, Zaidi A (2007) Impact of heavy metal toxicity on plant growth, symbiosis, seed yield and nitrogen and metal uptake in chickpea. Aus J Exp Agric 47:712–720

    CAS  Google Scholar 

  76. Woodies TC, Hunter GB, Johnson FJ (1977) Statistical studies of matrix effects on the determination of cadmium and lead in fertilizer and material and plant tissue by flame atomic absorption spectrophotometry. Anal Chem Acta 90:127–136

    Google Scholar 

  77. Xu KW, Penttinen P, Chen YX, Chen Q, Zhang XP (2013) Symbiotic efficiency and phylogeny of the rhizobia isolated from Leucaena leucocephala in arid–hot river valley area in Panxi, Sichuan, China. Appl Microbiol Biotechnol 97:83–793

    Google Scholar 

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Acknowledgments

We gratefully acknowledge Dr. Ali Ashraf Mehrabi, Ilam University, Iran, for analyzing the data. We gratefully acknowledge Prof. Ralf Oelmüller, Friedrich Schiller University Jena, Germany, for the critical reading of the manuscript.

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  1. Department of Soil Science, School of Agriculture, Shiraz University, Shiraz, Iran

    Mozhgan Sepehri

  2. Department of Agriculture, Food and Resource Sciences, University of Maryland Eastern Shore, Princess Anne, MD, USA

    Behnam Khatabi

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Sepehri, M., Khatabi, B. Combination of Siderophore-Producing Bacteria and Piriformospora indica Provides an Efficient Approach to Improve Cadmium Tolerance in Alfalfa. Microb Ecol 81, 717–730 (2021). https://doi.org/10.1007/s00248-020-01629-z

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