Studies On Etiology And Control Of Leaf Spot Of Coptis Chinensis

Coptis chinensis(Ranunculaceae)is a traditional Chinese medicinal herb distributed widely in China to show its detoxifying,heat drying and anti-tumor activities.Hubei province is the main production area of C.chinensis.In recent years,the leaf spot of C.chinensis incidence area and the degree of harm of the disease in Hubei province have increased rapidly,the incidence of disease in the field of about 30%.However,the etiology of the pathogen and the prevention and control technology research have not been reported in detail.In this study,the pathogen of this disease was identified,the effects of temperature,illumination,carbon and nitrogen sources on the growth of pathogens were studied,the effects of ten kinds of fungicides and one biocontrol strain on the pathogen were determined.The mail results are as follows:1.The strains of Phoma sp.were isolated from the diseased leaves of C.chinensis in nursery of Enshi Taishanmiao Coptis planting base.Pathogenicity were tested by inoculating with mycelial plugs and conidial suspension of Phoma sp.on leaves of C.chinensis.The result indicated that Phoma sp.was the pathogenic fungus of leaf spot of C.chinensis.The pathogen on the PDA medium colonies round,the edge of the rules,velvet like,and grow fast;but slower growth in the OA medium,mycelium submerged,branching;The fungal pycnidia were dark brown,100.5~120 ?m × 175.3~210 ?m in diameter;conidia were colorless,monosporous,sticky or long oval,2~2.5 ?m × 4.1~5.4?m in diameter.The phylogenetic analysis of the ITS sequence showed that the fungus shares 100% identity with Phoma aquilegiicola(HM222537.1).Therefore,according to the morphological characteristics,molecular identification results and the latest classification of phoma species,the strain was identified as Calophoma aquilegiicola.2.The biological characteristics of pathogens showed that the average diameter of the colonies was 6.72 cm after 5 days of growth on MA medium.The pathogens were grown in the range of pH 4.0-10.0,with the optimal pH of 6.0.The pathogen could grow at 4~25°C,the optimum temperature was 20°C;There was no significant difference in the growth of pathogens under different illumination conditions.In all the tested carbon sources,the effect of pathogens on maltose was the best,and the utilization of mannitolwas the worst.In all the tested nitrogen sources,the use of peptone was the best,and the use of ammonium nitrate was the worst.The lethal temperature of pathogen mycelial growth and conidial germination was 50°C water bath for 10 min.3.Laboratory screening experiments showed that all the fungicides had certain inhibitory effects on the pathogen of C.chinensis leaf spot,but there were obvious differences among the tested fungicides.The effect of 10% difenoconazole was the best,and the EC50 value was 0.0252 ?g/mL.Followed by 206.7 g/L of fenfluramazole,whose EC50 value was 0.8016 ?g/mL,the antifungal effect of 20% Tolclofos methyl,70%thiophanate methyl and other fungicides was relatively poor.The pot experiment results showed that the incidence of disease was only 3.8% after treatment with 10%difenoconazole for 5 days,the disease index was 0.96,and the control effect was 96.5%.The incidence of disease after treatment with 25% tebuconazole was 7.33%,the disease index was 1.83,and the control effect was 93.29%.The effect of the new organic synthetic fungicide of 206.7g/L fenfluramazole on the control of C.chinensis leaf spot was 92.05%.4.The biocontrol strain ES-89 was isolated from the rhizosphere soil of C.chinensis in Enshi.The strain ES-89 was identified as Burkholdria sp.by observing the culture traits and strain morphology combined with 16 S rDNA molecular identification method.The tablet confrontation experiment found that the inhibition rate of pathogens of C.chinensis leaf spot was above 57%.The fermentation supernatant of ES-89 could inhibit the mycelial growth of pathogen,and when the fermentation supernatant and PDA volume ratio was 1:3,the inhibition rate was over 95%.In vitro leaf control experiments showed that the inhibition effect of ES-89 fermentation supernatant was more than 80%.The results of pot experiment showed that the leaf spot of C.chinensis treated with ES-89 fermentation liquid show mild incidence,the incidence rate was only 12.5%,the average disease index was 3.1,and the control effect was 88%.The results showed that ES-89 had a good disease prevention situation and application prospects.