Screening Of High Pathogenicityon Strain Of Beauveria Based On Living Larvae Of Protaetia Brevitarsis

Beauveria are widely used in the biological control of insect pests in the world at present.They have been studied to have good control effects on many agricultural and forestry pests.Liquid-solid biphasic fermentation is commonly used in China to produce conidia of Beauveria.However,if Beauveria is transferred to artificial medium for many generations,its living ability will be reduced and its virulence will be affected.Protaetia brevitarsis larva is the saprophytic insect,which mainly feed on the waste materials such as straw,rotten vegetable leaf,kitchen waste.At present,the ecological cycle mode of the ecological transformation treatment of the Protaetia brevitarsis has been applicated into vegetables,melon and fruit planting bases and so on,using organic waste materials raise a large number of Protaetia brevitarsis.This study makes full use of this associated resource to screen and cultivate Beauveria strains with high pathogenicity,which is of great significance to solve the problem of strain degradation caused by long-term use of fermentative method to produce Beauveria.At the same time,it also improves the utilization value of farmland and kitchen waste.In this study,5 strains of Beauveria B01,B02,B03,B04 and B05,were isolated from different kinds of bombys batryticatu collected in the field.Through a series of screening,the highly pathogenic strains of the larvae were finally obtained.the main findings are as follows:1.Isolation,Purification and Identification of Beauveria From Different SourcesThe isolated and purified strains were inoculated on PDA medium for morphological observation and microscopic examination.Five strains were identified as Beauveria by observing the spore-producing structure and morphology conidia and hyphal.Molecular identification showed that b01-b04 was Beauveria bassiana and B05 was Beauveria brongniartii.2.Observation on Biological Characteristics of Beauveria StrainsThrough the comparison of colony diameter,spore yield and spore germination rate of 5strains of Beauveria,the results showed that there were significant differences in colony diameter and spore yield of 5 strains of Beauveria(P<0.05).The colony diameters of 5strains of Beauveria were as follows:B05>B04>B03>B01>B02.Sporulation was shown as:B04>B03>B01>B02>B05.The spore yield of strain B05 was significantly lower than that of the other four strains(P<0.05).The spore yield of strain B05 was only 0.59×10~7spores/cm~2,on the 15th day and 2.87×10~7 spores/cm~2 of strain B04(P<0.05),and the spore yield of strain B05 was only 0.59×10~7 spores/cm~2 at the 15th day,while that of the strain B04 was 2.87×10~7spores/cm~2.5 strains of Beauveria had no significant difference in spore germination rate(P<0.05),all of which were more than 90%,showing:B04>B03>B05>B01>B02.According to the three indexes of growth rate,spore yield and spore germination rate of five strains of Beauveria,the strain B04 had better biological characteristics.Although the growth rate of strain B05 was significantly higher than that of the other four strains,the spore yield of strain B05 was too little.3.Screening of Highly Virulent Strains From the Larvae of Protaetia brevitarsisFive strains of Beauveria were biotested on the larvae of Protaetia brevitarsis,and the results showed that the mortality and bombys batryticatus rate of the larvae will be affected by different strains and different spore concentration.All the five strains of Beauveria could cause the disease in the larvae of Protaetia brevitarsis,but there were significant differences in mortality and bombys batryticatus rates(P<0.05).The degrees of the mortality rates of the five strains of Beauveria were shown as follows:B05>B04>B03>B02>B01.There was no significant difference between B04 and B05(P>0.05).The degrees of the bombys batryticatus rates of the five strains of Beauveria were shown as follows:B04>B05>B03>B02>B01.The results of bioassay were combined with the biological characteristics of the strain,finally screened out the strain B04 that gained its propagation.4.Effect of two inoculation methods on the infection of BeauveriaDirect impregnation method and acupuncture immersion method were used to treat the larvae of Protaetia brevitarsis.The results showed that the mortality of larvae and the rate of fossilitis were significantly higher in the treatment of acupuncture immersion.The mortality rate was 86.67%and the rate of fossilitis was 83.33%.The mortality rate of larvae treated by direct immersion was 73.33%,and the rate of bombys batryticatus was 70%.5.Effects of Different Temperature and Humidity on the infection of BeauveriaThe mortality and infection rates of larvae caused by Beauveria were significantly different under different degrees of temperature and humidity.When the temperatures were25?and 30?,strain B04 showed strong virulence to the larvae of Protaetia brevitarsis Lewis.There was no significant difference in larval mortality between the two temperatures,but there was significant difference in bombys batryticatus rate between the two temperatures,The rate of bombys batryticatus was the highest at 25?.The rate of mortality and bombys batryticatus were lower when the temperature was 20?.With the increase of relative humidity,the mortality rate of the Protaetia brevitarsis larvae was also increasing.When the relative humidity was 95%,the mortality rate of larvae was the fastest and the infection rate was the highest.Finally,the selected environmental conditions are 25?and 95%relative humidity.6.Toxicity of Beauveria to Monochamus alternatus larvaeIn order to test the virulence of the screened strain to other pests,the virulence of strain B04 to Monochamus alternatus larvae was tested.The results showed that the mortality of larvae increased with the increase of time.After 30 days of treatment,the mortality of larvae increased.The larval mortality rate was 72%,the infection rate was 68%,and that of the control group was 0.5%.