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Ochrobactrum ciceri mediated induction of defence genes and antifungal metabolites enhance the biocontrol efficacy for the management of Botrytis leaf blight of Lilium under protected conditions

  • Rajendran Priyanka
  • Sevugapperumal NakkeeranEmail author
Original Article
  • 29 Downloads

Abstract

Biological control with bacterial bioagents is a cost-effective method for the management of foliar diseases of cut flowers under protected conditions. The antagonistic microflora in Lilium ecosystem was exploited and its potential against Botrytis cinerea was assessed for its management. However, the bacterial antagonist Ochrobactrum ciceri has not been explored for management of B. cinerea, a pathogen causing leaf blight of Lilium. In the present study, 42 bacterial antagonists were tested for their antifungal activity against B. cinerea. Among them, the growth of B. cinerea was suppressed up to 46% by O. ciceri (MM17) in vitro. GC/MS analysis of crude metabolites of O. ciceri (MM17) co-cultured with cell wall of B. cinerea produced four antifungal non-volatile metabolites when compared with the solely cultured bacterium. Similarly, gas chromatography/mass spectometry-thermal desorption (GC/MS-TD) analysis of the volatile metabolites of O. ciceri (MM17) indicated that the bacterium produced growth promoting compounds upon interaction with the cell wall of B. cinerea apart from the antibacterial compounds. However, no growth promoting compounds were produced when the bacterium was cultured separately. Further, qRT-PCR analysis revealed an increase in the expression profile of PAL (34.49 folds), PR 10 (4.02 folds) and ascorbate peroxidase (APX) (15.55 folds) transcripts when treated with O. ciceri (MM17), challenged against B. cinerea (SEL). Further, the efficacy of antagonist bacterial strains was assessed for the management of Botrytis leaf blight under protected conditions. Foliar application of O. ciceri (MM17) under protected conditions suppressed leaf blight by 77% and increased the stem yield. This study highlights the potential of O. ciceri (MM17) for the management of Lilium leaf blight under protected cultivation.

Keywords

Lilium Botrytis Ochrobactrum Volatile compounds Non-volatile compounds Defence genes 

Notes

Acknowledgements

The authors would like to give special thanks for the encouragement provided by Dr. V. G. Malathi, Dr. P. Renukadevi and Dr. T. C. K. Sugitha. The support provided by Professor and Head, Department of Plant Pathology and Dr. U. Sivakumar, The Dean, School of Post Graduate studies, Tamil Nadu Agricultural University are deeply acknowledged. The authors would also like to acknowledge DST FIST, UGC-SAP and ICAR for funding.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Statement of human and animal rights

This article does not contain any studies with human or animal subjects performed by the any of the authors.

Supplementary material

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ESM 1

Phenotypic characters of fifteen B. cinerea strains (PNG 1780 kb)

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High Resolution Image (TIF 575 kb)
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ESM 2 (DOC 46 kb)
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ESM 3 (DOC 36 kb)
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ESM 4 PCR amplification of Botrytis cinerea strains. M – 100 bp ladder, Lane 1 – PB, Lane 2 – BSl, Lane 3 – AR, Lane 4 – SEL, Lane 5 – SOR, Lane 6 – AC1, Lane 7 – ER, Lane 8 – SIB, Lane 9 – AC2, Lane 10 –JP, Lane 11 – SEV, Lane 12 – NV, Lane 13 – BR, Lane 14 – BO, Lane 15 – HN, Lane P – Positive control (JPG 447 kb)
42161_2018_212_MOESM5_ESM.jpg (653 kb)
ESM 5 PCR amplification of 16S rRNA of bacterial strains Lane L – 100 bp Ladder, Lane1 – MM1, Lane2 – MM 2, Lane 3- MM 3, Lane 4 – MM 4, Lane 5 – MM 5, Lane 6 – MM 6, Lane 7 – MM 7, Lane 8- MM 8, Lane 9 – MM 9, Lane10 – MM 10, Lane11 – MM 11, Lane 12- MM 12, Lane 13 – MM 13, Lane 14 – MM 14, Lane 15 – MM15, Lane16 – MM 16, Lane17 – MM17, Lane 18- MM 18, Lane 19 – MM 19, Lane 20 – MM 20, Lane 21 – MM 21 (JPG 652 kb)
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ESM 6 Suppression of mycelial growth of B. cinerea by the bacterial strains through dual culture plate test (JPG 4409 kb)
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ESM 7 Suppression of mycelial growth of B. cinerea by the bacterial strains using spot inoculation method. (JPG 1174 kb)
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ESM 8 (DOC 35 kb)
42161_2018_212_MOESM9_ESM.doc (42 kb)
ESM 9 (DOC 42 kb)
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ESM 10 (DOC 40 kb)
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ESM 11 (DOC 45 kb)
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ESM 12 Efficacy of bacterial strains on the suppression Lilium leaf blight and plant growth promotion under protected cultivation. (JPG 346 kb)
42161_2018_212_MOESM13_ESM.doc (42 kb)
ESM 13 (DOC 42 kb)

