筋骨草内昆虫蜕皮激素类似物对杨干象的杀虫作用

筋骨草内昆虫蜕皮激素类似物对杨干象的杀虫作用 筋骨草内昆虫蜕皮激素类似物对杨干象的 杀虫作用 JournalofForestr. \,Research,l3(3):l77—182(2002) PesticidalcharacterofphBunge(Labiatae)onlarvaeoecdysteroidsfromAjuga ofCryptorrhynchuslapathiL.(Coleop— tera:Curculionidae) CHIDe—fu’, SUNMing—xue,XIAWen—fu. (~CollegeofForestResourcesandEnvironment,NortheastForestryUniversity, Harbin150040,RR.China) (ForestManagementBureau,GreatXing’anMountain,JiagedaqL165000,P.R.China) ~ForestBureau,Lind~an,HeilongjiangProvmce,163000,P.R.China) Abstract:EightkindsofphytoecdysteroidsextractedfromdifferentpartsofAjugamultifloraBunge(Labiatae)thatwereco1. tectedfromdifferentptacesatdifferenttimeweretestedforkillingeffectsonthe2.instarlarvaeofCryptorrhvnchuslapathiLby addingthemtotheartificialdietoflarvae.Theexperimentalresultsindicatedthatadding1—3.mLDhytoecdysteroidstotheartifi. ciatdietcouldlead58%一100%of2一 instarlarvaeofC.1apathitodeathwithin24days.ThephytoecdysteroidsextractedfrOmthe wholeplantofA.multiflorawhichwascollectedbeforefloweringtimeweremuchmoreeffectivethanthoseextractedfrOmthe plantscollectedatfloweringandafterfloweringperiods,andthemodifiedmortalityrateoflarvaereached65. 22%,85.07%,and 98.11%atthedosagelevelof1一mL12一mL1and3一 mLextracts,respectively.TheextractmadefromrootofA.multifloraplant wasmoreeffectiveinkillingefficiencythanthosefromstemandleaves, andtheaveragedeathratesoflarvaewereuDtO1OO% 98.20%and98.32%atdosagelevelsof1一mL12一mL,and3一 mLextracts,respectivelyThekillingspeedoftheextractedphy. toecdysteroidswasslowerthanthatoftrjf}umur0n,hexaftumuronordettamethrinemulsifiableconcentrate. Themortatityrateof larvaeiscloselyrelatedtothefeedingdurationonthedietscontainingphytoecdysteroidsFeedingonthedietswithadditionof phytoecdysteroidsfor16days,morethan80%oftreated2-instarlarvaeofelap athiweretedtodeath.Thekillingeffectofthe extractswaslittteaffectedbythegrowthareasofA.multifloraplantandtheadd ingwaytoartificia}diet. Keywords:AjugamultiforaBunge;CryptorrhynchuslapathiL.Larva;Phyto ecdysteroid CLCnumber:$767.3:S763.38Documentcode:AArt.icleID:1007.662X(2002)03—0177.O6 lntroduction Phytoecdysteroidsareasortofactivenaturalcompounds thatcanregulatethegrowthofinsectpests.andtheir chemicalstructuresaresimilarwiththemoltingpheromone secretedbyinsects.PhytOecdvsterOidsarecontainingin plants(Koreedaetal,1972;Campsetal,1985a,1985b, 1990b;Nieeta1.1987;Qiueta1.1997).Morethan40years ago,scholarsbegantostudythecomposingandcontentof thosecompoundsindifferentplants.aswellastheir physiologicaleffectondifferentinsectsfShimomuraetal, 1970;Bandaraetal,1989:Bathoryeta?0:Campsetal 1990a:Chieta1.1997a:Shaoeta1.1987).Uptonow,more than13Okindsofphytoecdysteroidshavebeenfound.The mostcommoncompoundsare20一OHecdysone.cyaster— one,makisterone,ajugalactone,makisterone,etc.