Screening And Identification Of Antagonistic Bacteria Against Bacterial Leaf Spot Of Cucumber And Its Biological Characteristics

650 strains of bacteria were isolated from soil in hainan,anhui,zhejiang and other places,screening of antagonistic bacteria against the new pathogen of Pseudomonas cichorii(Swingle Stapp)by the inhibition zone method.Among them,the strain numbered JNL has the best antagonistic effect.The antagonistic bacteria were identified by morphological observation,physiological and biochemical characterization,16S rDNA homology sequence analysis and gyrA gene sequence analysis.JNL strain was positive in gram staining reaction,capable of hydrolyzing starch and liquefied gelatin,negative in methyl red test and negative in acetylmethyl methanol test.Malonate could not be used,but fructose,raffinose,galactose and mannitol could be used for fermentation to produce acid.Glucose,lactose,galactose,sucrose,raffinose and fructose can be used as carbon sources.Sorbitan,sorbitol,rhamnose and sweet alcohol cannot be used as carbon sources.Combining 16S rDNA homology sequence analysis and gyrA gene sequence analysis,the strain JNL was identified as Bacillus amyloliquefaciens(Wang et al.2008).Rifampicin was used to label antagonistic bacteria to explore the colonization rule of antagonistic bacteria on cucumber leaves.The test results showed that the antagonistic bacteria could colonize on cucumber leaves,at the beginning of inoculation,the amount of bacteria was the highest,and gradually decreased over the time.Under different light treatment conditions,we found that there was no significant difference in the colonization amount of the induced strain under 12 h light and 12 h darkness compared with that under 24 h light,but the colonization amount of the labeled strain was significantly reduced under 24 h darkness.Three methods were used to study the effect of JNL controls cucumber leaf spot disease.That is cucumber leaves first sprayed with antagonistic strain suspension and 24 hours later sprayed Pathogen suspension;The antagonistic strain suspension and the Pathogen suspension suspension were sprayed at the same time;First sprayed the Pathogen suspension and 24 hours later sprayed antagonistic strain suspension;Cucumber leaves with only the pathogen bacterial suspension as a control.The results showed that the control effects of of JNL controls cucumber leaf spot disease in the three treatment methods were 68.2%,65.07%and 47.6%,respectively.It is indicating that most effective control is observed when JNL strain is applied before pathogen application,while applying JNL strain after pathogen application is least effective.After inoculation of JNL strain on cucumber leaf,the activity of five defense enzymes in cucumber was improved compared with that before.POD activity increased by 1.64 times,PPO,SOD and CAT activity was nearly doubled of the control group,and PAL activity was 2.13 times of the control group.Therefore;it can be inferred that antagonistic bacteria JNL can induce the resistance of cucumber to diseases,so as to resist the infection of pathogenic bacteriaDetermination of optimum culture conditions of antagonistic bacteria by single factor variable method.The results showed that strain JNL optimum pH of 7,the optimal temperature of 30℃,the optimum carbon source as galactose,the optimum nitrogen source for potassium nitrate,optimum table speed of 150 r/min,the optimum training time is 24-36 h.