Influence of host preference, mating, and release density on the parasitism of Telenomus remus (Nixon) (Hymenoptera, Platygastridae)

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REVISTA

BRASILEIRA

DE

Entomologia

AJournalonInsectDiversityandEvolution

w w w . r b e n t o m o l o g i a . c o m

Biological

Control

and

Crop

Protection

Inﬂuence

of

host

preference,

mating,

and

release

density

on

the

parasitism

of

Telenomus

remus

(Nixon)

(Hymenoptera,

Platygastridae)

Ana

Paula

de

Queiroz

a

_,

_Adeney

_de

_Freitas

_Bueno

b,∗

_,

_Aline

_{Pomari-Fernandes}

c

_,

Orcial

Ceolin

Bortolotto

d

_,

_Adriana

_Yatiem

_Mikami

d

_,

_Lopes

_Olive

e

a_Instituto_Agronômico_do_Paraná,_Londrina,_PR,_Brazil

b_Empresa_Brasileira_de_Pesquisa_{Agropecuária,}_Londrina,_PR,_Brazil c_Universidade_Federal_da_Fronteira_Sul,_Laranjeiras_do_Sul,_PR,_Brazil

d_Universidade_Federal_do_Paraná,_Setor_de_Ciências_Biológicas,_Departamento_de_Biologia,_Curitiba,_PR,_Brazil e_Universidade_Federal_de_Rondônia,_Porto_Velho,_RO,_Brazil

a

r

t

i

c

l

e

i

n

f

o

Articlehistory:

Received6September2016

Accepted16December2016

Availableonline29December2016 Associateeditor:DanielSosaGómez Keywords: Arrhenotokousparthenogenesis Optimalnumber Phenologicalstages Preimaginalconditioning

a

b

s

t

r

a

c

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Weevaluatedtheinfluenceofhostpreference,mating,andreleasedensityonTelenomusremus(Nixon, 1937)(Hymenoptera:Platygastridae)parasitizingeggsofSpodopterafrugiperda(Smith,1797) (Lepi-doptera:Noctuidae).First,wetestedhostpreferenceofT.remus(freechoicetest)offeredachoicebetween eggsofCorcyracephalonica(Stainton,1865)(Lepidoptera:Pyralidae)andS.frugiperda.Parasitism capac-ityandhostpreference(S.frugiperda)ofT.remusrearedoneitherofthetwohostsdidnotdiffer.Secondly, weevaluatedtheinfluenceofmatingbehaviorofT.remusfemalesonitsparasitism.Onlytheoffspring sexratiodifferedbetweentreatments,indicatingthatthespeciesreproducesbyparthenogenesisofthe arrhenotokytype.Finally,weevaluatedtheinfluenceofreleasedensityonT.remusparasitism.Thiswas testedbyreleasingdifferentnumbersoftheparasitoidperS.frugiperdaeggusingT.remusrearedfor differentnumbersofgenerationsonC.cephalonicaeggs.Theregressionanalysisbetweenpercentageof parasitismanddensityofreleasedT.remusfemalesshowedaquadraticeffectforalltestedparasitoid generations(F35,F40,andF45)withmaximumparasitismfrom65.07%to71.69%.Ourresultsallowthe

conclusionthat(a)T.remusprefersS.frugiperdaeggs,regardlessofthehostonwhichthisparasitoidwas reared,showingnopreimaginalconditioning;(b)Matingdoesnotaffectthenumberofeggsparasitized byT.remusorthedevelopmentofitsoffspring;and(c)TheoptimalT.remusreleasedensitywhenreared onC.cephalonicaisbetween0.133and0.150females/S.frugiperda.

Introduction

Telenomusremusparasitizeseggsofvariousspeciesoftheorder Lepidoptera,many of whichare globalcroppests(Cave, 2000). Despitepossessingtraitsfavorablefortheiruseasbiological con-trol,thisparasitoidiscurrentlyonlyrearedonasmallscaledueto thedifﬁcultiesofrearingitonitsnaturalhostSpodopterafrugiperda (Pomari-Fernandes et al., 2015). Alternatively, T. remus can be rearedonafactitioushoststhatmaynotbetheparasitoid’s pref-erencebutisstilladequateforitssuccessfuldevelopment(Parra, 1997).Inthiscontext,Corcyracephalonica,whichcanberearedin thelaboratorymoreeasilyandatalowercostthanS.frugiperda (Kumaretal.,1986),hasbeensuggestedasapossiblefactitious hostofT.remus(Kumaretal.,1986;Pomarietal.,2012).However,

∗ Correspondingauthor.

