The Study On The Associative Nitrogen Fixation Activity Of Xiang Sigu Tailings Pioneer Plants

Nitrogen is extremely scare for plants in primary environment of tailings, however, with the increase of tailings disposal year by year, pioneer plants begin to settle in the tailings. Research found that in primary environment of tailings non-legume plants are the main pioneer plants superior to other plants, which indicates that in their growing process non-legume plants can still have stable and reliable source of nitrogen even in severe environment. Analysis suggests, pioneer plants in tailings can provide a stable source of nitrogen throughout associative nitrogen fixation in rhizosphere. However, knowledge about associative Nitrogen Fixation of non-legume plants in tailings is obviously lack up to now.This paper determined associative nitrogen fixation activity of Xiang sigu tailings pioneer plants in different seasons (summer and autumn) by acetylene reduction method, then separated and screened associative azotobacter from plants rhizosphere, and used acetylene reduction method to determine nitrogen fixation activity of the strains. On this basis, we made 16S rDNA sequence analysis of high nitrogen fixation activity strains. Conclusions are as follows:1. Rhizosphere soil column of pioneer plants in Xiang sigu tailing wasteland were all detected to have associative nitrogen fixation activity,but there were significant differences among different plants. In autumn, associative nitrogen fixation activity of 7 pioneer plants’ rhizosphere soil column were between 2.414 to 36.418 μmol·m-2·h-1.While in the next summer, we selected Cogon grass, cattail and reed as study objects, finding that average associative nitrogen fixation activity of these 3 plants’rhizosphere soil column were 36.832μmol·m-2·h-1 higher than 29.723μmol·m-2·h-1 in autumn. In different seasons (autumn and summer) associstive nitrogen fixation activity detected from pioneer plants’rhizosphere soil column were higher than that from blank and soil column2. Nitrogen fixation activity of pioneer plants’roots, rhizosphere soil and root soil were obviously different, and different plants conformed to the same trend:roots>rhizosphere soil >root soil. In summer, Cogon grass, cattail and reed those three pioncer plants’average associative nitrogen fixation activity in roots, rhizosphere soil and root soil were respectively 77.248μmol·kg-1·h-1,1.191μmol·kg-1·h-1 and 0.093μmol·kg-1·h-1.Then in autumn, average associative nitrogen fixation activity in roots, rhizosphere soil and root soil were detected lower than in summer, the data were 63.759μmol·kg-1·h-1, 0.792μmol·kg-1·h-1 and 0.078μmnol·kg-1·h-1 respectively.3.We separated azotobacter from different samples of xiangsi valley by nitrogen-free cultur e medium,finding that the average number of azotobacter in plant rhizosphere were 2.39×106/g, much more than in exposed field 8.96×105/g. From tailing samples we totally got 51 stains, of which 21 had nitrogen fixation activity, and the average value was between 12.24 to 3416.98 nmol·mL-1·h-1, 4 strains’ activity value was over 500 nmol·mL-1·h-1.4. We carned out 16S rDNA sequence analysis and comparison of those 4 high nitrogen fixation activity strains, the result suggested: Azoiobacter WKGD06 and Azorhizobium caulinodans bad the closest genetic relationship, the similarity was up to 100%. Azotobacter WKGD09 and Rhizobium miluonense had the closest genetic relationship, the similarity was up to 99.78%. Azotobacter WKGD14 and Klebsiella michiganensis had the closest genetic relationship, the similarity was up to 99.12% Azotobacter WKGD19 and Caulobacter segnis had the closest genetic relationship, the similarity was up to 98.35%.5. By comparison, we found that though the covering period of soil (5～10years) on Yang shanchong tailings was similar to that of Xiang sigu tailings, associative nitrogen fixation activity of pioneer plants’ rhizosphere soil column in Yang shanchong tailings was lower than in Xiang sigu tailings. Analysis suggested that the difference was due to lack of total nitrogen, humus carbon and other nutrients, and the low rate of water content. Even though covering soil can efficiently control environmental pollution caused by tailing dust, ecological restoration of tailings seems impossible if the nutritional status and the water content of soil are not improved.The study not only provides scientific evidence for the research on nitrogen source of tailings pioneer non-legume plants, and supports the ecological treatment of tailings as well. Moreover, the study result also expands the knowledge about associative nitrogen fixation to the pioneer plants in severe environment of tailings.