Biological aspects of immature stage of Nyssomyia whitmani (Antunes and Coutinho) (Diptera, Psychodidae, Phlebotominae) in laboratory conditions

RevistaBrasileiradeEntomologia59(2015)100–103

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

REVISTA

BRASILEIRA

DE

Entomologia

Medical

and

Veterinary

Entomology

Biological

aspects

of

immature

stage

of

Nyssomyia

whitmani

(Antunes

and

Coutinho)

(Diptera,

Psychodidae,

Phlebotominae)

in

laboratory

conditions

Letícia

Moraes

Ribeiro

_,

_Elisa

_Teruya

_Oshiro

_,

_Daiana

_Alovisi

_de

_Souza

_,

_Magda

_Freitas

_Fernandes

_,

Kleiton

Maciel

dos

Santos

_,

_Nathália

_Lopes

_Fontoura

_Mateus

_,

_Alessandra

_Gutierrez

_de

_Oliveira

a_{ProgramadePós-Graduac¸ãoemBiologiaAnimal,UniversidadeFederaldoMatoGrossodoSul(UFMS),CampoGrande,MS,Brazil}

b_{LaboratóriodeParasitologiaHumana,CentrodeCiênciasBiológicasedaSaúde,UniversidadeFederaldoMatoGrossodoSul(UFMS),CampoGrande,MS,Brazil}

c_{ProgramadePós-Graduac¸ãoemEntomologiaeConservac¸ãodaBiodiversidade,FaculdadedeCiênciasBiológicaseAmbientais,UniversidadeFederaldaGrandeDourados(UFGD),}

Dourados,MS,Brazil

a

r

t

i

c

l

e

i

n

f

o

Articlehistory:

Received13August2014 Accepted16February2015 Availableonline8April2015 AssociateEditor:MariaAniceM.Sallum Keywords:

Biologicalcycle Larvae

Phlebotominesandflies

a

b

s

t

r

a

c

t

Nyssomyiawhitmani(AntunesandCoutinho,1939)hasbeenconsideredasacomplexofcrypticspecies, andsomeofthepopulationsofthiscomplexplaysanimportantroleinthetransmissionofLeishmaniaspp. inBrazil.Thepresentstudyreportsthebiologicalaspectsconcerningtheproductivityoutofeggsandthe developmenttimeofthedescendantsoffemalesobtainedinDouradosmunicipality,MatoGrossodoSul state.Thefemaleswerecapturedwithmodifiedelectricaspirators,fedinhamstersandfurther individ-ualizedincontainersforbreeding.Attheinsectary,temperatureandrelativehumidityweremaintained onaverageof24.5◦Cand67.3%,respectively.From944females3737eggswereobtained,748(20.0%) evolvedtothestageoflarvae,and93(12.4%)ofthesereachedadultstage.Thelifecyclelasted80.6 daysandthelastlarvalinstarwasthelongest.Theuseofahigherproteindietrevealedasignificant improvementinlarvaldevelopment.

©2015SociedadeBrasileiradeEntomologia.PublishedbyElsevierEditoraLtda.Allrightsreserved.

Introduction

Nyssomyiawhitmaniisrecordedin720Brazilianmunicipalities anditisdistributedover25states,includingtheFederalDistrict (Queirozetal.,1994;Membriveetal.,2004;Teodoroetal.,2006; Galati,2014).InMatoGrossodoSulthisspecieshasbeenrecorded inmanymunicipalities(Galatietal.,1996,2006;Oliveiraetal., 2003,2006;Braga-Mirandaetal.,2006;Nascimentoetal.,2007; Nunesetal.,2008;Almeidaetal.,2010).

Thespecieswasfoundbyvisualizationofflagellateforms or detectionofLeishmania(V.)braziliensisDNAinsomestatesasCeará, Maranhão,SãoPaulo,ParanáandMatoGrossodoSul(Pessoaand Coutinho,1941;Azevedoetal.,1990;Queirozetal.,1994;Luzetal., 2000;Oliveira-Pereiraetal.,2006;Paivaetal.,2010)andtherefore, itwasimplicatedasaprobablevectoroftheparasiteatthestudied sites.ThefirstinformationaboutthespecieslifecycleNyssomyia whitmaniwasobtainedbyBarretto(1942)from specimens col-lectedfromthesoutheastregionofBrazil.However,moreresearch aboutpopulationdynamicsofthissandflyspeciesanditsimmature

∗ Correspondingauthor.