References

  1. Akram W, Anjum T (2011) Quantitative changes in defence system of tomato induced by two strains of Bacillus against Fusarium wilt. Indian Journal of Fundamental and Applied Life Sciences  1:7–13Google Scholar
  2. Al-Tameme HJ, Hadi MY, Hameed IH (2015) Phytochemical analysis of Urtica dioica leaves by fourier-transform infrared spectroscopy and gas chromatography-mass spectrometry. J Pharmacogn Phytother 7(10):238–252CrossRefGoogle Scholar
  3. Angelini RM, Rotolo C, Masiello M, Gerin D, Pollastro S, Faretra F (2014) Occurrence of fungicide resistance in populations of Botryotinia fuckeliana (Botrytis cinerea) on table grape and strawberry in southern Italy. Pest Manag Sci 70:1785–1796.  https://doi.org/10.1002/ps.3711 CrossRefGoogle Scholar
  4. Cano RJ, Borucki MK, Higby-Schweitzer M, Poinar HN, Poinar GO, Pollard KJ (1994) Bacillus DNA in fossil bees: an ancient symbiosis. Appl Environ Microbiol 60:2164–2167Google Scholar
  5. Cavalcanti ÉBVS (2014) Estudo fitoquímico e biológico dos frutos e raízes de Piper caldense C. DC. (Piperaceae). Universidade Federal da Paraíba, João PessoaGoogle Scholar
  6. Chadha P, Das RH (2006) A pathogenesis related protein, AhPR10 from peanut: an insight of its mode of antifungal activity. Planta 225:213–222CrossRefGoogle Scholar
  7. Chaiharn M, Chunhaleuchanon S, Lumyong S (2009) Screening siderophore producing bacteria as potential biological control agent for fungal rice pathogens in Thailand. World J Microbiol Biotechnol 25:1919–1928CrossRefGoogle Scholar
  8. Chakraborty U, Chakraborty BN, Basnet M, Chakraborty AP (2009) Evaluation of Ochrobactrum anthropi TRS-2 and its talc based formulation for enhancement of growth of tea plants and management of brown root rot disease. J Appl Microbiol 107(2):625–634CrossRefGoogle Scholar
  9. Chakraborty BN, Chakraborty U, Saha A, Dey PL, Sunar K (2010) Molecular characterization of Trichoderma viride and Trichoderma harzianum isolated from soils of North Bengal based on rDNA markers and analysis of their PCR-RAPD profiles. Global Journal Of Biotechnology & Biochemistry 5:55–61Google Scholar
  10. Chen LJ, Yin YY, Sun SK, Sun J (2017) First report of a gray mold on Lilium cernuum Komar leaves caused by Botrytis cinerea in Liaoning province of China. J Plant Pathol 99(1):301Google Scholar
  11. Chilvers MI, Du Toit LJ (2006) Detection and identification of Botrytis species associated with neck rot, scape blight, and umbel blight of onion. Plant Health Prog. 113Google Scholar
  12. Chiou AL, Wu WS (2003) Formulation of Bacillus amyloliquefaciens B190 for control of lily grey mould (Botrytis elliptica). J Phytopathol 151:13–18CrossRefGoogle Scholar
  13. Datar VV, Mayee CD (1981) Assessment of loss in tomato yield due to early blight. Indian Phytopathol 34:191–195Google Scholar
  14. Dennis C, Webster J (1971) Antagonistic properties of species group of Trichoderma production of non-volatile antibiotics. Trans Br Mycol Soc 57:25–39CrossRefGoogle Scholar
