Endophytic bacterial diversity in four rice cultivars grown in two soil-types, their plant-probiotic features and rhizospheric deployment under P-stress were investigated.
Oryza sativa cvs. TCN1, TCS10, TK8, and TN71 were cultivated in greenhouse using non-sterile acidic and near-neutral paddy soils for 60 days. Root, stem and leaf tissues were screened for culturable bacterial endophytes using nutrient agar. Endophytes were identified and profiled for plant-probiotic features. The richness, Shannon-Weiner diversity, evenness and Venn’s distribution in terms of endophytic strains were evaluated. Seed-borne endophytes were characterized through DGGE. The deployment of endophytes into the rhizosphere in TCN1 and TK8 under gnotobiotic P-stress was assessed.
A total of 52 distinct endophytic bacterial strains affiliated to 5 classes and 20 discrete genera exhibiting differential plant-probiotic features were isolated from various tissues of four different rice cultivars. The diversity and distribution of endophytes fluctuated with soil-type, tissue-type and rice genotype. Gnotobiotic insoluble P treatment revealed significantly enhanced deployment of P-solubilizing rhizobacteria in TCN1 as compared to soluble P and P-lacking control.
Rice endophytic bacteria are diverse, and their distribution within the plant and deployment as rhizobacteria were found to be influenced by host genotype, edaphic factors and nutrient stress.
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Barraquio WL, Revilla L, Ladha JK (1997) Isolation of endophytic diazotrophic bacteria from wetland rice. Plant Soil 194:15–24
Boddey RM, de Oliveira OC, Urquiaga S, Reis VM, de Olivares FL, Baldani VLD, Döbereiner I (1995) Biological nitrogen fixation associated with sugar cane and rice: contributions and prospects for improvement. Plant Soil 174:195–209
Botta AL, Santacecilia A, Ercole C, Cacchio P, Del Gallo M (2013) In vitro and in vivo inoculation of four endophytic bacteria on Lycopersicon esculentum. New Biotechnol 30:666–674
Bremner JM, Mulvaney CS (1982) Nitrogen‑total. In: Page AL, Miller RH, Kenney DR (Ed.) Methods of soil analysis. Part 2 – microbiological and biochemical properties. New York: Soil Science Society of America pp 595–624 (SSA book series, 5)
Chaudhary HJ, Peng G, Hu M, He Y, Yang L, Luo Y, Tan Z (2012) Genetic diversity of endophytic diazotrophs of the wild rice, Oryza alta and identification of the new diazotroph, Acinetobacter oryzae sp. nov. Microb Ecol 63:813–821
Chen YP, Rekha PD, Arun AB, Shen FT, Lai WA, Young CC (2006) Phosphate solubilizing bacteria from subtropical soil and their tricalcium phosphate solubilizing abilities. Appl Soil Ecol 34:33–41
Cheng YH, Cheng AH, Chen SS, Tu CC (1989) The outbreaks of pod rot of peanut and its control. J Agri Res China 38:353–364
Clark LL, Dajcs JJ, McLean CH, Bartell JG, Stroman DW (2006) Pseudomonas otitidis sp. nov., isolated from patients with otic infections. Int J Syst Evol Microbiol 56:709–714
Compant S, Clement C, Sessitsch A (2010) Plant growth-promoting bacteria in the rhizo- and endosphere of plants: their role, colonization, mechanisms involved and prospects for utilization. Soil Biol Biochem 42:669–678
Cottyn B, Debode J, Regalado E, Mew TW, Swings J (2009) Phenotypic and genetic diversity of rice seed-associated bacteria and their role in pathogenicity and biological control. J Appl Microbiol 107:885–897
Cuong ND, Nicolaisen MH, Sørensen J, Olsson S (2011) Hyphae-colonizing Burkholderia sp.- a new source of biological control agents against sheath blight disease (Rhizoctonia solani AG1-IA) in rice. Microb Ecol 62:425–434
