| Osier weevil(or willow beetle;willow weevil)(Cryptorhynchus lapathi L.)is considered to be an economically important pest of Salicaceae plants in most North American and European countries,as well as in some Asian countries such as China,Korea and Japan.Since the larval and pupal stages of Osier weevil living in the bark or xylem of trees,controlling of this pest is comparatively difficult.At present,in the forestry production,the prevention and control of the pests still mainly rely on chemical pesticides.Biological control of this pest was seldom been studied.In this paper,researches were carried out to find out the relationship between the resistance level and strains,source of parents,the physical properties of xylem and bark.The influence of plant volatile or plant essential oil on the behavioural responses of C.lapathi L was also concerned.The stronger virulent strains of Beauveria bassiana had strongest pathogenicity to C.lapathi L.larvae was selected,the process and pathogenicity of B.bassiana infecting C lapathi L.larvae was studies as well;Biological control was carried out by uesing Beauveria bassiana,Dastarcus helophoroides,Scleroderma guani in the field experiments;We expect to find the effective means to control the damage of C.lapathi L.1.In this study,7 entomopathogenic fungi strains were tested for pathogenicity to C lapathi L.larvae in the laboratory,and the pathogenic symptoms was observed.The results indicated that strains CFCC87323 of Beauveria bassiana had strongest pathogenicity to C lapathi L.larvae and the strains ACCC30830 took the second place.Using CFCC8732 conidiospore suspension at the of concentration for 1.0×108 conidia·mL-1 to infect larvae of C.lapathi L.in the lab,the mortality ratio of larvae was 93.83%,the adjusted mortality ratio of larvae was 93.38%,the quickest median lethal times(LT50)was 3.536 d,the median lethal concentration(LC50)was 7.0×103 conidia·mL-1;Using ACCC30830,ACCC3083,CFCC87327 conidiospore suspension at the of concentration for 1.0×108 conidia-mL-1 to infect larvae of C.lapathi L.in the lab,the mortality ratio and the adjusted mortality ratio were greater than 70%in the lab and in the field experiments.Results indicated that this four strains showed pathogenic effect to C lapathi L.larvae.In the observation test of pathogenic symptoms,from the strains CFCC87323 beginning to infect the larvae of C lapathi L.to infect obvious symptoms to their cadave,might take 74.67 h,from the strains CFCC87323 beginning to infect the larvae of C.lapathi L.to their hyphae covering the whole body,might take 93.67±3.21 h.This indicated that the strain CFCC87323 showed good pathogenic effect to C.lapathi L.larvae.2.Sanning electronic microscope(SEM)and transmission electron microscopy(TEM)were used to observe the process and ultrastructural changes of body wall of C.lapathi L.larvae infected by B.brongniartii(Sacc.)Petch.The results of SEM observation indicated that at 12 h after inoculation,spores adhered on different areas of body wall of the C.lapathi L.larvae.such as in the spiracles,around sensilla and on internode membrane.At 24 h after inoculation,hypha or germ tubes of B.brongniartii(Sacc.)Petch penetrated into the integuments of C.lapathi L.larvae,and no appressoria was found.At 90 and 96 h after inoculation,hypha penetrated out from the surface of C.lapathi L.and began to multiply rapidly.Results of TEM observation indicated that the penetration of hypha was the combination of mechanical pressure and histolysis.At 60 h after inoculation,cuticula was separated from epidermis,a lot of hypha penetrated into the epidermis,organelles near hypha was dissolved,and cell nucleuses was destroyed and flocculent of solutes appeared around the hypha.Those phenomena had never been reported before.It could be confirmed that enzymatic action was involved in the process of B.brongniartii(Sacc.)Petch intruding into C.lapathi L.larvae.At 84 h after inoculation,hypha massively self-reproduced by non-septa or one-septa or two-septa ways in blood cavity.3.The dynamic of total hemolymph cell and phenoloxidase activity of C.lapathi L.larvae infected by B.bassiana was studied.The results indicated strong immune response and increased for TCH concentration of C.lapathi