E-mail:[email protected](A.F.Bueno).

continuousrearingofaparasitoidonafactitioushostmayaffectits parasitismorhostpreference,andmaydirectlyinﬂuenceits efﬁ-ciencyagainstthetargetpest.Thisisprobablyduetopreimaginal conditioningoccurringduringlarvaldevelopment(Cobert,1985), abiologicalprocesswhichneedsfurtherstudyforT.remusreared onC.cephalonicaeggs.

Knowledgeofthebiologyandecologyofinsectsandtheir natu-ralenemiesisaprerequisiteforthesuccessofbiologicalcontrolin IntegratedPestManagement(IPM)(Cave,2000).Adultmatingcan impactparasitism,andshouldbetakenintoaccountwhentesting theuseofaneggparasitoidinmassiveﬁeldreleases(Pratissolietal., 2009).MalesofT.remushaveonelarvalinstarlessthanfemales (Cave,2000),andthereforeemergeearlierthanfemalesfromthe samehosteggmass.Thenewlyemergedmalesguardeggmasses toensuretheirmatingwithfemalesassoonastheyemerge(Cave, 2000).Becauseparasitismcapacitymaydifferbetweenmatedand unmatedT. remusfemales, theinﬂuence of matingatthe time ofemergenceshouldbeassessed priortoadoptingtherecently

http://dx.doi.org/10.1016/j.rbe.2016.12.004

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developedtechnologyofaerialreleaseofindividualpupaecloseto emergence.

Additionally,superparasitismcandecreasethenumberof par-asitized eggs, and may occur when an excessive number of parasitoidsperhosteggisreleased(Cave,2000).Researchisneeded totesttheinfluenceofreleasedensityonparasitismwiththe long-termgoalofdeterminingtheoptimalnumberofparasitoidstobe releasedintothefield(SáandParra,1993).Tothisend,thisstudy evaluatestheinfluenceofhostpreference,mating,andrelease den-sityonT.remusparasitismoneggsofS.frugiperda.Theresultsyield crucialinformationforthesuccessofrearingT.remusanditsrelease inthefield.

Materialandmethods

Thestudies(bioassay1and2)werecarriedoutundercontrolled laboratoryconditions (25◦C±2◦C, relative humidity 80%±10%, photoperiod14/10h[light/dark])andinasemifield(greenhouse) (bioassay3)atEmbrapaSoybean,Londrina,StateofParaná,Brazil. Thisworkinvolvedthreeindependentbioassays.Inthefirst bioas-sayweassessedhostpreferenceoftheparasitoidT.remusoffered achoicebetweeneggsofC.cephalonicaandS.frugiperda.Inthe secondbioassayweevaluatedtheinfluenceofmatingonT.remus parasitismofS.frugiperdaeggs.Inthethirdbioassaywedetermined theoptimalnumberofparasitoidstobereleasedforthe success-fulcontrolofS.frugiperdainmaize.Allhostsandparasitoidsused intheexperimentswereobtainedfromtherearinglaboratoryat EmbrapaSoybean.

Bioassay1:hostpreferenceofTelenomusremus

All T. remus colonies evaluated in the host preference test originatedfrominsectsreared onC.cephalonica eggsattheF40

generationandonS.frugiperdaeggsattheF350generation(Parra, 1997)inordertocomparethetwoinsectpopulations.The exper-imentwascarriedoutinacompletelyrandomized2×2factorial design(2parasitoidcoloniesby2hosteggs)and15replicates.Each replicateconsistedofanarenaaccordingtoThuleretal.(2007):a polyethylenebottle(4and2cmindiameter)forparasitoidrelease wasplacedinthemiddleofthearenaaroundwhichfourtubes (vol-ume1.5mL)containingthehostspecimenswerearrangedatequal distances.

Approximately 150 eggs of a single host (C. cephalonica or S. frugiperda) were glued to labeled, white cardboard cards (2.5×5cm)withwhiteglue (Tenaz®).Twocardsper hostwere individuallyintroducedintothetubeslocatedonoppositesides ofthearena.Next, fournewlyemerged (24h)T. remusfemales fedonhoneywerereleasedintothearenafromthecentral bot-tle.After24h,thecardswereremovedandplacedindividuallyin 1.5mLDurantubesuntiltheemergenceofadults.Atthisstep,the numberofparasitizedeggswasrecorded.

