Delivered by Ingenta to IP: 5.10.31.151 on: Tue, 07 Aug 2018 10:39:17 Copyright (c) American Society for Taxonomists. All rights reserved. © Copyright2009bythe American SocietyofPlantTaxonomists Systematic Botany sect. by Webster (1993 ) dividedthegenusinto40sections. Ee etal.2008 ). Julocroton era Astraea genus ismonophyleticaftertheexclusionofformersect. netic studiesof herbs are alsowellrepresented. Thefirstmolecularphyloge- shrubby habitpredominates across thegenus,buttrees and senescent leavesthatturnorangebefore absciscing.The flowers, clearorcolored sap, frequent petiolarglands,and narrow orcondensedthyrsoid inflorescences ofunisexual acters includingconspicuousstellateorlepidotetrichomes, species are easilyrecognized inthefieldbyasuiteofchar- logical synapomorphiesforitsconstituentsections. and subtropics worldwide), andthelackofdistinctmorpho- genus ( s. s.)haveproven tobeproblematic duetothelarge sizeofthe (2001) establishedafourthsubsection, subsect. (Griseb.) Müll. Arg., subsect. Cyclostigma and Madagascar( Table 1) . and , andafewspeciesfrom mainland Africa Webster (1993 ) initiallydivided tributed inlowlandandmontaneareas ofMexico,Central cumscription, thesectionconsistedofabout60speciesdis- and ashrubby toarborescent habit.FollowingWebster’s cir- nodes oftheinflorescence), stellatepubescence,reddish sap, nate flowersassociatedwiththepistillateonesatlower presence ofbisexualcymulesintheinflorescences (i.e.stami- sect. and expanded Grisebach’s sect. cymules). Theclassification of Webster (1993 ) incorporated ing themaincharacterdefining thesection(bisexuallower South America; thesespecies are smalltorobust shrubs lack- Webster, withthree speciesfrom Mexicoandthree from ( Baillon 1858 ; Grisebach 1859 ). Webster’s morphological dis- The mostrecent sectionalclassificationof of systematics and The Crotonopsis lineages inthegenus. has evolvedonmultipleoccasionsin gesting thatreticulate evolutionmighthaveplayedarole intheevolutionaryhistoryofthisgroup. Theseresults highlightt tions orinformalclades.TheITSandchloroplast phylogeniesare discordant regarding thepositionofredefined section with itsmembersappearinginninedifferent cladesthroughout group’s fullmorphologicalandgeographicrange.Thetwogenephylogeniesconcurred thatsection tid spacer of thespeciessampledisredefined here asareduced, monophyleticsection Palanostigma Cyclostigma Keywords— Abstract— (Klotzsch)Baill.,andinclusionofthesegregate gen- ∼

2 1,300 species),itswidegeographicalrange(tropics Palanostigma University ofMichiganHerbariumandDepartmentEcologyEvolutionaryBiology, 3600Varsity Drive, 1 Mart.,and Department ofBotany, UniversityofWisconsin-Madison, 430LincolnDr., Madison,Wisconsin 53706U.S.A. intothree subsections:subsect. oeua Pyoeeis f h Dao’ Bod Blood Dragon’s the of Phylogenetics Molecular (2009), 34(2):pp.360–374 trnL–F Michx., The phylogeneticrelationships of Griseb.( Webster 1993 ) wasdefinedbythe ofitn gn hsois, IS, Mdcnl lns, Norpc age e rg Drago , de Sangre Neotropics , , Medicinal ITS, histories, gene Conflicting , bothofwhichhavebasalbisexual cymules

3 Harvard UniversityHerbaria,22Divinity Avenue, Cambridge,Massachusetts02138U.S.A. sequencesfrom 48ofthe63speciesingroup, includingmembersofthefourrecognized subsectionsand representing the Moacroton Mart.exBaill.Subsequently, Webster andtribeCrotoneae indicatedthatthe Cubacroton (): A PolyphyleticAssemblageUnraveled iad Riina, Ricarda Croizat ( Berry etal.2005 ; van Sampatik Cyclostigma Aan Alain, Croton Croton

4 G.L.Webster, and Author forcorrespondence ( [email protected] ) Eremocarpus , andthatthere are othermorphologicalcharactersthatcaneffectively distinguishdifferent tree L.(Euphorbiaceae andBaillon’ssub- Xalapenses Croton Communicating Editor:AlanW. Meerow Croton Ann Arbor, Michigan48108U.S.A. 1 , 2 , 4 section al . Berry, E. Paul Cyclostigma proposed Benth., G.L. Croton Croton Cyclostigma Croton

360 . Thecladethatretains thetaxonomictypeofsectionandabouthalf sensuWebster were examinedusingnuclearribosomalITSandplas- 2 within species. tions isdubious duetothelackofconsistency ofthecharacter of bisexualcymules asadiagnosticcharacter for spread across thegenus. Webster (1993 ) admittedthattheuse Luntia

clear separationbetweenthesubsections( Table 1 ). of morphologicalcharacters,mostwhichdonotprovide a tinction ofthefour tions besides sect. the group asawhole.Forexample,othersectionsin could notidentifyanymorphologicalsynapomorphiesfor a doctoraldissertationbyB. Smith (2002 ). Interestingly, he was forasmallgroup of12speciesfrom , aspartof wound-healing ( Ubillas etal.1994 ; Jones 2003 ). marily fortreating cutstoprevent infectionandtoaccelerate domestic purposesinindigenousandrural communities,pri- on’s blood)becauseoftheblood-red latex,andare usedfor Müll. Arg.) are commonlyknownas“sangre dedrago”(drag- species of 1994 ; Meza 1999 ; are forthemedicinalpropertiesBorges oftheirlatex( andKing2000 ; Ubillas etal. Jones 2003 ). Several Smith 2002 ). However, themostpopularusesofthesetrees coal, andforcarpentrycabinetmaking( Lorenzi 1992 ; species isalsousedforboatandhouseconstruction, forchar- mental trees insomeColombian cities.Thewoodofseveral Kunth and and cityparksinsomeBraziliancities. 1992 ; Carrenho etal.1997 ), anditisalsousedforurbanareas toration ofdegradedriverbasinsinsouthernBrazil( Lorenzi For example, group idealcandidatesfor restoration ofdegradedforests. ecological characteristicsmakemostofthemembersthis (subsect. in moistforest habitats,but someofthesmallshrubby species mountain landslideareas. Mostmembersofthesectiongrow disturbed sitessuchasroadsides, stream andriveredges, are fast-growing trees andshrubs occurringpredominantly in n Bnai W vn Ee van W. Benjamin and Mmes f et sect. of Members The onlyrecent taxonomictreatment forsect. Cyclostigma Cleodora (Raf.)G.L.Webster), andstellatepubescence iswide- Xalapenses Croton Croton C. bogotanus , andtheremaining speciesare placedinothersec- (Klotzsch)Baill., Cyclostigma Croton urucurana

sect. trnL–F Cyclostigma ) grow inopenanddryvegetation.These Section Cyclostigma Cyclostigma . Cyclostigma Cuatrec. havesimilarusesasorna- havebasalbisexual cymules(sec- 1 , 3 subsectionsisbasedonasuite Baill.iscurrently usedinres-

sensuWebster ispolyphyletic, Eutropia snu Wbtr 19 2001) (1993, Webster sensu Cyclostigma (especially (Klotzsch)Baill.and hat thetree habit Cyclostigma Croton mutisianus Croton lechleri

Cyclostigma Croton sug- Croton sec-

Delivered by Ingenta to IP: 5.10.31.151 on: Tue, 07 Aug 2018 10:39:17 Copyright (c) American Society for Plant Taxonomists. All rights reserved. * * * * Leaves mostlypalmatelyveinedandcopiouslystellatebeneath;stamens10–100+;pistillateflowersdistinctlypedicellate,sepa South America * * * * South America Leaves pinnatelyveined,crenate, sparselyappressed-stellate; stamens10–20;pistillateflowerslong-pedicellate;sepalsofpi Mexico andC. America Leaves palmatelyveinedwithdensestellatepubescence;stamens12–35;basalcymulesfemaleflowersonly, femaleflowerss the monophyly andtherelationships of thefoursubsections port for in theanalyses. Ourgoalswere to(a)test themolecularsup- two non senting different hood, andBayesianapproaches. Severalotherspecies repre- Webster (1993 ) usingmaximumparsimony, maximum likeli- of manymore representatives of (1993 , 2001 ) and Smith (2002) . Speciesincludedinourmolecularsampling(48spp.)are indicatedwithasterisks. Leaves palmatelyorpinnatelyveined,subentire ordenticulate,copiouslystellatebeneath;stamens15–65;calyxofpistillate Monoecious trees orshrubs; indumentumstellate;stemsoftenexudingreddish sap;leavesalternate,mostlypalmatelyveinedor * * * * * Mexico andCentral America * Cascarilla sampled and cies aspartoftwosistercladesinacombinedanalysisITS of thegenus.Theirphylogeneticanalysisrecovered thesespe- Cyclostigma 361 RIINA ET AL.: DRAGON’SBLOODCROTON 2009] * * * C. fastuosus C.caldensis C.stenopetalus C. coriaceus C. chocoanus C. celtidifolius C. callicarpiifolius C. suberosus C. xalapensis C. cordiifolius C. sampatik C. anisodontus C. aequatoris C. abutiloides C. verapazensis C. draco C. ater C. echinocarpus C. draconoides C. densiflorus In thisstudyweanalyzeITSand Berry etal.(2005 ) included19speciesof Table styles multifid. cate-valvate; stylesbifid. styles bifid;petiolarglandssometimeslacking. valvate; stylesbifid. sepals entire, eglandular, valvateorreduplicate-valvate; stylesbifidtomultifid present; petalspresent instaminateflowers,usuallyreduced inpistillateflowers;stamens11–100 ormore; pistillateflowers nately veined,biglandulartomultiglandularatbase,glandssessileorstipitate,rarely lacking;inflorescence terminal,bise trnL–F Croizat Cham.&Schltdl. 1 Croton . . Croton Griseb.(includingformersect. Cyclostigma Ml. Arg. Müll. Ml. Arg. Müll. Kunth Baill. Croizat Croizat Kunth Kunth data;oneofthesecladesincluded severalofthe Kunth Pax&K.Hoffm. (= ebr o of Members sensu Webster (1993 ) inthefirstmolecular study Baill. Ml. Arg. Müll. Baill. Ml. Arg. Müll. G.L.Webster Donn.Sm.South America Müll. Arg. Vahl (= speciesastheoutgroup, were alsoincluded sect. Croton speciesintermixedwithofsect. Cyclostigma C. huberi Croton lineagesrelevant forourstudy, and sect. C. piluliferus Steyerm.) Croton Cyclostigma sensuWebster, particularly trnL–F Rusby) sect. Velamea s. l.(63spp.),subsectionalclassification,anddiagnosticcharactersofeachsubsectionsensu Webster sequencevariation Cyclostigma Section /SubsectionsDiagnosticcharacters subsect. Baill.). subsect. Croton Croton subsect. subsect. Palanostigma sensu sect. sect. Xalapenses Sampatik Cyclostigma Cyclostigma * * * * * * * * * * * * South America * * * * * * * * C.quadrisetosus C.nuntians C.killipianus C.huitotorum C.polycarpus C.boliviensis C. rimbachii C. palanostigma C. medusae C. lagoensis C. redolens C. purdiei C. perspeciosus C. mutisianus C. piptocalyx C. organensis C. magdalenensis C. pungens C. chilensis C. macrobothrys C. lechleri C. huberi C. hibiscifolius C. gracilipes C. gossypiifolius C. floccosus G.L.Webster to sect. proposed by Webster (1993 ); (b)identifytheclosestrelatives separate thisgroup from therest of stricto andidentifythemorphologicalcharactersthatbest cumscription ofamonophyletic species belonging to data matricescontain 48(37 we couldanalyze more thoroughly. Ourfinalreduced ITSand of the (1993 ). Thisapproach allowedustoselectonlyafewspeciesfrom most ( Table 1 ) andrepresentatives ofall40 sectionsof initial datasetincludedallthesampled membersofsect. mum parsimony(results notshownhere) inPAUP* ( Swofford 2002 ). This including alarger taxonsamplingacross G.L.Webster Mart.exBaill. Taxon SelectionandSampling Approach— Steyerm. Griseb. Ml. Arg. Müll. Ml. Arg. Müll. Croton Ml. Arg. Müll. Ml. Arg. Müll.