Dan M, Marien GJ, Goldsand G (1984) Endocarditis caused by Staphylococcus warneri on a normal aortic valve following vasectomy. Can Med Assoc J 131:211–213
Edwards U, Rogall T, Blocker H, Emde M, Bottger EC (1989) Isolation and direct complete nucleotide determination of entire genes. Characterization of a gene coding for 16S ribosomal RNA. Nucleic Acids Res 17:7843–7853
Elbeltagy A, Nishioka K, Suzuki H, Sato T, Sato Y, Morisaki H, Mitsui H, Minamisawa K (2000) Isolation and characterization of endophytic bacteria from wild and traditionally cultivated rice varieties. Soil Sci Plant Nutr 46:617–629
Engelhard M, Hurek T, Reinhold-Hurek B (2000) Preferential occurrence of diazotrophic endophytes, Azoarcus spp., in wild rice species and land races of Oryza sativa in comparison with modern races. Environ Microbiol 2:131–141
Felsenstein J (1981) Evolutionary trees from DNA sequences: a maximum likelihood approach. J Mol Evol 17:368–376
Feng Y, Shen D, Song W (2006) Rice endophyte Pantoea agglomerans YS19 promotes host plant growth and affects allocations of host photosynthates. J Appl Microbiol 100:938–945
Fujii T, Huang YD, Higashitani A, Nishimura Y, Iyama S, Hirota Y, Yoneyama Y, Dixon RA (1987) Effect of inoculation with Klebsiella oxytoca and Enterobacter cloacea on dinitrogen fixation by rice-bacteria associations. Plant Soil 103:221–226
Grimont PAD, Grimont F, Farmer JJ III, Asbury MA (1981) Cedecea davisae gen. nov., sp. nov. and Cedecea lapagei sp. nov., new Enterobacteriaceae from clinical specimens. Int J Syst Bacteriol 31:317–326
Hallmann J, Berg G (2006) Spectrum and population dynamics of bacterial root endophytes. In: Schulz BJE, Boyle CJC, Sieber TN (eds) Microbial root endophytes, vol 6. Springer, Berlin, pp 15–31
Hallmann J, Quadt-Hallmann A, Mahaffee WF, Kloepper JW (1997) Bacterial endophytes in agricultural crops. Can J Microbiol 43:895–914
Hardoim PR, van Overbeek LS, van Elsas JD (2008) Properties of bacterial endophytes and their proposed role in plant growth. Trends Microbiol 16:463–471
Hardoim PR, Andreote FD, Reinhold-Hurek B, Sessitsch A, van Overbeek LS, van Elsas JD (2011a) Rice root-associated bacteria: insights into community structures across 10 cultivars. FEMS Microbiol Ecol 77:154–164
Hardoim PR, Sessitsch A, Reinhold-Hurek B, van Overbeek LS, Van E (2011b) Assessment of rice root endophytes and their potential for plant growth promotion. In: Hardoim PR (ed) Bacterial endophytes of rice – their diversity, characteristics and perspectives. Groningen University, Groningen, pp 77–100
Hardoim PR, Hardoim CC, van Overbeek LS, van Elsas JD (2012) Dynamics of seed-borne rice endophytes on early plant growth stages. PLoS One 7:e30438
Hardoim PR, Nazir R, Sessitsch A, Elhottova D, Korenblum E, van Overbeek LS, van Elsas JD (2013) The new species Enterobacter oryziphilus sp. nov. and Enterobacter oryzendophyticus sp. nov. are key inhabitants of the endosphere of rice. BMC Microbiol 13:164–176
Hartmann A, Schmid M, Van Tuinen D, Berg G (2009) Plant-driven selection of microbes. Plant Soil 321:235–257
Hayward AC, Fegan N, Fegan M, Stirling GR (2010) Stenotrophomonas and Lysobacter: ubiquitous plant-associated gamma-proteobacteria of developing significance in applied microbiology. J Appl Microbiol 108:756–770
Heiner CR, Hunkapiller KL, Chen SM, Glass JI, Chen EY (1998) Sequencing multimegabase-template DNA with BigDye terminator chemistry. Genome Res 8:557–561
Higgins DG, Bleasby AJ, Fuchs R (1992) Clustal V: improved software for multiple sequence alignment. CABIOS 8:189–191
Hirota Y, Fujii T, Sano T, Iyama S (1978) Nitrogen fixation in the rhizosphere of rice. Nature 276:416–417
