| Currently, the increase of crop yields and protection from pests and diseases mainly rely on excessive application of chemical fertilizer and pesticide in agricultural production. However, it also arises a series of problems at the same time, such as environment pollution, disruption of ecological balance and food safety. Besides the reduction of the harmness of diseases and pests, microbial fertilizer has been demonstrated to improve crop production in previous research. Therefore, it is of great significance to carry out scientific research on screening beneficial microorganism and developing them into the microbial fertilizer. The selection of biocontrol strains with growth promotion, microbial ferteilizer development and their biological effects have been conducted in this study, and the main obtained results are as follows:1. To screen endophytic bacteria with a good application potential, IAA biosynthesis, nitrogen-fixing and phosphate solubilizing activity of maize endophytic bacteria isolated and stored in our lab as well as chlorophyll content in maize leaves after treated with bacteria had been determined. The results showed that Bacillus amyloliquefaciens Y19 strain has a better performance in IAA biosynthesis and nitrogen fixation and phosphate solubilization, and the chlorophyll content after inoculation with Y19 is higher than those of Y13 and Y10 strain. For these reasons, Y19 was used as test bacterium in the following test.2. The plasmid p HAPII carrying green fluorescent protein gene gfpmut3 a was transformed into Y19 cells by natural transformation, and a tagged strain Y19-GFPmut3 a with good fluorescence was obtained. Results showed that the introduction of exogenous plasmid has no significant effect on the antifungal and plant growth-promotion of Y19, according to the plate antagonistic bioassay and maize pot experiment in the greenhouse. In greenhouse maize pot bioassay, Y19’s colonization into roots, stems and leaves were observed on 9 DAI(days after inoculation) by using confocal laser scanning microscope. 100 DAI, the gfp-tagged strain still could be detected in the rhizospheric soil, roots, stems and leaves tissues of maize, eggplant and Chinese cabbage. These results indicated that Y19 owns a strong colonization capacity and good transduction ability in different plants.3. Y19-GFPmut3a(106CFU/m L) fermentation broth including xanthan gum with a final concentration at 0.03% was inoculated into tobacco by foliar spray and(or) root irrigation. And the results showed that the plant height, fresh weight and dry weight of tobacco in the double treatment with leaves-spraying and root-irrigating could significantly increase. 30 days DAI, the gfp-tagged strain was able to be detected in tobacco, which indicates that maize endophytic bacterium Y19 also owns a strong colonization ability in tobacco.4. In order to test the antagonism among beneficial bacteria, Y19 was mixed with a plant growth-promoting and disease control bacterium Bacillus amyloliquefaciens B9601-Y2, a phosphate-solubilizing bacterium Burkholderia cepacia 1-2 and YN201305, respectively. The result showed that there is no obvious inhibition effect between every two strains. On this basis, with the sole bacterium’s culture as control,the mixture of each two strains’ fermentation broth was mixed into the matrix with a volume-mass concentration at 1: 2, and the final matrix were subsequently used as cultivation substrates for corn, radish, cabbage and rape in warm house. The result found that Y19 fermentation broth mixed with matrix has the best growth promoting effect.5. The fermentation broth of Y19(106CFU/m L) was thoroughly mixed into matrixes with different concentrations of nitrogen, phosphorus and potassium chemical fertilizer, respectively. Subsequently, the complex matrixes were used for the growth promotion of floating rice seedlings and maize pot bioassay. It turned out that Y19’s liquid culture mixed with the matrix containing 3~4% chemical phosphate fertilizer has the best plant growth-promoting effect, which can increase the maize fresh weight by 448.05%-487.60% on 30 days after pot planting, compared with control. It indicates that Y19 could mix with the matrix carrying chemical fertilizer and the following complex matrix can significantly promote maize seedlings’ growth.6. The detection of indigenous microbial multi-function and mass in maize rhizospheric soil were performed. The result found that after applied with Y19 microbial fertilizer, the amount of bacteria and actinomycete in maize and its rhizosphere increased, but the amount of fungi reduced, especially for Fusarium sp.. Besides that, according to the average well color development(AWCD) in the Biolog assay, the fertilizer also increased the microbes efficiency of carbon source utilization the rhizosphere, mainly focusing on the use efficiency of carbohydrates, amino acids and carboxylates. These data implies that Y19’s microbial fertilizer might improve the micro-environment of rhizospheric soil to exert its growth-promoting and disease-controlling effect.7. The shelf life test of Y19 microbial fertilizer indicates that Y19 in pure matrix and the matrix including 3% phosphate fertilizer have the highest survival rate. After one year’s storage, the living cells in fertilizers could maintain more than 107cfu/g at 28℃ and 50% humidity. |
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