Pittier B.A.Smith Croizat Ml. Arg. Müll. Jacq. Baill. Croizat Cyclostigma Ml. Arg. Müll. Ml. Arg. Müll. Croizat Baill. Benth. Croizat Ml. Arg. Müll. Kunth Croizat Klotzsch Baill. Lam. Vahl Ml. Arg. Müll. lineagesascladeplaceholders andthushaveadataset

aeil n Methods and Material Croton within sect. trnL–F Croton Cyclostigma and47ITSsequences) outofthe63 Croton ; and(c)provide anewcir- Croton Croton stillate flowernotredupli- flower notreduplicate- * * * * * * * * * Africa andMadagascar * * snu Wbtr 19 2001; (1993, Webster sensu C sarcopetaloides C. C.wellensii C.pynaertii C.congensis C.draconopsis C.chypreae sect. xual basalcymulesusually C. smithianus C. urucurana C. sarcopetalus C. echioides C. ruizianus C. priscus C. goudotii C. oreades C. mongue C. warmingii C. speciosus wasanalyzedusingmaxi- ls ±reduplicate-valvate; distinctlypedicellate, A preliminary dataset Madagascar lobed, lessoftenpin- . essile orsubsessile; Croton Cyclostigma Croizat Leandri Baill. Leandri deWild. Baill. deWild. Ml. Arg. Müll. Baill. deWild. Ml. Arg. Müll. sensu Webster Ml. Arg. Müll. Baill. Croizat Ml. Arg. Müll. Ml. Arg. Müll. Cyclostigma S.Moore sensu trnL–F

Delivered by Ingenta to IP: 5.10.31.151 on: Tue, 07 Aug 2018 10:39:17 Copyright (c) American Society for Plant Taxonomists. All rights reserved. likelihood ratiotest ( Posada andBuckley2004 ). shown tobeasuperior methodofmodelselectionthan thehierarchical using MrModeltest v. 2.2( Nylander 2004 ) and AIC. The AIC hasbeen genetic analysis,theoptimalmodels ofsequenceevolutionwere inferred ate modelofDNA substitutionforeachdataset.FortheBayesianphylo- Modeltest 3.7( Posada andCrandall1998 ) toestimatethemostappropri- retained ineachrandomadditionsearch to100. tion TBRsearches, multrees ineffect, and limitingthenumberoftrees estimated using1,000bootstrapreplicates with100random taxonaddi- and multrees ineffect. MP bootstrappercentages (BP)foreachcladewere random taxonadditionsequencereplicates usingTBRbranchswapping detected mostparsimonioustrees. Thissearch wasconductedwith5,000 (1997 ) toensure thatthestrictconsensustree adequatelysummarizesall with thestrictconsensustopology, followingthestrategyof Catalán etal. bone constrainttosearch for trees ofequalorshorterlengthincompatible We usedtheconsensustree of1,000,000shortestretained trees as aback- random taxonadditionreplicate tosavenomore than1,000shortesttrees. ous trees exceededthecomputational capacity. We therefore limitedeach was notabletobecompletedbecausethelarge numberofmostparsimoni- as missingdata.Inthecaseof trees ineffect. All characterswere equallyweighted,andgapswere treated random taxonadditionreplicates usingTBRbranchswappingandmul- 4.0b10 ( Swofford 2002 ). We performedMP heuristicsearches with1,000 parsimony framework. gies, asimplementedinPAUP* 4.0b10( Swofford 2002 ) underamaximum Templeton test( Templeton 1983 ) wasusedtotestalternativetree topolo- Uninformative andgappedcharacterswere excluded.Inaddition,the sis usingsimpleaddition,heuristicsearches withTBRbranch-swapping. sand datapartitionreplicates were subjecttomaximum parsimonyanaly- Farris etal.1994 ) asimplementedinPAUP* ( Swofford 2002 ITS wasevaluatedusingtheincongruence lengthdifference test(ILD; ). Onethou- S2154), andsequencesare depositedinGenBank( Appendix 1 ). gram RDP3.Thedatamatricesare archived inTreeBASE (studynumber GENECONV ( Padidam etal.1999 ) usingthedefaultsettingsofpro- ods RDP ( Martin andRybicki 2000 ), MaxChi( Maynard Smith1992 The ITSsequenceswere analyzedwiththerecombination detectionmeth- ), and of 2001 ). Alignment ofthesedatasetswasfacilitatedbyourbroader sampling V2.0a11 (Rambaut1996–2002)orMacClade4.03( Maddison and ( Staden 1996 ), andthenmanuallyalignedusingSequence Alignment Editor were edited andassembledusingtheStadenPackageVersion 2003.0b1 at theUniversityofWisconsin-Madison BiotechnologyCenter. Sequences netic beadsandsequencedon ABI 377or ABI 3100automatedsequencers primers usedforPCR.Cyclesequencingproducts were cleanedwithmag- cycle-sequenced in10 at 72°C,followedbyafinalextensionof7min72°C.PCRproducts were 0.5 mindenaturationat94°C,1annealing48°C,and1.5extension PCR profile consistedofaninitial1.5minpremelt at94°Cand35cyclesof 6 SSEAI OAY [Volume 34 the using primers“c”and“d”forthe SYSTEMATIC BOTANY for herbariummaterial,amplificationwasperformedintwosegments, spacer were amplifiedusingprimers“c”and“f”of Taberlet etal.(1991 ); with herbariumsamples.ThecpDNA amplification failedwiththefirstpairofprimers,whichhappenedoften Internal primersITS3andITS4( White etal.1990 ) were usedwhenDNA using theprimersITS-I( Urbatsch etal.2000 ) andITS4( White etal.1990 ). Mini kits(Qiagen,Valencia, California).TheITSregion wasamplified ica-dried, andherbariumtissueofsingleindividualsusingDNeasyPlant 1. Appendix tion, GenBankaccessionnumbers,andsamplesources isprovided in of the83speciesusedinthisstudywithauthorities,voucherinforma- rent P.E. Berry&Cordeiro and holders, andtwotaxawere usedasoutgroups ( Croton Cyclostigma most ofthesection’sgeographicalrange,includingseveralOldWorld Table 1 ), withrepresentatives from allfoursubsections,andspanning 362 used 25 each primer(20mmol/L),15.8 each), 1.25 polymerase, 2.5 The Akaike Information Criterion(AIC)wasusedintheprogram Maximum parsimony (MP)analyseswere conductedusingPAUP* Analyses— Phylogenetic Molecular DA eunig n Alignment— and Sequencing DNA Polymerase chainreaction (PCR)amplificationoftheselectedmarkers Croton trnL–trnF Croton cladeswere selectedfrom theinitiallarge datasettoserveasplace- μ , othermembersoftribeCrotoneae, andotherEuphorbiaceae. L reactions containing0.2 molecularphylogenetichypothesis( Berry etal.2005 ). A list μ species.Inaddition,oneortworepresentatives ofmanyother L DMSO,0.25 spacer. μ L 10 μ × l reactions usingBigDyereaction kitsandthesame ExTaq Buffer, 3 Astraea lobata μ L 0.4%bovineserum albumin (BSA),0.5 μ L ddH trnL trnL–F μ L of5units/ DNA wasextractedfrom live,sil- intron andprimers“e”“f”for nogune ewe between Incongruence 2 trnL 0, and1 (L.)Klotzsch),followingthecur- dataset,theparsimonysearch μ L dNTP mixture (2.5mmol/L intron and μ Brasiliocroton mamoninha L oftemplateDNA.The μ L Takara ExTaq DNA trnL–F trnL–F intergenic and μ L