E-mail:alessandra.oliveira@ufms.br(A.G.deOliveira).

stagesofbiologyarestillneededtounderstandtheinsect’spattern ofabundanceanditsrelationshipwiththeenvironment.

Inspiteofthefewpublishedstudiesabouttheirimmatureforms (larvaeandpupae) andtheduration of thesestages, thisstudy aimedtoobtaininformationonovipositionandthelifecycleof Ny.whitmani,whichcomprisesthestagesofegg,larvaeandpupae, inordertoassisttheunderstandingofthisspeciesecology. Materialsandmethods

Thecaptureswereheldbetween2012and2013,from18:00h to21:00h,usinganelectricaspiratorattachedtoa6Vbatteryina chickencooplocatedattheFazendaCoqueirofarm,inthe munici-palityofDourados,MatoGrossodoSul(MS),HighwayMS162,Km 10Dourados-Ithaum(22◦12Sand54◦54W).Thespecimenswere transportedtotheLaboratóriodeParasitologiaHumanada Univer-sidadeFederaldoMatoGrossodoSul(UFMS),locatedatCampo Grande/MS,accordingtothemethodologyofRangeletal.(1985, 1986).

Apple slices were placed inside the cage to feed themales andfemalesuntiltheirarrivalatthelaboratory.Intheevening, ahamster(Mesocricetusauratus)previouslyanesthetizedwithan intramuscularinjection(ketamin75mg/kgandxylazine10mg/kg http://dx.doi.org/10.1016/j.rbe.2015.02.012

L.M.Ribeiroetal./RevistaBrasileiradeEntomologia59(2015)100–103 101

Table1

Analysisofcollectedfemales,totaloffemalesthatlaideggs,totaleggslaid,totalhatchedlarvae,totalemergedadultsandaveragetemperature(◦_{C)andrelativehumidity}

(RH%)duringtheeightcollectionperiodsbetween2012and2013.

Collectionmonth/year

(2012–2013)

Females Femalesthat

laideggs(%)

Numberofeggs

(eggs/females)

Totaloflarvae (%)

Totalofpupae Adults(%) Temperature(◦C)and

humidity(UR%) Collection1 215 13(6.0) 390(30.0) 52(13.3) – – 24.7◦_/63.9% Collection2 151 8(5.3) 324(40.5) 104(32.1) – – 25.6◦_/65.7% Collection3 187 18(9.6) 448(25.0) 102(22.7) – – 25.4◦_/64.0% Collection4 10 4(40.0) 84(21.0) 6(7.1) – – 24.2◦/63.0% Collection5 90 6(6.7) 86(14.3) – – – 23.9◦/68.3% Collection6a _{5 5(100.0) 88(17.6) 41(46.6) 13(31.7) 11(84.6) 23.7}◦_/71.7% Collection7 148 78(52.7) 1640(21.0) 431(26.3) 149(34.5) 136(91.2) 24.1◦_/69.3% Collection8 138 33(23.9) 677(20.5) 18(2.7) 3(16.6) 2(66.6) 24.6◦_/72.7% Total 944 165(17.48) 3737(22.6) 748(20.02) 165(22.0) 149(90.3) 24.5◦_/67.3%

a_{Beginningofanewlarvaldiet.}

IMin2:1rate)wasplacedinsidethecage,inorderthatthefemales couldbloodfeedonitduringanaveragetimeof40min.After48h, thefemales wereindividualizedina0.5cmplaster-lined plastic containerwith2.5cm diameterand 3.5cm height,which were cappedwithvoilefabric. Apiece ofcotton soakedinundiluted honeywasplacedontopofthevoilefabric,asasourceof carbohy-drateforthefemales(BrazilandBrazil,2003).

