Study On The Role Of Extracellular Enzyme And Effects On Phenoloxidase Of Hemiberlesia Pitysophila Takagi Infected By Entomogenous Pestalotiopsis Disseminata

Hemiberlesia pitysophila Takagi was the great harm insects of pine forests in southern China, With small-bodied, strong masking, and other characteristics,most of them parasiticed in pine’s needle sheath,formulated double protection of sheaths and shelly, Therefore its prevention more difficult than other forest pests. Entomogenous Pestalotiopsis disseminate was a kind of biocontrol by controlling Hemiberlesia pitysophila Takagi,it had the most virulent to Hemiberlesia pitysophila Takagi, it was a kind of fungi by making use of dead scale insects isolated from natural bodies.In this article,the acitivity changes of proteases,chitinase,lipase produced by the isolate GXSTYJ03of Entomogenous Pestalotiopsis disseminate was used to study the pathogenesis in Hemiberlesia pitysophila Takagi,including the effect on prophenoloxidase-activating system of Hemiberlesia pitysophila takagi infected by Entomogenous pestalotiopsis disseminate,in addition,analysised the pathogenic virulence factors and immune response of Hemiberlesia pitysophila takagi infected by Entomogenous pestalotiopsis disseminate,to explore infection mechanisms and pathogenesis, Provided scientific theoretical reference for further application prevention of Hemiberlesia pitysophila Takagi.The main results were as follows:1. Through the study of detecting the activities of protease, chitinase and lipase and virulence produced by different concentrations of Entomogenous Pestalotiopsis disseminate,1×10~7was the best concentration of spore of enzyme production,its result of corrected mortality was higher,the difference was significant from the lower concentrations of spore suspension,the same with1×10~8.That meant1×10~7was the best concentration of spore of enzyme production,its activities of protease, chitinase and lipase were highest,and its insecticidal was best.2. There were significant positive correlations between the virulence and activities of protease,chitinase and lipase produced by Entomogenous Pestalotiopsis disseminate spores, it meant the higher activities of protease, chitinase and lipase,the stronger virulence of strains, the effect of infection of Hemiberlesia pitysophila Takagi was also the best.3. Respectively Using the methods of Folin-phenol, DNS and Nitrophenol to detect the activities of protease,chitinase and lipase produced by Entomogenous Pestalotiopsis disseminate, the results showed that the peak of the protease’s enzyme production was earlier than chitinase and lipase,was day five; Enzyme production peak of chitinase was day seven;Lipase was day eight. The production of protease activities were generally higher than chitinase and lipase.4. The epidermal protein content of Hemiberlesia pitysophila Takagi was85%, chitin content was9%, fat content was6%. The epidermal protein content was higher than chitin content and fat content,such as protein> chitin> fat.5. The temperature optimum of phenoloxidase was35℃, in the range of20-40℃exhibited higher stability;The pH optimum of Phenoloxidase was7.5, in the range of pH6.5-7.5exhibited higher stability;Using L-DOPA as a substrate, when smaller concentration increaseed with the concentration in a concentration in the range; Phenoloxidase exhibited higher activities when in the range of the concentration of1-2mg/mL concentration,the highest when in the concentration of2mg/mL; Over this concentration, The enzyme reaction rate wasn’t increased with increasing substrate concentration,but subsequently decreased.6. Entomogenous Pestalotiopsis disseminate had a significant impact to the prophenoloxidase-activating system of Hemiberlesia pitysophila Takagi. Mycelium lysate had activation to phenoloxidase:Mycelium lysate maked phenoloxidase activity of plasma and blood cell lysates increased; Inhibition: after using laminarin activated phenoloxidase of plasma and blood cell lysates,then added mycelium lysate, phenoloxidase activity subsequently dropped.7. Hemiberlesia pitysophila Takagi appeared apparent increase in activity of phenoloxidase after infected by Entomogenous pestalotiopsis disseminate,it showed Hemiberlesia pitysophila Takagi had some immune response to Entomogenous pestalotiopsis disseminate.Infected Hemiberlesia pitysophila Takagi after injected by laminarin,its phenoloxidase activity level was significantly lower than those didn’t infected,it showed that Entomogenous pestalotiopsis disseminate was provided with suppression function to the prophenoloxidase-activating. system in the course of infection, In order to overcome the immune response of Hemiberlesia pitysophila Takagi.