Screening Of Antagonistic Bacteria Against Fusarium Fujikuroi In Rice And Preliminary Study On Its Biocontrol Mechanism

Rice is one of the main grain crops in China,and its cultivation play a important role of grain production safety.However,the occurrence of rice diseases seriously affects rice cultivation.The common rice fungal diseases include rice base rot,grain wilt,bakanae disease,rice blast,white leaf blight and so on.Bakanae disease is widely distributed in China,which is mainly caused by Fusarium fujikuroi and Fusarium proliferatum.After the outbreak of the disease,it generally causes a decrease in rice yield of 10%～20%,and as high as 50%in severe cases.Chemical pesticides are mainly used to control bakanae disease which have potential risks to the environment and human health,.Thus,it is important to develop biocontrol fungicides in agricultural cultivation.In this study,three antagonistic bacteria J2,J3 and F3 with prevention and control effects on bakanae disease were isolated and screened from the rhizosphere soil of rice-crab ecological cultivation by dilution method and plate confrontation method,and the species were identified according to morphological characteristics,physiological and biochemical characteristics and molecular biology analysis to clarify their biological status.The effect of antagonistic bacteria on the control of bakanae disease and its effect on the growth of rice were investigated by pot experiment.In addition,the culture conditions of antagonistic bacteria were also studied.The main results of this study are as follows:1.Screening and identification of antagonistic bacteria.Fusarium fujikuroi was isolated from diseased rice plants,and three antagonistic bacteria were screened from rice and crab co-cropping soil.The antagonistic bacteria were identified by morphological characteristics,physiological and biochemical tests and 16S r RNA sequence comparison.J2 was identified as Bacillus amyloliquefaciens.J3 is Bacillus sp.F3 belongs to the genus Bacillus subtilis.In the plate confrontation test,the inhibition rate of three strains against Fusarium fujikuroi reached 60%,48%and 50%,respectively.2.Effect of antagonistic bacteria on control of Fusarium fujikuroi and its effect on rice growth.Pot experiment showed that J2,J3 and F3 were inoculated on Fusarium fujikuroi and cultured for fifteen days.The three strains of antagonistic bacteria significantly reduced the incidence of Fusarium fujikuroi,and the control rates reached 69%,60%and 62%,respectively.Compared with CK,rice plant height was significantly increased after inoculation with antagonistic bacteria alone.The plant height and chlorophyll of rice were significantly increased after inoculation with rhizoma sinensis and antagonistic bacteria J2,J3and F3,compared with the treatment of rhizoma sinensis only.The results showed that inoculation of three antagonistic bacteria could promote the growth and chlorophyll synthesis of rice seedlings.3.Antagonism and vitality promotion:Under the treatment of 10%,20%and 50%inoculum of the sterile fermentation solution of antagonistic bacteria,Fusarium fujikuroi mycelia showed abnormal enlargement and shortening at the top.The antagonistic strains of PNC-J2,PNC-J3 and PNC-F3 with stable inheritance were screened by the antibiotic labeled strains,and they were still resistant to Fusarium fujikuroi.The rice plants inoculated with PNC-J2,PNC-J3,and PNC-F3 antagonistic bacteria were isolated,and it was found that PNC-J2,PNC-J3,and PNC-F3 antagonistic bacteria were present in the leaves of the roots and stems of the plants,indicating that the antagonistic bacteria PNC-J2,PNC-J3,and PNC-F3 could colonize in rice plants.All the three strains of antagonistic bacteria can secrete protease and cellulase,which can effectively act on pathogenic bacteria and cause their growth to be hindered.In addition,the three strains of antagonistic bacteria also had the ability to solubilize phosphorus and secrete ferrophilin and auxin,which could effectively promote the growth of rice.After inoculation,the activities of superoxide dismutase（SOD）,peroxidase（POD）,catalase（CAT）,soluble sugar and soluble protein bacteria in root and leaf of rice were significantly increased,and the content of malondialdehyde（MDA）was decreased,which could effectively resist the influence of some adverse factors in rice growth.4.Optimization of culture conditions of antagonistic bacteria:the optimal inoculum amounts of J2,J3 and F3 strains were 2%,4%and 3%,respectively.The optimal medium for J2 strain was 2%soluble starch,2.5%KNO₃and pH 7.5.The optimum medium for J3 strain was 1.5%soluble starch,2.5%KNO₃and pH 7.The optimal medium for strain F3 was 2.5%soluble starch,1.5%KNO₃and pH 7.