The containers were kept in covered plastic boxes (34.0cm×24.5cm×9.0cm) with filter paper in the base, or infoam boxes(31.0cm_×20.0cm_×22.0cm)withplaster onthe wallsandthebase,inordertoconservemoistureandtemperature ofthemicroenvironment.Theseparameterswerecontrolledwith adigitalthermo-hygrometer,whichmaintainedanaverage tem-peratureof24.5◦Candairrelativehumidityof67.3%.Afterdeath, females were dissected for species identification, through the observationofspermathecaemorphologyandidentifiedaccording Galati(2003).OncethefemalesofNy.whitmaniwereidentified, theireggsweregrouped,countedandtransferredwithfine-tipped brushestoa0.5cm plaster-linedPetridishes (8.5diameterand 1.5cm height) accordingRangeletal. (1985).Aftertransferring theeggs,soilcollectedfromthehenhousewheretheinsectswere oncecollectedwasadded. Afterthelarvaehatched,lyophilized liver(OXOID–dehydratedliver)wasintroducedintheirdiet.Daily observationsweremadeuntiltheemergenceofthewingedadults. Thelarvaeatallinstars(L1,L2,L3andL4)andthepupaewere counted.Larvaehatchingandecdysiswererecordedtoverifythe speciesbiologicalcycle.ToestimatethedurationofNy.whitmani lifecycle,includingincubationperiod,durationoflarvalinstarsand pupalperioduntiladultemergence,theminimumduration (mini-mumtime,inconsecutivedays,betweentheobservationofthefirst individualoflarvalperioduntiltheobservationofthefirst individ-ualofthesecondinstarandsoon);andthemaximumduration (maximumtime,inconsecutivedays,betweentheobservationof thefirstindividualoftheinstaruntiltheobservationofthelast individualofthesameinstar),andmedianwerecalculated.

Results

Fromthe944individualizedfemales,3737eggswereobtained fromtheovipositionof165females.Theminimumandmaximum timesofdevelopmentoftheimmatureformswere54.3and106.9 days,respectively.

Fromthetotaloffemalesfed,17.48%laideggs,withthelowest percentageinthesecondcollection(5.3%)andthehighestinthe60 (100%),followedbyseventhcollection(52.7%).Theaveragenumber ofeggsperfemalewas6.1(Table1).Theeggswereelliptical, pre-senteddarktonalityandwerelaidsinglyorinsmallclusters.The meanincubationperiodoftheeggswas9.5daysanditsmedian was12days.Athatching,firstinstarlarvae(L1)presentedasingle pairofcaudalsetaeandwhitishbodiesandheads,withacephalic

portionthatbecamedarkenedafterafewhours.Thisphaselasted aminimumof7.0andthemaximumof11.2days,withamedian of5days(Table2).

TheL2larvaepresentedtwopairsofcaudalsetaeandfedon thehenhousesoilsetinthecontainerswithmoreaviditywhen comparedtoL1larvae.Themaximumandminimumtimeswere 4.9and9.4days,respectively(medianof7days)(Table2).

Itwaspossibletocheckwiththenakedeyethatthethirdinstar larvaewasafewtimeslargerthanthesecondandmaintainedtwo pairsofcaudalsetae.Thisinstarlastedaminimumof6.1daysand maximumof18.0days,withamedianof10days(Table2).

Thefourthandlastlarvalinstar(L4)wascharacterizedbythe presenceofaspotinthelasttergite.Itwasthelongestperiodamong theimmatureformswithamedianof21days(range16.1–29.8), andafterthistime,thelarvaehadtheirlastecdysis.Itwasobserved thatinthisperiodthelarvaereduceditsfeedingandlocomotion. Atthebeginningofthepupalstage,thepupaepresentedyellowish andlightbrowneyes,whichchangedtoablackenedshadeasthe endofthephaseapproachedandremaineduntiltheyreachadult form.Thisperiodshowedarangebetweenaminimumof10.7days andamaximumof22.7days,withamedianof16.7days.The bio-logicalcyclepresentedamedianof80.6days(Table2).Therewas nodifferencebetweenemergedmalesandfemales.

