Screening of Multi-functional Nitrogen Fixing Bacteria and Their Application in Soil Ecological Restoration

doi:10.13560/j.cnki.biotech.bull.1985.2017-0678

Abstract

Abstract: The objective of this study is to screen multi-functional nitrogen-fixing bacteria based on nitrogenase and ACC（1-aminocyclopropane-1-carboxylate）deaminase activity，to develop inoculant and test its effect in wasteland soil ecological restoration. First，nitrogenase activity was determined by acetylene reduction assay，and ACC deaminase activity was quantitatively measured by colorimetry. Then，the 16S rRNA of the strains was amplified by PCR，and the phylogenetic status of the strains was investigated by sequence analysis and alignment. The identification of strains was carried out based on the morphology，physiology，biochemical test results，and 16S rRNA analysis. As results，total 47 strains with nitrogenase activity higher than 9 nmol C₂H₄/（h · mg）protein were screened，and the nitrogenase activities of the 43 strains were higher than that of positive control Azotobacter ACCC11103. And 20 strains producing ACC deaminase of 0.326 μmol/（h·mg）to 21.980 μmol/（h·mg）were obtained. Plant growth promotion test showed that the 47 strains increased the fresh weights of Chinese cabbage（Brassica campestris）. Based on 16S rRNA sequence，the 51 selected strains were identified as Agrobacterium albertimagni，Arthrobacter pascens，Bacillus circulans，Burkholderia vietnamiensis，Chitinophaga ginsengisegetis，Dyadobacter fermentans，Ensifer adhaerens，Enterobacter aerogenes，Lysobacter yangpyeongensis，Micrococcus luteus，Paenibacillus alginolyticus，Phyllobacterium ifriqiyense，Pseudomonas fluorescens，Rhizobium alamii，Sinorhizobium adhaerens，Sporolactobacillus laevolacticus of 31 species of 16 genera. After test，4 excellent strains of 7012，7134，7144，and 7164 were manufactured as inoculant termed as NFMF（Nitrogen Fixing and Multi Function）biofertilizer，with which the field experiment showed that soil microorganism quantity increased significantly after 3 months of NFMF biofertilizer inoculation，and soil activities of sucrase，catalase，urease，phosphatase，cellulase and xylanase significantly increased compared with the no inoculation control. In conclusion，NFMF can significantly increase wasteland soil bioactivity and it is valuable in the ecological restoration of wasteland soil.

Key words: biological nitrogen fixation, ACC deaminase, soil enzyme, ecological restoration

MAO Xiao-jie, WANG Xin-min, ZHAO Ying, ZHOU Yi-qing, SUN Jian-guang. Screening of Multi-functional Nitrogen Fixing Bacteria and Their Application in Soil Ecological Restoration[J]. Biotechnology Bulletin, 2017, 33(10): 148-155.

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