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Molecular characterization and identification of plant growth promoting endophytic bacteria isolated from the root nodules of pea (Pisum sativum L.)

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Abstract

Root nodule accommodates various non-nodulating bacteria at varying densities. Present study was planned to identify and characterize the non-nodulating bacteria from the pea plant. Ten fast growing bacteria were isolated from the root nodules of cultivated pea plants. These bacterial isolates were unable to nodulate pea plants in nodulation assay, which indicate the non-rhizobial nature of these bacteria. Bacterial isolates were tested in vitro for plant growth promoting properties including indole acetic acid (IAA) production, nitrogen fixation, phosphate solubilization, root colonization and biofilm formation. Six isolates were able to produce IAA at varying level from 0.86 to 16.16 μg ml−1, with the isolate MSP9 being most efficient. Only two isolates, MSP2 and MSP10, were able to fix nitrogen. All isolates were able to solubilize inorganic phosphorus ranging from 5.57 to 11.73 μg ml−1, except MSP4. Bacterial isolates showed considerably better potential for colonization on pea roots. Isolates MSP9 and MSP10 were most efficient in biofilm formation on polyvinyl chloride, which indicated their potential to withstand various biotic and abiotic stresses, whereas the remaining isolates showed a very poor biofilm formation ability. The most efficient plant growth promoting agents, MSP9 and MSP10, were phylogenetically identified by 16S rRNA gene sequence analysis as Ochrobactrum and Enterobacter, respectively, with 99 % similarity. It is suggested the potential endophytic bacterial strains, Ochrobactrum sp. MSP9 and Enterobacter sp. MSP10, can be used as biofertilizers for various legume and non-legume crops after studying their interaction with the host crop and field evaluation.

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References

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

    CAS  Google Scholar 

  • Angus A, Hirsch AM (2010) Insights into the history of the legume beta-proteobacterial symbiosis. Mol Ecol 19:28–30

    Google Scholar 

  • Aravind R, Kumar A, Eapen SJ (2012) Pre-plant bacterisation: a strategy for delivery of beneficial endophytic bacteria and production of disease-free plantlets of black pepper (Piper nigrum L.). Arch Phytopathol Plant Prot 45:1115–1126

    Article  Google Scholar 

  • Arnon DI, Hoagland DR (1940) Crop production in artificial culture solution sand in soil with special reference to factors influencing yields and absorption of inorganic nutrient. Soil Sci 50:463–483

    CAS  Google Scholar 

  • Bansal RK (2009) Synergistic effect of Rhizobium, PSB and PGPR on nodulation and grain yield of mung bean. J Food Legum 22:37–39

    Google Scholar 

  • Beijerinck MW, Delden AV (1902) Uber die Assimilation des freien Stickstoffs durch Bakerien. Centralbl Bakt Abt II 9:3–43

    CAS  Google Scholar 

  • Benhizia Y, Benhizia H, Benguedouar A, Muresu R, Giacomini A, Squartini A (2004) Gamma proteobacteria can nodulate legumes of the genus Hedysarum. Syst Appl Microbiol 27:462–468

    Article  CAS  Google Scholar 

  • Bhattacharyya PN, Jha DK (2012) Plant growth-promoting rhizobacteria (PGPR): emergence in agriculture. World J Microbiol Biotechnol 28:1327–1350

    Article  CAS  Google Scholar 

  • Birnboim HC, Doly A (1979) A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res 7:1513–1523

    Article  CAS  Google Scholar 

  • Dong Y, Iniguez LA, Ahmer BMM (2003) Kinetics and strain specificity of rhizosphere and endophytic colonization by enteric bacteria on seedlings of Medicago sativa and Medicago truncatula. Appl Environ Microbiol 69:1783–1790

    Article  CAS  Google Scholar 

  • Duangpaeng A, Phetcharat P, Chanthapho S, Boonkantong N, Okuda N (2012) The study and development of endophytic bacteria for enhancing organic rice growth. Procedia Eng 32:172–176

    Article  CAS  Google Scholar 

  • Ekin Z (2010) Performance of phosphate solubilizing bacteria for improving growth and yield of sunflower (Helianthus annuus L.) in the presence of phosphorus fertilizer. Afr J Biotechnol 9:3794–3800

    CAS  Google Scholar 

  • Fabre F, Planchon C (2000) Nitrogen nutrition, yield and protein content in soybean. Plant Sci 152:51–58

    Article  CAS  Google Scholar 

  • Fedorova E, Redondo FJ, Koshiba T, de Felipe MR, Pueyo JJ, Lucas MM (2005) Aldehyde oxidase (AO) in the root nodules of Lupinus albus and Medicago truncatula: identification of AO in meristematic and infection zones. Mol Plant Microbe Interact 18:405–413