Hoagland DR, Arnon DI (1938) The water-culture method for growing plants without soil. Univ. of Calif. Agr. Exp. Sta. Berkeley, Circular 347
Holt KE, Baker S, Weill FX, Holmes EC, Kitchen A, Yu J, Sangal V, Brown DJ, Coia JE, Kim DW, Choi SY, Kim SH, da Silveira WD, Pickard DJ, Farrar JJ, Parkhill J, Dougan G, Thomson NR (2012) Shigella sonnei genome sequencing and phylogenetic analysis indicate recent global dissemination from Europe. Nat Genet 44:1056–1059
Hurek T, Reinholdhurek B, Vanmontagu M, Kellenberger E (1994) Root colonization and systemic spreading of Azoarcus sp. strain BH72 in grasses. J Bacteriol 176:1913–1923
Jeong H, Jeong DE, Kim SH, Song GC, Park SY, Ryu CM, Park SH, Choi SK (2012) Draft genome sequence of the plant growth-promoting bacterium Bacillus siamensis KCTC 13613T. J Bacteriol 194:4148–4149
Ji X, Lu G, Gai Y, Gao H, Lu B, Kong L, Mu Z (2010) Colonization of Morus alba L. by the plant growth-promoting and antagonistic bacterium Burkholderia cepacia strain Lu10-1. BMC Microbiol 10:243–254
Ji SH, Gururani MA, Chun SC (2014) Isolation and characterization of plant growth promoting endophytic diazotrophic bacteria from Korean rice cultivars. Microbiol Res 169:83–98
Jones Junior JB (2001) Laboratory guide for conducting soil tests and plant analysis. CRC, Boca Raton, p 363
Kaga H, Mano H, Tanaka F, Watanabe A, Kaneko S, Morisaki H (2009) Rice seeds as sources of endophytic bacteria. Microbes Environ 24:154–162
Kämpfer P, Denner EB, Meyer S, Moore ER, Busse HJ (1997) Classification of “Pseudomonas azotocolligans” Anderson 1955, 132, in the genus Sphingomonas as Sphingomonas trueperi sp. nov. Int J Syst Bacteriol 47:577–583
Kim OS, Cho YJ, Lee K, Yoon SH, Kim M, Na H, Park SC, Jeon YS, Lee JH, Yi H, Won S, Chun J (2012) Introducing EzTaxon-e: a prokaryotic 16S rRNA gene sequence database with phylotypes that represent uncultured species. Int J Syst Evol Microbiol 62:716–721
Knauth S, Hurek T, Brar D, Reinhold-Hurek B (2005) Influence of different Oryza cultivars on expression of nifH gene pools in roots of rice. Environ Microbiol 7:1725–1733
Kumar G, Kanaujia N, Bafana A (2012) Functional and phylogenetic diversity of root-associated bacteria of Ajuga bracteosa in Kangra valley. Microbiol Res 167:220–225
La Scola B, Birtles RJ, Mallet MN, Raoult D (1998) Massilia timonae gen. nov., sp. nov., isolated from blood of an immunocompromised patient with cerebellar lesions. J Clin Microbiol 36:2847–2852
Lee S, Ka JO, Song HG (2012) Growth promotion of Xanthium italicum by application of rhizobacterial isolates of Bacillus aryabhattai in microcosm soil. J Microbiol 50:45–49
Lin SY, Shen FT, Young CC (2011) Rapid detection and identification of the free-living nitrogen fixing genus Azospirillum by 16S rRNA-gene-targeted genus-specific primers. Antonie Van Leeuwenhoek 99:837–844
Liu JR, Lin YD, Chang ST, Zeng YF, Wang SL (2010) Molecular cloning and characterization of an insecticidal toxin from Pseudomonas taiwanensis. J Agric Food Chem 58:12343–12349
Loaces I, Ferrando L, Scavino AF (2011) Dynamics, diversity and function of endophytic siderophore-producing bacteria in rice. Microb Ecol 61:606–618
Louden BC, Haarmann D, Lynne AM (2011) Use of blue agar CAS assay for siderophore detection. J Microbiol Bio Edu 12:51–53
Mano H, Morisaki H (2008) Endophytic bacteria in the rice plant. Microbes Environ 23:109–117
Mano H, Tanaka F, Watanabe A, Kaga H, Okunishi S, Morisaki H (2006) Culturable surface and endophytic bacterial flora of the maturing seeds of rice plants (Oryza sativa) cultivated in a paddy field. Microbes Environ 21:86–100
Mano H, Tanaka F, Nakamura C, Kaga H, Morisaki H (2007) Culturable endophytic bacterial flora of the maturing leaves and roots of rice plants (Oryza sativa) cultivated in a paddy field. Microbes Environ 22:175–185