Discussion

Barretto(1941,1942)reportedlowervalues forthelifecycle when studying populations of Ny. whitmani from Southeast of Brazil,withaveragesof50.5days.Whenstudyingthebiologyof NyssomyiaintermediaandNy.whitmani,Barretto(1941,1942) ver-ifiedtherapidandgradualdevelopmentofeggswhentemperature wassetbetween25◦ and27◦C.Rangeletal.(1985),reporteda rangeof26–56days(average41days)whenmonitoringthelife cycleofNy.intermediaat25◦Cand86%humidity.Andrade-Filho etal.(2004)notedalifecycleof34.9daysattemperaturesranging between25◦Cand26◦Cand80%humidity,whenstudyingthislast species.

Table2

Duration(days)ofegg,larvalandpupaldevelopmentofNy.whitmaniunder lab-oratoryconditionsattemperatureof24.5◦_C(±1◦_{C)andRH∼67%overtheeight}

collectionperiodsbetween2012and2013.

Instar Minimum(days) Maximum(days) Median(days)

Egg 9.5 15.9 12.0 Larva1 7.0 11.2 5.0 Larva2 4.9 9.4 7.0 Larva3 6.1 18.0 10.0 Larva4 16.1 29.8 21.0 Pupa 10.7 22.7 16.7 Total 54.3 106.9 80.6

102 L.M.Ribeiroetal./RevistaBrasileiradeEntomologia59(2015)100–103

Braziletal.(1997)studiedthebiologyofEvandromyialentiand foundthatitslifecyclelastedfor40.2daysattemperatures ran-gingbetween26◦Cand28◦Candrelativehumidityof80%.When comparingthecycleofthisspecieswithLu.longipalpisandNy. inter-media,thepreviousauthorsverifiedthatEv.lentihasthelongest cycle,followedbyNy. intermedia(32.8 days)andLu.longipalpis (29.5days). Evandromyiacarmelinoi, species colonized byAlves etal.(2000),showedanaveragetimeofdevelopment,fromegg toadultform,of42days,ataveragetemperatureof25–26◦Cand humidityof80%.GuzmánandTesh(2000)whenobserving devel-opmenttimeinrelationtotemperatureforthespeciesPhlebotomus papatasi,Ph.perniciosusandLutzomyialongipalpis,notedthatmost ofthelarvaeofthelastspeciesdiedattemperaturesunder18◦C; whichistheoppositeofwhathappenedwithPh.papatasiandPh. perniciosus.Thesetwospecieshadtheircyclesextended(150and 412days,respectively)atlowertemperatures,buthadit substan-tiallyshortenedwhenthetemperaturewaselevatedto28◦C.

Inthepresentstudy,temperatureremainedbetween23◦Cand 25◦C and theaveragehumidity didnot exceed65%in thefirst fivecollections.Duringthelastmonths,humiditywasabove70% andtemperaturewasbetween23◦Cand24◦C,whichwasa bet-terperiodforlarvaldevelopmentandadultemergence.Therefore, itispossiblethattemperatureandhumiditymayhaveinfluenced inthedevelopmenttimeofimmaturestages,andmayevenhave startedtheprocessofdiapause.

Barretto(1942)studiedthebiologicalcycleofNy.whitmaniwith specimenscollectedinthesoutheastregion.Heobservedthe occur-renceofdiapauseamongthelarvae,becausethelifecycleextended until176days.However,Brazil(personalcommunication)worked withthesamespecies,withindividualsoriginatedfromCeará,and reported37dayslifecycle,withnoobservationofthediapause phenomenon.

ReadyandCroset(1980)studiedPh.perniciosusandPhlebotomus ariasiandnotedthatenvironmentalstimulicaninducediapause, especiallyforthirdandfourthlarvalinstarsattemperaturesbelow 21◦C. AccordingtoChaniotis(1967),thediapauseisan adapta-tionperiod toenvironmental conditions.Carvalhoet al. (2011) suggestedthatdiapausecouldbeanevolutionarymechanismof thesandflytosurvivelowtemperaturesandadverseconditions, resumingmetabolicactivitieswhenconditionsbecomefavorable.

