Advertisement

World Journal of Microbiology and Biotechnology

, Volume 30, Issue 10, pp 2701–2709 | Cite as

Identification of bacteria associated with underground parts of Crocus sativus by 16S rRNA gene targeted metagenomic approach

  • Sheetal Ambardar
  • Naseer Sangwan
  • A. Manjula
  • J. Rajendhran
  • P. Gunasekaran
  • Rup Lal
  • Jyoti VakhluEmail author
Original Paper

Abstract

Saffron (Crocus sativus L), an autumn-flowering perennial sterile plant, reproduces vegetatively by underground corms. Saffron has biannual corm–root cycle that makes it an interesting candidate to study microbial dynamics in its rhizosphere and cormosphere (area under influence of corm). Culture independent 16S rRNA gene metagenomic study of rhizosphere and cormosphere of Saffron during flowering stage revealed presence of 22 genera but none of the genus was common in all the three samples. Bulk soil bacterial community was represented by 13 genera with Acidobacteria being dominant. In rhizosphere, out of eight different genera identified, Pseudomonas was the most dominant genus. Cormosphere bacteria comprised of six different genera, dominated by the genus Pantoea. This study revealed that the bacterial composition of all the three samples is significantly different (P < 0.05) from each other. This is the first report on the identification of bacteria associated with rhizosphere, cormosphere and bulk soil of Saffron, using cultivation independent 16S rRNA gene targeted metagenomic approach.

Keywords

Saffron Rhizosphere Metagenomics 16S rRNA gene Bacterial community 

Notes

Acknowledgments

Authors are grateful to Prof. Michel Aragno, Honorary professor University of Neuchatel, Switzerland for his scientific advice. SA is thankful to CSIR-UGC for Fellowship. We are also thankful to Mr Farooq Ahmad Joo and Mr C.L. Bhat (State Agriculture Department Government of J&K, India), for their help in sample collection and for sharing valuable inputs about Saffron cultivation in Kashmir valley.

