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UBC Theses and Dissertations

N2-fixation and seedling growth promotion of lodgepole pine by wild-type and GFP-labeled Paenibacillus polymyxa Tang, Qian


Paenibacillus polymyxa P2b-2R is a bacterium that originated from the internal stem tissues of lodgepole pine (Pinus contorta var. latifolia (Dougl.) Engelm.) seedlings growing in the interior of British Columbia. Several studies have suggested that P. polymyxa P2b-2R can fix nitrogen in association with lodgepole pine and can promote plant growth. To further evaluate this strain, P. polymyxa P2b-2Rgfp, a green fluorescent protein (GFP)-labeled derivative of P2b-2R, was generated and used in a long term inoculation study. To confirm its ability to fix N₂ and to contribute to the N nutrition of pine, as well as comparing the similarities or differences between GFP-labeled and wild-type P2b-2R, a one-year study on bacterial colonization and lodgepole pine seedling growth responses to inoculation with wild-type or GFP-labeled P. polymyxa P2b-2R, was performed. Surface-sterilized lodgepole pine seeds were sown in glass tubes containing an autoclaved sand/montmorillonite clay mixture that contained a nutrient solution with a small amount of nitrogen labeled with ¹⁵N as Ca(¹⁵NO₃)₂ (5 % ¹⁵N label). After sowing, seeds were inoculated with either P. polymyxa P2b-2R or P. polymyxa P2b-2Rgfp. Non-inoculated controls received phosphate-buffered saline. The results indicated that both P2b-2R and P2b-2Rgfp were: (i) able to form persistent rhizospheric and endophytic populations, (ii) capable of in situ N fixation and (iii) enhancing seedling growth continuously after four months. Seedlings inoculated with P2b-2R derived 13%-40 % of foliar N from the atmosphere, while those treated with P2b-2Rgfp derived 18%-47 % of foliar N from the atmosphere during seedling growth period. There was a temporary seedling growth reduction in P2b-2R treatments two months after sowing, but seedlings recovered from the growth reduction thereafter. However, P2b-2Rgfp treatments did not display such a decline. Seedlings treated with P2b-2Rgfp had generally higher endophytic populations, foliar N concentrations, %N derived from atmosphere and dry weights, but lower rhizospheric populations than those treated with P2b-2R. However, these differences were not significant. GFP did not affect N-fixation and growth promotion of wild-type P2b-2R in general, however, GFP-labeled P2b-2R might have short-term advantages in early pine growth stages but these are not significant in the longer term.

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