Theresults(Table1)wereanalyzedfornormality(Shapiroand Wilk,1965)andhomogeneityofvarianceoftreatments(Burrand Foster,1972)and,whenevernecessary,transformedtoperform ANOVA.Thenumberofparasitizedeggswastransformedbylog (x+1).ThetreatmentmeanswerecomparedusingTukey’stestat aprobabilitylevelof5%(SASInstitute,2009).

Bioassay2:inﬂuenceofparasitoidmatingonitsparasitism

MatedandunmatedT.remusfemalesrearedonC.cephalonica eggs(F40generation)wereofferedeggsofS.frugiperdafora24h

period. Each female was placed in a Duran type tube (1.5mL) containingadropletofhoneyasfood,andofferedapproximately 100S.frugiperdaeggs(upto24hold)gluedtowhitecardboard cards(2.5×5cm)(sixreplicatespertreatment).Afterwards,cards

Table1

Bioassay1:numberofeggs(SpodopterafrugiperdaandCorcyracephalonica) para-sitizedbyTelenomusremusfromdifferentcolonies(rearedonS.frugiperdaandC. cephalonicaeggs).Temperature25±2◦C,relativehumidityof80%±10%,andthe photoperiodof14/10h(light/dark).

Host Parasitoidcolony T.remusreared onC. cephalonica T.remusreared onS.frugiperda Mean C.cephalonica 2.29±1.03 0.00±0.00 1.19±0.57b S.frugiperda 94.36±12.71 81.08±13.53 87.96±9.18a Mean 48.32±10.85A 40.53±10.47A CV(%) 39.15 Fparasitoid 1.57 Fhost 259.46 Fparasitoid*host 1.73 pparasitoid 0.2161 phost <0.0001 pparasitoid*host 0.1950 DFparasitoid 1 DFhost 1 DFparasitoid*host 1 DFtotal 53

Means(±SE)followedbyidenticalupper-caseletterswithinarow(different par-asitoidcolonies),andlower-caseletterswithinacolumn(differentegghosts)did notstatisticallydifferaccordingtoTukey’sStudentizedrangetestat5%probability. Originaldatafollowedbystatisticsperformedondatatransformedinlog(x+1).

wereremovedandplacedindividuallyinDuran-typetubesuntil theemergenceofadults. Thenumberof parasitizedeggs, para-sitoidemergence(%),sexratio,andlongevityofparentalfemales wasrecorded. The results(Table2) wereanalyzed for normal-ity (Shapiro and Wilk, 1965) and homogeneity of variance of treatments(BurrandFoster,1972)and,whenevernecessary, trans-formedtoperformANOVA.Thenumberofparasitizedeggswas transformed by log (x+1) and parasitoid emergence (%) trans-formedbyarcsin

X/100.Then,treatmentmeanswerecompared byTukey’stestataprobabilitylevelof5%(SASInstitute,2009).

Bioassay3:inﬂuenceofT.remusreleasedensityonitsparasitism

Theinﬂuence ofthe releasedensity of T.remus onits para-sitismwasdeterminedbyreleasingdifferentnumbersofT.remus inrelationtoagivennumberofeggsofS.frugiperda,usingT.remus rearedforadifferentnumberofgenerations(F35,F40,andF45)onC.

cephalonicaeggs.WechoseT.remusrearedonC.cephalonicaeggs becausethisparasitoidisplannedtobereleasedintheﬁeldona largescaleforbiologicalcontrolprograms.Anindependenttrial wascarriedoutforeachparasitoidgenerationundergreenhouse conditions,usingafullyrandomizedexperimentaldesignwithten treatments(0,0.017,0.033,0.050,0.067,0.083,0.100,0.117,0.133, and0.150T.remusfemalesperS.frugiperdaegg)andﬁvereplicates. Eggs of the pest species were obtained from laboratory rearing and exposed to treatments inside iron-framed cages (40cm×40cm×120cm)coveredwithvoilefabricineach repli-cate. A pot of 40-cmdiameter withtwo hybrid IPR114maize plants wasplaced inside eachof thesecages,attachingawhite cardboardcardwith150eggstothewhorlofeachplant.Different numbers(5,10,15,20,25,30,35,40and45)ofpreviouslymated females(Pomarietal.,2013)werereleased,representingthe pro-portionsof0.017,0.033,0.050,0.067,0.083,0.100,0.117,0.133,and 0.150T.remusfemalesperS.frugiperdaegg.Noparasitoidswere releasedinthecontroltreatment.Parasitismwasallowedfor24h, andtemperatureandhumiditywererecordedusingadigitaldata logger(InstruthermHT-500)(Table3).Next,eggswerecollected andmaintainedinPetridishesat25◦Cuntiltheydarkenedand parasitoidsemergedforsubsequentevaluation.Werecordedtotal percentofparasitismperreplicate.Thisprocedurewasrepeated

