River dynamics shape clonal diversity and genetic structure of an Amazonian understorey herb

14 Pages·2011·0.46 MB·English

Journalof Ecology2011, 99, 373–382 doi: 10.1111/j.1365-2745.2010.01775.x
River dynamics shape clonal diversity and genetic
structure of an Amazonian understorey herb
Matthias Schleuning1*, Thomas Becker1,2, Giovana P. Vadillo3,4, Thomas Hahn4,5,
Diethart Matthies1 and Walter Durka4
1Plant Ecology,Departmentof Biology, Universityof Marburg,Karl-von-Frisch Str. 8,35043Marburg, Germany;
2Departmentof Ecology andEcosystemResearch, Albrecht von HallerInstitute forPlant Sciences,Universityof
Goettingen,UntereKarspuele 2,37073 Goettingen,Germany;3Facultyof BiologicalSciences, NationalUniversityof
SanMarcos(UNMSM), Venezuela Av.34, Lima 1,Peru; 4DepartmentofCommunity Ecology (BZF), UFZ-Helmholtz
CentreforEnvironmental Research, Theodor-Lieser Str. 4,06120 Halle,Saale,Germany; and5ITESEcosystem
Management,ETH Zurich,Universitaetsstr.16, 8092Zurich,Switzerland
Summary
1. ClonalherbsareanimportantfeatureoftheunderstoreyofAmazonianforests.However,little
isknownabouttheenvironmentaldriversdeterminingthepopulationgeneticsoftheseherbsand
aboutthespatialscaleofgeneflow.
2. With amplified fragment length polymorphism markers, we analysed the clonal diversity and
genetic structure of a clonal Amazonian herb (Heliconia metallica) in south-eastern Peru at two
spatial scales. First, we sampled leaves in 24 patches differing in flooding frequency and canopy
opennessin2 km2ofafloodplainforest,andsecondin21riverinepopulationsfromtheAndean
foothillstotheAmazonlowlandsalonga550-kmstretchofriver.
3. Atthesmallspatialscaleinthefloodplainforest,theclonaldiversityofH.metallicawashigher
atfloodedthanatnon-floodedsites,butclonaldiversitydidnotincreasewithcanopyopenness.
4. Atthelargespatialscale,clonaldiversitywasverylowinriverinepopulationsatup-anddown-
riversites,suggestingthatseedlingrecruitmentwashigheratmid-altitudeswherethefloodinginten-
sity isintermediate.Geneticdiversityofriverinepopulationsmonotonouslyincreased downriver,
indicatingunidirectionalgeneflowmediatedbyhydrochory.
5. Geneticdifferentiationamongriverinepopulationswasverylow(F = 0.06)andfollowedan
ST
isolation-by-distance pattern, indicating a stepping-stone type of gene flow by seeds. Despite the
muchsmallerspatialscale,geneticdifferentiationamongpatchesinthefloodplainforestwashigher
(F = 0.16),duetospatiallyrestrictedgeneflowintheforestunderstorey.
ST
6. Synthesis.ThegeneticstructureofH.metallicaistheresultofseedlingrecruitmentbeinglargely
limited toflooded sites and of hydrochoricseed dispersal between populations growing on river-
banks.WeconcludethatriverdynamicsarethemajordeterminantofthegeneticstructureofAma-
zonian plants and that largely undisturbed river systems, such as the Amazon, provide a crucial
vectorforgeneflow,evenatlargespatialscales.
