Study On Optimization Of Fermentation Process,Preservation Technology And Application For Production Of Microbial Agent Of Phosphate-solubilizing Bacteria PC06 By Using Biogas Slurry From Pig Farm

Biogas slurry is the liquid remaining after fermentation of biogas.It is not only rich in a large number of nutrients that can be quickly absorbed,but also contains other active substances for the growth of microorganisms.With the continuous expansion of the scale of biogas projects in China,water pollution could be caused by direct discharge of biogas slurry from poultry or livestock breeding farm without efficient treatment.The treatment of biogas slurry has become one of the main concern of the people.Phosphorus is one of the necessary nutrient elements for plant growth.At present,a large amount of chemical phosphate fertilizer is applied in China,which has led to the continuous increase of phosphorus fixation in soil,while the lack of available phosphorus in soil is more serious,and it will also aggravate agricultural non-point source pollution and other problems.Phosphate-solubilizing bacteria could improve soil phosphorus utilization efficiency and improve soil nutrient structure.Bacillus amyloliquefaciens contains a variety of physiological habits and is easy to be isolated and cultured.And because it can produce endogenous spores with diverse resistance,it is considered as one of the effective microbial strains for producing microbial agents.The present research took biogas slurry,Bacillus amyloliquefaciens PC06?phosphate-solubilizing bacteria?,as test objects,took the spore quantity as indicator,investigated the biogas liquid spore culture medium of phosphate-solubilizing bacteria based on biogas slurry through single factor and othogonal test design.On the basis of optimizing of the medium and obtaining the high-yield phosphate-solubilizing bacteria PC06,the carrier and protective agent of the microbial agent were screened for the limitations of the living body dosage form and the shelf life,and it was prepared as a high-efficiency,long-term agricultural microbial agent while using the biogas slurry efficiently.Finally,through the field experiment,the self-developed microbial agent of phosphate-solubilizing bacteria PC06 was used to verify the effect of growth promoting of rape.In summary,the use of biogas slurry for the fermentation of phosphate-solubilizing bacteria PC06 can not only realize the high-value utilization of biogas slurry,but also obtain high-quality microbial agent of phosphate-solubilizing bacteria,which can achieve effective phosphating of poorly soluble phosphates.This provides a new way for the utilization of biogas slurry resources and provides a scientific basis for the large-scale application of strain PC06.The main conclusions are as follows:?1?The phosphate-solubilizing bacteria PC06 is Bacillus amyloliquefaciens,the cells are rod-shaped,the spores are oval-shaped,and the colonies are round,flat and white.The growth curve of strain PC06 was measured and reached the peak of growth at 24 h.?2?The results of screening components and concentration of biogas slurry medium showed that the best concentration of biogas slurry for PC06 cultivation was 70%.The best carbon source,nitrogen source and inorganic salt of biogas slurry medium were corn flour,soybean meal and K₂HPO_4.The concentrations of the corn flour and soybean meal were 1.5%and 0.5%,respectively.Through orthogonal test,the optimum ingredients of culture medium were 3%corn flour,1%soybean meal and 0.05%K₂HPO₄,under these conditions,the spore quantity of PC06 could reach 6.1×10⁹ cfu/mL.?3?Screening the carrier and protective agent for the high-yield phosphate-solubilizing bacteria PC06 after optimizing the biogas slurry medium.The results showed that the best carrier was modified corn stalk and the best protective agent was liquid paraffin?adding one percent of the span-80?for liquid microbial agent of strain PC06.After preservation for 180 days,the survival rate was 68.24%and 65.71%,respectively,and the viable count was 5.8×10⁹ cfu/mL and 2.3×10⁹ cfu/mL,respectively.The best carrier for solid microbial agent was corn stalk.After preservation for 180 days,the survival rate was 84.14%,and the viable count was 1.45×10¹⁰ cfu/g.?4?The experimental results of the dissolution ability of microbial agent of phosphate-solubilizing bacteria PC06 on insoluble phosphate showed that the microbial agent had good phosphorus solubilizing ability after storage for 180 days at room temperature,and the highest phosphorus solubilizing amount from hard dissolved calcium phosphate Ca₃?PO₄?₂ was 471 mg/L.?5?The results of rape field trial showed that the application of microbial agent of phosphate-solubilizing bacteria PC06,compared with the blank group,increased the available phosphorus content in the soil,and the content increased by 84.6%.Moreover,the phosphorus content in the aboveground and underground parts of rapeseed increased by 59.8%and 51.0%,respectively.At the same time,the plant height,root length,dry weight of the shoot,dry weight of the underground and the biomass yield of rape were significantly increased by 51.5%,30.9%,35.0%,82.2%and 50.1%,respectively.