Screening Of A High Virulent Isolate Of Beauveria Bassiana And The Pathogenesis To Apriona Germari Larvae

In this study, a high virulent isolate Beauveria bassiana Bb00 to Apriona germari larvae was screened from the isolates collected from the soil using the "Tenebrio molitor Bait Method". To this isolate, the biological characteristics, the pathogenicity against A.germari and factors affecting the pathogenicity, infection process and defense response of the host to the pathogen, the pathological changes of A.germari larvae infected by B.bassiana, as well as the compatibility of some pesticides with B.bassiana were studied. The purposes of this study were to establish the pathogenesis of B.bassiana to the A.germari and interaction between B. bassiana and A.germari, to evaluate the potential of the fungus as a biocontrol agent for A.germari, which can provide the basis for the application of the fungus in the integrated control of A.germari. The results showed that:1,The screening of a high virulent isolateNine B.bassiana were isolated from different soil using the "Tenebrio molitor Bait Method". The isolation rate was 45%. The pathogenicity of different strains to A.germari larvae was tested. The result showed that Bb00 was the most virulent one. The correct mortality was up to 100% and the infection rate was 86.3%. LT₅₀ was 4.13 days, and LC₅₀ on the A.germari larvae was determined to be 3.05×10⁶/mL.2,The study of biological characteristics of Bb00The most suitable media for the mycelia growth and conidia production of Bb00 were PPDA and Czapek, respectively. For the nutrient growth and the conidia germination, the optimal temperature and humidity were 25℃and 100%, respectively. Conidia in the nutrient liquid of 2%glucose＋1%peptone germinated rapidly, with high germination rate. The ultraviolet ray has no significant impact on the nutrient growth, but highly restrained the conidia germination. In addition, the illumination showed unimportant effect on the conidia germination.3,The study of pathogenicity to A.germari and factors affecting the pathogenicity The Bb00 can not infect the A.germari eggs, and has a low infection of 24% to adults. At 25℃and RH 100%, the pathogenicity reached the highest. In the test range, the pathogenicity of the successive reisolates obtained by repeatedly passing through the Czapek reduced, while obtained by repeatedly passing through the A.germari larvae increased.4,The study of infection process and defense response of the host to the pathogenPenetration process of B.bassiana include adherence, germination, appressoria and penetration. The conidia on the cuticle of A.gerrnari larvae germinated in 16h after inoculation. The hyphae penetrated the cuticle and propagated in the coelom. Then it infected and destroyed the cutex, hemloymph lipoid and muscles, alimentary tract and malpighian tubule, etc. The phenoloxidase activity in hemolymph of A.gerrnari larvae infected by B. bassiana increased within the early days of infection, but followed by decreasing because of the adaptability of B.bassiana. The phenoloxidase activity in hemolymph of A.germari larvae infected by successive reisolates obtained by repeatedly passing through the Czapek medium was higher than that by repeatedly passing through the A.gerrnari larvae. There was some correlation between the maximal time producing phenoloxidase activity and LT₅₀ of different reisolates. At the same time, a high relationship between the maximal activity and the LC₅₀ was also found. All this indicated that the phenoloxidase activity in the hemolymph of A.germari larvae can not only weigh the immunity ability of the host in the process of B.bassiana infection, but also act as an important reference index of pathogen. The hemocytes of A.germari larvae can resist the invasion of B.bassiana. The immune response mainly include adhesion, phagocytosis, aggregation, encapsulation and nodule formation. The peak response times were 2～2.5d after invasion of B.bassiana. The defense responses were just temporarily inhibition for the pathogen but could not clear them completely.5,The study of pathological changes of A.germari larvae infected by B.bassiana The food consumption, weigh gain and efficiencies of conversion of ingested food (ECI) of A.germari larvae infected by B. bassiana were significantly lower than the control. To resist the B. bassiana, the respiration increased 3 days after inoculation, and 5 days later, the resist system of the insect was destroyed and respiration metabolism level was decreased. When the larvae were near to dead at 7 days, the respiration decreased mostly. The content of protein in infected A.gerrnari larvae increased in the early days of infection, and decreased with the hypha growth in insect haemolymph while some protein was consumed or organization organ of protein was destroyed. Before the dead of insect, the change trend of free amino acids content in A.germari larvae was decreasing followed by increasing. But the different amino acids changed differently, indicating that different growth phases of isolates needs different amino acids. The activity of SOD, CAT and POD of A.gerrnari rised in the infected initial stage while dropped in the infected later stage. The activity of POD was always higher than the control, implied an important acting of POD in the resisting against B.bassiana. Meantime the activity of carboxylesterase, glutathione-S-transferase and acetylcholinesterase in A.germari larvae infected by B.bassiana were changed. Carboxylesterase and glutathione-S-transferase activity increasing followed by decreased. But the change of acetylcholinesterase activity was more complex and expressing restraining and activation alternatively.After spores penetrated into the hemocoele, as the hyphae propagated and grew in the coelum, pathological changes occurred on all infected tissues and organs, including cuticle separating from epidermis, incompaet of fat body, disorganized digestive tract, crack in the muscle tissue and the disintegrated trachea tissue.6,The study of the compatibility of some pesticides with B.bassianaFive widely used pesticides were assessed for compatibility with B.bassiana. The effects of these pesticides on spore germination and mycelial growth were quantified. The result showed that Mieyouniao No.3 has the lowest restraining rate to the spore germination and insignificantly effect on the myeelial growth at the low concentration. So Mieyouniao No.3 was thought as the best pesticide to blend with B.bassiana.