chains. credibility valueswasobtainedbycombiningthetrees from bothMCMC for evidenceofgoodmixing,amajorityrule consensustree withclade visual examinationofalikelihood-by-generationplot. After inspection and samplingevery50generations.Theburn-inperiodwasestimatedby four linkedchains(sequentialheat=0.2)run for1,000,000generations, two MarkovchainMonteCarlo(MCMC)analyses,eachconsistingof program MrBayes3.1.2( Huelsenbeck andRonquist2001 ). We conducted topologies forconsistency. Bayesiananalyseswere conductedusingthe pendent heuristicsearches inGARLIandcompared theresulting tree gram GARLIversion0.951( Zwickl 2006 ). We performedfourinde- F evidence ofintragenicrecombination. Thelengthofthe RPD, MaxChiandGENECONVmethodsdidnotdetectany sequences overall.TheanalysisoftheITSdatasetusing was 2.7%. Almost noambiguoussitesare foundintheITS tive. Thepercentage ofmissingcharactersfortheITSmatrix 349 are variablecharacters,and261are parsimony informa- aligned lengthoftheITSdatasetis771positions,which (outgroup) to734basesin of theITSsequencesvariedfrom 713basesin for ITS( trnL–F not beamplifiedsuccessfullyforoneofthetwomarkers.The from samplesextractedfrom herbariummaterialthatcould trnL–F their taxonsampling,with83accessionsforITSand75 for the are parsimonyinformative. Thepercentage ofmissingdata is 1,222bplong,andhas344variablecharactersofwhich178 to 1080basesin included intheITSdataset: Arg., sis Leandri, Cyclostigma circumscription of Webster (1993 ; see From here on,“sect. Table 1 ), and“sect. eral different Cyclostigma on thisdataset(MP, ML,BA)recovered apolyphyleticsect. Bayesian analyses. All three analyticalapproaches performed I + tial strictconsensus. the shortestlengthfoundthatwasincompatiblewithini- constraint asdescribedinthemethods,norwasanytree of the strictconsensustree from thesearch aboveasabackbone ters). Noshortertrees were foundafterasecondsearch using were calculatedincludingparsimonyuninformativecharac- Length (L)=540(CI0.752,andRI0.830;bothCI RI search ofthe thus theindelcodingwasdropped from thefinalanalyses. already highlysupported whenusingthenongapcodeddata, using parsimony, butthecladesthattheysupportedwere informative. We codedtheindelsandanalyzedmatrix have numerous indelsand someofthemwere parsimony cies, including thetypeofsection( the monophyletic cladethatretained the majorityofthespe- sequencesvariedfrom 909basesin Maximum Likelihood(ML)analyseswere conductedusingthepro- Size andStructure ofMolecularDataSets— For thisdatasetthegeneraltimereversible model(GTR+ Region— trnL–F Chloroplast of Analysis Donn.Sm.,and Γ ) modelwasselectedforbothmaximumlikelihoodand C. magdalenensis datamatrixincludesonlyonespeciesnotsequenced ( Appendix 1 ). Themissing sequencesare primarily trnL–F Croton draconoides C. redolens s.s.,”orsimply“the s.l.withalmosthalfofitsspeciesresolved insev- Croton matrixwas8.4%.Thesedatasetsdiffered in trnL–F C. huberi Pittier, C. xalapensis lineages,asshownintheML tree ( Fig. 1 ). Cyclostigma datasetretained 1,000,000trees with Müll. Arg., Steyerm.Thealigned Müll. Arg.), andlacksninespecies Results C. smithianus C. pachypodus C. ater Kunth.The s.l.”refers tothetaxonomic Cyclostigma C. mongue Croizat, Astraea lobata C. gossypiifolius Croizat, G.L.Webster. The trnL–F C. lagoensis Baill., clade,”refers to h parsimony The trnL–F Astraea lobata C. verapazen- (outgroup) h length The sequences C. oreades matrix Vahl). Müll. trnL–

Delivered by Ingenta to IP: 5.10.31.151 on: Tue, 07 Aug 2018 10:39:17 Copyright (c) American Society for Plant Taxonomists. All rights reserved. with grayvertical bars(inmostcasescladenamescorrespond toWebster’s namesforsectionsorsubsections). posterior probabilities (PP)are below branches.Specieswithagraybackground are membersofsection 363 RIINA ET AL.: DRAGON’SBLOODCROTON 2009] Fig. 1 . . Maximum likelihoodtree obtainedfrom analyses of trnL – F data;parsimony bootstrappercentages (BP) Cyclostigma > 50%are abovebranchesandBayesian s.l.Main Croton cladesare indicated Delivered by Ingenta to IP: 5.10.31.151 on: Tue, 07 Aug 2018 10:39:17 Copyright (c) American Society for Plant Taxonomists. All rights reserved. C.oreades that group. clades, onlyin thiscasethe tion sistertothegroup formedby trnL–F corroborate theresults ofthe represented here bythree species( et al.(2005 , Fig. 5 , cladesC-5toC-11). as showninthecombinedITSand of otherNewWorld sectionsthatincludesthesamegroups also recovered ahighlypolyphyleticsect. ses. Theresults oftheMP, ML,and BA oftheITSdata matrix was selectedforthemaximumlikelihoodandBayesiananaly- characters). Thegeneraltimereversible model(SYM+I indexes were calculatedincluding parsimonyuninformative parsimonious trees (L =1,179,CI0.465,andRI0.758;both discussion). are distributedamong seven different Other membersofsect. the etic group sistertothelarge NewWorld cladethatincludes ( Fig. 1 ). All theOldWorld speciessampledformamonophyl- C.goudotii included inthe of theNewWorld clades,including large cladecontainingtheOldWorld speciesandthemajority Fig. 1 ). The lineages ofNeotropical as partofthesect. less than2or3mtall.Thisphylogenyrecovers thesespecies sis, C.sarcopetalus i. . 2). ( Fig. the sistercladeof depicted intheML tree ( Fig. 2 ). [Volume 34 SYSTEMATIC BOTANY Müll. Arg., clade ( three different analyses(MP, ML,BA). least three clades,andthisconfigurationwasrecovered bythe The reduced sect. Lamprocroton Griseb., Croton is sistertoahighlydiversifiedgroup includingatleastseven parsimony BP and1.00BayesianPP. Along with bility (PP).The percentage (BP)supportand1.00Bayesianposteriorproba- group ofsect. The 364 Cyclostigma trnL–F clade anditsclosestrelatives within logeny regarding thephylogeneticpositionof shows amajordifference intopologyfrom the s.l. includingthetypespecies.However, the ITSphylogeny one ofthesecladesretained halfofthesampled species comingoutinninedifferent clades( Fig. 2 ). Similarly, phyletic sect. (MP, ML,BA)performedontheITSmatrixrecovered apoly- The OldWorld taxaassignedtosect. Aayi o ITS— of Analysis The OldWorld representative ofsect. ie h the Like Half ofthespeciessampledthatfalloutside Cyclostigma trnL–F cladesorsections( phylogeny, the OldWorld cladeoccupiesthesameposi- phylogeny, inwhich C. abutiloides Croton cupreatus Priscus ), andtheirpositionsoutsidethe Baill.,alsofelloutsideofthe C. pungens trnL–F datasupports , theITSdatastrongly supportssect. Cascarilla (Müll. Arg.) Pax,and Cyclostigma Cyclostigma cladeandtenother Cyclostigma clade, Müll. Arg., and trnL–F phylogeny, allthree analyticalapproaches Cyclostigma Cascarilla Cyclostigma Kunth, The MP search resulted in24,992most clade(95%BP, 1.00PP)issistertothe Jacq., dataset,theMadagascanendemic Croton isrecovered sistertoanassemblage s.l.withmore thanhalfthesampled Cyclostigma Geiseleria s.with84%parsimonybootstrap Barhamia cladeiswellsupportedwith79% clade,oneofthemost species-rich C. cupreatus C. ruizianus C. aequatoris s.consistsofapolytomyat Cyclostigma s.s.,with100%BP and1.00PP C. cupreatus (C-3cladein Berry etal.2005 ; trnL–F C. gracilipes (Klotzsch)Baill., Argyroglossum C. goudotii trnL–F (Klotzsch)Baill., Cyclostigma Croton s.l.sampledfor Cupreatus–Argyroglossum Croton phylogeny. As inthe Müll. Arg., Croizat asthesister cladeisnotpartof Croizat, phylogenyin Berry Cyclostigma wassistertosect. Croton cladesorsections. Cyclostigma Cyclostigma Cyclostigma . Contrarytothe Baill.)are shrubs Cyclostigma , , C. mongue s.( Fig. 1 ). C. cupreatus Baill.; Fig. 1 ). clades(see trnL–F Cascarilla Cyclostigma Cyclostigma Cyclostigma C. chilensis C. xalapen- s. l. are Eluteria s.l.,as Luntia, trnL–F clade clade phy- s.l. , and , as Γ it )