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Table2

Bioassay2:biologicalparametersofmatedandunmatedTelenomusremusfemales(fromCorcyracephalonicaeggs)thatparasitizedSpodopterafrugiperdaeggs.[Temperature of25±2◦C,relativehumidityof80%±10%,andthephotoperiodof14/10h(light/dark).].

Treatment Numberofparasitizedeggs1 _Parasitoid_emergence_(%)2 _Sex_ratio _Longevity_of_parental_females_(days)

Matedfemales 53.33±5.68ns _99.13_±_0.26ns _0.61_±_0.04_a _5.63_±_0.34ns Unmatedfemales 46.00±5.27 98.78±0.81 0.00±0.00b 6.08±0.18 CV(%) 11.84 4.46 19.88 10.52 p 0.3506 0.9264 <0.0001 0.2835 F 0.98 0.01 253.13 1.32 DFerror 8 8 8 8

1 _Means_(±SE)_followed_by_identical_upper-case_letters_within_a_row_(different_parasitoid_colonies),_and_lower-case_letters_within_a_column_(different_egg_hosts)_did_not

statisticallydifferaccordingtoTukey’sStudentizedrangetestat5%probabilityOriginaldatafollowedbystatisticsperformedondatatransformedinlog(x+1).

2 _Original_data_followed_by_statistics_performed_on_data_transformed_by_arcsin

_X/100. ns _ANOVA_not_{signiﬁcant.}

Table3

WeatherdatarecordedbyadataLoggerduringbioassay3(inﬂuenceoftheT.remusreleasedensityonitsparasitism).

Generation–stadium Temperature Relativehumidity

Phenological Minimum Maximum Average Minimum Maximum Average

F35–V2/V3 19.8 27.1 23.45 47.3 95.5 71.4 F35–V4/V5 23.2 36.8 30 54.3 85.1 69.7 F35–V8/V9 25.6 38.4 32 49.1 90.2 69.6 F40–V2/V3 23.3 29.3 26.3 53.6 84.2 68.9 F40–V4/V5 21.4 36.4 28.9 40.1 90.1 65.1 F40–V8/V9 19.3 26.7 23 77.1 95.8 86.4 F45–V2/V3 21.8 38.8 30.3 52.1 96.1 74.1 F45–V4/V5 24.9 37.3 31.1 55.2 80.6 67.9 F45–V8/V9 21.8 38.8 30.3 52.1 96.1 74.1

atvariousdevelopmentalstagesoftheplants(V2/V3,V4/V5,and V8/V9accordingtoMagalhãesandDurães,2006)becausedifferent leafsurfacesmightprovidedifferentsizesofareaforaparasitoid toﬁndeggsandthereforemayimpactitsparasitism.Theaverage parasitismobservedforeach parasitoiddensityduringdifferent plantstages wasusedintheanalyses(Fig.1).Thenumber ofT. remusfemalespereggofS.frugiperdaandpercentageofparasitism wereusedinaregressionanalysis(SASInstitute,2009).Parasitism datawas analyzedfor normality(Shapiro and Wilk, 1965) and homogeneityofvarianceoftreatments(BurrandFoster,1972),and whenevernecessarytransformedtoperformANOVA.Parasitism valuesofT.remusgenerationsF40 andF45weretransformedby

(x+1)0.5_._The_treatment_means_were_then_compared_by_Tukey’s_test

ataprobabilitylevelof5%(SASInstitute,2009).