Othermajorfactor forthelarvaetoreachtheadultstage is thefeeding.Inthepresentstudy,amix ofrabbitfecesandfish food(1:1)wasinitiallyused,butitresultedinproliferationofa largeamountoffungiandconsiderablemortality,especiallyamong thefirstinstarlarvae.Therefore,analternativedietconsistingof lyophilizedliverandsoilwastestedanditenhancedlarval devel-opmentandtheemergenceofthewingedforms.Theadditionofsoil fromthecapturesitewastriedbecauseitwasbelievedthatbesides stimulatingtheexistingmemory(KellyandDye,1997).This mate-rialcouldcontainorganicmatterofhighnutritionalvalue,dueto thepresenceofchickenfeces,anditcouldworkasafoodsource fortheimmatureforms(Forattinietal.,1976).

WhenRangeletal.(1985)studiedtheestablishmentofacolony ofNy.intermedia,theyusedcommercialfishfoodandachieved aratioof46.6% adultsregardingthenumber ofhatchedlarvae. Rangeletal.(1986)accompaniedthedevelopmentofNy.intermedia andLu.longipalpislarvaefedwithfoodofanimalandvegetal ori-gin.Thecommercialfishfoodwaswellacceptedbybothspecies anddidnotfavor fungidevelopment.Gemetchu(1971)studied thelarvaldietofPhlebotomuslongipesandobservedtheir prefer-enceforlyophilizedliverratherthanrabbitfeces,meatorblood. GuzmánandTesh(2000)analyzedtheeffectoftwodifferentdiets onLu.longipalpis(fecesmixedwithliverpowderanddecomposing leaves)andobservedthatthelarvaefedwithfecesandliver pow-derhadafasterandsynchronousdevelopmentthanthosefedwith leaves.ThisresultssupportsthefindingsofChaniotis(1975),who

fedLutzomyiatrapidoilarvaewithlyophilizedliverandnotedthe samebehaviorinlarvaldevelopment,insteadofwhenthelarvae werefedwithcookedlettuce.

Besidesfood,temperatureandhumidity,anotherfavorable fac-torforsuccessinpostureandadultemergencewasthegroupingof femalesafterblood feedingandpriortooviposition.Tengroups of35 females onaveragewereplaced togetherin plaster-lined glasscontainersof10cmdiameter.Throughthisexperimentwas observedthatthegroupedfemaleslaidmoreeggsonaverage(6.1) thantheindividualizedones(3.9).However,thesedatawerenot usedbecausefrom30emergedfemales(F1),sevenbelongedtothe speciesPsathyromyiabigeniculata(Sábioetal.,2014).

Thisfactmaybeassociatedwithchemicalsignalsreleasedby sandflies tolocatehosts and ovipositionsites, asproven inthe studyconductedbyHamilton(2008)withthespeciesLu. longi-palpis.Itisnoteworthythatattheposturemoment,femalesdeposit dodecanoicacidintheireggsthroughtheaccessoryglands,thus attractingotherpregnantfemalestoperformposture(Dougherty etal.,1994;Doughertyand Hamilton,1997).Alvesetal.(2003) studiedLutzomyiareneiandfoundthatfemalesalsodepositfatty acids in their eggs, which could probably act as oviposition pheromones.

AccordingtoWard(1977),sandflieslifecyclecouldalsobe influ-encedbydifferentpopulationsandmethodsofbreeding,because, asalreadyknown,sandfliesspeciesrequireanadaptationtothe laboratoryenvironment.Andyet, adultsofcertainspecies have differentfeedingpreferencesofothers(Mutingaetal.,1989). There-fore, further studies on sandflies biology are needed to better understand the behavior, feeding habits and life cycle of each species.

Conflictsofinterest

Theauthorsdeclarenoconflictsofinterest. Acknowledgments

WearethankfultoCAPESandFUNDECTN◦10/2011-UNIVERSAL –Protocol:23157.345.913.08032012forthefinancialsupport,and totheGraduatePrograminAnimalBiologyforsupport.Tothe Ani-malEthicalCommitteeoftheUniversidadeFederaldoMatoGrosso doSul(projectnumber442/2012).

References

Almeida,P.S.,Minzão,E.R.,Minzão,L.D.,daSilva,S.R.,Ferreira,A.D.,Faccenda,O., Andrade-Filho,J.D.,2010.EcologicalaspectsofPhlebotomines(Diptera: Psy-chodidae)intheurbanareaofPontaPorãmunicipality,StateofMatoGrossodo Sul,Brazil.Rev.Soc.Bras.Med.Trop.43,723–727.

