Abstract
Six diazotrophic bacteria were isolated from surface-sterilized roots of rice variety HUR-36, which is grown with very low or no inputs of nitrogen fertilizer. Out of six bacteria one isolate, RREM25, showed appreciable level of nitrogenase activity, IAA production, and Phosphate solubilization ability, and was further characterized with a view to exploiting its plant growth promoting activity. Based on 16S rRNA gene sequence analysis, this isolate was identified as Burkholderia cepacia. Diazotrophic nature of this particular isolate was confirmed by Western blot analysis of dinitrogenase reductase and amplification of nifH. Microscopic observation confirmed colonization of gfp/gusA-tagged RREM25 in the intercellular spaces of cortical as well as vascular zones of roots. Inoculation of RREM25 to rice plants resulted in significant increase in plant height, dry shoot and root weight, chlorophyll content, nitrogen content and nitrogenase activity. Plant growth promoting features suggest that this endophytic bacterium may be exploited in rice cultivation after a thorough and critical pathogenicity test.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Altschul SF, Madden TL, Schafeer AA et al (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acid Res 25:3389–3402
Bhatia R, Dogra RC, Sharma PK (2002) Construction of green protein (GFP)-marked strains of Bradyrhizobium for ecological studies. J Appl Microbiol 93:835–839
Boddey RM, de Oliveira OC, Urquiaga S et al (1995) Biological nitrogen fixation associated with sugar cane and rice: contributions and prospects for improvement. Plant Soil 174:195–209
Caballero-Mellado J, Martynez-Aguilar L, Paredes-Valdez G et al (2004) Burkholderia unamae sp. nov., a N2-fixing rhizospheric and endophytic species. Int J Syst Evol Microbiol 54:1165–1172
Chen WM, James EK, Coeny ET et al (2006) Burkholderia mimosarum sp. nov., isolated from root nodules of Mimosa spp. from Taiwan and South America. Int J Syst Evol Microbiol 56:1847–1851
Chiarini L, Bevivino A, Dalmastri C et al (2006) Burkholderia cepacia complex species: health hazards and biotechnological potential. TRENDS in Microbiol 14:277–286
Estrada-de los Santos P, Bustillos-Cristales R, Caballero-Mellado J (2001) Burkholderia, a genus rich in plant-associated nitrogen fixers with wide environmental and geographic distribution. Appl Environ Microbiol 67:2790–2798
Fahraeus A (1957) The infection of clover root hairs by nodule bacteria studied by simple glass slide technique. J Gen Microbiol 16:374–381
Glick BR (2005) Modulation of plant ethylene levels by the bacterial enzyme ACC deaminase. FEMS Microbiol Lett 251:1–7
Govindarajan M, Balandreau J, Kwon SW et al (2008) Effects of the inoculation of Burkholderia vietnamensis and related endophytic diazotrophic bacteria on grain yield of rice. Microbial Ecol 55:21–37
James EK, Olivares FL (1998) Infection and colonization of sugarcane and other gramineous plants by endophytic diazotrophs. Crit Rev Plant Sci 17:77–119
Jefferson RA, Kavanagh TA, Bevan MW (1987) GUS fusions: β- glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J 6:3901–3907
Katherine AM, Vania LMP, Alexandre R et al (2008) Endophytic colonization of rice (Oryza sativa L) by diazotrophic bacterium Burkholderia kururiensis and its ability to enhance plant growth. Annal Brazi Acad Sci 80:477–493
Ladha JK, de Bruijn FJ, Malik KA (1997) Assessing opportunities for nitrogen fixation in Oryza sativa: a frontier project. Plant Soil 194:1–10
Manulis S, Shafrir H, Epstein E et al (1994) Biosynthesis of indole-3-acetic acid via the indole-3-acetamide pathway in Streptomyces spp. Microbiol 140:1045–1050
Quanhrani-Bettache F, Porte J, Teyssier J et al (1999) pBBR1-GFP: a broad-host-range vector for prokaryotic promoter studies. Bio Tech 26:620–622