(Darvas efa/19971.Thesecompoundsnotonlycandisturbthe growthandmoltingoftreatedinsects.butalsocanbring themtosterilityeventodeath.Someofthoseph~oecdy— sones,atacertaindosage,actinglikeinsectiuvenilehor- Foundationitenl:ThispaperwassupportedbyNationa1Natura1Science FoundationofChinaI39970620)andthe”IRAP0YT” Biogra1)h. ‘,:CHIDe.tu(1962.J,iliaIc,PhD,professorl11NOllI1castEorestr~ Li1l,ersit\HarbinI50040.RRChinaE.mail’chidefu@0451con1 Receive(Idate:2002—06:O Res1)onsibleeditor:ChalRuiha1 mone,canmakelarvaedevelopintooverageone,and someofthemcaninduceinsectintodiapausefCamps.etal 1990c,1991;Darvas,eta1.1991).Thepesticidalcharacter ofthosephytoecdysteroidsonsomeinsectpests,suchas Closteraanastmosis(Linnaeus),StilpnofiacandidafStaud— inger),Tuberolachnussalignus(Gmelin),Hyphantriacunea (Drury),AporiacrataegiLinneaus,Malacosomaneusteria testaceaMotschulsky,Lymantriad/sparL.,Aphrophora/l门一 termediaUhler.ParthenolecaniumcomiIBouche).Myzus persicae(Sulzer)etc,werealsobeenstudied(Chieta1. 1997a,1997b:Darvaseta1.1997,Shao1987).Highcon— centratedphytoecdysteroidscanleadpeststodeath,while lowerconcentrationusuallyleadinsectstopupation.soitis widelyusedinChinatomakelarvaeofBombyxmorifL.) pupatingsynchronouslyinIaterautumnfNieeta1.1987). Manyplantscontainphytoecdysteroids.Morethan1OO Ajugaspeciesliveintheworldand18speciesdistributein China.MostofthoseAjugaspeciescontainphytoecdy— sones,forexample,AjugadecumbensThunb,A.n~ponen— sisMakino,A.multifloraBungeandA.inearifofiaParmD. Amongthosespecies,A.multifloraBunge,containscom— parativelyhigherconcentrationofphytoendysones(Darvas efa/.1997).T0understandtheusingpotentialofthose PhytOecdysterOidscontainedinA.multiflorainthesustain— ablecontrollingoftrunkborer,thispapermadestudieson thekillingeffectsofthosephytoecdysteroidsonlarvaeof 178CHIDe.fuet?f CryptorrhynchuslapathiL.(Coleoptera:Curculionidae) Materialsandmethods Materials Indianmeal,soybeanflour,powderofpopularbark, mixedvitaminIiquor,Wessonsalt,sodiumbenzoate,dis— tilledwater,andglassplate(15cmx15cm)wereusedin rearingIarvaofC.LapathfL.2一instarIarvaeofC.1apath,L. werecollectedonApriI24.1999.frompoplarstandsin HongqiForestFarm,Daqingcity,HeilongjiangProvince. A.multifloraBunge(Labiatae)plantsusedformakinpow— derandextractsofphytOecdvsterOdswerecollectedfrOm QianshaninLiaoningProvince.ZhalantuninInnerMongolia andJiagedaqiinGreatXing’anMountainareaofHeilongji— angProvince,beforeflowering,atfloweringtime,andafter flowering.Thecollectedplantsweredriednaturallyinthe shade. FivepercenthexafIumuronemulsifiableconcentrate (HEC)wasprovidedbyDalianRazerPesticidesCo.,Ltd; 5%fIumuronemulsifiableconcentrate(TEC)wasPro— videdbvinTOnghuaPesticide&ChemicaIStockCo.,Ltd. and2.5%deltamethrinemulsifiableconcentratefDEC)was boughtfrOmmarket. Methods Reannglarvae. C.Lapa~iLisakindofboringpest.Youngerlarvabores betweenphloemandxylemandeatscambiumandolder lalvadigstunnelsntothexylem.Itsalmostimpossibleto rearthispestonasegmentoftrunkandobserveitsreac— tiontopesticidesindoor.Therefore.anartificiallyrearing