Screening Of Mixed Strain Resistant To Tomato Bacterial Wilt And Study On Biological Fertilizer

Tomato bacterial wilt is one of the most serious soil-borne plant diseases in the world and has quite a bit of damage.Under the premise of environmental protection,biocontrol is one of the effective ways to reduce the incidence of the disease and yield loss.Actinomycetes as BCAs attention resources,the application of the BCAs also play an increasingly important role in agriculture,and chemical reagents such as synthetic fungicides have limitations and harmfulness in biocontrol.Therefore,using antagonistic microorganisms present in the environment,development of environmental protection and efficient type of probiotics as an alternative to chemical pesticides,so this disease is a subject of great concern at present.Because of a single strain has many shortcomings,such as shorter duration,not economic,etc,therefore this study from the perspective of the mixed strains,opposite to carbendazim resistant strain distribution,build mixed strain.In the later stage,a series of studies were carried out,such as pot experiment,colonization experiment and preparation of biological organic fertilizer.On the basis of studying the preparation,provide composite researches in the aspects of controlling soil-borne disease provide theoretical basis for the feasibility.In this study,actinomycetes were isolated and purified from tomato rhizosphere soil in Linyi city,Shandong Province.The antagonistic strains were screened by using filter paper with Ralstonia solanacearum as the indicator.Six strains of Streptomytes NEAU-Z2,NEAU-D18,NEAU-88,NEAU-174,NEAU-C12 and NEAU-66 had strong antibacterial activity.Through compatibility test,the strains NEAU-D18 and NEAU-88 had good compatibility,and the biocontrol effect of mixed strain(NEAU-D18+NEAU-88)on tomato bacterial wilt was studied through seedling disease resistance test,in vitro test and pot experiment,and the mechanism of biological control of mixed strain(NEAU-D18+NEAU-88)was discussed from the perspective of nutritional competition.In the later stage,the mixed strains were taken as the research object and cultivated by solid fermentation to make the compound bioorganic fertilizer,so as to clarify its stability and mechanism and effect of biocontrol action,and to provide reference for the development and utilization of the compound bioorganic fertilizer in the actual production of tomato bacterial wilt as soon as possible.In addition,the biocontrol bacteria strain NEAU-66 was identified.The main experimental results are as follows:(1)Six separate media(CPA,DPA,SSA,HV,GS,AAG)were used to isolate actinomycetes from soil.After morphological observation and molecular level analysis,the bacteria were combined,and a total of 108 actinomycetes were screened.Streptomyces,Kribbella,Nocardia,Micromonospora and Methylobacterium are included in the total.Streptomyces is the dominant bacterium.(2)The activity of 108 actinomycetes against bacterial wilt was detected.NEAU-88,NEAU-Z2,NEAU-C12 and NEAU-66 had strong antimicrobial activity against R.Solanacearum QL-Rs1115,with diameter of 25.16,15.32,17.78 and 13.00 mm,respectively.The other two strains,NEAU-174 and NEAU-D18,were provided by the laboratory with antibacterial diameters of 24.46 mm and 24.51 mm.(3)The compatibility of NEAU-Z2,NEAU-D18,NEAU-88,NEAU-174,NEAU-C12 and NEAU-66 strains were tested to screen out the non-inhibitory strains.The results show that NEAU-D18 and NEAU-88 are mutually compatible.The antibacterial diameter of the compound strain was 28.98 mm.(4)In the seedling stage experiment and pot experiment,the biocontrol effect of the mixed strain(NEAU-D18+NEAU-88)was significantly better than that of the single biocontrol strain(P<0.05).In the seedling stage experiment,when the concentration of 10~8 CFU/m L and the ratio of NEAU-D18:NEAU-88 was 1:1,the spore suspension of the mixed strain had the best control effect.In the pot experiment,the control effect of the mixed strains(NEAU-D18+NEAU-88)(70.10%)was significantly higher than that of single strains NEAU-D18(47.30%)and NEAU-88(51.29%).(5)Nutrition competition experiments showed that NEAU-D18,NEAU-88 and the mixed strain had nutritional similarities with R.Solanacearum QL-Rs1115,with NOI values of 0.71,0.82and 0.88,respectively.And it showed that the mixed strains are more competitive than the single strain in theory.(6)Through solid fermentation matrix of screening,to determine the optimal solid fermentation of two strains of bacteria matrix for vermicompost and wheat bran,and the ratio between the two late to solid mixed strains fermentation culture conditions optimization,get the best fermentation conditions,respectively is the quantity of 10%,60%water content,temperature28°C and 7 days in fermentation,mixed strains reached the maximum amount of spores.Then the biological fertilizer made from the fermentation products was potted,and the biological control effect was 76.41%.(7)Polyphasic taxonomic identification of strain NEAU-66.According to 16S r RNA gene sequence alignment,physiological and biochemical characteristics,culture characteristics of different media,morphological characteristics and chemical composition analysis,the strain was identified as a potential new species in Streptomyces.