L.larvae and adult infected by the trains CFCC87323 of Beauveria bassiana.The greater TCH concentration of C.lapathi L.the stronger immune response.The strongest immune response and increasing of phenoloxidase activity of C.lapathi L.larvae and adult infected by the trains CFCC87323 of Beauveria bassiana,were occurred at 24-28 h and 32-36 h after infection respectively.4.Electroantennogram(EAG),Y-tube olfactometer bioassays,and wind tunnel bioassays were conducted to test the electrophysiological and behavioral responses of C.apathi L.to 13 kinds of plants volatiles to select effective individual plant volatiles or essential oils.Results indicate that male and female C lapathi L.showed significantly strong EAG responses to different concentrations of linoleic acid,a-pinene,(Z)-3-hexen-l-ol,geraniol,turpentine,and salicylaldehyde.In Y-tube olfactometer bioassays,salicylaldehyde and a-pinene elicited significantly high repellent effects on the female and male C.lapathi L.Linoleic acid and(Z)-3-hexen-l-ol showed significantly greater luring effects on the male and female C lapathi L.compared with the control.Turpentine showed only significantly higher repellent effects and geraniol only showed significantly greater luring effects on the male C lapathi L compared with the control.In wind tunnel bioassays,different concentrations of salicylaldehyde,a-pinene,and turpentine elicited significantly higher repellent effects on both male and female C.lapathi L.compared with the control.Different concentrations of linoleic acid,(Z)-3-hexen-l-ol,and geraniol showed significantly greater luring effects on both male and female C.lapathi L.compared with the control.These results have provided a theoretical basis for the further development of C.lapathi L.luring and repellent agents.5.The population density of C lapathi L.on different poplar strains,percentage of infested poplar strains,as well as the hardness of xylem and phloem,thickness of phloem,diameter at breast height(DBH),the source of parent of 200 different 2 to 3,5 to 6,10 to 11 and 11 to 12 years old poplar strains was studied,and following conclusions were gotten:the phloem and xylem hardness was harder,their resistance level to C.lapathi L.were higher.The phloem and xylem hardness were softer,their pest resistance level was lower.If the diameter at breast height(DBH)of 2 to 3,5 to 6 years old poplar strains was larger,then its resistance ability might be lower.If the DBH of 10 to 11,11 to 12 years old poplar strains was larger,then its resistance ability might be higher.According to comprehensive analysis of relationship between source of parents and all poplar strains at different resistance level,P.pseudo-simonii,P.cathayana,P.simonii,p.suaveolens strains were resistance to C.lapathi L.P.deltoids strains were susceptible.6.Smearing Fungal spore suspension on trunk and non-woven fabric band methods was used in the field experiments on the control of C.lapathi L.The adjusted death rate of C.lapathi L.larvae caused by infection of strains CFCC87323 by smearing on the trunk was 77.62%.The adjusted death rate of C.lapathi L.larvae caused by infection of strains CFCC87323 by non-woven fabric band method was 63.21%;The adjusted death rate of C lapathi L.larvae caused by infection of strains ACCC30830 by smearing on the trunk was 66.06%.The adjusted death rate of C.lapathi L.larvae caused by infection of strains ACCC30830 by non-woven fabric band method was 53.7%.The adjusted death rate of C.lapathi L.larvae caused by infection of strains ACCC30831 and CFCC87327 by smearing on the trunk were more than 50%.The adjusted death rate of C.lapathi L.larvae caused by infection of strains ACCC30831 and CFCC87327 by non-woven fabric band method were more than 35%.The results indicated that strains ACCC30831 and CFCC87327 showed certain pathogenic effect to C.lapathi L.larvae in the field experiments.The mortality rate of C.lapathi L.larvae caused by reaeasing of Dastarcus helophoroides adults in the forest by 1:2 ratio was more than 40%.Those results indicated that those entomopathogenic fungi strains and Dastarcus helophoroides may serve as a potential source of biocontrol agents for C.lapathi L.. |
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