References

  1. Alstrom S, Gerhardson B (1987) Charracterisation of a Serratia plymuthica isolate from plant rhizospheres. Plant Soil 103(2):185–189CrossRefGoogle Scholar
  2. Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search program. Nucleic Acid Res 25:3389–3402CrossRefGoogle Scholar
  3. Ambardar S, Vakhlu J (2013) Plant growth promoting bacteia from Crocus sativus. World J Microbiol Biotechnol 29(12):2271–2279CrossRefGoogle Scholar
  4. Amman RL, Ludwig W, Schleifer KH (1995) Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol Rev 59:143–169Google Scholar
  5. Andersen SM, Johnsen K, Sørensen J, Nielsen P, Jacobsen CS (2000) Pseudomonas frederiksbergensissp. nov., isolated from soil at a coal gasification site Int. J Syst Evol Microbiol 50:1957–1964CrossRefGoogle Scholar
  6. Araujo JF, de Castro AP, Costa MMC, Togawa RC, Pappas Júnior GJ, Quirino BF, Bustamante MMC, Williamson L, Handelsman J, Krüger RH (2012) Characterization of soil bacterial assemblies in brazilian savanna-like vegetation reveals acidobacteria dominance. Microb Ecol. doi: 10.1007/s00248-012-0057-3
  7. Arjun JK, Kumarapillai H (2011) Metagenomic analysis of bacterial diversity in the rice rhizosphere soil microbiome. Biotechnol Bioinf Bioeng 1(3):361–367Google Scholar
  8. Ashrafuzzaman M, Hossen FA, Ismail MR, Hoque MA, Islam MZ, Shahidullah SM, Meon S (2009) Efficiency of plant growth-promoting rhizobacteria (PGPR) for the enhancement of rice growth. Afr J Biotechnol 8(7):1247–1252Google Scholar
  9. Berendsen RL, Pieterse CMJ, Bakker PAHM (2012) The rhizosphere microbiome and plant health. Trends Plant Sci 17(8):1360–1385Google Scholar
  10. Brady SF (2007) Construction of soil environmental DNA cosmid libraries and screening for clones that produce biologically active small molecules. Nat Protoc 2(5):1297–1305CrossRefGoogle Scholar
  11. Brady CL, Venter SN, Cleenwerck I, Engelbeen K, Vancanneyt M, Swings J, Coutinho TA (2009) Pantoea vagans sp. nov., Pantoea eucalypti sp. nov., Pantoea deleyi sp. nov. and Pantoea anthophila sp. nov. Int J Syst Evol Microbiol 59:2339–2345CrossRefGoogle Scholar
  12. Buée M, De Boer W, Martin F, van Overbeek L, Jurkevitch E (2009) The rhizosphere zoo: an overview of plant-associated communities of microorganisms, including phages, bacteria, archaea, and fungi, and of some of their structuring factors. Plant Soil 321:189–212CrossRefGoogle Scholar
  13. Chao A, Bunge J (2002) Estimating the number of species in a stochastic abundance model. Biometrics 58:531–539CrossRefGoogle Scholar
  14. Chryssanthi DG, Dedes PG, Karamanos NK, Cordopatis P, Lamari FN (2011) Crocetin inhibits invasiveness of MD-MB-231 breast cancer cells via downregulation of matrix mettaloproteinases. Planta Medica 77(2):146–151Google Scholar
  15. Cohen SN, Chang ACY, Leslie HSU (1972) Nonchromosomal antibiotic resistance in bacteria: genetic transformation of escherichia coli by r-factor DNA. Proc Nat Acad Sci USA 69(8):2110–2114CrossRefGoogle Scholar
  16. Elena P, Ivanova EP, Christen R, Bizet C, Clermont D, Motreff L, Bouchier C, Zhukova NV, Crawford RJ, Kiprianova EA (2009) Pseudomonas brassicacearum subsp. neoaurantiaca subsp. nov., orange-pigmented bacteria isolated from soil and the rhizosphere of agricultural plants. Int J Syst Evol Microbiol 59:2476–2481CrossRefGoogle Scholar
  17. Esmaeili N, Ebrahimzadeh H, Abdi K, Safarian S (2011) Determination of some phenolic compounds in Crocus sativus L. corms and its antioxidant activities study. Pharm Mag 7(25):74–80CrossRefGoogle Scholar
  18. Esmaeili N, Ebrahimzadeh H, Abdi K, Mirmasoumi M, Lamei N, Shamami MA (2013) Determination of metal content in Crocus sativus L. corms in dormancy and waking stages. Iran J Pharm Res 12(1):31–36Google Scholar