data. Although mostofthe bone ofthe (100% BS,1.00PP). As withthe strained sect. data setssupportthesetwoconflictingtopologies.We con- available ( including alltaxaforwhichboth (ILD) test( Farris et al.1994 ) appliedtothecombineddataset confirmed bytheresults oftheincongruence lengthdifference the positionof topological difference betweenthetwogenephylogenies is two phylogenies. ogies donotalwayscorrespond exactlywhencontrastingthe by ITSwere alsosupportedbythe dence— C.cupreatus content isinfluencingthephylogeneticassociationofeither sets andwedidnotfindanypatternsuggestingthattheGC ined AT:GC nucleotideratiosacross allspeciesinourdata similar tothetopologyofITSphylogeny. We alsoexam- the tree topologyresulting from thecombineddatasetismost support inothers(duetotheconflictingclades).Ultimately, resolution insomepartsof thephylogeny, andadecrease in from different discordant generegions, there isanincrease in not shown). As wouldbeexpectedwhencombiningdata matrix toobservetheresulting phylogeneticpattern(results Nonetheless, weconductedanalysesonthecombineddata answer ourinitialquestionsaboutthisgroup within sets becausetheseparategenephylogeniesare sufficient to Cyclostigma ITS datadoesnotreject a hypothesisinwhich test wasnotsignificantinthiscase( Webster’s (1993)sectional nomenclature wheneverhis view sion ( Fig. 2 ). We assignedprovisional namestocladesusing topologies, andweselectedthe ML tree toguide thediscus- sary. Thetrees derivedfrom MP, ML, andBA havecompatible Cyclostigma phylogenetic relationships amongthedifferent taxaofsect. Webster’s sections,including Croton here, togetherwithothermolecularphylogeneticstudiesof that are largely homoplasiousacross thegenus.Ourresults the sectionsare definedbysuitesofmorphologicalcharacters ous synapomorphiestosupporttheindividualsections. All cal similaritiesamongspecies,buthedidnotfindanyobvi- of thegenuswasbasedongeneralpatternsmorphologi- ing ofthiscomplexandlarge genus.Webster’s subdivision without doubtanenormouscontributiontotheunderstand- see ifthe do nothavea the ITSdataset,wemadeaconstrainttosearch fortrees that significant usingtheTempleton test( was ninestepslongerthantheunconstrainedtree, whichwas relationship. Theshortesttree compatiblewiththis constraint Te The We usedtheTempleton testtoevaluatehowtheseparate Congruency— Data Croton SectionCyclostigmainLightofMolecularEvi- For thisstudywedidnotcombinethe We usetheITSphylogenytoreview themainpatternsof ( Berry etal.2005 ; van Eeetal.2008 ), showthatmanyof Cyclostigma The sectionalsynopsisof p trnL–F s.l.,referring tothe are notsistertoeachother. =0.007). orsect. Cyclostigma Cascarilla Cascarilla datasignificantlyrejects thisphylogenetic cladeishighlysupportedbytheITSdata Cascarilla Cyclostigma From ourresults itisclearthatthemain + tobesisterthe cladeisnotfullyresolved bytheITS Cyclostigma Discussion Cyclostigma withthe Cyclostigma trnL–F clade.Thisobservationwas Croton trnL-F trnL–F trnL–F s.clade.TheTempleton p =0.0499–0.0974),sothe p phylogenywhenneces- Cyclostigma =0.0201).Then,using andITSsequencesare b ese (93) was (1993) Webster by phylogeny, theback- subcladesrecovered dataset,theirtopol- , are polyphyletic. trnL–F Cyclostigma Cascarilla andITSdata clade. cladeto Croton and . Delivered by Ingenta to IP: 5.10.31.151 on: Tue, 07 Aug 2018 10:39:17 Copyright (c) American Society for Plant Taxonomists. All rights reserved. most casescladenamescorrespond toWebster’s namesforsectionsorsubsections).Webster’s (PP) are belowbranches.Specieswithagraybackground are memberofsect. symbols totherightofeachspecies name. geographic distribution. Someoftheseclades were already clades. TheOld World cladewas theonlyonenamedafter used thenameofanincludedspecies tonameadditionalnovel of asectionwascompatiblewith ourphylogeneticresults. We 365 RIINA ET AL.: DRAGON’SBLOODCROTON 2009] Fig. 2 . . Maximum likelihood tree from ITSdata;parsimonybootstrappercentages (BP) Cyclostigma Neotropical, with theexceptionofOldWorld clade. Every lineage in thisphylogeny, includingtheoutgroup, is other studieswithin identified by Berry etal.(2005 ), andotherswere identifiedby s.l.Main > 50%are abovebranches,andBayesianposteriorprobabilities Cyclostigma Croton Croton cladesare indicatedwithgrayvertical bars(in subsections( Table 1 ) are alsoindicatedwith (vn e 06; vn e t l 2008). al. et Ee van 2006; Ee (van Delivered by Ingenta to IP: 5.10.31.151 on: Tue, 07 Aug 2018 10:39:17 Copyright (c) American Society for Plant Taxonomists. All rights reserved. of is produced. Inanextensive surveyonthewoodanatomy secondary xylemoflaticifersorspecializedcellswhere latex covered characterforsect. stipules, andstellatepubescence( Figs. 3 , 4 a cordate base),twoormore petiolarglands,welldeveloped ). A recently dis- more than150),broad, palmately-veinedleaves(usuallywith nate flowerswithmore than 16stamens(afewspecieswith ment-like petalsare present), bifidtoquadrifidstyles,stami- apetalous pistillateflowers(sometimeshighlyreduced, fila- the basalbisexualcymuleswithpedicellatepistillateflowers, grado” species.Othercharactersshared bythesespeciesare include thewidelyknown“sangre dedrago”or“sangre de to large-sized trees thatproduce red latexinthetrunks and s. l.Mostofthespeciesin species from oursamplingof48speciessect. and 1.00PP, Fig. 1 ). Thecore trnL–F by theITSdata(70%BP, 0.70PP, Fig. 2 ), results from the etic trnL–F C.piptocalyx tion oflaticifers,are unique tothe Although noneoftheseindividualcharacters,withtheexcep- bear laticifersandthatthecharactercouldbepolymorphic. s. s.,howeveritappearsthatnoteveryspeciesinthesection et al.(2009 ) foundthatlaticifersare uniquetosect. created anew subsectionwithinsect. most species-rich lineageswithin seven placeholdersfrom sect. Cyclostigma ary relationship shownbyITSand Webster’s subsectionaldivisiondoesnotreflect theevolution- related clades.Thisphylogeneticpatternclearlyshows that [Volume 34 SYSTEMATIC BOTANY Cyclostigma acters thatseparatetheselineageswithputativemembersof have misguidedWebster andindicatemorphologicalchar- sense. We alsodiscussthemorphologicalpatternsthatcould ing putativemembersofsect. shown bythesubsectionsandanalyzeeachcladecontain- Cyclostigma defined subsectionsfortwoofhis40sections,namelysect. shows asimilarpatternofdiscordance ( Fig. 2 ). Webster only sectional subdivisionofWebster mappedonthephylogeny spread outinninedifferent clades( Fig. 2 ). Likewise,thesub- sensu Webster islargely polyphyletic,withitsmembers 366 and whereas membersofsubsect. All speciesofsubsect. to sect. side ofthe as wellseveralothermembersofthissubsection,fallout- only partofsubsect. especially montaneandriparianforests. species inthiscladealsohaveapreference formoisthabitats, ence ofthissuitecharactersisuniquetotheclade.The Cleodora However, thetypeofsubsect. clade, besidesthenominalsubsect. The CyclostigmaClade(Sections.s.)— Scin Cascarilla— Section Our phylogeneticresults indicatethatsect. Of thefoursubsectionsofsect. Croton Cyclostigma C. sampatik analysisgivebettersupportforthisclade(79%BP andITSdataallowustoidentifyacore monophyl- , and Luntia andofsect. s.l.were placed inawellsupportedcladewith s.l.from membersofsect. andsect. Cyclostigma Müll. Arg., , but C. goudotii Müll. Arg.) are distributedinfourdistantly clade. Although itismoderatelysupported C. warmingii Palanostigma Cyclostigma Luntia Ten ofthe48sampledspeciessect. clade.Mostofthesespeciesbelong Xalapenses C. organensis belongstotheOldWorld clade. Cyclostigma . Belowwediscussthepattern Cyclostigma Palanostigma Cascarilla Sampatik Cyclostigma Cyclostigma isretained inthe Müll. Arg. belongstosect. inparticular, Wiedenhoeft Cyclostigma belongtosect. trnL-F Cyclostigma Baill., Croton s.isthepresence in Cyclostigma ( Cyclostigma , which isoneofthe Cyclostigma . C. cordiifolius cladeare medium claderetained 21 inthetraditional ( C. priscus . Wbtr (2001) Webster . C. palanostigma clade,thepres- s.l.( Table 1 ), Cyclostigma Cyclostigma Cyclostigma Cyclostigma s. (Fg 2). (Fig. (subsect. Cascarilla Croizat, Baill., Both ), ,

( with fivespeciesformerlyplacedinsubsect. and matourensis Müll. Arg. (includedinthisanalysis),wehave sampled Matourenses section (the Cuneati sect. Cascarilla acters present inmembersofsect. subsect. copetalus members ofthe of thecolumella,mostspeciesinsect. dispersal. Besidestheshrubby habitandthedistinctshape unclear, butitmaybeinvolvedinpollentubegrowth orseed ing thesetwolineages.Thefunctionalrole ofthischaracteris then thischaracterisakeymorphologicaldifference separat- sect. tion. Inaddition,ifothergeneticmarkersbesidesITSconfirm synapomorphy isusefulasadiagnosticcharacterforthesec- Section ing, withsmooth,rounded andinflatedventralsides( Fig. 5 ). minal appendagesonthecolumellathatare usuallyascend- synonymized in Berry etal.2005 ) havethree prominent ter- Cascarilla or specialfeatures. However, allmembersexaminedinsect. and inmostspeciesof species usuallyremains afterthedehiscenceofthree cocci, cumscription. Thecentralcolumellaofthecapsule phy forsect. Cascarilla Ee 2006 ). Mostmembersofsect. sect. subsect. posed ofamixture ofspeciesformerlyfrom sect. tion the typeofsection( sect. we canpredict thatthefollowing specieswillalsobepartof the unsampledspeciesofsect. environments. From theexaminationofmorphology rower pistillatesepals,and theirpreference fordryorarid cymules, sessilepistillateflowers,muchsmallerandnar- rant withinsect. Xalapenses subsect. pistillate flowers.Oursamplingincludedfourspeciesfrom ual cymulesatthebaseofinflorescence andhavesessile included twosubsections( and Cyclostigma (100% BS,1.00BP)containingthree putativespeciesofsect. ogy andare likelytobemembersofthe Croton distal appendages.Manytaxapreviously assignedtoother trichomes, andtheyallhavethecolumellawithinflated small shrubs withsessilepistillateflowers,bifidstyles,stellate L. Webster, and C. abutiloides While examiningmorphologicalfeatures insections eto Luntia— Section C. pungens Cuneati C. palanostigma Cyclostigma Cascarilla Matourenses Matourenses Cascarilla sectionsorunclassifiedspeciesexhibitthismorphol- ( Matourenses Cascarilla ) andonespecies( Sampatik C. cuneatus Palanostigma Xalapenses speciesshare thesamesuiteofcharactersdefining (includingmembersofWebster’s sect. and ) thatincludedseveralspeciesheconsidered aber- Aubl. and s.l.( ( Webster 1993 ). In additionto cladein Figs. 1 and 2 ) thatisnotsistertosub- , , Cascarilla C. aequatoris Cyclostigma astheclosestrelative ofthe : : ), andtheyallemerge insect. C. sarcopetaloides s. C. boliviensis Cyclostigma . Thetypespeciesofsect. , andtheyform amonophyleticclade( van Cyclostigma isthelargest cladewithin . Although someofthese specieshavechar- C. chocoanus ( ) isshownhere tobelonganovelclade . Consequently, sect. This group isawellsupportedclade ) ( Fig. 2 ). Section C. xalapensis andmembersofsect. C. skutchii Croton thatsupportsitsnewmolecularcir- C. matourensis , wediscovered anovelsynapomor- , , C. gracilipes C. echioides cladeintheabsenceofbisexual s.l.becausetheylackthebisex- Matourenses Müll. Arg. All thesespeciesare itdoesnotshowanydistinctive Croizat, Cyclostigma Standl.from sect. S.Moore, , , Luntia C. suberosus Cyclostigma , , ) belongingtosubsec- C. ruizianus Müll. Arg.) placedin Luntia C. smithianus Cascarilla Cascarilla s. exhibitlaminar and Luntia s.l.from Table 1 , Cyclostigma C. stenopetalus C. megalodendron Croton sensuWebster Cascarilla Luntia Luntia Cuneati , noneofthe , , s.iscom- differ from clade. C. chilensis Cyclostigma Cyclostigma , and Velamea Luntia , andthis subsect. subsect. Croizat, ) with ), clade, Croton along C. sar- sub- , as , G. C. ,