methodmightbeadopted.Inourexperiments.anartificiaI dietusedbyHouAiju(1991)wasadoptedbutsomemodi— ficationsweremade.whichisthemostsuitableforthe qrowthofthoseIarvae(seeTable1). Table1.Maincomponentsintheartificialdietforthelarvae MaincomponentsDosageMaincomponentsDosage Indianmeal(g)20Wessonsalt(g)O.5 Soybeanflour(g)4Sodiumbenzoate(m1)0.5 Powderofpoplarbark(g)8Distilledwater(m1)20 Mixedvitaminliquor(g)0.5 Note:1)Mixedvitaminsolutionare1OO?mgnicotinicacid1OO—mg padothenicacid.50一mgriboflavin,25一mgpyridoxine,25一 mgthiamire. and25一mgfolocin,dissolvedin1000一mLwater;2)Wessonsalt:1O.OOg CaCo3,0039gCuSO4?5H2O,1.47gFe3(PO4)2?2H2O,0.02gMnSO4, 500gMgSO4.O009gKAI(SO4)’2H20,6.00gKCI,16.00gKH2PO4, 0005gKI.0.057gNaFand6.00gNaCldissolvedin1000.mLwater;3) Sodiumbenzoatesolutionis0.6sodiumbenzoatedissolvedin1O00m1 water;4)Powderofpoplarbarkisthedriedfreshpopularbarkintheair, thencrushitintofinepowder. Afterblendingingredientsuniformityinaglasscontainer, thisartificialdietwasputbetweentwopiecesofglassplate. Thisfoodisabout0.4—0.5cminthickness.Itwassteamed inasteamerandcoolednaturally.Aftercooling.asmalI holewasmadeinonesideofapieceofdiet.afterthenwe putIarvaeintotheseholesfeachholeforoneIarva.oreach pieceofdietonlyrearedonelarva)andIetIarvaefeedon theartificiaIdiet.BreedingplateswerekeptnananimaI rearingroomatconstanttemperatureof25—28.C.under relativehumidityof6O%一7O%anddarkcondition.Whenever thesedietsweredecomposedorthesetunne1werealmost transfixion.dietswererenewedandtheIivingstatusofthe Iarvaewasrecorded.DletswereusuallYrenewedeveryfive days. MakingphytoecdysteroidsextractsfromAjugapplants NaturallydriedA.multifloraplantwasgroundedtopow der.20一gpowderwasweighedandputintoaglasscon tainer.and15O—mL1OO%methano1wasadded.afterthenit wassonicatedfor5minandcentrifugedfor15minat 2800rpm.Supernatant(methanolphase)wascollected.The sedimentwasextrattedagainwith15O—mLmethano1.The supernatantsobtainedfrOmthesecondextractionwas mixedwithfirstone.Themixedmethanolphasewas evaporatedundervacuum.Aftersolventwasremove~. 20一mLethanolwasaddedtothosedriedextracts.and1.mL methanoIextractscouldbeobtainedfrOmonearamdried materials.Theseextractswerestoredatatemperatureof 一 50.C.Eightkindsofdifferentextractswereobtainedbythis method.Wegiveeachofthemanabbreviationsothatwe canuseiteasilyinthediscussionfseeTable2). Table2.AbbreviationofextractedphytOecdysterOldsmade frOm五iuoamultlflora. Collecting;PartofCollectingtime aCelants AbbreviatiOn Ofextfacts Qianshan Beforeflowering wholeplantFlowering …………曼—_g_ QWBE QWFE QWAE QRFE QSFE QLFE ;rootFlowering istemFlowering ……………一一 i!e_ave_s_…………一E!…一 _ a!a_ntun…一—!曼越…一,Q….. JiagedaqijWholeplantFlowering………. 