Alves,J.C.M.,Pedras,M.J.,AndradeFilho,J.D.,Brazil,B.G.,Souza,A.A.,Brazil,R.P., 2000.BiologyofutzomyiacarmelinoiRyan,Fraiha,Lainson&Shaw(Diptera: Psychodidae)underlaboratoryconditions.Entomol.Vectores7,281–286.

Alves,J.C.M.,Hamilton,J.G.C.,Brazil,R.P.,2003.OvipositionresponseofLutzomyia (Lutzomyia)renei(Martins,Falcão&Silva)(Diptera:Psychodidae)toextractsof conspecificeggsinlaboratorybioassays.Entomotropica18,121–126.

Andrade-Filho,J.D.,Galati,E.A.B.,Falcão,A.L.,2004.Biologyofthefirstgeneration ofalaboratorycolonyofNyssomyiaintermedia(Lutz&Neiva,1912)and Nys-somyianeivai(Pinto,1926)(Diptera:Psychodidae).Mem.Inst.OswaldoCruz99, 597–601.

Azevedo,A.C.R.,Rangel,E.F.,Costa,E.M.,David,J.,Vasconcelos,A.W.,Lopes,U.G., 1990.NaturalinfectionofLutzomyia(Nyssomyia)whitmani(Antunes&Coutinho, 1939)byLeishmaniaofthebraziliensiscomplexinBaturité,Cearástate, north-eastBrazil.Mem.Inst.OswaldoCruz85,251.

Barretto,M.P., 1941. Observac¸õessobre a biologiadoPhlebotomus whitmani AntuneseCoutinho,1939(Diptera,Psychodidae)emcondic¸õesexperimentais. Pap.AvulsosdeZool.(SãoPaulo)1,87–100.

Barretto,M.P.,1942.Observac¸õessobreabiologiaemcondic¸õesnaturaisdos flebó-tomosdoEstadodeSãoPaulo(Diptera:Psychodidae),TesedeLivre-Docência. FaculdadedeMedicinadaUniversidadedeSãoPaulo,SãoPaulo.

Braga-Miranda,L.C.,Miranda,M.,Galati,E.A.B.,2006.Phlebotominefaunainarural areaoftheBrazilianPantanal.Rev.SaúdePúbl.40,324–326.

L.M.Ribeiroetal./RevistaBrasileiradeEntomologia59(2015)100–103 103 Brazil,R.P.,Brazil,B.G.,2003.BiologiadeflebotomíneosNeotropicais.In:Rangel,E.F.,

Lainson,R.(Eds.),FlebotomíneosdoBrasil.EditoraFiocruz,RiodeJaneiro,pp. 257–270.

Brazil,R.P.,Carneiro,V.L.,AndradeFilho,J.D.,Alves,J.C.M.,Falcão,A.L.,1997.Biology ofLutzomyialenti(Mangabeira)(Diptera:Psychodidae).An.Soc.Entomol.Bras. 26,1.

Carvalho,G.M.L.,Vasconcelos,F.B.,Silva,D.G.,Botelho,H.A.,Andrade-Filho,J.D., 2011.Diversityofphlebotominesandflies(Diptera:Psychodidae)inIbitipoca StatePark,MinasGerais,Brazil.J.Med.Entomol.48,764–769.

Chaniotis,B.N.,1967.ThebiologyofCaliforniaPhlebotomus(Diptera:Psychodidae) underlaboratoryconditions.J.Med.Entomol.4,221–233.

Chaniotis,B.N.,1975.AnewmethodforrearingLutzomyiatrapidoi(Diptera: Psycho-didae),withobservationsonitsdevelopmentandbehaviorinthelaboratory.J. Med.Entomol.12,183–188.

Dougherty,M.,Hamilton,G.,1997.Dodecanoicacidistheovipositionpheromone ofLutzomyialongipalpis.J.Chem.Ecol.23,2657–2671.

Dougherty, M.J., Hamilton, J.G.C., Ward, R.D., 1994. Isolation of oviposition pheromonefromtheeggsofthesandflyLutzomyialongipalpis.Med.Vet. Ento-mol.8,119–124.