Ramos HJ, Roncato-Maccari LD, Souza EM et al (2002) Monitoring Azospirillum-wheat interactions using the gfp and gusA gene constitutively expressed from a new broad-host range vector. J Biotechnol 97:243–252
Reinhold-Hurek B, Hurek T, Gillis M et al (1993) Azoarcus gen. nov., nitrogen-fixing proteobacteria associated with roots of Kallar grass (Leptochloa fusca (L) Knuth), and descriptions of two species, Azoarcus indigens sp. nov. and Azoarcus communis sp. nov. Int J Syst Bacteriol 43:574–584
Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425
Sandhu GR, Aslam Z, Salim M et al (1981) The effect of salinity on the yield and composition of Diplachne fusca (Kallar grass). Plant Cell Environ 4:177–181
Schloter M, Bode W, Hartmann A (1995) Characterization and application of a strain-specific monoclonal antibody against the rhizosphere bacterium Azospirillum brasilense Wa3. Hybridoma 16:183–187
Sharma PK, Sarita S, Prell J (2005) Isolation and characterization of an endophytic bacterium related to Rhizobium/Agrobacterium from wheat (Triticum aestivum L.) roots. Curr Sci 89:608–610
Simon R, Priefer UB, Puehler A (1983) Vector plasmids for in vivo and in vitro manipulations of Gram-negative bacteria. In: Puehler A (ed) Molecular genetics of the bacteria—plant interactions. Springer-Verlag KG, Berlin, pp 98–106
Singh MK (2008) Genetic basis of colonization by endophytic bacteria and their role in plant growth promotion of rice. Ph.D. thesis, Banaras Hindu University, Varanasi, India, pp 95–99
Singh RK, Mishra RPN, Jaiswal HK et al (2006) Isolation and identification of natural endophytic rhizobia from rice (Oryza sativa L.) through rRNA gene PCR-RFLP and sequence analysis. Curr Microbiol 52:345–349
Singh MK, Kushwaha C, Singh RK (2009) Studies on endophytic colonization ability of two upland rice endophytes, Rhizobium sp. and Burkholderia sp., using green protein reporter. Curr Microbiol 59:240–243
Son H, Park G, Cha M et al (2005) Solubilization of insoluble inorganic phosphates by a novel salt and pH tolerant Pantoea agglomerans R-42 isolated from soyabean rhizosphere. Bioresource Technol 97:204–210
Sturz AV, Christie BR, Nowak J (2000) Bacterial endophytes: potential role in developing sustainable system of crop production. Crit Rev Plant Sci 19:1–30
Tamura K, Dudley J, Nei M et al (2007) MEGA4: molecular evolutionary genetics analysis (MEGA) software version 4.0. Mol Biol Evol 24:1596–1599
Ueda T, Suga Y, Yahiro N et al. (1995) Remarkable N2- fixing bacterial diversity detected in rice roots by molecular evolutionary analysis of nifH gene sequences. J Bacteriol 177: 1414–1417
Yanni YG, Rizk RY, Corich V et al (1997) Natural endophytic association between Rhizobium leguminosarum bv trifolii and Oryza sativa roots and assessment of its potential to promote Oryza sativa growth. Plant Soil 194:99–114
Yanni YG, Rizk RY, Fatah FKA et al (2001) The beneficial plant growth-promoting association Rhizobium leguminosarum bv trifolii with Oryza sativa plants. Aust J Plant Physiol 28:845–870
Acknowledgments
We are thankful to HJO Ramose (Curitiba, PR, Brazil), for providing plasmid pHRGFPGUS as gift. MKS is thankful to UGC, India for the award of Research Fellowship, to carry out this study. Research works were partly supported by the grant received from the DST Gov. of India, New Delhi.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Singh, M.K., Singh, D.P., Mesapogu, S. et al. Concomitant colonization of nifH positive endophytic Burkholderia sp. in rice (Oryza sativa L.) promotes plant growth. World J Microbiol Biotechnol 27, 2023–2031 (2011). https://doi.org/10.1007/s11274-011-0664-z
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11274-011-0664-z