Delivered by Ingenta to IP: 5.10.31.151 on: Tue, 07 Aug 2018 10:39:17 Copyright (c) American Society for Plant Taxonomists. All rights reserved. ( 09 RIAE L:DAO’ LO RTN 367 RIINA ET AL.: DRAGON’SBLOODCROTON 2009] I. Pistillateflowers withelongate,green bifid styles( sae C. coriaceus Fig. ). G.Echinate-muricatefruits withbifidstyles ( 3 . . Floral andfruit morphology of ). D.Quadrifidstyles( C. perspeciosus Croton ). E.Fruits andbifid styles ( C. echinocarpus sect. C. floccosus Cyclostigma ). ). H.Long-pedicellatepistillate flowers/fruits withbifidstyles( s. A. Thyrsoidinflorescence ( C. piluliferus ). F. Oneofthefewexamplesshort inflorescences ( C. lechleri from ). B–C.Bisexualcymulesand fruit C. lechleri from Ecuador). C. medu- Delivered by Ingenta to IP: 5.10.31.151 on: Tue, 07 Aug 2018 10:39:17 Copyright (c) American Society for Plant Taxonomists. All rights reserved. hrys piluliferus 6 SSEAI OAY [Volume 34 SYSTEMATIC BOTANY 368 Fig. . H.Oneofseveral typesofstellatetrichomesinsect. 4 . . respectively. D–F. Petiolar glands( Vegetative features shared bymembersofsect. C. lechleri from Ecuador, Cyclostigma Cyclostigma . . C. piluliferus s. A. Arborescent habit( , and C. rimbachii C. echinocarpus ). G.Redlatex bleedingoutofthetrunk of ). B–C.Stipulesin C. echinocarpus C. macrobot- and C. Delivered by Ingenta to IP: 5.10.31.151 on: Tue, 07 Aug 2018 10:39:17 Copyright (c) American Society for Plant Taxonomists. All rights reserved. mella of fruit dehiscenceshowingthe persistent columella( 369 RIINA ET AL.: DRAGON’SBLOODCROTON 2009] Fig. 5 . . C. ruizianus Structure ofthedistal endofthecolumellamemberssect. . E–F. Columellaof C. floccosus (sect. C. abutiloides Cyclostigma ). B–C.Two views ofthedistalendcolumella of s.s.)lacking thepronounced andinflated distalappendagestypicalofsect. Cascarilla (A–D), andsect. Cyclostigma s.(E–F). A. Mature inflorescence after C. pungens . D.Distalendofthecolu- Cascarilla . . Delivered by Ingenta to IP: 5.10.31.151 on: Tue, 07 Aug 2018 10:39:17 Copyright (c) American Society for Plant Taxonomists. All rights reserved. Argentina. TheMP analysisof shrubs, alldistributedinsoutheasternBrazilandnorthern Lima (pers.comm.),whostudiessect. as thesisterspeciestoentire clade(unpubl.data).Letícia mately 20 analyzed inadifferent datasetincludingalloftheapproxi- sister tosect. and BA analysesof the phylogeneticpositionof Section not shared withanyother speciesof (most Croton cordiifolius ing strongly support(99%BS,and1.00BP)anovelcladeinclud- new samplesof short axillaryinflorescences. Additional sequencedatafrom habit, pinnatelyveinednarrow leaves,absenceoflatex,and However, itdiffers from sect. iolar glands,stellatetrichomes,andpinnatelyveinedleaves. ence ofinflorescences with bisexualcymules,bifidstyles,pet- (1993 ) insect. branch attraction. from herbariummaterial), oritmaybetheresult oflong poor sequenceduetolowqualityDNA (DNA wasextracted this relationship isanartifact ofinadequatetaxonsampling,a and membersofsect. find anyclearmorphologicalsimilaritybetween tion, the yloides Lamprocroton , andourphylogeneticanalysesplaceitsistertosect. diifolius and cate withstellate trichomesonthetopoftall andthickened cate sepals.In addition, theirfruits are conspicuously muri- the apexofteeth,andpistillate flowerswithlarge redupli- veined leaves,crenate-dentate leafmargins withglandson fer from membersofthe ern Braziliandistribution.The speciesinthe arborescent habit,reddish tocolorlesslatex,andasoutheast- veined leaves,petiolarglands, stellate-appressed trichomes, bisexual cymules,quadrifidstyles,muricatefruits, pinnately Brazil (R.Riinaunpubl.data).Thesethree speciesshare basal cies inthiscladeisanundescribedspeciesfrom southeastern assigned toanyoftheWebster’s sections( Fig. 2 ). Thethird spe- in sect. logenetic positionofthisspecies. [Volume 34 SYSTEMATIC BOTANY cal similaritiesare predict willbemembersofthiscladebasedonmorphologi- 6–12-fid styles.Otherspeciesofsect. trichomes, reduplicate calyces,fewerthan16stamens,and Cyclostigma more branches.Membersofsect. ing outwards intoanobviousflange),andstyleswith6–12or lobes thatare strongly valvate,withtheadjacent edges fold- reduplicate calycesinbothstaminateandpistillateflowers(i.e. spicuous bracts(3–4)subtendingeachcymule,10–15stamens, dote trichomes,petiolarglands,basalbisexualcymules,con- share thearborescent habit,acombinationofstellate andlepi- glands scattered onbothsurfacesoftheleaves.Theyalso 370 Croton cordiifoliusandSection Lamprocroton— Te rsu Clade— Priscus The C. priscus C. nuntians Baill., Croton isashrubby speciesfrom thecoastofsoutheastern Cyclostigma Lamprocroton trnL–F Lamprocroton s.inthepresence oflaminarglands,lepidote (represented inourphylogenyby specieshaveterminalinflorescences). Inaddi- andanotherspecies( C. pallidulus Lamprocroton Cyclostigma Croizat ( Table 1 ). sequenceforthisspecieshasa33bpdeletion C. cordiifolius isunusualinhavingaxillaryinflorescences C. huitotorum Croton cordiifolius subsect. trnL–F consistsofaround 20speciesofsmall Lamprocroton Croton priscus speciessampled todate,itcomesout Baill.,and subsect. Cyclostigma (Fg. . hn When 2). 1, (Figs. , andallanalysesofITS,placeit Sampatik are neededtoconfirmthephy- C. cordiifolius trnL–F Cyclostigma Croizat, Luntia . Thepossibilityexists that Sampatik C. eichleri wasincludedbyWebster wasplacedby Webster dataisunabletoresolve ( Table 1 ), andourdata Cyclostigma cladeinthepinnately C. troncosoi Lamprocroton Croton s.differ from sect. C. killipianus s.s.initsshrubby , butboththeML , duetothepres- Priscus sampled sofar. Müll. Arg.) not C. cordiifolius C. cordiifolius s.l.thatwe Ahumada). C. erythrox- , couldnot Croton cor- cladedif- Croizat, is

species ofsect. stalks. However, thisfruit characterhasalsoevolvedinone included byWebster insect. and stellateindumentum.However, theMalagasy species cences, bisexualcymules,multifidstyles,petiolarglands, cies ofsect. s. l.haveanoverallresemblance totheNeotropical spe- of sect. exclusively aNewWorld lineage. into accounttodefinitelyconfirm thatsect. phological studieswillneed totakethese African species branches ofsect. in theMalagasyspecies,contrast totheusuallylongstyle Another difference isthe reduced lengthofthestyle branches ers orhavethemreduced toglandularfiliformappendages. sect. spicuous petalsinthepistillateflowers,whereas speciesof from southeasternBrazil). shown here. TheOldWorld membersofsect. World clade,followingthepatternofMalagasytaxa De Wild., and sect. Old World clade. Thissuggeststhatthe African speciesof pled (unpubl.data)andtheyallbelongtoamonophyletic Approximately 40more OldWorld specieshavebeensam- Croton et al.(2005 ) recovered all21oftheOldWorld speciesof sect. showing thatthe African species related tosect. logeny indicatesthattheMalagasyspeciesare notclosely and oneoftheminbothmatrices( Section or three fourthsoftheirlengthformingatubularstructure. times fusedpistillatesepals,quadrifidstylesonehalf cymules, 15–30stamens,imbricate,accrescent, andsome- habit, red tocolorlesslatex,petiolarglands,basalbisexual acters shared bymembersofsect. C.cajucara members ofWebster’s sect. supported clade(78%BS,1.00PP)thatincludesmostofthe Our results supportthesetwospeciesbeingpartofawell whereas theleavesof venation (acharacterdefiningsubsect. into different subsections. probably onecharacterusedbyWebster toseparatethem Palanostigma were placedbyWebster insubsect. and C.oreades sampling waslimitedtothree Malagasyspecies( continental Africa andfourfrom Madagascar( Table 1 ). Our Croton tubular structure. sepals, andthecharacteristicallyconnatestylesforminga trichomes, pinnatevenation,imbricateorfusedpistillate separated from sect. pinnately andpalmatelyveinedleaves.Thisgroup canbe ineous lepidoteindumentum.There are alsospecieswith well asspecieswithadensesilveryand/ordarkferrug- Te aptk Clade— Sampatik The Scin Cleodora— Section Te l Wrd Clade— World Old The C. pynaertii Cyclostigma Cyclostigma Cyclosigma includedintheirsamplingasamonophyleticgroup. from theOldWorld insect. Cleodora Cyclostigma , and Benth.and respectively ( Table 1 ). Leafvenationwas Cyclostigma C. pynaertii Cyclostigma C. goudotii , whichwere assignedby Webster (1993 ) to s.l.,donotbelongthere. However, Berry Cyclostigma containsspecieswithstellatetrichomesas s.eitherlackpetalsinthepistillateflow- s.l.( Cyclostigma subsect. Cyclostigma C. draconopsis Croton organensis C. hoffmannii C. warmingii : theyare trees withlong inflores- This clade includestwomembers ), allincludedintheITSdataset Webster includedeightspeciesof DeWild.) are apartoftheOld . We donothave moleculardata ( s.Future molecularandmor- Croton organensis C. echinocarpus Sampatik Cleodora Cyclostigma bythepresence oflepidote C. draconopsis C. goudotii Cyclostigma Cleodora Müll. Arg., Müll. Arg. Somechar- are palmatelyveined. , here represented by Sampatik , namely ad and Sampatik s.l.allhavecon- Müll. Arg. also are arborescent ). OurITSphy- Cyclostigma andsubsect. haspinnate s.l.,sixfrom , , C. warmingii C. piptocalyx C. congensis C. congensis Cyclostigma C. mongue , Tbe 1), Table , is , ,

Delivered by Ingenta to IP: 5.10.31.151 on: Tue, 07 Aug 2018 10:39:17 Copyright (c) American Society for Plant Taxonomists. All rights reserved. sis French Guiana). Secco etal.(2001 ) described colombianus C.trombetensis ous morphologicalsimilaritieswiththe unsampled speciesfrom the Amazon region thatshowobvi- Croton sister tosect. while intheITSanalyses( Fig. 2 ) itisrecovered asaclade Corylocroton Sampatik Corylocroton C-1 in Berry etal.2005 ), represented inoursamplingbysect. 2005 ), oratthebaseof base ofthecore Müll. Arg. asagradeintwoalternativepositions,eitheratthe from southeasternBrazil. ole. Another speciesinthiscladeis since theydiffer inshape, size,andpositiononthepeti- be homologoustothepetiolarglandsofsect. olar glandsinthespeciesof needs tobeexaminedmore broadly within might beasynapomorphyforthisclade,butthecharacter spots onadarkbrown background. Thehornlike nectaries and thelaterallycompressed seedswithregular whitish minate flowers,15–20stamens,deeplytrilobedcapsules, the leafmargins, presence ofhorn-likenectariesin thesta- stipitate glandsontheabaxialsurfaceoflaminanear trichomes, pinnatelyveinedleaves,dentateleafmargins, and resemble eachotherinthestellate(appressed)-lepidote petiolar glands.Theydiffer from sect. inflorescences withbasalbisexualcymules,bifidstyles,and also share somecharacterspresent insect. Atlântica forests ofsoutheasternBrazil( casanus northeastern Argentina andsoutheasternBolivia, our samplingby phylogenetic hypothesis. this speciesare neededtotestthismorphologicallybased sect. that itismore closelyrelated tothe this species,butanexaminationofitsmorphologysuggests sequence datafrom thefewherbaiumspecimensknownof C. megistocarpus ber of 371 RIINA ET AL.: DRAGON’SBLOODCROTON foothills ofthe Andes ( less latexandare widespread inthe Amazon basinuptothe phyletic clade( Fig. 2 ). Theyare alllarge trees withcolor- recovered thesethree speciesasastrongly supportedmono- subsect. and 2009] tral American-centered group ofabout13specieswithonly Standl. from Mexico.Section with Croton et al.(2008 ) usedabroader samplingofthebasallineages leaf margins, absenceof red latex,andbifidstyles. van Ee stellate-lepidote trichomes,pinnatelyveinedleaves,dentate with dentate-glandularmargins, epipetiolarglands,similar members ofthe ers andbilocularcapsules.However, itresembles thethree nearly uniquewithin Scin Corylocroton— Section andplaceditinsect. C. sampatik Cyclostigma C. pachypodus needsadditionalmolecularworkbecausethere are andrecovered thespeciesof Croton Pittierfrom northernSouth America, and group isrecovered asagradeleadinguptosect. Cyclostigma J.Murillo-A.,andanundescribedspeciesfrom , thetopologyfavored by van Eeetal.(2008 ), G.L.Webster. Inthe sect. Corylocroton R.deS.Secco,P. E.Berry&N. A. Rosa, , andonemember( Croton J. A. González&Poveda,and Sampatik . Molecularphylogeneticanalysesincluding C. beetlei Quadrilobus (includedhere ascladeplaceholder), ( Table 1 ). Ourphylogeneticresults clade(CladesC-2 toC-11 in Berry etal. Croton C. sampatik cladeinthealmostglabrous leaves Croton . However, thisparticulargroup of Cyclostigma Croizat from theChacoregion of Croton sapiifolius This section isrepresented in inhavingtetramerous flow- Corylocroton sensu Webster (1993 ), andis subgenus trnL–F C. ater , , Sampatik C. ater C. sapiifolius s.l.We couldnotget Sampatik Cyclostigma, analyses( Fig. 1 ), the Sampatik Sampatik ) ofsect. C. piptocalyx ), andintheMata isamostlyCen- istheonlymem- Moacroton clademightnot Cyclostigma Croton cladethanto C. trombeten- cladealong Cyclostigma group (e.g. Cyclostigma C. olivaceus Ml. Arg. Müll. C. lundellii . Thepeti- however, C. cara- (Clade ) They ). : long : C. ,