Forattini,O.P.,Rabello,E.X.,Galati,E.A.B.,1976.Novosencontrosdeflebotomíneos noEstadodeSãoPaulo,Brasil,comespecialreferênciaaLutzomyialongipalpis. Rev.SaúdePúbl.10,125–128.

Galati,E.A.B.,2003.MorfologiaeTaxonomia.In:Rangel,E.F.,Lainson,R.(Eds.), Fle-botomíneosdoBrasil.EditoraFiocruz,RiodeJaneiro,pp.23–175.

Galati,E.A.B.,2014.Bioecologia eIdentificac¸ãodePhlebotominae,availableat:

www.fsp.usp.br/egalati(accessed03.11.14).

Galati,E.A.B.,Nunes,V.L.B.,Dorval,M.E.C.,Oshiro,E.T.,Cristaldo,G.,Cristaldo,M.A.E., daRocha,H.C.,Garcia,W.B.,1996.Estudodosflebotomíneos(Diptera, Pychodi-dae),emáreadeleishmaniosetegumentar,noEstadodeMatoGrossodoSul, Brasil.Rev.SaúdePúbl.30,115–128.

Galati,E.A.,Nunes,V.L.,Boggiani,P.C.,Dorval,M.E.C.,Cristaldo,G.,Rocha,H.C.,Oshiro, E.T.,Damasceno-Júnior,G.A.,2006.Phlebotomines(Diptera:Psychodidae)in forestedareasoftheSerradaBodoquena,stateofMatoGrossodoSul,Brazil. Mem.Inst.OswaldoCruz101,175–193.

Gemetchu,T.,1971.Liverandyeastaslarvaldietsincolonizationofasandfly (Phle-botomuslongipes).Trans.R.Soc.Trop.Med.Hyg.65,682–684.

Guzmán,H.,Tesh,R.B.,2000.Effectsoftemperatureanddietonthegrowthand longevityofphlebotominesandflies(Diptera:Psychodidae).Biomédica20, 190–199.

Hamilton,J.G.C.,2008.Sandflypheromones:theirbiologyandpotentialforusein controlprograms.Parasite15,252–256.

Kelly,D.W.,Dye,C.,1997.Pheromones,kairomonesandtheaggregationdynamics ofthesandflyLutzomyialongipalpis.Anim.Behav.53,721–731.

Luz,E.,Membrive,N.,Castro,E.A.,Dereure,J.,Pratlong,F.,Dedet,J.A.,Pandey,A., Thomaz-Soccol,V.,2000.Lutzomyiawhitmani(Diptera:Psychodidae)asvector ofLeishmania(V.)braziliensisinParanástate,southernBrazil.Ann.Trop.Med. Parasitol.94,623–631.

Membrive,N.A.,Rodrigues,G.,Membrive,U.,Monteiro,W.M.,Neitzke,H.C., Lonar-doni,M.V.C.,Silveira,T.G.V.,Teodoro,U.,2004.Flebotomíneosdemunicípiosdo NortedoEstadodoParaná,suldoBrasil.Entomol.Vectores11,673–680.

Mutinga,M.J.,Kamau,C.C.,Kaddu,J.B.,Kyai,F.M.,Omogo,D.M.,Mwandandu,J., Ndambuki,J.,1989.Thebiologyandcolonizationofsomekenyanphlebotomine sandflyspecies(Diptera,Psychodidae).Int.J.Trop.InsectSci.10,677–683.

Nascimento, J.C., Paica, B.R., Malafronte, R.S., Fernandes, W.D., Galati, E.A.B., 2007.Naturalinfectionofphlebotomines(Diptera:Psychodidae)ina visceral-leishmaniasisfocusinMatoGrossodoSul,Brazil.Rev.Inst.Med.Trop.SãoPaulo 49,119–122.

Nunes,V.L.B.,Galati,E.A.B.,Cardozo,C.,Rocca,M.E.G.,Andrade,A.R.O.,Santos,M.F.C., Aquino,R.B.,Rosa,D.,2008.Estudodeflebotomíneos(Diptera,Psychodidae) emáreaurbanadomunicípiodeBonito,MatoGrossodoSul,Brasil.Rev.Bras. Entomol.52,446–451.