Results

Bioassay1:hostpreferenceofTelenomusremus

TherewasnointeractionbetweenT.remuscolonies(T.remus reared on C. cephalonica or S. frugiperda eggs) and parasitized hosts (C. cephalonica or S. frugiperda) (pparasitoid*host=0.1950,

Fparasitoid*host=1.73) (Table 1). Telenomus remus reared on C.

cephalonicaeggsandS.frugiperdaeggsexhibitedsimilarparasitism capacityregardlessofthehostspeciestheywererearedon.The numberofparasitizedeggswassimilarforbothparasitoidcolonies (48.32 and 40.53 eggs for parasitoids reared on C. cephalonica andS.frugiperda,respectively,pparasitoid=0.2161,Fparasitoid=1.57)

(Table 1). In contrast, higher numbers of S. frugiperda eggs (87.96)wereparasitizedcomparedwithC.cephalonicaeggs(1.19) (phost<0.0001,Fhost=259.46)(Table1),indicating aclear

prefer-enceforS.frugiperdaeggsregardlessofthehosttheparasitoidwas rearedon.

Bioassay2:inﬂuenceofparasitoidmatingonitsparasitism

No differences were found between mated and unmated femaleswithrespecttothenumberofparasitizedeggs,parasitoid

emergence(%)andlongevityofparentalfemales(days)(Table2).In contrast,offspringsexratiodifferedbetweentreatments(0.61for matedfemalesand0.00forunmatedfemales,Table2),indicating thatT.remusreproducesparthenogeneticallywithcharacteristics ofthearrhenotokytype.

Bioassay3:inﬂuenceofthereleasedensityofT.remusonits parasitism

For S. frugiperda eggsattachedto maize leaves, we found a quadraticeffectfortheregressionanalysisbetweenthepercentage ofparasitismanddensityofreleasedT.remusfemales(numberof parasitoidperpestegg)inallparasitoidgenerationstested(Fig.1). Maximumparasitismwas71.69%,71.69%,and65.07%forF35,F40,

andF45,respectively.Themaximumparasitismratewasreached

atdensitiesbetween0.133and0.150ofT.remusfemalesperS. frugiperdaegg(Fig.1).Throughoutthestudy,climaticdatawere recordedwithadataloggeratgreenhouseconditions.Differences intemperatureandrelativehumiditybetweenallevaluatedgrowth stagesareshowninTable3.

Discussion

Parasitoidrearingonfactitioushostsisessentialforthesuccess ofitsmassrelease inaugmentative biologicalcontrolprograms (Buenoetal.,2008)andrequirestheestablishmentofprocedures tomonitorthequalityoftheproducedinsect.AccordingtoCobert (1985),continuousrearingof parasitoidsonfactitioushostscan causethelossoftheirabilitytorecognizeandchooseahostand thereforereducetheirefficiencyagainstthetargetedpestspecies. Inourstudy,thisnegativeeffectwasnotobserved forT.remus rearedonC.cephalonicaeggs.Parasitismofeggsofthetargetfield pest (S.frugiperda)wassimilar in parasitoidfemales reared on both C.cephalonicaand S.frugiperda. However,theinfluenceof thefactitioushostonparasitoidqualitymightalsodependonthe numberofgenerationstheparasitoidisrearedonthesamehost.

Pratissolietal.(2004)reportedthatparasitismcapacityofspecies rearedonfactioushostswasinverselyproportionaltothenumber

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100 A: F₃₅ B: F40 C: F45 90 80 70 60 50 40 30 20 10 19.5 23.32 35.36 47.99

Parasitism (%)=4.5 + 675.19 (parasitoid number) - 1753.92 (parasitoid number)2

R2_=0.8131

Parasitism (%)=1.40 + 90.20 (parasitoid number) - 302.55 (parasitoid number)2

R2_=0.9226

Parasitism (%)=1.47 + 72.82 (parasitoid number) - 209.47 (parasitoid number)2 R2_=0.8997 53.73 65.02 47.92 47.07 71.69 0 0 0.017 0.033 0.050 0.067 0.083 0.100 0.117 0.133 0.150 0 0.017 0.033 0.050 0.067 0.083 0.100 0.117 0.133 0.150 0 0.017 0.033

T. remus density/ S.frugiperda eggs

0.050 0.067 0.083 0.100 0.117 0.133 0.150 100 90 80 70 60 50 40 30 P a rasitism, % 20 10 0 100 90 80 70 60 50 40 30 20 10 8.02 13.16 23.28 32.17 34.18 42.99 38.62 53.46 65.07 0 7.15 18.34 35.11 41.16 51.18 62.11 71.69 64.75 27.62

