Beauveria Bassiana İzolatlarından Kitinaz Enzimi Eldesi ve Bazı Zararlı Böceklere Etkisinin Araştırılması

Abstract

Today, the use of biological control agents in combating agricultural pests has gained serious importance. Microorganism preparations, fungus spores and predator insects are used as biological control agents. The use of entomopathogenic organisms for agricultural pests is costly and difficult to maintain. Today, chitinolytic enzymes are obtained from entomopathogenic microorganisms. Since the enzyme kitinase is the metabolite of living organisms, it is technically useful for storage and use. Beauveria bassiana, the entomopathogenic fungus, has previously been used as a biological control agent and is a proven organism as a biological agent. In this study, partial and advanced purification of chitinolytic enzymes in the various Beauveria bassina strains which has been isolated in the Turkey soil was performed. The experiments were started with 50 different isolates and studies were continued with 10 isolates which were found to be strong in chitinolytic enzymes with shrimp shells on solid medium. Various liquid media have been tested for enzyme purification and Cobalt Medium has been produced by the highest activity. Lüleburgaz-1 (Lül-1) strain showed the highest activity in cobalt medium and the studies were continued with Lüleburgaz-1 (Lül-1) strain. The best production conditions for the partially purified enzyme were 28°C, 72 hours, pH 3.5. The optimum inoculum amount of 1000 μl and the optimum waiting temperature for maximum enzyme activity was determined as 50°C. The purified enzyme was found to be resistant to a wide range of temperature and pH. Ammonium sulfate precipitation, dialysis and column chromatography methods were used for further purification of the enzyme. Decrease in enzyme activity was observed after purification. The molecular weights of partial and advanced purified enzyme extracts were determined by SDS-PAGE method and found to be ~ 44 kDA. Partial and advanced purified enzyme extracts were tested by using dipping and spraying methods on the Galleria mellonella (Lepidoptera: Pyralidae) and Tenebrio molitor (Coleoptera: Tenebriodeae) larvaes, which are warehouse pest model organisms. The dipping method was found to be more effective than the spraying method. As a result of statistical analysis, each method in which the enzyme was applied was statistically different from the control group. As a result of the study, it was concluded that the enzyme extracts applied by dipping method were more effective by looking at the average death time. It is expected that the study will lead to further purification stages and the use of chitinolitic enzymes will be introduced into biological control.