Rogerjovani Æ Luisaamoæ Elenaarriero

RogerJovani Æ LuisaAmoÆ ElenaArriero

OliverKroneÆ AlfonsoMarzal Æ Peter Shurulinkov

GustavoTomasÆ DanielSolÆ Jana Hagen Pilar LopezÆ Jose Martın Æ Carlos Navarro Jordi Torres

Doublegametocyteinfectionsinapicomplexanparasitesofbirds andreptiles

Abstract The simultaneousoccurrenceof maleand femalegametocytesinsideasinglehostbloodcellhas beensuggested toenhanceapicomplexantransmission [’’doublegametocyteinfection(DGI) hypothesis’’].We didabibliographicsearchandadirectscreenofblood smearsfromwildbirdsandreptilestoanswer,forthe firsttime,howcommonaretheseinfectionsinthewild. Takingthesetwoapproachestogether,wereporthere casesofDGIs inPlasmodium, Haemoproteus,Leucocy- tozoonandHepatozoon, andcasesofmale–femaleDGIs inHaemoproteus ofbirdsandreptilesandinLeucocy- tozoonofbirds.Thus,wesuggestthatDGIsandmale–

female DGIs are more widespreadthan previously thought,openinga newresearchavenueon apicom- plexantransmission.

InPlasmodium andrelatedapicomplexanparasites,the occurrenceoftwoormoregametocytesinsideasingle hostbloodcellrecently attractedtheattentionof researchers fortheirpotentialto enhanceparasite transmissionbetweenvertebratehosts(Jovani2002).In thesemicroparasites,transmission occurswhenthe gametocytescontainedinthehostbloodcellsarein- gestedwithinthebloodmealofasuitablevector.Oncein thevector,maleandfemale gametesdifferentiatefrom

maleandfemalegametocytes,respectively,andatleast

R.Jovani()

DepartmentofAppliedBiology,

EstacionBiologicadeDonana(CSIC),Avda.

MariaLuisas/n,41013Sevilla,Spain

E-mail:

Tel.:+34-954-232340

Fax:+34-954-621125

L.AmoÆ E.ArrieroÆ G.TomasÆ P.LopezÆ J.Martın

DepartmentofEvolutionaryEcology,

Museo Nacional deCienciasNaturales(CSIC),

JoseGutierrezAbascal2,28006Madrid,Spain

O.KroneÆ J.Hagen

InstituteforZooand WildlifeResearch,

P.O.Box601103,10252Berlin,Germany

A.MarzalÆ C.Navarro

DepartmentofAnimalBiology,

FacultaddeCiencias,UniversidaddeExtremadura,

06071Badajoz,Spain

P.Shurulinkov

InstituteofZoology,1TzarOsvoboditelBlvd.,

1000Sofia,Bulgaria

D.Sol

DepartmentofBiology,McGillUniversity,

1205avenueDocteurPenfield,Montreal,Quebec,

H3A1B1,Canada

J.Torres

LaboratoryofParasitology,FacultatdeFarmacia,

UniversitatdeBarcelona,Av.JoanXXIIIs/n,

08028Barcelona,Spain

oneofthemobilemalegametesmustsearchforand fertilizeoneoftheimmobilefemalegametescontainedin thesamebloodmeal.Gametocytesarenormallyalone withinsinglehostbloodcells(e.g.erythrocytes),butthey arealsofoundincouples(doublegametocyteinfections, DGIs)orintrios(TGIs)withinagivenbloodcell.

Femalegametesareverysmallcomparedwiththe bloodmealvolumeingestedby the vector; andmale gametesdonotseemtohaveanysearchingmechanism tofindthefemalegamete(Gaillargetal.2003).Thus,it seemslogicalthatsomemechanismoperatingwhilein thevertebrate hostcouldhaveevolvedtoincreasethe proximityofgametocytesandthusgameteswhenonthe bloodmeal;andtwonon-mutually exclusivehypotheses havebeenproposed.Jovani(2002)proposedthatmale– femaleDGIs (i.e.onebloodcellinfectedbyonemale andonefemalegametocyte) mightreducethetime neededformalegametestoencounterfemale gametes, facilitatingparasitetransmission betweenvertebrate hosts.Gaillardetal.(2003)recentlyproposedthatery- throcytesinfectedbygametocytescouldadhereamong theminperipheral hostcapillaries,enhancingthe probabilityofgameteencounter onceinthevector bloodmeal.