    Article  CAS  Google Scholar 

  • Fraile B, Paniagua R, Rodriguez MC (1988) Long day photoperiods and temperature of 20 °C induce spermatogenesis in blinded and non-blinded marbled newts during the period of testicular quiescence. Biol Reprod 39:649–655

    Article  CAS  Google Scholar 

  • Fujishige NA, Kapadia NN, De Hoff PL, Hirsch AM (2006) Investigations of Rhizobium biofilm formation. FEMS Microbiol Ecol 56:195–206

    Article  CAS  Google Scholar 

  • Garbeva P, Overbeek LS, Vuurde JW (2001) Analysis of endophytic bacterial communities of potato by plating and denaturing gradient gel electrophoresis (DGGE) of 16S rDNA based PCR fragments. Microb Ecol 41:369–383

    CAS  Google Scholar 

  • Gordon SA, Weber RP (1951) Calorimetric estimation of indoleacetic acid. Plant Physiol 26:192–195

    Article  CAS  Google Scholar 

  • Gough C, Vasse J, Galera C, Webster G, Cocking E, Denarie J (1997) Interactions between bacterial diazotrophs and non-legume dicots: Arabidopsis thaliana as a model plant. Plant Soil 194:123–130

    Article  CAS  Google Scholar 

  • Hameed S, Yasmin S, Malik KA, Zafar Y, Hafeez FY (2004) Rhizobium, Bradyrhizobium and Agrobacterium strains isolated from cultivated legumes. Biol Fertil Soils 39:179–185

    Article  Google Scholar 

  • Hardy RWF, Holsten RD, Jackson EK, Burns RE (1968) The acetylene-ethylene assay for nitrogen fixation: laboratory and field evaluation. Plant Physiol 43:1185–1207

    Article  CAS  Google Scholar 

  • Hirsch AM (2010) How rhizobia survive in the absence of a legume host, a stressful world indeed. In: Seckbach J, Grube M (eds) Symbiosis and stress: cellular origin, life in extreme habitats and astrobiology. Springer 17, pp 375–391

  • Hirsch AM (2010b) Insights into the history of the legume beta-proteobacterial symbiosis. Mol Ecol 19:28–30

    Article  Google Scholar 

  • Ibanez F, Angelini J, Taurian T, Tonelli ML, Fabra A (2009) Endophytic occupation of peanut root nodules by opportunistic Gammaproteobacteria. Syst Appl Microbiol 32:49–55

    Article  CAS  Google Scholar 

  • Kobayashi DY, Palumbo JD (2000) Bacterial endophytes and their effects on plants and uses in agriculture. In: James CW, White JF (eds) Microbial endophytes. Marcel Dekker Inc, New York, pp 199–233

    Google Scholar 

  • Ma W, Guinel FC, Glick BR (2003) Rhizobium leguminosarum biovar viciae 1-aminocyclopropane-1-carboxylate deaminase promotes nodulation of pea plants. Appl Environ Microbiol 69:4396–4402

    Article  CAS  Google Scholar 

  • Marsudi NDS, Glenn AR, Dilworth MJ (1999) Identification and characterization of fast- and slow-growing root nodule bacteria from South-Western Australian soils able to nodulate Acacia saligna. Soil Biol Biochem 31:1229–1238

    Article  CAS  Google Scholar 

  • Mishra PK, Mishra S, Selvakumar G, Bisht JK, Kundu S, Gupta HS (2009) Coinoculation of Bacillus thuringeinsis-KR1 with Rhizobium leguminosarum enhances plant growth and nodulation of pea (Pisum sativum L.) and lentil (Lens culinaris L.). World J Microbiol Biotechnol 25:753–761

    Article  Google Scholar 

  • Nair A, Juwarkar A, Singh S (2007) Production and characterization of siderophores and its application in arsenic removal from contaminated soil. Water Air Soil Pollut 180:199–212

    Article  CAS  Google Scholar 

  • Okan YS, Albercht L, Burris RH (1977) Methods of growing Spirillum lipoferum and for counting the pure culture in association and with plant. Appl Environ Microbiol 33:85–88

    Google Scholar 

  • Patel HA, Patel RK, Khristi SM, Parikh K, Rajendran G (2012) Isolation and characterization of bacterial endophytes from Lycopersicon esculentum plant and their plant growth promoting characteristics. Nepal J Biotechnol 2:37–52

    Article  Google Scholar 

  • Patten CL, Glick BR (2002) Role of Pseudomonas putida indole acetic acid in development of the host plant root system. Appl Environ Microbiol 68:3795–3801

