Study On The Screening,identification And Biocontrol Mechanism Of Antagonistic Actinomycetes Of Mango Stemend Rot

Mangifera indica L.is a famous tropical fruit.Stem-end rot is one of the main postharvest diseases of mango,which causes huge economic losses and seriously restricts the storage,transportation of mango postharvest.To enrich antagonistic resources of mango stem-end rot,find a non-toxic and pollution-free method for postharvest control instead of chemical fungicides,with Botryodiplodia theobromae Pat.and Phomposis mangiferae Ahmad as the indicator pathogen,isolation and screening of actinomycetes antagonistic to stem-end rot from mango rhizosphere soil by plate dilution method and double culture method.The classification status of antagonistic actinomycetes was clarified,the fermentation conditions were optimized,the in vivo control effect was tested,the antibacterial mechanism was preliminarily explored,and the ways to improve the biological control effect were explored.The results are as follows:（1）A total of 223 actinomycetes were isolated from soil samples collected in Hainan University Experimental Teaching Base,Sanya Mar Garden and Hele-Garden.Antagonistic actinomycetes A2,B85 and D16 were screened by double culture method and had good genetic stability against Botryodiplodia theobromae Pat.and Phomposis mangiferae Ahmad.And they have inhibitory effect on a variety of fungal pathogens,with a wide antibacterial spectrum.Antagonistic strains A2,B85 and D16 had certain inhibitory effect on stem-end rot.In the early stage,they could well inhibit the occurrence of stem rot and the expansion of the disease spots,which was equivalent to the effect of 250 mg·L^-1 Prochloraz treatment.（2）By observing the morphological characteristics,culture characteristics,physiological and biochemical characteristics of the strains,and carrying out 16S r DNA sequence determination and phylogenetic analysis,the strains A2 and B85 were identified as Streptomyces malaysiensis.And D16 was identified as Streptomyces lydicus.（3）By optimizing the fermentation conditions,it is determined that the liquid fermentation medium of strain A2 is millet juice medium.The composition and culture conditions of the medium were as follows:glucose 3.0%,corn flour 3.0%,Na Cl 0.25%,Ca CO₃ 0.1%,initial p H 7.0,culture temperature 28℃,inoculation amount 10%,shaking speed 200 r·min^-1,culture time 5 days.The liquid fermentation medium of strain B85 was determined to be millet juice medium.The medium composition and culture conditions were as follows:glucose 3.0%,corn flour 2.5%,Na Cl 0.2%,Ca CO₃0.3%,initial p H 7.0,culture temperature 28℃,inoculation amount 10%,shaking speed200 r·min^-1,culture time 5 days.The liquid fermentation medium of strain D16 was glucose yeast extract medium.The composition and culture conditions of the medium were as follows:glucose 2.5%,yeast extract 1.5%,Na Cl 0.25%,Ca CO₃ 0.3%,initial p H 6.0,culture temperature 28℃,inoculation amount 10%,shaking speed 200r·min^-1,culture time 5 days.After optimization,the diameters of the inhibition zone in fermentation broth of strains A2,B85 and D16 were increased from 8.33 mm,7.67 mm and 6.67 mm to 24.33 mm,23.67 mm and 17.67 mm,respectively.（4）The strain A2 and B85 inhibited the occurrence of stem-end rot by producing active substance with antibacterial effect,inhibiting the spore germination of Botryodiplodia theobromae Pat.,nutritional competition,and improving the activities of SOD,POD and CAT in fruits.The antimicrobial mechanisms of strain D16 include:the production of active substance and volatile organic compounds with antimicrobial effects,parasitism,inhibit the spore germination of Botryodiplodia theobromae Pat.,nutrient competition and increased the activities of SOD,POD and CAT.（5）The mixed treatment of 100 mg·L^-1 SA and 50 g·L^-1 Ca Cl₂ with strain A2,50mg·L^-1 SA and 50 g·L^-1 Ca Cl₂ with strain B85,and 50 mg·L^-1 SA and 20 g·L^-1 Ca Cl₂with strain D16 could effectively inhibit the occurrence of stem-end rot and improve the activities of SOD,POD and CAT in fruits.And the effect was better than that of SA,Ca Cl₂ and antagonist alone.In conclusion,antagonistic actinomycetes A2,B85 and D16 have good inhibitory effect on the mango stem-end rot.They are genetically stable and have a wide antibacterial spectrum.And they can inhibit action of Botryodiplodia theobromae Pat.through a variety of mechanisms.Antagonistic actinomycetes can be used in the postharvest treatment of mango fruits.The effect is equivalent to that of chemical fungicides.It is used to control the occurrence of stem-end rot and has good biocontrol potential.