Muthukumarasamy R, Kang UG, Park KD, Jeon WT, Park CY, Cho YS, Kwon SW, Song J, Roh DH, Revathi G (2007) Enumeration, isolation and identification of diazotrophs from Korean wetland rice varieties grown with long-term application of N and compost and their short-term inoculation effect on rice plants. J Appl Microbiol 102:981–991
Nelson DW, Sommers LE (1982) Total carbon, organic carbon, and organic matter. In: Page AL, Miller RH, Kenney DR (Ed.) Methods of soil analysis. Part 2 – microbiological and biochemical properties. New York: Soil Science Society of America pp 539–579. (SSA book series, 5)
Niu DD, Liu HX, Jiang CH, Wang YP, Wang QY, Jin H, Guo J (2011) The plant growth-promoting rhizobacterium Bacillus cereus AR156 induces systemic resistance in Arabidopsis thaliana by simultaneously activating salicylate- and jasmonate/ethylene-dependent signaling pathways). Mol Plant Microbe Interact 24:533–542
Okunishi S, Sako K, Mano H, Imamura A, Morisaki H (2005) Bacterial flora of endophytes in the maturing seed of cultivated rice (Oryza sativa). Microbes Environ 20:168–177
Olivares FL, Balandi VLD, Reis VM, Baldani JI, Dobereiner J (1996) Occurrence of the endophytic diazotrophic Herbaspirillum spp. in roots, stem, and leaves, predominantly of Gramineae. Biol Fertil Soils 21:197–200
Ongena M, Jacques P (2008) Bacillus lipopeptides: versatile weapons for plant disease biocontrol. Trends Microbiol 16:115–125
Panhwar QA, Radziah O, Zaharah AR, Sariah M, Razi IM (2011) Role of phosphate solubilizing bacteria on rock phosphate solubility and growth of aerobic rice. J Environ Biol 32:607–612
Panhwar QA, Naher UA, Jusop S, Othman R, Latif MA, Ismail MR (2014) Biochemical and molecular characterization of potential phosphate-solubilizing bacteria in acid sulfate soils and their beneficial effects on rice growth. PLoS One 9:e97241
Patten CL, Glick BR (1996) Bacterial biosynthesis of indole-3-acetic acid. Can J Microbiol 42:207–220
Ruiza D, Agaras B, de Werra P, Wall LG, Valverde C (2011) Characterization and screening of plant probiotic traits of bacteria isolated from rice seeds cultivated in Argentina. J Microbiol 49:902–912
Sessitsch A, Hardoim P, Döring J, Weilharter A, Krause A, Woyke T, Mitter B, Hauberg-Lotte L, Friedrich F, Rahalkar M, Hurek T, Sarkar A, Bodrossy L, Van Overbeek L, Brar D, Van Elsas JD, Reinhold-Hurek B (2012) Functional characteristics of an endophyte community colonizing rice roots as revealed by metagenomic analysis. Mol Plant Microbe 25:28–36
Shivaji S, Chaturvedi P, Suresh K, Reddy GS, Dutt CB, Wainwright M, Narlikar JV, Bhargava PM (2006) Bacillus aerius sp. nov., Bacillus aerophilus sp. nov., Bacillus stratosphericus sp. nov. and Bacillus altitudinis sp. nov., isolated from cryogenic tubes used for collecting air samples from high altitudes. Int J Syst Evol Microbiol 56:1465–1473
Stoltzfus JR, So R, Malarvithi PP, Ladha JK, de Bruijn FJ (1997) Isolation of endophytic bacteria from rice and assessment of their potential for supplying rice with biologically fixed nitrogen. Plant Soil 194:25–36
Sun L, Qiu F, Zhang X, Dai X, Dong X, Song W (2008) Endophytic bacterial diversity in rice (Oryza sativa L.) roots estimated by 16S rDNA sequence analysis. Microb Ecol 55:415–424
Suresh K, Prabagaran SR, Sengupta S, Shivaji S (2004) Bacillus indicus sp. nov., an arsenic-resistant bacterium isolated from an aquifer in West Bengal, India. Int J Syst Evol Microbiol 54:1369–1375