Fig.1.Bioassay3:parasitism(%)inSpodopterafrugiperdaeggsafterthereleaseof differentdensitiesofTelenomusremusperhosteggusingparasitoidsfromdifferent colonies.T.remusrearedonCorcyracephalonicafor35generations(A),40 gener-ations(B)and45generations(C).Averagevaluesfromtrialsrepeatedatmaizeat V2/V3,V4/V5andV8/V9developmentstage.

ofgenerationsforwhichtheywerekeptonthesamehostspecies inthelaboratory.Therefore,qualitycontrolmustassessparasitism capacity of all generations throughout parasitoid rearing. We observedsimilarparasitismforT.remusonS.frugiperdaregardless oftheoriginoftheparasitoid(rearedoneitherC.cephalonicaorS. frugiperdaeggs)evenafterrearingitonC.cephalonicaeggsfor45 generations(F45generation).Theseresultshighlightthepotential

ofC.cephalonicaasafactitioushostforT.remusrearing.

Inaddition,S.frugiperdaeggscontinuedtobepreferredbyT. remusrearedonC.cephalonica eggsinthefree-choicetest sup-portingthehypothesisthatitconstitutesanadequatefactitious host.Thisresultalsosuggeststhatthehostacceptancebehavior ofT.remusfemalescannotbeattributedtoexperienceacquired during larvaldevelopment (preimaginalconditioning) as previ-ously described by Cobert (1985) and Kaiser et al. (1989) and morerecentlybyPomari-Fernandesetal.(2015).Ourﬁndingalso excludeslearningor_{␣-conditioning,}wherefemalesassociatenew stimuli (acquired)withinnate stimuliand thus canadapt their behaviortotheenvironmentinwhichtheylivedasadultsoryoung

individuals(Kaiseretal.,1989;Nurindahetal.,1999).Theabsence ofpreimaginalconditioningorlearningwaspreviouslyreportedfor T.remusrearedonS.frugiperdaeggsand,whentestedfordifferent hosts(S.frugiperdaversusS.cosmioides,S.frugiperdavs.S.albula, andS.cosmioidesvs.S.albula)thisspeciespreferredS.cosmioides asahost(Goulartetal.,2011).ThissupportstheuseofT.remusin biologicalcontrolbecauseofthepossibilityofrearingitona sin-glehostforseveralgenerationswithoutreducingitseffectiveness againstdifferenttargetpestsintheﬁeld.

This host preference of T. remus is probably related to the nutritionalqualityofthehost.Addingmorecomplexitytothis phe-nomenon,Molinaetal.(2005)claimthatnotonlythenutritional qualityofthefuturehostbutalsooftheprevioushostonwhichthe parasitoidwasrearedmightinﬂuencehostpreference.Eggsurface, colorandothertraitsofthehostcanalsoinﬂuencehostpreference inamorecomplexdecision-makingprocess(Cônsolietal.,1999). However,thepreferenceofT.remusforS.frugiperdaeggswhen rearedonC.cephalonicamaybemorecloselyrelatedtothesuperior nutritionalvalueofS.frugiperdacomparedtoC.cephalonica.

Theimportanceofadultmatingmustalsobeconsideredwhen rearingT.remus,becauseitcancompromisethemaintenanceofthe parasitoidinthefield(Pratissolietal.,2009;Farrokhietal.,2010). Arecentstrategytoreleaseeggparasitoidsinthefieldhasbeento useisolatedpupaethataresprayedonplants,forexampleof Tri-chogrammaspp.inBrazil.However,malesofT.remuswhichemerge earlierthanfemalesduetotheirshorterlifecycleawaitfemalesto emergefromthesameparasitizedeggmass(Cave,2000), allow-ingforhighmatingratesdirectlyafteremergence.Therefore,the sprayingofT.remusindividualpupaeinthefieldcouldnegatively impactthereproductivebehavioroftheparasitoid,highlightingthe importanceofevaluatingmatingdeprivation.

Theabsenceofmatingdirectlyinﬂuencedoffspringsexratio, leadingtotheproductionofmalesonly.However,otherbiological parameterswerenotaffected.Theobservedreproductionmodeof T.remusviaparthenogenesisofthearrhenotokytype(i.e.fertilized femalesgiverisetodiploidfemales,whereasunfertilizedfemales giverisetohaploidmales),hasbeendescribedasthemostcommon reproductivetypeforinsectsoftheorderHymenoptera(Pratissoli etal.,2014).