partitions are incongruent). acters were removed. (*A significant combined datasetofITSand Cascarilla 0 (fulldataset) ugop0.007 Cyclostigma Cyclostigma C.cupreatus Cascarilla Outgroup this species. is underwaytoobtainadditionalplastidsequencedatafrom tid datasetsdidnotinclude genetic pattern,asexpected.Unfortunately, theseotherplas- 2008 ), and Sanderson andDonoghue1989 ). character statejustbychance( Givnish andSytsma1997a , b there ismore chancethattwoormore taxawillshare aderived ; homoplasy increases asthe numberoftaxaincreases because gene transfer( Planet 2006 ). There isalsostrong evidencethat evolution orhomoplasyonatree, inadditiontohorizontal that significantincongruence canbecausedbyconvergent data, andthat are themaingroups responsible fortheincongruence inthe et al.2008 ). TheGuatemalan evidence asamonophyleticgroup (see van Ee2006 ; van Ee Section C.beetlei the wellsupportedsect. sis bisexual cymules.Ourresults clearlyshowthat stellate trichomes),thelackofstipules,andabsence presence oflepidotetrichomes(sometimesalongwithsparse rated from sect. placement. Speciesfrom sect. s. l.by Donnell Smith(1912) , and Webster (1993 ) followedthis known from thetypespecimen,wasplacedinsect. ( Fig. 2 ), whereas the supports sect. Cyclostigma trnL–F Gene Histories— ILD testwiththepresent data setandthe One possibleexplanationforthedifferences inbehaviorofthe these twocladesintheirseparate and theyrecovered thesameconflictingtopologiesbetween s. (sixspecies)and results showedthatsect. this ideabyconductingaseriesofILDtests( Table 2 ), andthe of taxaincludedfrom agiven conflictingclade.We explored Berry etal.(2005 ) isthatthetestsensitivetonumber included severalrepresentatives ofbothsections ( passed theILDtestonlyafterremoving twoconflictingtaxa et al.(2005 ), usingacombineddatasetofITS and ter clade( Fig. 1 ). C. setiger Table Tre te clrpat ein regions chloroplast other Three The SisterCladeofSectionCyclostigmaandConflicting The first doesnotbelongtosect. Taxa excluded trees are discordant regarding thesistercladeofsect. clade clade+ Corylocroton 2 reaching farsouthintowesternSouth America. . . Hook.and ss 0.950 s.s.+ s.s. 0.018 matK anditsphylogeneticpositionwithin Incongruence lengthdifference tests(ILD)applied tothe Croton C. cupreatus Cascarilla C. cupreatus C. cupreatus ( Riina 2006 ) showthesameconflicting phylo- Cyclostigma Our results showthattheITSandchloroplast molecularphylogenypublishedby Berry Cascarilla isverywellsupportedbythemolecular C. insularis trnL–F asthesistergroup ofsect. 0.277 0.881 Corylocroton trnL-F Cascarilla onlyplaysaminorrole. Itisknown Cyclostigma dataplace (13species),butnot bytheabsence ofred latex,the Croton verapazensis Corylocroton . All uninformative andgappedchar- C. cupreatus Baill.).Nonetheless,thatwork p valueindicatesthatthetwodata andthe trnL–F rbcL, ndhF . , andisinsteadpartof p C. cupreatus value(significantat0.05level)* Croton canbeeasilysepa- , butcurrent work andITSanalyses. Cyclostigma ( van Eeetal. , whichisonly 0.269 0.007 datasetfrom Cyclostigma C. verapazen- C. cupreatus Cyclostigma Cyclostigma Croton. asthesis- trnL–F clade ITS s. , ,

Delivered by Ingenta to IP: 5.10.31.151 on: Tue, 07 Aug 2018 10:39:17 Copyright (c) American Society for Plant Taxonomists. All rights reserved. ent insect. three charactersabsentin members ofsect. and hasbisexualcymuleswithpedicellatepistillateflowers, [Volumefind that 34 cally, itishard totellwhichoneiscloser two alternativephylogeniesandcompare themmorphologi- focus onthe detailed morphologicalsurveyacross thegenus.Evenifwe is more concordant withmorphologysinceitwillrequire a work withthisgroup. ogy, ecology, andgeographyshouldbeconsidered infuture orate thispattern,andinformationfrom morphology, karyol- will needtoexplore multipleindependentmarkerstocorrob- ter understandtheevolutionaryhistoryof caused byanyofthefactorsmentionedabove.Inorder tobet- event. Butthephylogeneticpatternshownhere couldalsobe SYSTEMATICevolution andthatitoriginatedbyanancienthybridization BOTANY netic incongruence ( Soltis andKuzoff 1995 ). tion) havealsobeensuggestedascausalfactorsforphyloge- and heterogeneous evolutionaryrates(longbranchattrac- et al.2005 ; Meerow etal.2006 ). Taxon density, samplingerror, 1995 ; Comes and Abbott 2001 ization andplastidcapture; ( Sytsma 1990 Doyle etal.2003 ; ; Soltis andKuzoff Okuyama ary convergence orhomoplasy, andancientorrecent hybrid- polymorphisms inchloroplast andnucleargenes,evolution- uted toeventssuchasdifferential lineagesortingof ancestral Bakker 2005 ). Thesediscrepancies amonggenesare attrib- angiosperms (see Rieseberg andSoltis1991 ; Vriesendorp and inherited) isacommonpatterninphylogeneticstudies (uniparentally inherited)andnucleargenes(biparentally 372 here showthatthesisterposition ofsect. ( overlap instamennumber, whereas tive andreproductive organs. However, cupreatus because bothofthemhavestellatepubescence,whereas sequences isnecessarytodetermineif of thephylogeny, andnewtaxonsampling usingchloroplast tion from othermarkersisneededtobetterresolve thispart might changewhenothertaxaare included.More informa- so theassociationof ity ofthebackbonenodeshaveverylowbootstrapsupport, tion, with in bothphylogenies,butitispossiblethattherelationship either cies (e.g. Finally, giventhemedicinal usesofthedragon’sblood spe- on thequestionofitsphylogenetic placementwithin Cyclostigma ers isfocusingonthephylogenetic relationships within sect. et al.2007 ). Ongoingworkwithadditionalmolecularmark- Webster’s (1993)classification,includingnewspecies ( Riina new membersofsect. the group. Thissuiteofcharactersisguidingusinfinding identify asuiteofmorphologicalcharactersthatcharacterize and gene history. as theimmediatesisterofsect. < 10). Ontheotherhand,molecularcharactersevaluated It isdifficult todecidewhichgenephylogenyinthiscase It ispossiblethat Conflict amongphylogenetictrees basedonplastidgenes In conclusion,thephylogeneticpatternresulting from ITS trnL–F C. cupreatus C. cupreatus C. cupreatus hasametallic lepidoteindumentumbothonvegeta- C. lechleri Cascarilla Cyclostigma allowustorecircumscribe sect. s.s.,includingthesenewmembers, aswell Cyclostigma is anartifactofthetaxonsampling.Inaddi- isinaregion ofthetree where themajor- ( seemsmore similarto ), ourfindingsare relevant tofuture phar- trnL–F Cyclostigma C. cupreatus Cyclostigma s.Sections cladeanditssistercladesfrom the ) or Cascarilla s.istheproduct ofreticulate Cyclostigma s.previously unplacedin withthe Cascarilla (ITS)iswellsupported C. cupreatus C. cupreatus Cyclostigma C. cupreatus Cyclostigma Cyclostigma Cascarilla inthechloroplast Cyclostigma and Cyclostigma Cyclostigma Cyclostigma doesnot andpres- s.with remains isatree mainly s.we Croton clade and . We . C. .