  15. Dheepa R, Vinodkumar S, Renukadevi P, Nakkeeran S (2016) Phenotypic and molecular characterization of chrysanthemum white rust pathogen Puccinia horiana (Henn) and the effect of liquid based formulation of Bacillus spp. for the management of chrysanthemum white rust under protected cultivation. Biol Control 103:172–186.  https://doi.org/10.1016/j.biocontrol.2016.09.006 CrossRefGoogle Scholar
  16. Foyer CH, Noctor G (2005) Redox homeostasis and antioxidant signaling: a metabolic interface between stress perception and physiological responses. Plant Cell 17:1866–1875CrossRefGoogle Scholar
  17. Gomez KA, Gomez AA (1984) Statistical procedures for agricultural research, 2 Edn. JohnWiley and Sons, New York, NYGoogle Scholar
  18. Guinebretiere MH, Berge O, Normand P, Morris C, Carlin F (2001) Identification of bacteria in pasteurized zucchini purées stored at different temperatures and comparison with those found in other pasteurized vegetable purées. Appl Environ Microbiol 67:4520–4530CrossRefGoogle Scholar
  19. Gupta D, Kumar M (2017) Evaluation of in vitro antimicrobial potential and GC–MS analysis of Camellia sinensis and Terminalia arjuna. Biotechnol Rep (Amst) 13:19–25Google Scholar
  20. Hahm MS, Sumayo M, Hwang YJ, Jeon SA, Park SJ, Lee JY, Ahn JH, Kim BS, Ryu CM, Ghim SY (2012) Biological control and plant growth promoting capacity of rhizobacteria on pepper under greenhouse and field conditions. J Microbiol 50:380–385CrossRefGoogle Scholar
  21. Ham MS, Park YM, Sung HR, Sumayo M, Ryu CM, Park SH, Ghim SY (2009) Characterization of rhizobacteria isolated from family Solanaceae plants in Dokdo island. Korean J Microbiol Biotechnol 37:110–117Google Scholar
  22. Hassan MN, Afghan S, ul HZ, Hafeez FY (2014) Biopesticide activity of sugarcane associated rhizobacteria: Ochrobactrum intermedium strain NH-5 and Stenotrophomonas maltophilia strain NH-300 against red rot under field conditions. Phytopathol Mediterr:229–239Google Scholar
  23. Hou PF, Chen CY (2003) Early stages of infection of lily leaves by Botrytis elliptica and B. cinerea. Plant Pathol Bulletin 12:103–108Google Scholar
  24. Huber B, Scholz HC, Kämpfer P, Falsen E, Langer S, Busse HJ (2010) Ochrobactrum pituitosum sp. nov., isolated from an industrial environment. Int J Syst Evol Microbiol 60:321–326CrossRefGoogle Scholar
  25. Jayaraj J, Anand A, Muthukrishnan S (2004) Pathogenesis- related proteins and their roles in resistance to fungal pathogens. In: Punja ZK (ed) Fungal disease resistance in plants: biochemistry, molecular biology, and genetic engineering. Haworth Press, New York, pp 139–177Google Scholar
  26. Kai M, Haustein M, Molina F, Petri A, Scholz B, Piechulla B (2009) Bacterial volatiles and their action potential. Appl Microbiol Biotechnol 81:1001–1012CrossRefGoogle Scholar
  27. Kämpfer P, Buczolits S, Albrecht A, Busse HJ, Stackebrandt E (2003) Towards a standardized format for the description of a novel species (of an established genus): Ochrobactrum gallinifaecis sp. nov. Int J Syst Evol Microbiol 53:893–896CrossRefGoogle Scholar
  28. Knapp JE, Chandlee JM (1996) RNA/DNA mini-prep from a single sample of orchid tissue. BioTechniques 21:54–56CrossRefGoogle Scholar
