Study On The Screening Of Excellent Strains Of Entomopathogenic Fungi And Their Control For Curculio Chinensis Chevrolat(Coleoptera:Curculionidae)

Camellia oleifera,Elaeis guineensis,Olea europae and Cocos nucifera are the four largest woody oil species in the world.C.oleifera is a unique woody oil plant in China.Because of their high economic value and strong market competition potential,they are widely cultivated in south of China.At present,the pests and diseases of C.oleifera are becoming more and more serious,and researches on green prevention and control have been received increasing attention.Curculio chinensis is one of the most serious pests of C.oleifera fruit and one of the forestry dangerous pest in China,which greatly affects the yield and quality of C.oleifera fruits.This study screened excellent strains of Metarhizium spp.and Beauveria bassiana for the prevention and control of C.chinensis adults and 5th instar larvae.The main results are as follows:1.At five temperature gradients of 15℃,20℃,25℃,30℃and 35℃,there were significant differences in the biological characteristics of growth rate,sporulation and spore germination rate among different temperatures for each fungal strain（P<0.05）.Ma09 had the fastest growth rate at 30°C and 25℃;The growth rates of Bb01,Ma05,Ma25 and Mac985 were highest at 30℃;Bb02 had the fastest growth rate at 25℃.The sporulation and spore germination rate of the six strains were highest at 25℃and lowest at 15℃.These results indicate that temperature has important influence on the biological characteristics of the six strains.There were also significant differences in the biological characteristics of growth rate,sporulation and spore germination rate among the six strains at specific temperature（P<0.05）.The growth rate,sporulation,and spore germination rate of Bb01 and Ma09 were relatively higher than other strains at five temperature gradients.Different strains showed significant differences in the pathogenicity against C.chinensis adults and 5th instar larvae（P<0.05）.Among them,Bb01 and Ma09 had stronger virulence to both adults and 5th instar larvae of C.chinensis than other strains.The LT₅₀0 of Bb01 and Ma09 to adults were 4.64d and 4.93d,respectively.The accumulative corrected mortality of adults for Bb01 and Ma09 reached 100%on the15th day,and the infection rates of adults were 76.00%and 78.67%,respectively.For the 5th instar larvae,the LT₅₀0 of Bb01 and Ma09 were 2.15d and 3.49d,respectively,the accumulative corrected mortality of both strains reached 100%on the 10th day,and the infection rates were 81.00%and 86.31%,respectively.These results revealed that both Bb01 and Ma09 had the characteristics of high growth rate,sporulation and high spore germination rate as well as strong pathogenicity against C.chinensis,showing potential application values for the biocontrol of C.chinensis.2.The time-dose-mortality model was used to fit the bioassay data,and the lethal dose and lethal time of Ma09 and Bb01 strains to adults and 5th instar larvae of C.chinensis were estimated.With the increase of inoculation time,the LD₅₀0 and LD₉₀of the adults and 5th instar larvae of C.chinensis gradually decreased,and the dose effect increased gradually.After C.chinensis were inoculated with Ma09,the estimated values of Log(LD₅₀)on the 10th day to the 5th instar larvae and adults were3.62 and 5.55,respectively,and the estimated values on the 10th day of Log(LD₉₀)were 5.75 and 8.11,respectively.After the 5th instar larvae and adults of C.chinensis were inoculated with Bb01,the estimated values of Log(LD₅₀)on the 10th day were3.78 and 5.66,respectively,and the estimated values on the 10th day of Log(LD₉₀)were 5.77 and 7.77,respectively.The lethal time of Ma09 and Bb01 to C.chinensis adults and 5th instar larvae all reduced with increasing of conidial dose.When C.chinensis were treated with Ma09 strain of 1.0×10⁵¹.0×10⁸ spores·mL^-1,the 5th instar larvae LT₅₀0 decreased from 6.63d to 1.81d,and the LT₅₀0 of adults decreased from 7.56d to 3.15d.When C.chinensis were treated with Bb01 strain of 1.0×10⁵¹.0×10⁸ spores·mL^-1,the 5th instar larvae LT₅₀0 decreased from 6.89d to 1.09d,and the LT₅₀0 of adults decreased from 8.23d to 2.78d.3.Bb01 and Ma09 were used to control C.chinensis adults in forests.Results showed that the population density of the fungal treated area was not significantly different from that of the control before the spraying of fungi（April）（P>0.05）.There is a certain difference in the density of insects between the treated areas and controls from June to August.In May and September,there was no significant difference of insect population density between fungal treatd and control areas（P>0.05）.The population density of Ma09 and Bb01 treated areas were significantly lower than that in the controls from June to August（P<0.05）.From May to September,the relative control effects of Ma09 on C.chinensis were 34.32%,71.24%,53.14%,60.54%,and13.61%,respectively,the relative control effects of Bb01 were 38.29.%,61.98%,58.87%,72.65%,23.83%,respectively.In the forest,using Bb01 and Ma09 to control5th instar larvae of C.chinensis,the results showed that the death rate of C.chinensis5th instar larvae gradually increased with time after the spraying of fungi,the control effect of introducing the 5th instar larvae after spraying the fungi is better than the 5th instar larvae introduced before spraying the fungi.30 days after spraying bacteria,the death rates in the treatment groups were significantly higher than that in the control groups（P<0.05）.In the Ma09 treatment groups,except that the death rate was 44.67%in the treatment group of 5 days after the introduction of the 5th instar larvae,the mortality rates of the other treatment groups were all over 70%;the mortality rates was above 50%in the Bb01 treatment groups.After 60 days of spraying the fungi,the mortalities of C.chinensis 5th instar larvae in fungal treatment groups were significantly higher than that of the controls（P<0.05）,the death rate was about 50%in the treatment group of the 5th instar larvae introduction 5 days before spraying the fungi,and about 80%in the other treatment groups.After 90 days of spraying the fungi,the mortalities of C.chinensis 5th instar larvae in fungal treatment groups were significantly higher than that of the controls（P<0.05）,the treatment groups of 5th instar larvae introduced on the same day after spraying fungi and larvae introduced 5days after spraying fungi,the mortality of the 5th instar larvae were all over 90%.the mortality rate of the group that introduced the 5th instar larvae 20 days after the spraying of the fungi was about 80%,the mortality rate in the group of the 5th instar larvae introduction 5 days before spraying the fungi was about 60%.The above results showed that spore suspension of Ma09 and Bb01 could continuously control adults and larvae of C.chinensis in the forest.Conclusion:compared with other strains,Ma09 and Bb01 not only had fast growth rate,high sporulation and spore germination rate in a relatively wide range of temperature,but also had a relatively stronger pathogenicity against adults and 5th instar larvae of C.chinensis.Ma09 and Bb01 can effectively reduce the population densities of C.chinensis adults and 5th instar larvae in the forest.Therefore,Ma09and Bb01 have important application values for prevention and control of C.chinensis in mountainous regions where large diurnal temperature difference existed.