Key-words: AFLP markers, ecological genetics and ecogenomics, flooding regime, Heliconi-
aceae, hydrochory, Peru, population genetics, seed dispersal, tropical rain forest, unidirec-
tional gene flow
(Rusterholz,Kissling&Baur2009;Vandepitteetal.2010).In
Introduction
Amazonianforests,naturaldisturbancessuchastreefallsand
Environmental disturbance is a crucial factor shaping flooding strongly affect the demography of bothtrees (King
the demography and genetic structure of plant populations 2003;Stevenson2007)andunderstoreyplants(Svenning2002;
Schleuning, Huama´n & Matthies 2008). During flooding
events,watercandisperseseedsofAmazonianplantsoverlong
*Correspondence author. Biodiversity and Climate Research
Centre (BiK-F), Senckenberganlage 25, 60325 Frankfurt (Main), distances(Moegenburg2002),andthuspotentiallyshapesthe
Germany.E-mail:[email protected] genetic structure of Amazonian plants. However, we still do
(cid:2)2011TheAuthors.JournalofEcology(cid:2)2011BritishEcologicalSociety
374 M.Schleuninget al.
not understand how these important environmental factors flowcanlinkplantpopulationsoverlongdistancesduetodis-
determinethegeneticstructureofplantpopulationsintropical persal of seeds by water (Kudoh & Whigham 2001; Fe´r &
floodplainforests. Hroudova´ 2008;Kondo,Nakagoshi&Isagi2009).Riparian
Flooding regimes are important for many Amazonian plantscanexhibitastepping-stonetypeofgeneflowbetween
plants:about12%ofAmazonianforestsarecurrentlyaffected neighbouringpopulations(seeTeroetal.2003)thatcanlead
byriverdynamicsandabout25%havebeeninfluencedinthe topatternsofisolationbydistanceatdrift–dispersalequilib-
recentpast(Saloetal.1986).Intheinteriorofthefloodplain rium(Hutchison&Templeton1999)andtolowgeneticdiver-
forests,riverdynamicshavecreatedamosaicofmicrohabitats gencealongtheriver(Huetal.2010).Becauseseeddispersal
atdifferentelevationsabovetheriver(King2003;Schleuning, bywaterisunidirectional,thegeneticdiversityinpopulations
Huama´n&Matthies2008).Differencesinfloodingfrequencies ofriparianplants isexpectedtoincreasedownriver(Barrett,
among microhabitats strongly affect patterns of seedling Eckert&Husband1993).Incontrasttogeneflowalongrivers,
recruitmentintheforestunderstorey(King2003;Schleuning, gene flow in the temperate forest understorey is usually low
Huama´n&Matthies2009).Asecondimportantprocessthat anddivergenceamongpopulationshigh,whichismostlydue
influencesdemographicprocessesintheforestunderstoreyis tospatiallyrestrictedseeddispersal(Augeetal.2001;Jacque-
the formation of canopy gaps after treefalls (Svenning 2002; mynetal.2006a;Schmidtetal.2009).Otherthantemperate
Stevenson2007).Increasedsolarradiationontheforestfloor forestherbs,mostunderstoreyplantsfromtropicalforestsare
results in a higher probability of flowering (Svenning 2002; bird-dispersed,buttropicalunderstoreybirdshavesmallhome
Schleuning,Huama´n&Matthies2008)andofseedlingrecruit- rangesandusuallydisperseseedsovershortdistances(West-
ment(Montgomery&Chazdon2002;Schleuning,Huama´n& cott&Graham2000).
Matthies 2009). The effects of flooding and canopy gaps on Inthisstudy,weapplydominantamplifiedfragmentlength
plant demography suggest that both types of natural distur- polymorphism (AFLP) markers to study the effects of two
bance strongly affect the population genetics of Amazonian importantenvironmentalfactors(floodingandcanopygaps)
plants.Intemperateforests,environmentalgradientsoflight and of seed dispersal by water and by birds on the genetic
orwateravailabilityhavebeenshowntocausedifferencesin structureofaclonalAmazonianunderstoreyherb(Heliconia
the genetic diversity of populations (e.g. Kudoh et al. 1999; metallica Planchon & Linden ex Hooker, Heliconiaceae).