In contrasttosexratio,thenumbersofparasitized eggsper matedand unmated femalewere similar,as wellas parasitoid emergence(%)andlongevityofparentalfemales.Itshouldbenoted thatinourstudythelongevityoftheparentalfemaleswassimilarto thatobservedforT.remusofgenerationF19rearedonC.cephalonica

eggsasreportedbyPomari-Fernandesetal.(2015).Theseresults differfromPratissolietal.(2014)whoobservedaninfluenceof matingonfemalelongevityoftheeggparasitoidTrichogramma pretiosum (Riley, 1879) (Hymenoptera: Trichogrammatidae). In addition,Stouthamer (1993) reportsthat field releasesof para-sitoids(genusTrichogramma)reproducingbyparthenogenesisof thethelytokytypeweremoreefficientcomparedwithparasitoids withthose reproducing by parthenogenesis of the arrhenotoky type,illustratingtheimportanceofstudyingthereproductivemode ofeachparasitoidspecies.

OurresultssuggeststhatalthoughparasitismofT.remus,and thereforeitscontrolefficiency,wasnotaffectedbymating, par-asitoidpermanenceinthefieldmaybeimpactedwhenunmated femalesarereleased.Somestudiesreportedthefailuretoestablish T.remusinthefieldafteritsrelease(VanWaddillandWhitcomb, 1982;Figueiredoetal.,1999).Since additionalrelease mightbe alwaysnecessarywhenpestpopulationincrease,thereleasingof matedfemalesmightnotbeimportantconsideringthatthe para-sitismoftheavailableeggsinthefieldwouldbethesame(Carneiro etal.,2009).

Additionally, the success of T. remus as biological control depends ontheappropriate parasitoiddensityper S.frugiperda

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eggwhichcanbeevaluatedbyreleasingdifferentnumbersof par-asitoidsinrelationtoagiven numberof pesteggs. Westudied parasitoidreleasein maize,usingdifferentdensitiesofT.remus femalesoftheF35,F40,andF45generation.Parasitismoffemales

of all tested generations was positively related to the density offemales per S. frugiperdaegg, reachingmaximum parasitism between 0.133 and 0.150 T. remus/S. frugiperda egg (40–45 T. remusfemalesper300eggsof S.frugiperda).In asimilarstudy, theoptimaldensitywasalmost50% less thanin ourstudy(25 T.remusfemales/300 S.frugiperdaeggsofT. remusrearedonS. frugiperdareportedbyPomarietal.,2013).Giventhatparasitism canstronglydependontheparasitoidspeciesand/orstrains(Sá andParra,1993),thesediscrepanciesmaybeduetodifferent par-asitoidcoloniesusedinbothtrials.TherearingofT.remusonC. cephalonicaeggsfor manygenerations seemstogenerate para-sitoidswithlowerparasitismcapacitycomparedwiththosereared onS.frugiperdaeggs,requiringahigherdensityofT.remusfemales toeffectivelycontrolthepest.Althoughagreaternumberof par-asitoidsperpesteggareneededwhentheparasitoidisrearedon afactitioushost,severalotherfactors,suchastheeaseofrearing andlowcostoftheparasitoid,shouldbeconsidered.Overall,our resultsallowtheconclusionthat(a)T.remuspreferstoparasitizeS. frugiperdatoC.cephalonicaeggs,despitetheabsenceof preimagi-nalconditioning;(b)matingdoesnotaffectparasitismcapacityand developmentofT.remus;(c)theoptimalreleasedensityofT.remus rearedonC. cephalonicais between0.133and 0.150females/S. frugiperdaeggs,whichishigherthantheoptimalreleasedensityof T.remusrearedonitsnaturalhostS.frugiperda.Theseresults impor-tantlycontributetoexistingknowledgeonthesuccessfulrearing ofandﬁeldreleasestrategiesforT.remus.

Conﬂictofinterest

Theauthorsdeclarenoconﬂictofinterest.

Acknowledgments

TheauthorswouldliketothankEmbrapaSojaandthesponsor agenciesCAPESand CNPqfortheirﬁnancialsupportand schol-arships.ThispaperwasapprovedforpublicationbytheEditorial BoardofEmbrapaSoja.

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