兰…… JWFE EffectofphytoecdysteroidsonlarvaeofC.1apathfL Thepesticidalactivityofthosephytoecdysteroidswas testedwithconsiderationoftheinfluencesofcollectingtime ofplants,growingplace,ordifferentpartoftheplant.Those extracts,QWBE,QWFE,QWAE,QRFE,QSFE.QLFE. ZWFE.andJWFE.weretakenoutfrOmtherefrigeratorand sonicatedtwotimes(30seach)tomaketheextractssolve sufficiently.TheartificiaIdietwasaddedwithoneoftheex. tractsatadosageof1mLl2mL.and3mLincompanywith addingwater.Atthesametime.weset1一mLl2一mL.and 3?mL5%HEC.5%TEC.and2.5%DECtreateddietsas comparison.Thedietwithadditionof3一mLdistilledwater wastakenascontrastgroup.Weaddedtheextractstothe dietsateachtimethedietswererenewed.Threerepetitions JournalofForestryResearch.13(3):177—182(2002) weresetforeachtreatment.FifteentotwentyIarvaewere treatedineachrepetition. FortestingthedifferenceinmortalityrateofIarvae causedbytreatingperiods.3一mLsonicatedQWFE.3一mL 5%HEC.3一mL5%TEC.and3一mL2.5%DECwasaddedto theartificiaIdietincompanywithaddingwater.andthediet withadditionof3一mLdistilledwaterwassetascontract group.Sixdifferenttreatedperiods,4,8,12,16,2O,and24 days,werearranged.Threerepetitionsweresetforeach treatmentand15—2OheadsofIarvaeweretreatedineach repetition.LivingstatusofIarvaewasobservedandre— cordedevery2days.Totally24dayswasobserved. Thedifferenceinthemortalityratemodifiedbytheways ofaddingphytOecdysterOidstodietswasstudied.Three wayswereusedintermsofaddingthosephVtOecdysterOids Thefirstway,incompanywithaddingwater,wastoadd 1一mL12一mL.and3一mLQWFEtodiet.thenthosedietswere steamedandIarvaewereinoculatedjntoit.Thesecondway wastodilute1一mL.2一mLor3一mLQWFEin1O—mLwater andseparatelyinjectedthemintoacookeddietslowlyusing injector.ThesemoistdietswereputjnashadyroomtoIet waterevaporatenaturally.Whenthewatercontentofdiet droppeddowntoiustcookedIevel,Iarvawasintroduced intoit.Thethirdwaywastoadd1,2,and3gofAjugapow— dertothedietbeforejtwascooked.Thispowderwasmade frOmthewholeplantthatwascollectedfrOmQianshanarea atfloweringtime.ThenthisdietwascookedandIarvawas inoculatedintoit.Dietwithadditionofdistilledwaterwas usedascontrast.Threerepetitionsweresetforeachtreat— mentand15tO2OIarvaeweretreatedjneachrepetition. Experimentwasobservedandrecordedevery2days.Diet wasreplacedbynewlycookedoneevery5days. Resultsandanalysis Influenceofgrowingplace,collectingtimeorpartof plant OnApriI24.1999.trunksegmentswerecarriedbackto IaboratoryofNortheastForestryUniversity.LarvaeofG lapath,L.weretakenoutfrOmthosedissectedtrucksand putjntotheholesmadejntheartificiaIdiet.Livingstatusof thoseIarvaewascheckedevery48h.Dletswererenewed 4timeswithin24experimentaIdays.DatarecordedfrOm thisexperimentwascountedandIistedjnTable3. FromthosedataIistedjnTable3.wefoundthatthephy— toecdvsterojdsextractedfromAjugaplantshavepesticidal activities.ORetothreemillilitersofthoseextractsIed58% tO1OO%oftheC.1apathiL.Iarvaetodeath.Amongthe8 kindsofextractsused,QRFEwasthemostefficientone. Themodifiedmortalityratesofthe2一instarIarvaethatfed onthedietswithadditionof1一mLl2一mL.and3一mLQRFE wereupto1OO%.98.2O%and98.32%respectively.This phytOecdysterOids