The Functional Bacteria Tracing And Its Influence Mechanism During Bio-organic Fertilizer Preparation,Processing And Storage

Bio-organic fertilizer(BOF)is an emerging fertilizer with both effects of microbial fertilizer and organic fertilizer,serving as an important producing mean in sustainable agriculture.However,the domestic BOF industry is still in the initial stage,and the weekness and insuffcient of basal research always results in spotty and instable quality consequently.The quality of BOF and its effect as the microbial fertilizer depends on the quantity and activity of its functional bacteria.Hence,it is very critical for the improvement of BOF quality to study the influence mechanism of BOF preparation,processing technology and storage on the population of functional bacteria.Based on the optimization of liquid fermentation medium,Bacillus subtilis F2,as the functional bacteria of BOF,was traced in this research with plate counting and molecular biology methods,while the microbial community structure was also studied with the molecular biology methods,in order to explore the mechanism of the variation of functional bacteria population and microbial community structure of BOF which prepared by secondary composting or adsorption followed by different processing and packaging technologies.Furthermore,a strawberry pot experiment was conducted to investigate the growth-promoting effect of BOF.Here are the main achievements.1.The contrast experiment of shaking flask culture suggested that the CMGW medium,the 80 fold-dilution of concentrated monosodium glutamate waste(CMGW),can substitute for nutrient broth(NB)medium to culture strain F2.To further improve the F2 density in medium,the CMGW medium was optimized by the orthogonal experiment as follows:CMGW 12.5 g/L,beef extracts 1.0 g/L,peptone 4.0 g/L,MnSO4·2H2O 0.5 g/L,H3BO3 0.02 g/L,FeSO4·7H2O 0.1 g/L and MgSO4·7H2O 0.5 g/L.The viable cells density of F2 suspension in the optimized CMGW medium was 2.9 and 6.3 times as high as the CMGW medium and NB medium,respectively.2.The orthogonal secondary composting experiment for preparing BOF indicates that the 1%inoculation and the application of CMGW increased F2 spores population in BOF,and the addition of CMGW and mulberry branch sawdust and the adjustment of moisture content to 60%significantly increased the material temperature and alleviated the excessively high pH.3.The contrast experiment of CMGW application or not in secondary composting for preparing BOF inferred that CMGW as nutrient additive significantly promoted the F2 multiplication and enhanced its dominance in BOF.Results of denaturing gradient gel electrophoresis(DGGE)and illumine-MiSeq sequencing revealed that the microbial community structure of BOF varied obviously after secondary composting,and the addition of CMGW contributed to the change by improving species richness and enhancing the stability of microbial community structure of BOF.4.Two packaging technologies were implemented on BOF prepared by absorption in storage experiment.The results showed that the population of F2 viable cells was significantly larger in ordinary plastic package than that in vacuum package,and the variable coefficient of F2 viable cells abundance was smaller in ordinary package.5.Consequences of drying technology experiment of BOF prepared by absorption implied that raising heating temperature and decreasing heating time properly could increase the quantity of F2 viable cells,F2 spores and their stability during storage.6.Experiments of adding chemical fertilizer to BOF prepared by absorption indicated that the addition of urea significantly raised the population of F2 when the nitrogen content of BOF was 5%,and decreased it when the nitrogen content of BOF was no less than 7.5%.Besides,the addition of urea would enhance the loss of nitrogen content,result in excessively high pH and reduce the germination index of BOF during storage.7.The strawberry pot experiment showed that,BOF prepared with F2 as functional bacteria improved the plant biomass and the N,P,K nutrient content and accumulation;both the plate counting and terminal restriction fragment length polymorphism(T-rflp)analysis results indicated F2 colonization in strawberry rhizosphere;and the rhizosphere microbial community structure of BOF treatment was different with organic fertilizer(OF)treatment,while its Margalef,Pielou,Simpson and Shannon-wiener index were all lower than OF.