Lrni H 19 1992. H. Lorenzi, ( Drago de Sangre of Review 2003. of K. inference Jones, Bayesian MrBayes: 2001. Ronquist. F. and P. J. Huelsenbeck, Gieah A H R 15 1859. R. H. A. Grisebach, like- the and characters, Consistency, 1997b. Sytsma. J. K. and J. T. Givnish, mor- vs. molecular in Homoplasy 1997a. Sytsma. J. K. and J. T. Givnish, Dye J J , . . ol , . . ashr ad . . . rw 03. Diploid 2003. Brown . D. H. A. and Rauscher , T. J. Doyle, other L. and J. J., Guatemala J. from Doyle, plants Undescribed 1912. J. Smith, Donnell Fri, . . M Kleso A G Kue ad . ut . 19 etn signifi- Testing 1994. Bult. C. and Kluge, G. A. Kallersjo, M. S., J. Farris, reticu- phylogeography, Molecular 2001. Abbott. J. R. and P. H. Comes, Poaceae of Phylogeny 1997. Olmstead. G. R. and Kellogg, A. E. P. , Catalán, áreas em Glomales 1997. Barbosa. M. L. and Bononi, R. L. V. R., Carrenho, Bre, . . n S R Kn 00. 2000. King. R. S. and R. J. Borges, Meo, . . J FacsoOtg , . . ut , n R J Shel . 2006. Schnell . J. R. and Kunth, N. D. Francisco-Ortega, J. W. , A. Meerow, Bry P E , . . ip K J Wrak B W vn e ad . in 2005. Riina. R. and Ee, van W. B. Wurdack , J. K. Hipp, L. A. E., P. Berry, 1858. H. Baillon, SP, SPF, USM,USZ,andVEN). GUAY, HOXA,IAN,INPA, LOJA,LPB,MG,MYF, PEUFR,QAP, QCNE, herbaria whohelpedherduringfieldworkinSouth America (AMAZ, Documentation. Theseniorauthorisgratefultothestaff ofthefollowing (Grant No.DEB-0212481toPEB),andtheHuntInstituteforBotanical the UniversityofWisconsin-Madison, theNationalScienceFoundation University ofWisconsin-Madison (LACIS),theDepartmentofBotany (ASPT), theLatin American, Caribbean,andIberianStudiesatthe Plant Taxonomists (IAPT),the American SocietyofPlantTaxonomists support forthisstudywasprovided bytheInternational Association of two anonymousreviewers forusefulcommentsandcorrections. Financial Madison. We thankDavidBaum,KennethSytsma, Alan W. Meerow, and dissertation intheDepartmentofBotanyUniversityWisconsin- Myad mt, . 92. Aayig h msi srcue f ee genes. of structure mosaic the Analyzing 1992. J. Smith, Maynard amongst recombination of detection RDP: 2000. Rybicki . E. and 2001. D. Martin, Maddison. P. W. and R. D. Maddison, guide taxonsampling. macological studiesondragon’sbloodspeciesasatoolto Acknowledgments. Plantarum . plantas arbóreas nativas do Brasil N.Y.) research . bites, viralinfections,andwounds:traditionalusestoclinical American tree sapinthe treatment ofdiarrhea, inflamation,insect phylogeny . ins n K J Stm e Yr abig Uiest Press . University Cambridge York : New Sytsma. J. K. and Givnish Islands Evolution lihood ofcorrect phylogeneticinference . in 55–101 Pp. phological data:thelikelihoodofcorrect phylogeneticinference . nial soybeans( and polyploidreticulate evolutionthrough thehistory oftheperen- Central American Republics.XXXV . (Asteraceae) cance onincongruence . 121–132. lation, andlineagesortinginMediterranean MolecularPhylogeneticsandEvolution subfamily Pooideaebasedonchloroplast 107–113 . 24: de recomposi 24–26. of an Amazonian pioneerspecies . 42–60. sequences: Lineage sortinginareticulate area? clade of Amaryllidaceae basedon plastid Phylogenetic relationships andbiogeographywithintheEurasian aa. data . (Euphorbiaceae sensustricto)usingITSand Molecular phylogeneticsofthegiantgenus Masson . of MolecularEvolution aligned sequences . Phylogeny andCharacterEvolution 877–896. 9: American JournalofBotany . Lno . Reeve. L. London: . 320–330. 7: Journal ofAlternativeandComplementaryMedicine(NewYork, Bioinformatics (Oxford,England) . Evolution Étude généraledugroupe desEuphorbiacées ç Árvores brasileiras:manual deidentifica ão demataciliarMoji-Gua Molecular evolutionandadaptiveradiation Glycine Bioinformatics (Oxford,England) Croton Ph.D. This paperispartoftheseniorauthor’s 4: 126–129. 34: 5: 1943–1962. 55: ieaue Cited Literature subgenus Cladistics P. 3 2 i in 37–42 Pp. . Croton lechleri 2: 1520–1534. 92: . Sneln iae Associates. Sinauer Sunderland: . 0: 315–319. 10: Glycine , vl 1. Nv Oes Editora Odessa: Nova 1. vol. , 150–166. 8: Botanical Gazette Medicinal PlantConservation Flora oftheBritishWest Indian ). ). Molecular Phylogeneticsand MacClade 4.03:Analysisof Croton 7: 754–755. 17: Croton lechleri The NewPhytologist trnL-trnF , sustainableutilization ndhF ndhF ç u, SP, Brasil . Systematic Botany Senecio andtribeCrotoneae 6: 562–563. 16: andnrDNA ITS gn sequences. gene 4: 235–244. 54: ç DNA sequence ão ecultivode . Prs: Victor Paris: . sect. ) –A South , d. T J. T. eds. , Hoehnea Senecio Journal 161: 31: 6 :

Delivered by Ingenta to IP: 5.10.31.151 on: Tue, 07 Aug 2018 10:39:17 Copyright (c) American Society for Plant Taxonomists. All rights reserved. Sco R S , . . er , n N A Rs 01. 2001. Rosa. A. N. and Berry , E. P. S., R. Secco, Sih B A 20 2002. A. B. Smith, Snesn M J ad . . oohe . 18 atrs f aito i levels in variation of Patterns 1989. Donoghue. J. M. and J. M. Sanderson, Tbre, . L Gel , . atu ad . ovt . 19 nvra pies for primers Universal 1991. Bouvet. J. and Pautou, G. Gielly , L. P. Taberlet, , Package. Analysis Sequence Staden The 1996. R. Staden, chlo- and nuclear between Discordance 1995. Kuzoff . K. R. and E. D. Soltis, Uils R , . . oa , . . reig M R Kra , . . ig D F. D. King, R. S. Kernan, R. M. Bruening , C. R. Jolad, D. S. R., Ubillas, endonu- restriction from inference Phylogenetic 1983. R. A. Templeton, Ria R , . onj , n P E Bry . 20 nw pce o “age de “sangre of species new A 2007. Berry . E. P. and Cornejo , X. R., Riina, 2006. R. Riina, Stm, . . 90. DA n mrhlg: neec o pat phylogeny . plant of inference morphology: and DNA 1990 . J. K. Sytsma, (*and Parsimony Using Analysis Phylogenetic PAUP* 2002. L. D. Swofford, vn e B W , . . er , . in , n J E Gtérz mr 2008. Amaro . Gutiérrez E. J. and Riina, R. Berry , E. P. W. , B. Ee, van 2006 . W. B. Ee, van 09 RIAE L:DAO’ LO RTN 373 emergence Possible 1999. Fauquet. M. C. and Sawyer , S. M., Padidam, M. Ito, RIINA ET AL.: DRAGON’SBLOODCROTON M. Kawakita, A. Wakabayashi , M. Fujii, N. the Y. by , distributed Okuyama, Program v2. MrModeltest 2004. A. A. J. Nylander, de especies las de peruanos aborígenes Nombres 1999. N. E. Meza, 2009] Reeeg L H ad . . ots . 19 hlgntc osqecs of consequences Phylogenetic 1991. Soltis. E. D. and Oxford : H. v2.0a11 . L. Editor Rieseberg, Alignment Sequence 1996–2002. A. Rambaut, averaging model and selection Model 2004. Buckley . R. T. and D. Posada, incongruence testing and measuring disagreement: Tree 2006. J. P. Planet, Ubtc, . . B G Blwn ad . . oohe . 20 hlgn o the of Phylogeny 2000. Donoghue . J. M. and Baldwin, G. B. E., L. Urbatsch, Psd, . n K A Cadl 98. MDLET tsig h mdl of model the testing MODELTEST: 1998. Crandall. A. K. and D. Posada, 119 –123. betensis cao Ecuador . of homoplasy . California . amplification ofthree non-coding regions ofchloroplast DNA . 727–742. 49: roplast phylogeniesinthe (Euphorbiaceae) inEcuador Molecular Biology humans andtheapes . clease cleavagesitemapswithparticularreference totheevolutionof 65–84. drago” ( Wisconsin. of University Croton sect.Cyclostigma(Euphorbiaceae) Biotechnology Trends inEcology&Evolution Associates. Sinauer Sunderland: 4.0b10. version methods), other centered Molecular phylogeneticsandbiogeographyoftheCaribbean- s. s.) Review 285–296. of newgeminiviruses byfrequent recombination . Asian gression patternsinthree moleculardatapartitionphylogeniesof concerted Nonuniform 2005. Kato. evolution andchloroplast capture: M. heterogeneity ofobservedintro- and Murakami, N. Watanabe , University . author . Uppsala Universidad NacionalMayordeSanMarcos: . Department ofSystematic Zoology, Uppsala de suproducción enelPerú in que producen ellátexdenominado“Sangre deGrado” . Pp. 25 – 44 218–225. cytoplasmic geneflowinplants . Oxford . of University Bayesian approaches overlikelihoodratiotests . in phylogenetics:advantagesof Akaike InformationCriterionand in phylogenies . . ufa , . me R Sdel K Sie A Bair S Sfi , R. Safrin, SP-303, S. 1994. lechleri Brazier Nakanishi. , K. A. and Soike, Berova , an antiviraloligomericproanthocyanidin from thelatexof K. N. Kenny , Sidwell, Barnard , M. T. D. R. P. Tempesta , Orlando, Smee, Meza, S. E. D. M. Ayala , Rozhon, Huffman F. , Kuo , E. J. J. McMeekin, Flaster , D. T. Castañel, Stoddart, M. A. C. Barret , M. Sesin, DNA restriction sitedata . rDNA internal transcribedspacer(ITS)sequencesandchloroplast coneflowers andrelatives (Heliantheae: Asteraceae) basedonnuclear 793–808. DNA substitution . Desarrollando nuestradiversidadcultural:“Sangre deGrado”yelreto . Ph.D.Thesis. Madison : University ofWisconsin . (sangre dedrago) . 4: 132–165. 74: Mitella , twonewEuphorbiaceaefrom Amazonian Brazil . Croton Brittonia Croton Molecular systematicsoftheneotropical dragon’s bloodtrees 233–241. 5: (airgaee (Saxifragraceae). Evolution section subgenus Journal ofBiomedicalInformatics A systematicrevision ofCroton sectionCyclostigma Molecular phylogeneticswithinCroton (Euphorbiaceae 7: 1105 –1109 . 17: 9: 97–101. 59: Bioinformatics (Oxford,England) Evolution 3: 1781–1795. 43: Cyclostigma Phytomedicine Systematic Botany , d . . ea, Lm od Editorial, Fondo Lima: Meza, N. E. ed. , Heuchera Moacroton . Ph.D.Thesis. Davis : University of 104–110 . 5: 7: 221–244. 37: Molecular BiologyandEvolution group (Saxifragaceae) . Evolutionary Trends inPlants , Euphorbiaceae)from coastal (Euphorbiaceae s.s.) . 77–106. 1: Croton diasii . Ph.D.Thesis. Madison : 5: 539–565. 25: 9: 86–102. 39: Systematic Biology 4: 817–818. 14: and Virology Croton trom- Novon Molecular Evolution Botanical Croton Croton 265: Plant 22: : : 1 1 53: 5:

