Pathogenicity Of Isaria Cicada To Plutella Xylostella

As a precious traditional Chinese medicine,Isaria cicadae not only has high nutritional value and pharmacological activity,but also can be used in biological control of pests as a pathogenic fungus with strong pathogenicity.In this paper,the Cicada flower samples collected from Guiyang Forest Park were isolated.Morphological characteristics and the gene analysis of tef-1??rpb1 and nr LSU sequence indicated that the strains were Isaria cicadae.To explore the influence of different culture conditions and environmental conditions on the virulence of the strains;to study the difference of metabolites between the two strains by using UPLC-MS/MS detection technology;and to study the insecticidal activity of the crude toxin and metabolites of I.cicadae to Plutella xylostella.In order to provide a theoretical basis for the further study of metabolites of I.cicadae and the mechanism of their insecticidal action.The main results are as follows:1.Identification of strains and insecticidal activity to P.xylostella.Through morphological observation and molecular biological identification,the strains 3716 and GL were identified as I.cicadae.The results of insecticidal activity test showed that the toxicity of metabolized solution of the strain to P.xylostella was stronger,and the mortality of P.xylostella increased with the prolongation of treatment time.At 72h,the lethal rate of 3716 metabolized solution to P.xylostella was 70.00%,that of GL metabolized solution was 78.33%.The toxicity of the mycelia extract of I.cicadae to P.xylostella was relatively small.2.Screening of high toxin production medium and culture conditions.Czapek was selected as the culture medium which was suitable for the growth of strains and had strong pathogenicity to P.xylostella.Through the screening of different nutrients and culture conditions,the optimal culture medium formula of 3716 strain was obtained::sucrose 30 g,beef extract 2 g,K₂HPO₄1 g,Mg SO₄·7H₂O 0.5 g,KCl 0.5 g,Fe SO₄0.01 g,trace elements Na H₂PO₄30 g,vitamin E 0.02 g,distilled water 1 L.he crude toxin of the filtrate cultured under this nutrient condition treated P.xylostella after 72 h,the mortality of P.xylostella reached 88.33%.The optimal medium formula for GL strains is:glucose 30 g,peptone 2 g,K₂HPO₄1 g,Mg SO4·7H₂O 0.5 g,KCl 0.5 g,Fe SO₄0.01 g,trace element Mn SO₄30 g,vitamin C 0.02 g,distilled water 1 L The crude toxin of the filtrate cultured under this nutrient condition treated P.xylostella after 72 h and the mortality of P.xylostella reached 85.00%.Under the condition of 200 ml,20?,p H 8 and liquid culture for 8days,the crude toxin of filtrate had the highest pathogenicity to P.xylostella.The mortality of3716 strain to P.xylostella was 56.67%,and that of GL strain to P.xylostella was 45%.3.The mode of action of strain crude toxin on P.xylostella.Among the 4 extractants selected,the insecticidal active substances in the 3716 strain extracted by dichloromethane had the strongest pathogenicity to diamondback moth,and the mortality rate of diamondback moth was65.00%at 72 h;Under the extraction of carbonized carbon,the pathogenicity of the carbon tetrachloride extract obtained against P.xylostella was 68.33%at 72 h.The results showed that the metabolized crude toxin of the strain had obvious stomach toxicity and repellent effect on P.xylostella.At 72 h,the gastric toxicity of the crude toxin of strain 3716 against the diamondback moth reached 100%,and the avoidance rate was 85.52%;the GL strain crude toxin has a gastrotoxic activity on diamondback moth of 67.145%,the avoidance rate is 84.63%.It can be seen that the crude toxin metabolized by I.cicadae has stomach toxicity and avoidance effects on P.xylostella,but the effect of strain 3716 on P.xylostella is significantly greater than that of GL strain.4.Analysis of metabolites and insecticidal activity to P.xylostella.The results of metabonomics analysis showed that there were 48 metabolites in 5 categories,including amino acids and their derivatives,alkaloids,phenolic acids,lignans,legumes and other categories,among which 17 metabolites were different between the two strains,mainly alkaloids and phenolic acids.In the analysis of metabolic pathway,13 pathways were selected,mainly metabolic pathway.The results showed that the alkaloids indole-3-ethanol and n-methylnicotinamide had high pathogenicity to P.xylostella.The mortality of P.xylostella was 60%and 51.67%respectively at 72 h.5.Physiological and biochemical mechanism of the action of metabolizing crude toxin and metabolizing compound on P.xylostella.The results showed that the activities of glutathione S-transferase?GST?and acetylcholinesterase?ACh E?increased and the activities of carboxylesterase?care?were inhibited with the increase of treatment time,and the protein content of the treated group was significantly lower than that of the control group.The results showed that the activity of ache decreased,the activity of care and GST increased,and the content of protein decreased significantly.