  29. Kumar S, Stecher G, Tamura K (2016) MEGA7: molecular evolutionary genetics analysis version 7.0. For bigger datasets. Mol Biol Evol 33:1870–1874.  https://doi.org/10.1093/molbev/msw054 CrossRefGoogle Scholar
  30. Lee S, Hung R, Yap M, Bennett JW (2015) Age matters: the effects of volatile organic compounds emitted by Trichoderma atroviride on plant growth. Arch Microbiol 197:723–727CrossRefGoogle Scholar
  31. Marrez D, Sultan Y (2016) Antifungal activity of the cyanobacterium Microcystis aeruginosa against mycotoxigenic fungi.  Journal of Applied Pharmaceutical Science 6:191–198.  https://doi.org/10.7324/japs.2016.601130
  32. Ngoma L, Mogatlanyane K, Babalola OO (2014) Screening of endophytic bacteria towards the development of cottage industry: an in vitro study. J Hum Ecol 47(1):45–63CrossRefGoogle Scholar
  33. Okwu DE, Ighodaro BU (2009) GC-MS evaluation of the bioactive compounds and antibacterial activity of the oil fraction from the stem barks of Dacryodes edulis (G. Don) H. J Lam. International Journal of Drug Development and Research 1:117–125Google Scholar
  34. Onate-Sanchez L, Vicente-Carbojosa J (2008) DNA-free RNA isolation protocols for Arabidopsis thaliana, including seeds and siliques. BMC Research Notes 1:93.  https://doi.org/10.1186/1756-0500-1-93.
  35. Rao J, Liu D, Zhang N, He H, Ge F, Chen C (2014) Differential gene expression in incompatible interaction between Lilium regale Wilson and Fusarium oxysporum f. sp. lilii revealed by combined SSH and microarray analysis. J Mol Biol 48:802–812CrossRefGoogle Scholar
  36. Rigotti S, Gindro K, Richter H, Viret O (2002) Characterization of molecular markers for specific and sensitive detection of Botrytis cinerea Pers.: Fr. In strawberry (Fragaria ananassa Duch.) using PCR. FEMS Microbiol Lett 209:169–174Google Scholar
  37. Sowndhararajan K, Marimuthu S, Manian S (2013) Biocontrol potential of phylloplane bacterium Ochrobactrum anthropi BMO-111 against blister blight disease of tea. J Appl Microbiol 114(1):209–218CrossRefGoogle Scholar
  38. Srinivasan GV, Sharanappa P, Leela NK, Sadashiva CT, Vijayan KK (2009) Chemical composition and antimicrobial activity of the essential oil of Leea indica (Burm. F.) Merr. Flowers. Indian Journal of Natural Product and Resources 8(5):488–493Google Scholar
  39. Sumayo M, Hahm MS, Ghim SY (2013) Determinants of plant growth-promoting Ochrobactrum lupini KUDC1013 involved in induction of systemic resistance against Pectobacterium carotovorum subsp. carotovorum in tobacco leaves. Plant Pathol J 29(2):174CrossRefGoogle Scholar
  40. Teyssier C, Marchandin H, Jean-Pierre H, Diego I, Darbas H, Jeannot JL, Gouby A, Jumas-Bilak E (2005) Molecular and phenotypic features for identification of the opportunistic pathogens Ochrobactrum spp. J Med Microbiol 54:945–953CrossRefGoogle Scholar
  41. Teyssier C, Marchandin H, Jean-Pierre H, Masnou A, Dusart G, Jumas-Bilak E (2007) Ochrobactrum pseudintermedium sp. nov., a novel member of the family Brucellaceae, isolated from human clinical samples. Int J Syst Evol Microbiol 57:1007–1013CrossRefGoogle Scholar