Jacquemynetal.2005),buttoourknowledge,nosuchstudies H. metallica grows patchily inthe understorey and on river-
havebeencarriedoutintropicalforests. banksofAmazonianfloodplainforests(Berry&Kress1991;
The effects of environmental disturbance on the genetic Schleuning,Huama´n&Matthies2008).Clonalpropagationis
diversityofplantpopulationsareparticularlystronginclonal muchmoreimportantthanseedlingrecruitmentinestablished
plants (Rusterholz, Kissling & Baur 2009; Vandepitte et al. populationsofH.metallicathatcanbeverylong-lived(Schleu-
2010)aslowprobabilitiesofsexualreproductionandseedling ning, Huama´n & Matthies 2008). Ramets of H. metallica
recruitmentreducetheratioofsexuallytoclonallyproduced flower mainlyincanopy gaps(Schleuning,Huama´n& Mat-
offspringandthustheclonaldiversityofapopulation(Wat- thies2008) thatarevisitedby their trap-lining hummingbird
kinson&Powell1993).Althoughlowratesofrepeatedseed- pollinators(Schleuningetal.2010).Theflowersarepartially
lingrecruitmentaresufficienttomaintaintheclonaldiversity self-incompatible(Schleuningetal.2010).Mostofthefleshy
inapopulation(Watkinson&Powell1993),manyclonalplant fruits of H. metallica are eaten by frugivorous understorey
specieslackseedlingrecruitmentaftercolonization,leadingto birds that regurgitate and thus disperse the seeds. However,
adecreaseinclonaldiversityovertime(Honnayetal.2006). seed dispersal in the forest understorey is spatially restricted
Becausethegenetsofclonalplantsareusuallyverylong-lived andabout35%oftheripefruitsfalltothegroundwithoutbird
(Ehrle´n&Lehtila¨ 2002),evenunderenvironmentalstress(Eri- handling (Schleuning, Huama´n & Matthies 2009). Thus, in
ksson1996),populationsconsistingofasingleorafewgenets populationsgrowingclosetostreamsandrivers,primaryand
can survive for a long time (Honnay & Bossuyt 2005). secondaryseeddispersalbywatermayalsobeimportant(see
Althoughmanyunderstoreyplantsintropicalforestspropa- alsoStiles1979).Thefleshyfruits,butnottheseedsofH.met-
gateclonally (Sagers 1993; Schleuning, Huama´n & Matthies allica, are able to float. Seeds are able to germinate without
2008), there are very few population genetic studies of these birdgutpassage(Schleuning,Huama´n&Matthies2009),and
plants(butseeMelendez-Ackermanetal.2005;Cuartas-Her- seedlingrecruitmentfromtheshort-livedseedsismuchhigher
na´ndez&Nu´n˜ez-Farfa´n2006),andnostudyhasinvestigated atfloodedthanatnon-floodedsites(Schleuning,Huama´n&
theeffectsofenvironmentaldisturbance. Matthies 2009). With the detailed knowledge of the natural
Dynamic river systems may not only influence thegenetic history of H. metallica, weaimed at identifying theenviron-
diversity within populations, but also the geneticdifferentia- mentaldriversofthepopulationgeneticsofthisherbattwo
tion among populations (Kondo, Nakagoshi & Isagi 2009; spatial scales. At the small scale, we sampled leaves in 24
Huetal.2010),andareexcellentmodelsystemstostudythe patches in a floodplain forest along a gradient of flooding
geneticstructureofplant metapopulations(Teroetal. 2003; frequencyandcanopy openness,andat thelargescalein21
Jacquemynetal.2006b).However,fewstudieshaveanalysed riverinepopulationsalonga550-kmstretchofriver.Wetested
thegeneticstructureofAmazonianplants,andallthesestudies four specific hypotheses: (1) At the small spatial scale,
havedealtwithtimbertrees(Russelletal.1999;Lemesetal. frequentlyfloodedpatchesofH.metallicahaveahigherclonal
2003;delaTorreetal.2008).Intemperateriversystems,gene diversitythannon-floodedpatches.(2)Theclonaldiversityin
(cid:2)2011TheAuthors.JournalofEcology(cid:2)2011BritishEcologicalSociety,JournalofEcology,99,373–382
Population genetics of anAmazonian herb 375
apatchincreaseswithcanopyopenness.(3)Atthelargespatial Atthesmallspatialscale,wesampled24patcheswithH.metallica
scale, the genetic diversity of riverine populations increases along a gradient of flooding frequency and canopy openness in
downriver because of unidirectional seed dispersal by water. c. 2km2ofafloodplainforestnearManuWildlifeCenter(260m
(4)Geneflowbyhydrochoryismoreeffectivethanbyornith- a.s.l.,12(cid:3)21¢20¢¢S,70(cid:3)42¢44¢¢W;site8inFig.1).Sampledpatches
wereseparatedbyatleast70mfromeachotherwithonlyindividual
ochoryandresultsinlowergeneticdifferentiationamongriver-
rametsinbetween(meandistancebetweensampledpatches:650m).
