Study On Metarhizium Pathogenic Factors Against Ticks And Their Preliminary Application

Ticks are ectoparasites classified in the class Acari and parasitize on the surface of animals, which are widely spread in all over of China,constricting the development of animal industry. The parasites not only cause degrading of animal production by bloodsucking, but also transmit various pathogens, including viruses, bacteria and parasites such as Theileria and Babesia. Ticks and tick borne diseases usually cause heavy economic loss in husbandry industry. Currently, tick control is predominantly by chemical therapy. However, due to the issues concerning resistance, environmental pollution and public sanitation, the wide application of this method has been limited. Therefore, more and more scientists shed their eyes on immune prophylaxis and biocontrol.As potential factors for biocontrol,entomopathogenic fungi have been widely applied in the control of insect pests in agriculture. However, its use has just started in laboratory in control of nematodes and ectoparasites.This study is taking pathogenic strains as target, focuses on their virulence factors----extracellular protease and chitinase, and their correlation to pathogenicity. Then the individual effects of the extracellular protease and chitinase was evaluated by induction of enzyme yields using selective cultural media. The optimized conditions for chitinase production were determined and crude enzyme was prepared. And the synergy effects in compatible with the microbial pesticide Bt was studied,which provides the basis of development for new biological agents. The major achievements are as following:1. Through study on the pathogenecity of 10 isolates of Metarhizium against unfed larvae of B. microplus and bioassay, Ma002 strain was assessed to be the highest in pathogenecity with lethality of 82.1% and the shortest LT50 of 4.83d.The fermentation of 10 Metarhizium isolates protease and chitinase, the virulence of the strain with different yields of these enzymes was assessed by biometrics.The coefficient correlation of r1,r2 and r0.05 were 0.8183,0.6828,respectively.The results showed that there was a significant correlation, in which the virulence of the strain with high yield of chitinase is significantly higher than that of the protease (r1> r₂).2. The characteristic of Ma002 for producing chitinase was studied by fermentation. The method showed optimal fermentation conditions was that colloid chitin was used as carbon source and peptone as nitrogen source; the inoculating periods was 24 h.The optimized cultural conditions were in flask with 12% volume medium of pH7.5-8.0 at 26℃, culturing for 96 h on 180 r/min shaker. Under these conditions, the chitinase activity increased 57% comparised to the original basal medium from 1.67 U/mL to 2.63 U/mL.3. The effection of chitinase as biological additive was observed.By combination with Bt pesticides under a certain percentage, its synergistic effect was observed. With the increase of the concentration of chitinase, the insecticidal effect also increased. Adding a final concentration of 2 U/mLchitinase to 1.6×102 mg/kg Bt solution the cumulative mortality from 52% to 85.8% and the insecticidal effect could increase by 62.2%, indicating that chitinase could be a synergist to enhance microbial pesticides effect of significantly.