Identifieation Of Bacillus Siamensis Antagonistic To Fruit Ripe Rot Of Kiwifruit And The Tentative Study Of Its Antimicrobial Substances

Fruit ripe rot of kiwifruit caused by Botryosphaeria dothidea is an important plant fungal disease during the period from fruit ripening to postharvest stage.The disease occurs seriously and results in a great loss in kiwifruit industry in Fengxin connty,Jiangxi province.At present,the disease is controlled mainly by agricultural and chemical measures.However,chemical control measures revealed lots of disadvantage.Therefore,it is necessary to study the disease from the perspective of biological control.This study aimed at screening a Bacillus with strong antagonism effect,laying a good foundation for biological control of fruit ripe rot of kiwifruit in the future.From the 30 soil samples of healthy kiwifruit rhizosphere in kiwifruit orchard with high incidence of fruit ripe rot Fengxin connty,267 strains of Bacillus were isolated by soil Bacillus separation methods.In plate confrontation culture test,9 strains of Bacillus with strong antimicrobial activity against Botryosphaeria dothidea were screened,in which strain JY-1 had the strongest activity with the antibacterial rate as 74.89%.Species of strain JY-1 was identified according to morphological,physiological and biochemical characteristics,and 16 S rDNA-ITS sequence analysis,In morphological,physiological and biochemical tests,JY-1 showed similar characteristics with Bacillus,and in sequence phylogenetic analysis,it shared 100% homology with the sequence of Bacillus siamensis in GenBank database.Based on above results,strain JY-1 was identified as B.siamensis.Methods of single-factor experiment and orthogonal experiment were adopted to optimize the fermentation medium and cultural conditions for JY-1 growth and its antagonistic substance production.The results showed that the most appropriate medium components were 2% sucrose,0.3% Yeast extract,3% Peptone,0.5%NaCl,and 0.09%MgSO4·7H2O.The optimal fermentation conditions were initial pH 7.0,temperature 30?,medium volume 50 mL/250 mL,inoculum volume 1%,rotating speed 220 r/min and incubation time 72 h.Preliminary research on the active metabolites of JY-1 was conducted in terms of antimicrobial spectrum,stability of the active metabolites,control effect of different fermentation components against fruit ripe rot of kiwifruit on detached fruits and the crude extraction of antibacterial substances.The results showed JY-1 was a biocontrol strain producing surfactants,which effectively control all nine pathogenic fungi and four pathogenic bacteria tested.The metabolites of strain JY-1 can not bear strong acid and alkali,but can bear high temperature,and insensitive to ultraviolet rays.Antimicrobial effect of different fermentation components under optimized conditions was better than those under no optimization conditions in disease control test of detached fruits,and control effect of the aseptic supernatant from three day's fermenting under optimized conditions was 92.21%.Protein in fermented broth was precipitated with ammonium sulfate and when the saturation of the ammonium sulfate is 30%,content of the active components was the highest.To summarize,strain JY-1 is a microorganism with biocontrol potential.It will have potential application value in controlling fruit ripe rot of kiwifruit.