    Article  CAS  Google Scholar 

  • Phetcharat P, Duangpaeng A (2012) Screening of endophytic bacteria from organic rice tissue for indole acetic acid production. Procedia Eng 32:177–183

    Article  Google Scholar 

  • Pikovskaya R (1948) Mobilization of P in soil in connection with vital activity by some microbial species. Microbiologica 17:362–370

    CAS  Google Scholar 

  • Selvakumar G, Kundu S, Gupta AD, Shouche YS, Gupta HS (2008) Isolation and characterization of nonrhizobial plant growth promoting bacteria from nodules of Kudzu (Pueraria thunbergiana) and their effect on wheat seedling growth. Curr Microbiol 56:134–139

    Article  CAS  Google Scholar 

  • Seneviratne G, Weerasekara MLMAW, Seneviratne KACN, Zavahir JS, Kecskes ML, Kennedy IR (2011) Importance of biofilm formation in plant growth promoting rhizobacterial action. Microbiol Monogr 18:81–95

    Article  Google Scholar 

  • Steel RGD, Torrie JH (1980) Analysis of covariance. In: Principles and procedures of statistics: a biometrical approach. McGraw-Hill, New York

  • Sturz AV, Christie BR, Matheson BG, Nowak J (1997) Biodiversity of endophytic bacteria which colonize red clover nodules, roots, stems and foliage and their influence on host growth. Biol Fertil Soils 25:13–19

    Article  Google Scholar 

  • Tan Z, Xu X, Wang E, Gao J, Martinez-Romero E, Chen W (1997) Phylogenetic and genetic relationships of Mesorhizobium tianshanense and related rhizobia. Int J Syst Bacteriol 47:874–879

    Article  CAS  Google Scholar 

  • Tariq M, Hameed S, Yasmeen T, Ali A (2012) Non-rhizobial bacteria for improved nodulation and grain yield of mung bean [Vigna radiata (L.) Wilczek]. Afr J Biotechnol 11:15012–15019

    CAS  Google Scholar 

  • Taurian T, Ibanez F, Angelini J, Tonelli ML, Fabra A (2012) Endophytic bacteria and their role in legumes growth promotion. Bact Agrobiol Plant Probiotics, Springer-Verlag, Berlin, Heidelberg, pp 141–168

  • Tien TM, Gaskins MH, Hubbell DH (1979) Plant growth substances produced by Azospirillum brazilense and their effect on the growth of pearl millet. Appl Environ Microbiol 37:1016–1024

    CAS  Google Scholar 

  • Tripathi AK, Verma SC, Chowdhury SP, Lebuhn M, Gattinger A, Schloter M (2006) Ochrobactrum oryzae sp. nov., an endophytic bacterial species isolated from deep-water rice in India. Int J Syst Evol Microbiol 56:1677–1680

    Article  CAS  Google Scholar 

  • Vincent JM (1970) A manual for the practical study of root nodule bacteria. Blackwell Science Publisher, Oxford

    Google Scholar 

  • Xu Y, Yokota A, Sanada H, Hisamatsu M, Araki M, Cho HJ, Morinaga T, Murooka Y (1994) Enterobacter cloacae A105, isolated from the surface of root nodules of Astragalus sinicus cv. Japan, stimulates nodulation by Rhizobium huakuii bv. Renge. J Ferment Bioeng 77:630–635

    Article  CAS  Google Scholar 

Download references

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Authors and Affiliations

  1. Microbial Physiology Laboratory, National Institute for Biotechnology and Genetic Engineering (NIBGE)/ PAEC, Islamabad, Pakistan

    Mohsin Tariq, Sohail Hameed, Tahira Yasmeen, Mehwish Zahid & Marriam Zafar

  2. Government College University Faisalabad (GCUF), Allama Iqbal Road, Faisalabad, Pakistan

    Mohsin Tariq & Tahira Yasmeen

  3. Centre of Agricultural Biochemistry and Biotechnology (CABB), University of Agriculture Faisalabad, Faisalabad, Pakistan

    Marriam Zafar

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  1. Mohsin Tariq
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  2. Sohail Hameed
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  3. Tahira Yasmeen
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  4. Mehwish Zahid
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Correspondence to Mohsin Tariq.

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Tariq, M., Hameed, S., Yasmeen, T. et al. Molecular characterization and identification of plant growth promoting endophytic bacteria isolated from the root nodules of pea (Pisum sativum L.). World J Microbiol Biotechnol 30, 719–725 (2014). https://doi.org/10.1007/s11274-013-1488-9

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