Oliveira,A.G.,Andrade-filho,J.D.,Falcão,A.L.,Brazil,R.P.,2003.Estudode flebotomí-neos(Diptera,Psychodidae,Phlebotominae)nazonaurbanadacidadedeCampo Grande,MatoGrossodoSul,Brazil,1999–2000.CadernodeSaúdePública19, 933–944.

Oliveira,A.G.D.,Galati,E.A.B.,Oliveira,O.D.,Oliveira,G.R.D.,Espindola,I.A.C.,Dorval, M.E.C.,Brazil,R.P.,2006.AbundanceofLutzomyialongipalpis(Diptera: Psychodi-dae:Phlebotominae)andurbantransmissionofvisceralleishmaniasisinCampo Grande,stateofMatoGrossodoSul,Brazil.Mem.Inst.OswaldoCruz101, 869–874.

Oliveira-Pereira,Y.N.,Rebêlo,J.M.M.,Moraes,J.L.P.,Pereira,S.R.F.,2006.Diagnóstico moleculardataxadeinfecc¸ãonaturaldeflebotomíneos(Psychodidae,tzomyia) porLeishmaniasp.naAmazôniamaranhense.Rev.Soc.Bras.Med.Trop.39, 540–543.

Paiva,B.R.,Oliveira,A.G.,Dorval,M.E.M.C.,Galati,E.A.B.,Malafronte,R.S.,2010.

Species-specificidentificationofLeishmaniainnaturallyinfectedsandflies cap-turedinMatoGrossodoSulState,Brazil.ActaTrop.115,126–130.

Pessoa,S.B.,Coutinho,J.O.,1941.Infecc¸ãonaturaleexperimentaldoflebótomospela LeishmaniabraziliensisnoEstadodeSãoPaulo.Hospital20,25–35.

Queiroz,R.G., Vasconcelos,I.A.B.,Vasconcelos,A.W.,Pessoa,F.A.C.,Sousa,R.N., David,D.R.,1994. CutaneousLeishmaniasisinCearaStateinNortheastern Brazil:incriminationofLutzomyiawhitmani(Diptera:Psychodidae)asa vec-torofLeishmaniabraziliensisinBaturiteMunicipality.Am.J.Trop.Med.Hyg.50, 693–698.

Rangel,E.F.,Souza,N.A.,Wermelinger,E.D.,Barbosa,A.F.,1985.Estabelecimentode colônia,emlaboratório,deLutzomyiaintermediaLutz&Neiva,1912(Diptera, Psychodidae,Phlebotominae).Mem.Inst.OswaldoCruz80,219–226.

Rangel,E.F.,Souza,N.A.,Wermelinger,E.D.,Barbosa,A.F.,Andrade,C.A.,1986.

BiologiadeLutzomyiaintermediaLutz&Neiva,1912eLutzomyialongypalpis Lutz&Neiva,1912(Diptera,Phychodidae),emcondic¸õesexperimentais.I. aspectosdaalimentac¸ãodelarvaseadultos.Mem.Inst.OswaldoCruz81, 431–438.

Ready,P.D.,Croset,H.,1980.DiapauseandlaboratorybreedingofPhlebotomus per-niciosusNewsteadandPhlebotomusariasiTonnoir(Diptera:Psychodidae)from southernFrance.Bull.Entomol.Res.70,511–523.

Sábio,P.B.,Andrade,A.J.,Galati,E.A.B.,2014.Assessmentofthetaxonomicstatus ofsomespeciesincludedintheshannonicomplex,withthedescriptionofa newspeciesofpsathyromyia(diptera:psychodidae:phlebotominae).J.Med. Entomol.51,331–341.

Teodoro,U.,Santos,D.R.,Santos,A.R.,deOliveira,O.,Poiani,L.P.,Silva,A.M.,Neitzke, H.C., Monteiro,W.M.,Lonardoni,M.V.C.,Silveira, T.G.V.,2006. Informac¸ões preliminaressobreflebotomíneosdonortedoParaná.Rev.SaúdePúbl.40, 327–330.

Ward,R.D.,1977.ThecolonizationofLutzomyiaflaviscutellata(Diptera: Psychodi-dae),avectorofLeishmaniaMexicanaAmazonensisinBrazil.J.Med.Entomol.14, 469–476.