Development Of Beauveria Bassiana Emulsifiable Formulation

Formulation development is the last but hard step for a fungal insecticide.Different formulae from the same funus culture may have very different shelf life,virulence and use convenience.A typical successful formulation is Green Muscle developed from Metarhizium anisopliae var.acridum,which has been widely use against desert locust in very dry grassland in west Africa.Similar formulations have been registered in China such as the oil suspensions by Tianren,Jiangxi,and Chongda,Chongqing.In development of fungal insecticides,emulsifiable formulation and wettable powder are among the hardest due to additive selection,especially selection of surfactants,most of which damage fungal spores.A good surfactant is not only well compatible to the spores, but also well disperseable.Currently,a few of emulsifiable products have been registered outside China,but no Chinese product has been registered in China.The objective of the present study is to develop such a stable emulsifiable formulation of Beauveria bassiana with high performance and long shelf life.In primary sieving,gelatin plates were used for 28 strains isolated from leafhoppers from RCEF Culture Collection.Four of them,Bb897,Bb922,Bb1001,Bb1094,were found to be of excellent extracellular proteinase activity.Among them,Bb1094 had the highest activity,with D/b up to 4.36±0.20.A bioassay with Bb897 against the tea lesser leafhopper showed a extremely significant dose-time effect.Time effect parameter increased from the 5th～9th day after inoculation while the 8th day after inoculation,mortality of the leafhopper reached the peak.Under the concentrations of 10⁷,10⁸ and 10⁹ conidia/ml,LC₅₀'s were at 10^8.25,10^7.19å’Œ10^10.56 conidia/ml,respectively on the 7^th,8^th and 9^th day.As compared to control,soybean oil,corn oil,sunflower oil,camellia oil,peanut oil, 15# white camphor oil,except rape seed oil,3# and 10# white camphor oil,exerted not only no damage on Bb897,but also provided obvious protection.Among 6 non-ion type surfactants with HLB value within 10～13,which were mixed with B.bassiana conidia for 3 month storage,only A had no significant impact on conidium germination,with excellent dispersancy:dispersal index I at 0.25 for a concentration of 0.5%,and at 0.27 for 1%.Therefore,the key technique was solvedAll UV protectants did not exert significant influence,and showed a certain degree of UV protection.Among them,nano class TiO₂ had obviously higher protection of spores than nano class ZnO,clay,and defatted milk powder.Irradiated by UV light for 30min, germination of pure conidial powder without TiO₂ declined by 46.5%,while that of conidia mixed with 1%TiO₂ did not decline significantly,and that of conidia mixed with 2%TiO₂ did not decline.V_B,V_C and V_E did not exert obvious impact.V_C at either 0.1mg/ml or 0.2mg/ml showed excellent free radical scavenger activity,i.e.,anti-oxidation ability.In the combination test of all factors involved,all combinations did not exert significantly different effect on germination and exerted almost no effect on anti-UV,but exerted significantly different effect on anti-oxidation.The amount of A and V_C affected substantial anti-oxidation,but TiO₂ almost did not affect.As for dispersancy,combination 8 showed stable dispersing effect at either 0 min,or 10 min or 30 min,displaying an optimal combination at A=1.225,VC=8.040,TiO₂=0.804, with germination at 82.8%,UV protection at 87.5%,anti-oxidation at 93.0%,and dispersion＜1.