The hypothesisproposedbyGaillardetal. (2003) agreeswiththediscoveredpatternofaggregateddistri- butionoferythrocytesinfectedbymalariagametocytes,

Table1 Someexamplesofpublisheddataontheoccurrenceofmultiplegametocyteinfectionsinapicomplexanparasites.DGIDouble gametocyteinfection

HostParasiteObservationReference

ColumbaliviaHaemoproteus columbae

DGIsin17of30hostindividualsinfected, with14–97DGIsineach individualhost

Graczyketal.(1994)

H.columbaeAphotographofamale–femaleDGIBennettand Peirce(1990)

H.columbaeAphotographofamale–femaleDGI, aphotographofaTGI

Mushietal.(1999)

Haemoproteus sppAphotographofafemale–femaleDGITsaietal.(1986)

Geochelone denticulate

H.geochelonisDrawingsandphotographsofmale–female

DGIs;text:‘‘quitecommon’’

occurrenceofDGIs

Lainsonand Naiff (1998)

BirdsHaemoproteus, Plasmodium, Leucocytozoonspp

Drawingsof:immatureDGIsfrom

20Haemoproteus spp,

oneimmatureTGIinH.kairullaevi,

onemale–femaleDGI

inH.bennetti,onemale–maleDGIinH.ortalidum,

twoimmatureDGIsinP.octameriumand P.columbae,

oneimmatureDGIandonemale–femaleDGI

inL.caulleryi

Valkiunas(1997)

Caimanc.crocodilesHepatozoon caimaniPhotographsofDGIsandTGIs, sexunknown

Lainsonetal.(2003)

HomalopsisbuccataHepatozoon sppPhotographofaDGI, sexunknownSalakijetal.(2002)

bothinhumanblood(Gaillardetal.2003)andinthe vectorbloodmeal (Pichonetal.2000).The‘‘DGI hypothesis’’reliesontheassumptionthat DGIs, and especiallymale–femaleDGIs,arewidespreadinnature, butthisiscurrentlyunconfirmed(Jovani 2002).Thegoal ofthisnoteistoshowthatDGIsarewidespreadamong invivoapicomplexanparasitesofwildpopulationsof birdsandreptiles.Moreover,wealsoreporttheoccur- renceofothermultiplegametocyteinfections(MGIs,i.e. morethanonegametocyteinsideasingleerythrocyte), becausethey informusthattheneededprocesstopro- duceaDGIdoesoccurinagivenhost–parasite system, i.e.theentranceand growthintoagivenerythrocyteof morethanonegametocyte.

Wefirstsystematicallysearchedintheparasiteliter- atureforcasesofMGIs.Informationregardingthiskind ofinfectionis littlelikelytoappearinthetitle,abstract, orkeywordsofapublication.Consequently, wewere forcedtosearchourfilesforpapersonapicomplexan infections;andthenweaccurately revisedeachpaperfor anymentionofMGIs. Oursurveyfound anyreference onthepossiblerelevanceofMGIsforanyaspectofthe biology oftheparasite.Indeed, somepapersevencon- tainedphotographsofDGIs, butwithoutanymention inthetext(e.g.Mushietal.1999).Despitethisevident lack ofattentionofresearchersforDGIsandMGIsasa whole,wefoundeightworkswithinformationonDGIs andTGIs(Table1).

Moreover,fromourownbloodsmearcollection,we selectedthosebloodsamplesof11bird andfourreptile speciesinfectedbysomeapicomplexanparasites.These bloodsmearswerecollectedfromwildhostsinGermany andSpainduringtheperiod1996–2002(see Table1).On eachsmear,we activelysearchedforMGIsonaportion equivalenttoca.2,000 erythrocytesforHaemoproteus andHepatozoon infectionsandca.25,000erythrocytes forLeucocytozoon infections,recordingwhenpossible

the sexfor maturegametocytesinvolvedin a MGI. MGIswerefoundinmostoftheparasite–hostassocia- tionsweexamined(Table2).

