In situ localization of Paenibacillus polymyxa HKA-15 in roots and root nodules of soybean ( Glycine max. L.)
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In situ localization of Paenibacillus polymyxa HKA-15 in roots and root nodules of soybean (Glycine max.L.) Kannepalli Annapurna & Dhanushkodi Ramadoss & Pranita Bose & Lakkineni VithalKumar
Received: 4 January 2013 / Accepted: 24 June 2013 # Springer Science+Business Media Dordrecht 2013
Abstract Background and aims Bacterial endophytes can colonize various plants and organs. However, endophytic bacteria (other than rhizobia) colonizing root nodules in legumes have been rarely analyzed. The present study aimed to examine the colonization and spread of gfptagged Paenibacillus polymyxa in soybean plants under gnobiotic conditions. Methods Inoculation with gfp-tagged Paenibacillus. polymyxa HKA −15 alone and in combination with Bradyrhizobium japonicum were done on soybean seedlings. In situ localization was detected through confocal microscopy and PCR. Results Inoculation with P. polymyxa-gfp strain alone and in combination with B. japonicum DS-1 had a stimulatory effect on the plant growth. There was an increase in shoot (7.2 %) and root dry weights (14.5 %) when the two strains were co - inoculated over that of B. japonicum inoculation alone. In vivo simultaneous visualization using Confocal Laser Scanning Microscopy (CLSM) showed the localization of the gfp-tagged P. polymyxa cells in the root nodules and its spread in the root tissue, both tap as well as lateral roots. Systemic spread into aerial tissue did not occur as indicated by the absence of bacteria. CLSM observations of the presence Responsible Editor: Peter A.H. Bakker. K. Annapurna (*) : D. Ramadoss : P. Bose : L. VithalKumar Division of Microbiology, Indian Agricultural Research Institute, New Delhi 110 012, India e-mail: [email protected]
of gfp-tagged P. polymyxa in the nodule and roots tissues was corroborated with PCR amplification of the gfpgene from these tissues. Conclusions CLSM and PCR methods confirmed that P. polymyxa invades roots and root nodules of soybean, but the spread is restricted to root tissue only. The strain improves plant growth when inoculated singly or in combination with B. japonicum. Keywords P. polymyxa . Endophyte . gfp . Root nodule . Soybean
Introduction Plants are constantly involved in interactions with a wide range of bacteria. Rhizobia are an example of a highly evolved mutualistic plant-bacterium interaction as they form nitrogen fixing nodules on roots of legumes. Rhizobia profit from plants because of the enhanced availability of C-, whereas the legume hosts receive benefits from the bacterium by growth enhancement (N-fixation) or stress reduction. Till recently rhizobia, members of α-proteobacteria were the exclusive group nodulating legumes but recent publications have shown β- proteobacteria also to form nitrogen fixing nodules on legumes. For long these were the only occupants of the nitrogen fixing root nodule. However, studies have reported the identification of a range of endosymbiotic but non-nitrogenfixing bacteria (commonly including species of the genus Pseudomonas) from root
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