  42. Van Baarlen P, Woltering EJ, Staats M, Van Kan JA (2007) Histochemical and genetic analysis of host and non-host interactions of Arabidopsis with three Botrytis species: an important role for cell death control. Mol Plant Pathol 8:41–54.  https://doi.org/10.1111/j.1364-3703.2006.00367.x CrossRefGoogle Scholar
  43. Van den Ende JE, Pennock-Vos MG, Bastiaansen C, Koster ATJ, Van der Meer LJ (2000) BoWaS: a weather-based warning system for the control of Botrytis blight in lily. Acta Hortic (519):215–220Google Scholar
  44. Vinodkumar S, Nakkeeran S (2017) Characterization and management of Botrytis cinerea inciting blossom blight of carnation under protected cultivation. J Environ Biol 38:527CrossRefGoogle Scholar
  45. Vinodkumar S, Nakkeeran S, Renukadevi P, Malathi VG (2017) Biocontrol potentials of antimicrobial peptide producing Bacillus species: multifaceted antagonists for the management of stem rot of carnation caused by Sclerotinia sclerotiorum. Front Microbiol 8:446CrossRefGoogle Scholar
  46. Wang Y, Hou YP, Chen CJ, Zhou MG (2014) Detection of resistance in Sclerotinia sclerotiorum to carbendazim and dimethachlon in Jiangsu Province of China. Australas Plant Pathol 43:307–312.  https://doi.org/10.1007/s13313-014-0271-1 CrossRefGoogle Scholar
  47. Weiberg A, Wang M, Lin FM, Zhao H, Zhang Z, Kaloshian I et al (2013) Fungal small RNAs suppress plant immunity by hijacking host RNA interference pathways. Science 342:118–123.  https://doi.org/10.1126/science.1239705 CrossRefGoogle Scholar
  48. White TJ, 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 315–322Google Scholar
  49. Wu XY, Walker MJ, Hornitzky M, Chin J (2006) Development of a group-specific PCR combined with ARDRA for the identification of Bacillus species of environmental significance. J Microbiol Methods 64:107–119CrossRefGoogle Scholar
  50. Xu L, Zhu L, Tu L, Liu L, Yuan D, Jin L, Long L, Zhang X (2011) Lignin metabolism has a central role in the resistance of cotton to the wilt fungus Verticillium dahliae as revealed by NA_Seq_dependent transcriptional analysis and histochemistry. J Exp Bot 62:5607–5621CrossRefGoogle Scholar
  51. Zhang JX, Xue AG (2010) Biocontrol of sclerotinia stem rot (Sclerotinia sclerotiorum) of soybean using novel Bacillus subtilis strain SB24 under control conditions. Plant Pathol 59:382–391.  https://doi.org/10.1111/j.1365-3059.2009.02227.x CrossRefGoogle Scholar
  52. Zou CS, Mo MH, Gu YQ, Zhou JP, Zhang KQ (2007) Possible contributions of volatile-producing bacteria to soil fungistasis. Soil Biol Biochem 39(9):2371–2379CrossRefGoogle Scholar
  53. Zou C, Li Z, Yu D (2010) Bacillus megaterium strain XTBG34 promotes plant growth by producing 2-pentylfuran. J Microbiol 48:460–466CrossRefGoogle Scholar
  54. Zurdo-Pineiro JL, Rivas R, Trujillo ME, Vizcaino N, Carrasco JA, Chamber M, Palomares A, Mateos PF, Martinez-Molina E, Velazquez E (2007) Ochrobactrum cytisi sp. nov., isolated from nodules of Cytisus scoparius in Spain. Int J Syst Evol Microbiol 57:784–788CrossRefGoogle Scholar

Copyright information

© Società Italiana di Patologia Vegetale (S.I.Pa.V.) 2018

Authors and Affiliations

  1. 1.Department of Plant PathologyTamil Nadu Agricultural UniversityCoimbatoreIndia

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