  19. Frankova L (2006) Colchicum autumnale L.: an ancient medicinal plant and its hysteranthousgeophytic life strategy. www.fyziologia.sav.sk/geophyte-colchicum
  20. Garrido JFA, Lugo DM, Rodríguez CH, Cortes GT, Millán V, Toro N, Abarca FM, Ramírez-Saad HC (2012) Bacterial community structure in the rhizosphere of three cactus species from semi-arid highlands in central Mexico. Antonie van Leeuwenhoek. doi: 10.1007/s10482-012-9705-3
  21. George IF, Hartmann M, Liles MR, Agathos SN (2011) Recovery of as-yet-uncultured soil Acidobacteriaon dilute solid media. Appl Environ Microbiol 77(22):8184–8188CrossRefGoogle Scholar
  22. Gottel NR, Castro HF, Kerley M, Yang Z, Pelletier DA, Podar M, Karpinets T, Uberbacher Ed, Tuskan GA, Vilgalys R, Doktycz MJ, Schadt CW (2011) Distinct microbial communities within the endosphere and rhizosphere of Populusdeltoides Roots across contrasting soil types. Appl Environ Microbiol 77(17):5934–5944 Google Scholar
  23. Haining M, Hua Y, Tu C, Yuan L, Wei P (2012) Analysis of monosaccharides in the saffron corm glycoconjugate by capillary electrophoresis. Chin J Chromatogr 30(3):304–308Google Scholar
  24. Hall TA (1999) Bioedit: a user friendly biological sequence alignment editor and analysis program for windows 95/98/NT. Nucleic Axid Symposium series No 41 95–98Google Scholar
  25. Hamady M, Lozupone C, Knight R (2010) Fast UniFrac: facilitating high-throughput phylogenetic analyses of microbial communities including analysis of pyrosequencing and PhyloChip data. ISME J 4(1):17–27CrossRefGoogle Scholar
  26. Hamza MA (2008) Understanding soil analysis data. Resource Management Technical Report 327, Western Australian Agriculture AuthorityGoogle Scholar
  27. Handelsman J (2004) Metagenomics: application of genomics to uncultured microorganisms. Microbiol Mol Boil Rev 68(4):669–685CrossRefGoogle Scholar
  28. Hillis DM, Bull JJ (1993) An empirical test of bootstrapping as a method for assessing confidence in phylogenetic analysis. Syst Biol 42(2):182–192CrossRefGoogle Scholar
  29. Hiltner L (1904) Überneuere Erfahrungen und Probleme auf dem Gebiete der Bodenbakteriologieunterbesonderer Berücksichtigung der Gründüngung und Brache. Arbeiten der Deutschen Landwirtschaftlichen Gesellschaft 98:59–78Google Scholar
  30. Hong S, Bunge J, Leslin C, Jeon S, Epstein SS (2009) Polymerase chain reaction primers miss half of rRNA microbial diversity. ISME J 3:1365–1373CrossRefGoogle Scholar
  31. Hultberg M, Bengtsson T, Liljeroth E (2010) Late blight on potato is suppressed by the biosurfactant-producing strain Pseudomonas koreensis 2.74 and its biosurfactant. Bio Control 55:543–550Google Scholar
  32. Inceoglu O, Al-Soud WA, Salles JF, Semenov AV, van Elsas JD (2011) Comparative analysis of bacterial communities in a potato field as determined by pyrosequencing. PLoS One 6(8):e23321. doi: 10.1371/journal.pone.0023321 CrossRefGoogle Scholar
  33. Johansen A, Olsson S (2005) Using phospholipid fatty acid technique to study short-term effects of the biological control agent PseudomonasfluorescensDR54 on the microbial microbiota in barley rhizosphere. Microb Ecol 49:272–281CrossRefGoogle Scholar
  34. Joshi P, Bhatt AB (2011) Diversity and function of plant growth promoting rhizobacteria associated with wheat rhizosphere in North Himalayan region. Int J Environ Sci 1(6):1135–1143Google Scholar
  35. Kamalipour M, Akhondzadeh S (2011) Cardiovascular effects of saffron: an evidence-based review. J TehUniv Heart Ctr 6(2):59–61Google Scholar
  36. Kim BK, Chung J, Kim SY, Jeong H, Kang SG, Kwon SK, Lee CH, Song JY, Yu DS, Ryu CM, Kim JF (2012) Genome sequence of the leaf-colonizing Bacterium Bacillus sp. strain 5B6, isolated from a cherry tree. J Bacteriol 194(14):3758–3759CrossRefGoogle Scholar