Moreover,froma parasitesurvey conductedby ShurulinkovandGolemansky(2002,2003),P.S.revised theirownnotestakenduringbloodsmearinspectionina searchforrecordsofMGIs. Althoughnotcomparable withtheothersamplesbecauseitwasnotasystematic searchforMGIs,itisofqualitativeimportanceandalso denotesthelowreportingoftheseinfectionsinpubli- cations,althoughpresentintheresearchers’notes.A totalof15DGIs wererecordedfrom11birdsinfected byHaemoproteus offivebird–parasite assemblages (bird–parasite[numberofbirdswithDGIs, totalnum- berofDGIs]:Sylviaborin–H. belopolskyi[1,3],Lanius minor–H.lanii[2, 2],Laniuscollurio–H.lanii[2,2], Acrocephalusarundinaceus–H.payevskyi[2, 3],Musci- capastriata–H.pallidus[4, 5]).MostoftheseDGIswere constitutedbyfemalegametocytes.However,H. pay- evskyi infectingA. arundinaceus showeda case of a male–femaleDGI. Moreover,twocasesofDGIs were recordedinPlasmodium polare infectingoneindividual ofHirundo rustica. SeeShurulinkovandGolemansky (2002,2003)forthetotalnumbersofspeciesandindi- vidualsexamined.

Ourresultssupporttheviewthat DGIs arewide- spread inavarietyofhost–parasiteassociations, includingPlasmodium, Haemoproteus, andLeucocyto- zooninfectionsofbirdsandHaemoproteus andHepa- tozoon parasitesofreptiles. Moreover,wefound male–femaleDGIs inHaemoproteus parasitesofbirds andreptilesandinLeucocytozoon ofbirds,validating themajor assumptionof the‘‘DGI hypothesis’’.The widespreadoccurrenceof DGIs, and particularlyof male-femaleDGIs, opensa promising directionfor futureresearchon thetransmissionstrategiesof api- complexanparasites.

Table2 OccurrenceofDGIs, male–female(m-f)DGIs,and triplegametocyteinfections (TGIs)inbloodsmears inspectedforthisstudy.

NNumberofinfected individualhostsinspected,or numberofindividualhostswith atleastoneD/TGI. Numbersin

HostCountryNNDGIsNm-fDGIs NTGIs

Haemoproteus parasites

ButeobuteoGermany83(1,1,2)00

FalcotinnunculusGermany30–0

F.subbuteoGermany70–0

TytoalbaSpain41(3)00

BubobuboSpain213(1,2,2)00

parenthesesindicatethe numberofD/TGIsineach

Strix alucoGermany,

Spain

2110(1,2,2,2,3,4,10,11,13,27) 1(1)4(1,1,3,15)

infectedhost.–Sexofparasite couldnotbedetermined

Parus caeruleusSpain4610(1,1,1,1,1,2,2,3,4,6)00

DelichonurbicaSpain577(1,1,1,1,1,1,2)1(1)0

LaniuscollurioSpain110–1(1)

Passer domesticus Spain3911(1,1,1,1,2,2,2,2,2,5,6)4(1,1,1,1) 0

Leucocytozoonparasites

B.buboSpain190–0

AthenenoctuaSpain20–0

Strix alucoSpain50–0

Parus caeruleusSpain152(1,2)–0

Hepatozoon parasites

P.caeruleusSpain90–0

Lacerta lepidaSpain380–0

L.monticolaSpain105 5(1,1,1,2,3)–0

Podarcis muralisSpain280–0

P.hispanicaSpain190–0

AcknowledgementsTheauthors thankJuanA.Fargallo,Jesus Martınez,MariloRomero,AlbertoLizarraga,FranciscoCampos, SantiagoMerino,JuanMoreno,JudithMoralesfortheirfieldwork andJavierCalderaforallowingthecollectionofbloodsmearsat theWildlifeRecuperationCentreof‘‘LosHornos’’.Grantsand funds areasfollows:R.J. (TelefonicaMobilesS.A.), L.A. (C.S.I.C.–Ventorrillo,BOS2002–00598, BOS2002–00547), E.A. (C.S.I.C.–Ventorrillo,07M/0137/2000),A.M.(Juntade Extrema- dura,FIC01A043), G.T. (ResidenciadeEstudiantes;FPI from Comunidad deMadrid,BOS2000–1125, BOS2001–0587). This studyhasbeendoneaccordingtothecurrentlaws ofeachcountry.

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