Swings J, Van Den Mooter M, Vauterin L, Hoste B, Gillis M, Mew W, Kersters K (1990) Reclassification of the Causal Agents of Bacterial Blight (Xanthomonas campestris pv. oryzae) and Bacterial Leaf Streak (Xanthomonas campestris pv. oryzicola) of Rice as Pathovars of Xanthomonas oryzae (ex Ishiyama 1922) sp. nov., nom. rev. Int J Syst Bacteriol 40:309–311
Taghavi S, van der Lelie D, Hoffman A, Zhang YB, Walla MD, Vangronsveld J, Newman L, Monchy S (2010) Genome sequence of the plant growth promoting endophytic bacterium Enterobacter sp. 638. PLoS Genet 6:e1000943
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. Mol Biol Evol 28:2731–2739
Tian X, Cao L, Tan H, Han W, Chen M, Liu Y, Zhou S (2007) Diversity of cultivated and uncultivated actinobacterial endophytes in the stems and roots of rice. Microb Ecol 53:700–707
Van Overbeek LS, Van Elsas JD (2008) Effects of plant genotype and growth stage on the structure of bacterial communities associated with potato (Solanum tuberosum L.). FEMS Microbiol Ecol 64:283–296
Vandamme P, Moore ER, Cnockaert M, De Brandt E, Svensson-Stadler L, Houf K, Spilker T, Lipuma JJ (2013) Achromobacter animicus sp. nov., Achromobacter mucicolens sp. nov., Achromobacter pulmonis sp. nov. and Achromobacter spiritinus sp. nov., from human clinical samples. Syst Appl Microbiol 36:1–10
Vanlaere E, Lipuma JJ, Baldwin A, Henry D, De Brandt E, Mahenthiralingam E, Speert D, Dowson C, Vandamme P (2008) Burkholderia latens sp. nov., Burkholderia diffusa sp. nov., Burkholderia arboris sp. nov., Burkholderia seminalis sp. nov. and Burkholderia metallica sp. nov., novel species within the Burkholderia cepacia complex. Int J Syst Evol Microbiol 58:1580–1590
Veitch J, Kansara A, Bailey D, Kustos I (2014) Severe systemic Bacillus anthracis infection in an intravenous drug user. BMJ Case Rep 2014
Vincent JM (1970) A manual for the practical study of root-nodule bacteria. Blackwell Scientific Publications Ltd., Oxford, UK, ISBN: 0632064102
Wang LT, Tai CJ, Wu YC, Chen YB, Lee FL, Wang SL (2010) Pseudomonas taiwanensis sp. nov., isolated from soil. Int J Syst Evol Microbiol 60:2094–2098
Watts D, MacBeath JR (2001) Automated fluorescent DNA sequencing on the ABI PRISM 310 genetic analyzer. Methods Mol Biol 167:153–170
Yang JH, Liu HX, Zhu GM, Pan YL, Xu LP, Guo JH (2008) Diversity analysis of antagonists from rice-associated bacteria and their application in biocontrol of rice diseases. J Appl Microbiol 104:91–104
You C, Zhou F (1989) Non-nodular endorhizospheric nitrogen fixation in wetland rice. Can J Microbiol 35:403–408
Young LS, Hameed A, Peng SY, Shan YH, Wu SP (2013) Endophytic establishment of the soil isolate Burkholderia sp. CC-Al74 enhances growth and P-utilization rate in maize (Zea mays L.). Appl Soil Ecol 66:40–47
Zhang X, Sun L, Qiu F, McLean RJ, Jiang R, Song W (2008) Rheinheimera tangshanensis sp. nov., a rice root-associated bacterium. Int J Syst Evol Microbiol 58:2420–2424
Authors would like to thank the editor and anonymous reviewers for their constructive comments on this manuscript. Authors acknowledge Dr. Wei-An Lai for technical support. This research was funded by grants (Grant No. 100-2313-B-150-001-MY3 and 102-2321-B-150-001) from Ministry of Science and Technology, Taiwan.
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Responsible Editor: Choong-Min Ryu.
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Hameed, A., Yeh, MW., Hsieh, YT. et al. Diversity and functional characterization of bacterial endophytes dwelling in various rice (Oryza sativa L.) tissues, and their seed-borne dissemination into rhizosphere under gnotobiotic P-stress. Plant Soil 394, 177–197 (2015). https://doi.org/10.1007/s11104-015-2506-5
- Rice endophytes
- Shannon-Weiner index
- P-solubilizing bacteria
- Nutrient stress