  37. Kirk JL, Beaudette LA, Hart M, Moutoglis P, Klironomos JN, Lee H, Trevors JT (2004) Method of studying soil microbial diversity. J Microbiol Method 58:169–188CrossRefGoogle Scholar
  38. Kumar K, Amaresan N, Bhagat S, Madhuri K, Srivastava RC (2010) Isolation and characterization of rhizobacteria associated with coastal agricultural ecosystem of rhizosphere soils of cultivated vegetable crops. doi: 10.1007/s11274-010-0616-z
  39. Leveau JHJ (2007) The magic and menace of metagenomics: prospects for the study of plant growth-promoting rhizobacteria. Eur J Plant Pathol 119:279–300CrossRefGoogle Scholar
  40. Luster J, Göttlein A, Nowack B, Sarret G (2009) Sampling, defining, characterising and modeling the rhizosphere—the soil science tool box. Plant Soil 321:457–482CrossRefGoogle Scholar
  41. Ma A, Lv D, Zhuang X, Zhuang G (2013) Quorum quenching in culturablephyllosphere bacteria from tobacco. Int J Mol Sci 14:14607–14619CrossRefGoogle Scholar
  42. Mahaffee WF, Kloepper JW (1997) Temporal changes in the bacterial communities of soil, rhizosphere and endorhiza associated with field-grown cucumber (CucumissativusL.). Microb Ecol 34:210–223CrossRefGoogle Scholar
  43. Mazumdar T, Goswami C, Talukdar NC (2007) Characterization and screening of beneficial bacteria obtained on King’s B agar from tea rhizosphere. Indian J Biotechnol 6:490–494Google Scholar
  44. Mehta P, Chauhan A, Mahajan R, Mahajan PK, Shirkot CK (2010) Strain of Bacillus circulans isolated from apple rhizosphere showing plant growth promoting potential. Curr Sci 98(4):538–542Google Scholar
  45. Melnyk JP, Wang S, Marcone MF (2010) Chemical and biological properties of the world’s most expensive spice: saffron. Food Res Int 43:1981–1989CrossRefGoogle Scholar
  46. Mishra A, Chauhan PS, Chaudhry V, Tripathi M, Nautiyal CS (2011) Rhizosphere competent Pantoeaagglomerans enhances maize (Zea mays) and chickpea (Cicerarietinum L.) growth, without altering the rhizosphere functional diversity. Antonie van Leeuwenhoek 100:405–413CrossRefGoogle Scholar
  47. Nehvi FA, Yasmin S (2010) Saffron farming in India the Kashmir connection. Financ Agric 42(5):9–15Google Scholar
  48. Okamoto H, Sat M, Miyat Y, Yoshikawa M, Isaka M (2000) Biocontrol of root rot of angelica trees by enterobacter cloacae and Serratiaficaria strains. J Gen Plant Pathol 66:86–94CrossRefGoogle Scholar
  49. Pang MF, Abdullah N, Lee CW, Ng C–C (2008) Isolation of high molecular weight dna from forest topsoilfor metagenomic analysis. Asia Pacific J Mol Biol Biotechnol 16(2):35–41Google Scholar
  50. Peiffer JA, Spor A, Koren O, Jin Z, Tringe SG, Dangl JL, Buckler ES, Ley RE (2013) Diversity and heritability of the maize rhizosphere microbiome under field conditions. PNAS 110(16):6548–6553CrossRefGoogle Scholar
  51. Rahmani A, Seighali N, Ebrahimzadeh H, Zarei JH (2012) Partial purification of peroxidase in corms of Saffron (Crocus sativus L) during dormancy and waking. New Cell Mol Biotechnol J 2(8):95–99CrossRefGoogle Scholar
  52. Riesenfeld CS, Schloss PD, Handelsman J (2004) Metagenomics: genomic analysis of microbial communities. Annu Rev Genet 38:525–552CrossRefGoogle Scholar
  53. Sachdeva D, Nemab P, Dhakephalkarb P, Zinjardec S, Chopadea B (2010) Assessment of 16S rRNA gene-based phylogenetic diversity and promising plant growth-promoting traits of Acinetobacter community from the rhizosphere of wheat. Microbiol Res 165:627–638CrossRefGoogle Scholar
  54. Saharan BS, Nehra V (2011) Plant Growth Promoting rhizobacteria: a Critical Review. Life Sci Med Res 21:1–29Google Scholar
  55. Shannon CE, Weaver W (1949) The mathematical theory of communication. University of Illinois Press, UrbanaGoogle Scholar