inepopulationsthanamongpatchesinthefloodplainforest.
We chose patches both under closed canopy and in small canopy
gaps,andsampledatdifferentelevationsabovetheriver(maximum
differenceinelevationc.6m),rangingindistancetotheriverfrom
Materials and methods
100to1100m.MostpatchesofH.metallicainthefloodplaininterior
areprobablyratherold(Schleuning,Huama´n&Matthies2008).
STUDY SYSTEM
Inthecentreofeachpatch,weestablishedaregulargridof6·9m
Amongtheover200Heliconiaspp.(Heliconiaceae),whicharewide- withcordsandsampledleavesfrom12rametsatequaldistancesof
spreadintheunderstoreyofneotropicalforests(Berry&Kress1991), 3m.Ineachgrid,wecountedthenumberofvegetativeandreproduc-
H.metallicaoccursfromHondurastoBolivia(Berry&Kress1991) tive ramets of H. metallica as a measure of patch density (range:
andiscommoninbothriverineandleveehabitatsinthefloodplain 0.80–4.96rametsm)2, see TableS1a in Supporting Information),
forestsofsouth-easternPeru.Likemostofitscongeners,H.metallica andcalculatedtheproportionofreproductiveramets(range:0–28%).
spreadsclonallybylateralrhizomes(Schleuning,Huama´n&Mat- Ineachpatch,wetookfourhemisphericalphotographsabovethe
thies2008).Rhizomesproduceerectabove-groundshoots(ramets)of H.metallicaramets(c.1.8maboveground)withacamerawithfish-
upto3.5mheightthatcandevelopasingleterminalinflorescence. eye-converter(NikonCoolpix4500withFC-E8converter,Nikon,
Rametseventuallydieafterreproduction,butnewshootsareformed Tokyo,Japan)mountedonalevelledtripod.WiththesoftwareGap
fromtherhizome(Schleuning,Huama´n&Matthies2008).Thestud- LightAnalyzer(Frazer,Canham&Lertzman1999),wedeterminedthe
iedpopulationsofH.metallicawerelocatedinthefloodplainofthe percentageofopenskyineachphotograph(i.e.canopyopenness)
MadredeDiosRiverinsouth-easternPeru(Fig.1).Precipitationin and calculated mean canopy openness (range: 2.8–6.8%, Table
thestudyareais2200–3000mmyear)1.Precipitationincreaseswith S1a).Duringfloodingeventsintherainyseasonsof2005,2006and
altitude and decreases in the lowlands towards the East (Amazon 2007,wevisitedthepatchestorecordwhethertheywerefloodedor
ConservationAssociation,unpublisheddata). not.Wedistinguishedpatchesthatwerefrequentlyflooded(inall
years,n=8),occasionallyflooded(inoneortwoyears,n=8),or
neverflooded(n=8,TableS1a).Fromthe24patchessampled,we
SAMPLING PROCEDURE
chosealarge,denselypopulatedpatchforanin-depthanalysis.We
Tostudy the small-scale effects of environmental disturbance in a markedaplotof6·36mwithcords,andindividuallymappedand
floodplainforestandthelarge-scalepatternsalongtheriver,wecol- sampledeachrametofH.metallicagrowingintheplot(n=264
lectedleafsamplesofH.metallicainpopulationsatthesmallandthe ramets).Tocomparethegeneticsimilarityoframetswithinclones
largespatialscale.InJanuary2007,wecollectedc.2·2cmofleaf withthatamongsiblings,wefurthersampled79seedsfrom21fruit-
tissuefromthemostrecentlyformedleafofeachramet.Beforeleaf ingrametsgrowinginsixofthe24patches.