inducedhighermortalityrate.Forexample,whenthediets wereseparatelytreatedwith1一mL,2一mLand3一mLQWBE. cOrrespOndjngIy66.22%,85.O7%and98.11%oftreated Iarvaewerecausedtodeath. IncomparisonwithQWBE,QWFEandQWAE.which werealImadefrOmwholeplantbuttheplantswerecol— Iectedjndifferenttime(seeTable2),QWBE(beforeflower— ing)wasmosteffectivejnkilling2-instarIarvaeofC.1apathi L,andtheQWAE(afterflowering)wascomparativelyinef— ficacy.TheextractsQWAE.intermsofdeathrateoftreated Iarvae,was9-78%,25.45%and23.59%IowerthanQWBE. ThekillingeffectivenessofQWFE(atfloweringtime)wasin themiddleamongthethreekindsofextracts.Thisresult showedthatextractsmadefrOmA.multifloraplantscol— Iectedbeforefloweringwasmoreeffectivethanthoseex— tractsmadefrOmplantsatfloweringtimeorafterflowering. ComparisonofQRFE(root),QSFE(stem)andQLFE (1eaf)treatedgroupsshowedthatQRFEwasmostefficient inkillingIarvaeofGlapath,L.withamortalityrateof treatedIarvaeof1OO%.98.2O%.and98.32%atacorre— spondingdosageof1mL,2mL,and3mL.Themortality rateofthoseIarvaetreatedwithQSFEorQLFEatthecor— respondingdosagewas8.41%tO22.75%Iowerthanthat treatedbyQRFE.Thisresultindicatedthattheextracts madefrOmtherootofA.mulfifloraatfloweringtimewas moreeffectivethanthosemadefrOmstemsorIeaves. TheIarvaetreatedwithextractsQWFE.ZWFEandJWFE whichwerealImadefromwholeplantbutcollectedfrom differentareasatfloweringtime.had66.67%.57.88%and 62.17%mortalityrespectivelyatadosageof1mLand 73.04%.77.13%and70-25%mortalityrespectivelyata dosageof2mL.Morethan88_24%ofIarvaediedafterfed ondietwithadditionof3mLofthoseextracts.Theresults ofthiscomparisonshowedthatthoseextractsmadefrOm. mulfifloraL.collectedfrOmQianshan.Zhalantun.orJiage— daqiatfloweringtimehavesimilarlypesticidalactivityon the2一instarIarvaeoflapath,L. Killingrapidityofthosephytoecdysteroids Inthisexperiment.3一mLQWFE,3一mL5%HEC.3一mL5% TEC.and3一mL2.5%DECwereaddedjntothedietofC. 1apathfL.andtheirkillingeffectswereseparatelytested. Twelvetimeswasobservedwithin24days.TheIiving statusofIarvaewasrecordedevery48h.Thedatare— cordedwascalculatedandplottedasFig.1.Fig.1showed thatthekillingspeedofQWFEwasmuchslowerthanthat ofHEC.TECorDEC.ThemortalityrateofthOseIan,ae treatedbVQWFEwasOinthefirst6daysandwasnOtOver 5O%in16d.ThedeathrateOfIarvaeqOtuptO98.44%jn 24daftertreatment.ThedeathratesOfIan,aewereOver 5O%in1OdintheHEC,TECOrDECtreatedqrOup.Inthe DECtreatedqrOup.32.88%Iarvaediedwjthjn2d,mOre than5O%diedin4dand1OO%djedjn6d. 18OCHIDe.rLIP,”/ Table3.ThemortalityrateoflarvaetakenartificiaIdietcontainingphytOecdv sterOjdsfrOmA.multiflora 1OO 8O 6O 董 4O 2o E 0 24681O121416182O2224 Fig1.Thecomparisonofthemortalityspeedof2-instarlar- vaeofc.1apathitreatedbydifferentchemicals TimeofperiodofthoseIarvaefedontreateddiet Thisexperimentwasinvestigatedevery48hin24days Themodifiedmortalityrateofthoselarvaewascalculated andshownasFig.2.Themortalityratevariedevidently withthetime(days)larvaefedon?