  56. Sharaf-Eldin M, Elkholy S, Fernandez J, Junge H, Cheetham R, Guardiola J, Weathers P (2008) Bacillus subtillis FZB24 affects quantity and quality of saffron (Crocus sativus L.). Planta Medica 74:1316–1320Google Scholar
  57. Shoebitz M, Ribaudo CM, Pardo MA, Cantorec ML, Ciampi L, Cura´b JA (2009) Plant growth promoting properties of a strain of Enterobacterludwigii isolated from Loliumperenne rhizosphere. Soil Biol Biochem 41:1768–1774CrossRefGoogle Scholar
  58. Singh BK, Munro S, Potts JM, Millard P (2007) Influence of grass species and soil type on rhizosphere microbial community structure in grassland soils. Appl Soil Eco 36:147–155CrossRefGoogle Scholar
  59. Singh HP, Uma S, Selvarajan R, Karihaloo JL (2011) Micropropagation for production of quality banana planting material in Asia-Pacific. Asia-Pacific Consortium on Agricultural Biotechnology (APCoAB), New Delhi, India, p 92Google Scholar
  60. Steinitz B, Cohen A, Goldberg Z, Kochba M (1991) Precocious gladiolus corm formation in liquid shake cultures. Plant Cell Tiss Org 26(2):63–70CrossRefGoogle Scholar
  61. Sturz AV, Nowak J (2000) Endophytic communities of rhizobacteria and the strategies required to create yield enhancing associations with crops. Appl Soil Eco 15:183–190CrossRefGoogle Scholar
  62. Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. MolBiolEvol 28:2731–2739Google Scholar
  63. Teixeira LCRS, Peixoto RS, Cury JC, Sul WJ, Pellizari VH, Tiedje J, Rosado AS (2010) Bacterial diversity in rhizosphere soil from Antarctic vascular plants of Admiralty Bay, maritime Antarctica. ISME J 4:989–1001CrossRefGoogle Scholar
  64. Thomas T, Gilbert J, Meyer F (2012) Metagenomics—a guide from sampling to data analysis. Microb Inform Exp 2:3CrossRefGoogle Scholar
  65. Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG (1997) The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res 25(24):4876–4882CrossRefGoogle Scholar
  66. Tuimala J (2004) A primer to phylogenetic analysis using Phylip package, 2nd edn. Center for Scientific Computing, EspooGoogle Scholar
  67. Tyson GW, Chapman J, Hugenholtz P, Allen EE, Ram RJ, Richardson PM, Solovyev VV, Rubin EM, Rokhsar DS, Banfield JF (2004) Community structure and metabolism through reconstruction of microbial genomes from the environment. Nature 428(4):37–43CrossRefGoogle Scholar
  68. Venter JC, Remington K, Heidelberg JF, Halpern AL, Rusch D (2004) Environmental genome shotgun sequencing of the sargasso sea. Science 304:66–74CrossRefGoogle Scholar
  69. Wahyudi AT, Astuti RP, Widyawati A, Meryandini A, Nawangsih AA (2011) Characterization of Bacillus sp strains isolated from rhizosphere of soybean plants for their use as potential plant growth for promoting Rhizobacteria. J Microbiol Antimicrobials 3(2):34–40Google Scholar
  70. Wani BA, Hamza AKH, Mohiddin FA (2011) Saffron: a repository of medicinal properties. J Med Plants Res 5(11):2131–2135Google Scholar
  71. Wechter P, Williamson J, Robertson A, Kluepfel D (2003) A rapid, cost-effective procedure for the extraction of microbial DNA from soil.World. J Microbiol Biotechnol 19:85–91CrossRefGoogle Scholar
  72. Zhou J, Bruns MA, Tiedje JM (1996) DNA recovery from soils of diverse composition. Appl Environ Microbiol 62:316–322Google Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2014

Authors and Affiliations

  • Sheetal Ambardar
    • 1
  • Naseer Sangwan
    • 2
  • A. Manjula
    • 3
  • J. Rajendhran
    • 3
  • P. Gunasekaran
    • 4
  • Rup Lal
    • 2
  • Jyoti Vakhlu
    • 1
    Email author
  1. 1.School of BiotechnologyUniversity of JammuJammuIndia
  2. 2.Department of ZoologyUniversity of DelhiDelhiIndia
  3. 3.Department of GeneticsMadurai Kamraj UniversityMaduraiIndia
  4. 4.Thiruvalluvar UniversityVelloreIndia

Personalised recommendations