Kunth; Ecuador, Tunguragua; Espirito Santo; EU586945. accession numbers( lector numberandherbariumwhere specimenisdeposited),andGenBank 2006. J. D. Zwickl, Müll. Arg.; ; gracilipes goudotii C.gossypiifolius C.glandulosus C.elegans Visnop B ad . . akr . 20 eosrcig atrs of patterns Reconstructing 2005. Bakker . T. F. and B. Vriesendorp, EU478029. EU586904. C.aequatoris lodendron Arg.; Brazil; Rio deJaneiro; EU586955; EU586902. Riina1449 Warming 1653 coideus Müll. Arg.; Brazil,RiodeJaneiro; glandulosepalus cosus erythroxyloides emporiorum Wbtr G L 19 poiinl yoss f h scin o te genus the of of Synopsis sections 2001. L. the G. Webster, of synopsis provisional A 1993. L. G. Webster, Wie T J , . rys S Le ad . alr . 19 mlfcto and Amplification 1990. Taylor . J. and Lee, S. Bruyns , T. J., T. White, Weehet A C , . in , n P E Bry . 20 Ryituie latic- “Ray-intrusive” 2009. Berry . E. P. and Riina, R. C., A. Wiedenhoeft, Paulo; Esser20011 99386 (WIS); EF408090;EU586916. (TEX); —;EU586952. Yucatán; (WIS); AY971305; AY971217. EF408111; EF421773. EU478070. EU478127; EU477889. EU586982; EU586925. hibiscifolius Brazil, MinasGerais; Saül; (WIS); EU586977;EU586954. EU497711; EU497736. EU586960; EU586906. Quezada313 DQ227557; DQ227525. EU586968; EU586913. EU586974; EU586919. EU497735. Esmeraldas; C.cuneatus ceus Baill.; Brazil,Bahia; Kunth; Brazil; Astraealobata Appendix Kunth;Ecuador, Pichincha; Ph.D. Thesis. Austin : The UniversityofTexas . large biologicalsequencedatasetsunderthemaximumlikelihoodcriterion reticulate evolutioninangiosperms:whatcanwedo? o-riuae non-articulated . Herbarium western tropical Mexico . Croton 593–604. p 1 2 n in 315–324 Pp. direct sequencingoffungalribosomalRNA genesforphylogenetics . M A Ins, D H Glad J J Sisy ad . . ht a Diego : San White. J. T. and Sninsky , J. J. Gelfand, Press . H. Academic D. Innis, A. M. ifers inspeciesof B.A.Smith;Ecuador; (MU);EU497707;EU477932. Phillippe 27019 Caruzo 32 Müll. Arg.; ; Baill.;Madagascar; Duran 3367 Baill.;Bolivia; Müll. Arg.; , Miranda; Kunth;Ecuador; (WIS);EU586979;EU586927. (uhrica). (Euphorbiaceae). (M); AY971282; AY971191. Brasiliocroton mamoninha C.cupreatus Kl.;Venezuela; C.arboreus C.hoffmannii C.astroites Croizat; Bolivia; (M);EU586959;EU586905. KunthexSpreng.; Ecuador, Imbabura; 1 Riina 1415 . Croizat; Ecuador, Guayas; (F);—;EU586926. Carvalho 3789 L.;USA,Wisconsin; 3: 353–388. 23: Pirani 4947 Lucena 598 Baill.;Brazil; Riina 1522 (SP);EU586920;EU586975. Millsp.;Belize; Species sampled,voucherinformation(origin;collector, col- Vahl; Venezuela; Genetic algorithmapproaches forthephylogeneticanalysisof (L.)Kl.;Brazil,Bahia; trnL-F PCR protocols: aguidetomethodsandapplications ; EF408101;EF421735. Thomas 13601 (NY);EU586978;—. C.huberi IAWA Journal C.caracasanus Riina 1371 C.beetlei C.draco C.hirtus Croton C.discolor C.curiosus C.ater C.cascarilla C.hircinus Arias 211 Millsp.;Mexico; (WIS); EU586995; EU586941. Croizat; Ecuador, Pichincha; Dryand.;PuertoRico; ; ITS,respectively; —=sequencemissing). Nee 47412 (SPF);EU586998;EU586944. (WIS);EU586984; EU586928. Ml. Arg. Müll. (WIS);EU586963;EU477957. Berry 7589 Ceron 6909 (CEPEC);EU586988;EU586933. C.lagoensis Wood 14086 section Caruzo 74 Taxon Hoffmann 248 Berry 7675 C.cajucara C.draconoides Wood 15844 Contributions From theUniversityofMichigan Croizat; Colombia,Boyacá; Steyerm.; Venezuela, Miranda; Schltdl.&Cham.;CostaRica; L’Her.; Brazil; Croton Croizat; Bolivia,SantaCruz; Riina 1391 Vincent 6058 (WIS);EU586979;EU586922. (NY);—;EU586929. C.lechleri Riina 1403 Willd.; PuertoRico; Riina 1525 Croizat; Argentina; Mota sn (NY);EU586971;EU586917. 0: 135–148. 30: 2: 793–823. 42: Vent.; Venezuela; van Ee512 (NY);EU586962;EU586909. C.cascarilloides (L.)L. Pittier;Venezuela; Cyclostigma (WIS), and (SP);EU586992;EU586938. (QCNE);EU586980;EU586923. ; P.E. Berry& CostaRica; C.celtidifolius (WIS); AY971296; AY971207. (LPB); EU586956;EU586902. Müll. Arg.; Brazil,MinasGerais; C.chocoanus (LPB);EU586961;EU586908. Benth.;Brazil; (WIS);EU586981;EU586924. (WIS);EU586957;EU586903. van Ee472 Phyllanthus (K);EU587000;EU586946. C.macrobothrys van Ee486 C.lundellii ; Müll. Arg.; Peru, SanMartin; Riina 1431 (WIS);EU586976;EU586921. Bahamas, CatIsland; C.chilensis (WIS);EU587001;EU586949. C.echinocarpus Riina 1290 (MU);EU478126;EU477888. (WIS);EU497713;EU478066. Müll. Arg.; French Guiana, Riina 1491 Lima 345 (Euphorbiaceae)maybe van Ee537 Raeusch;Thailand; van Ee598 (Euphorbiaceae)in I. Cordeir (WIS);EU497701; (WIS);EU586956; (WIS);EU586999; Croton abutiloides Cozt Ecuador, Croizat; (WIS);EU586996; Baill.;Brazil,Sao Riina 1291 Riina 1413 C.medusae Standl.;Mexico, Zuloaga 8438 (SPF);EU478160; van Ee547 Riina 1408 Müll. Arg.; Chile; (I) AY794698; (WIS); C.conduplicatus Riina 1288 C.magdalenesis Caruzo 95 C.cordiifolius Baill.;Brazil,

Lawrance 593 Ml. Arg.; Müll. van Ee344 Riina 1512 Berry 7590 C.eichleri C.mega- Taxon o; Brazil, C.coria- (WIS); C.heli- (WIS); (WIS); (VEN); (WIS); (WIS); C.floc- (WIS); Müll. Richey (SP); , eds. , (SI); 54: C. C. C. C. C. C. .

Delivered by Ingenta to IP: 5.10.31.151 on: Tue, 07 Aug 2018 10:39:17 Copyright (c) American Society for Plant Taxonomists. All rights reserved. 7 SSEAI OAY [Volume 34 SYSTEMATIC BOTANY EU587004; EU586953. EU586942. 374 Brazil, SaoPaulo; EU586993; EU586939. (WIS); EU586997;EU586943. (WIS); EU586969;EU586914. (K); —;EU586948. EU478155; EU478046. EU586985; EU586930. (MO); —;EU586947. Paz; (WIS); EU586986;EU586931. bachii C.redolens C.purdiaei gens Croizat; Brazil,SaoPaulo; C.roxburghii C.ruizianus Jacq.;Venezuela, Aragua; Riina 1500 Croizat; Ecuador, Loja; Müll. Arg.; Colombia;Becerra20(NY);EU586989;EU586934. C.pachypodus Pittier;Venezuela; N.P. Balakr.; Thailand; Müll. Arg.; Peru, Cusco; (WIS);EU586987;EU586932. C.organensis Riina 1533 C.mutisianus C.oreades C.perspeciosus C.mongue C.niveus G.L.Webster; CostaRica; Riina 1535 C.palanostigma C.piluliferus C.pallidulus (WIS);EF408148;EF421790. Gonzalez 4 Riina 1422 Riina 1272 Leandri;Madagascar; Baill.;Brazil,RiodeJaneiro; Jacq.;CostaRica; Baill.;Madagascar; Kunth;Ecuador; Esser 9911 (WIS);EU587002;EU586950. Croizat; Peru, Pasco; (VEN);—;EU586935. Riina 1487 (WIS);EU586990;EU586936. BrittonexRusby;Bolivia,La Baill.;Brazil; (WIS); AY971326; AY971241. (WIS); AY971329; AY971244. Kl.;Peru, Loreto; C.piptocalyx Ceron 6188 Valverde 1043 (WIS);EU586964; van Ee284 Rakotomalaza 462 Randrianairo 279 Caruzo 31 Ml. Arg.; Müll. Riina 1435 C.priscus Riina 1492 Caruzo 90 (QCNE); (WIS); C.rim- C.pun- (MO); (SP);

C.yavitensis C.xalapensis AY971260. gratissimus Müll. Arg.; Brazil,SaoPaulo; Botanical Garden; Guatemala; Gerais; Belgrano423 EF421792. 1658 EU478151; AY971251. (MO); AY971331; AY971246. (SI); EU586912;EU586966. EF408150; EF421754. EU587003; EU586951. 2263 EU586910. EU586911. (IEB);EU478144;EU477979. (MO);—;EU478101. Riina 1317 C.sampatik C.sapiifolius C.saltensis Burch.; Zambia:SongweGorge; Turkheim 2297 (SI);EU586994;EU586940. Croizat; Bolivia; Kunth;Mexico,Veracruz; (WIS);EU586991; EU586937. Berry 7683 C.sarcopetalus C.speciosus C.suberosus Müll. Arg.; Peru, Pasco; Griseb. Ml. Arg. Müll. (GH);—;EF421750. C.schiedeanus C.sp.nova; C.smithianus (WIS); AY971338; AY971257. Beck 5710 ; Agnia Argentina; Caruzo 56 Müll. Arg.; Venezuela; Kunth;Mexico,Michoacan; C.troncosoi Müll. Arg.; Argentina; ; Brazil,Bahia; Brazil,Bahia; C.urucurana (LPB);EU586973;EU586918. Schltdl.;CostaRica; Nee 29704 (WIS);EU586970;EU586915. Croizat Zuloaga 8478 Zimba 901 Riina 1447 C.verapazensis C.verreauxii Ahumada;Argentina; (NY);—;EU477987. ; CostaRica; Baill.;Brazil,Minas Lima 667 Lima 654 (O; AY971341; (MO); (WIS);EF408133; Berry 7590 (SI);EU586965; Baill.;Sidney C.warmingii Zuloaga 8600 Aguilar 886 Donn.Sm. (CEPEC); (CEPEC); Steinmann Aguilar (WIS); C. ;