Development Of Bio-organic Fertilizer For Inhibiting Red Root Disease Of Rubber Tree And Its Effect On Soil Microbial Flora

Natural rubber is an important raw material to maintain industrial production,and the degree of import dependence is more than 75%every year.Root disease is one of the three major diseases that hinder the production of natural rubber.Red root disease of rubber tree caused by Ganodermaphilippii is the most serious root disease in China,and causes varying degrees of harm in various rubber planting areas in the world.The reason for the outbreak of red root disease is the imbalance of soil microbial community,coupled with unfavorable prevention and control,the disease has become increasingly prominent,which has caused huge losses to the natural rubber industry.Soil condition,microbial diversity and structure affect the occurrence of diseases.In recent years,bio-organic fertilizer plays an important role in maintaining soil health.Therefore,the development of bio-organic fertilizer for inhibiting rubber tree red root disease,the analysis of soil microbial community characteristics of rubber tree red root disease and the study of its regulation means are the important basis for the control of red root disease of rubber tree.We used the plate counting,real-time PCR and high-throughput sequencing method to explore 1)solid fermentation formula and conditions of endophytic Bacillus subtilis Czk1.2)the effect of bio-organic fertilizer combined with fungicide on soil microflora in a short time,and 3)soil microbial flora characteristics of red root disease of rubber trees located at"Xinjin" farm in Qiongzhong,Hainan province,China.This study attempts to develop bioorganic fertilizer to inhibit rubber tree red root disease,reveal the characteristics of soil microflora of red root disease of rubber tree and the potential soil microbial ecological mechanism of applying bio-organic fertilizer combined with fungicides to control red root disease of rubber tree.The main results are as follows:1.Development of Bio-organic Fertilizer for inhibiting Red Root Disease of rubber treeThrough the study on the solid fermentation formula and conditions of endophytic Bacillus subtilis Czk1,it is concluded that:1)the proportion of mature chicken manure adding mushroom straw and corn straw is 38%,and the effect of adding mushroom straw is higher than corn straw.The promoting effect of coconut straw on fermentation was small,and the seed germination index of different combinations was more than 80%.2)the best parameters of solid fermentation in mushroom straw and chicken manure composting are as follows:10%of inoculation,10%of soybean powder,culture temperature of 37℃,40%of water content,and once 72 hours of turning over.Under this condition,the Czkl number and spore number could reach to 8.63×108CFU·g-1 and 8.37×108CFU·g-1 respectively,and 3)the best parameters of solid fermentation in corn straw and chicken manure composting are as follows:10%of inoculation,5%of soybean powder,culture temperature 37℃,40%of water content,and once 72 hours of turning over.under this condition,the Czk1 number and spore number could reach to 6.53×108CFU·g-1 and 5.97×108CFU·g-1 respectively.When 30%mushroom straw is added in this formula,the number of Czk1 and spores can reach 8.70 × 108CFU·g-1,and the number of spores can reach 6.90× 108CFU·g-1.2.Effects of bio-organic fertilizers and fungicides on soil microflora in the root zone of red root disease of rubber treeCompared with the conventional control and other controls,the total copy number of fungi treated with bio-organic fertilizer combined with fungicides was the lowest and the ratio of bacteria to fungi was the highest in the red root disease of rubber trees under field conditions.The results of MiSeq high-throughput sequencing showed that the application of fungicide or bio-organic fertilizer alone was unstable to the control of pathogenic fungi Ganoderma in soil.The relative abundance of Ganoderma was the least and the number of bacteria was the highest in the treatment of bio-organic fertilizer combined with fungicides.The beneficial bacteria Burkholderia,Trichoderma and Bacillus were significantly the highest.Furthermore,the correlation between soil physical and chemical properties,soil enzyme activity and soil microbial community was studied.it was found that the mean value of pH,ammonium nitrogen,nitrate nitrogen and polyphenol oxidase were significantly highest in bio-organic fertilizer combined with fungicides,while urease,sucrase and acid phosphatase were less.Trichoderma had significant positive correlation with pH and polyphenol oxidase,and Burkholderia,Ganoderma had significant positive and negative correlation with ammonium nitrogen.Burkholderia had significant negative correlation with urease and sucrase,while Ganoderma had positive correlation.3.The characteristics of soil microbial Community in the red root disease of rubber treesBy comparing the red root disease soil and healthy soil located at "Xinjin" Farm in Qiongzhong,Hainan Province,it was found that the soil nitrate nitrogen and acid phosphatase of diseased plants decreased significantly,while pH,water content,total nitrogen,catalase and sucrase increased significantly.The results of real-time PCR and MiSeq high-throughput sequencing showed that compared with healthy soil,the abundance of bacteria and fungi in diseased soil decreased significantly,and the effect of fungal community was greater than that of bacteria,and the Actinobacteria,Aciditerrimonas and Phenylobacterium in diseased soil decreased significantly,while those of Gemmatimonadetes,Rhizomicrobium and Gemmatimonas increased significantly.In the soil of diseased plants,Basidiomycota and Termitomyces decreased significantly,while Ascomycota,Archaeorhizomyces,Hygrocybe,Peniophora and Ganoderma increased significantly.Further study on the correlation between soil physical and chemical properties,soil enzyme activity and soil microbial community showed that the pathogen Ganoderma had significant positive correlation with pH,water content,total nitrogen,catalase and sucrase.