removal,wethoroughlycleanedtheleafsurfacewithtissuepaperto At the large spatial scale, we sampled 21 riverine sites with
reduce contaminationbyepiphyllic algae and fungi. Sampleswere H. metallica from 500ma.s.l. (12(cid:3)52¢08¢¢ S, 71(cid:3)22¢29¢¢ W) to
driedandstoredinplasticbagsfilledwithsilicagel. 165ma.s.l. (12(cid:3)31¢46¢¢S,68(cid:3)54¢04¢¢W)along a 550-km stretch of
Peru
Study
area
Manu river
6 7 0 500
23 4 ad5re de Dios river 8 9 10 11 12 13 14 1M5ad1re6 de Dios river Kilo1m9etr2es0 21
1 M Colorado river Inambari river 17 18mbopata river Altit02u 0–d02 e–0 03[m00 a.s.l.]
Ta 300 –400
400 –500
500 –600
0 50 100 N 600 –1000
Kilometres >1000
Fig.1.Mapofthestudyregionwiththelocationofthe21studysitesalongtheMadredeDiosRiverinsoutheasternPeru.Populationsinthe
interiorofthefloodplainforestwerestudiedatsite8,where24patchesofH.metallicaweresampledinanareaof2km2.
(cid:2)2011TheAuthors.JournalofEcology(cid:2)2011BritishEcologicalSociety,JournalofEcology,99,373–382
376 M.Schleuninget al.
river (Fig.1). The Upper Madre de Dios River is the altitudinal 0.019,wheremeanandSDarethemeanandstandarddeviationof
range limit ofH. metallica in south-eastern Peru. Tothe East, H. thepairwiseJaccardsimilarities,respectively.Theformularesulted
metallica is less abundant because of a reduction in precipitation inathresholdofT=0.901,correspondingtosevenmismatches.
towards the savannah areas in adjacent Bolivia and Brazil. We WeusedthefunctioncloneofAFLPDAT(Ehrich2006)todefine
determined the geographic coordinates and altitude a.s.l. of each clonal phenotypes and detected 190 genets in floodplain patches
site, and tracked the boat journey on the river with a GPS recei- andriverinepopulations.Eachgenetoccurredonlyinasinglepop-
ver. Track distances between sample sites were used to determine ulation,andmorethanhalfofthegenets(57%)occurredonlyonce.
the distance between sites along the river (mean distance between
neighbouring sites: 27.7km, range: 11.6–135.0km, Fig.1). Dis-
CLONAL AND GENETIC DIVERSITY
tancesalongtheriverwereverycloselycorrelatedwithdirectgeo-
graphic distances (r=0.997, Mantel-P<0.001). In contrast to Toestimateintrapopulationclonaldiversity,wedeterminedclonal
thefloodplainpatches,riverinepopulationsgrewnexttotheriver, diversityRasR=(G-1)⁄(n-1),whereGisthenumberofgenets
were linear in shape, and usually had an extension of more than andnisthenumberofsampledramets(Dorken&Eckert2001),as
50m; many of these populations might have established rather wellasthecomplementoftheSimpsonindexD,andtheevenness
recently on the riverbanks. In each population, leaf samples from E based on the Simpson index (Arnaud-Haond et al. 2007). The
12 ramets were collected at equal distances (c. 5m) along a tran- differentmeasuresofclonaldiversity(G,R,D)werecloselycorrelated
sectthroughthepopulation.Wecountedthenumberofvegetative (r>0.89),andspatialvariationinclonaldiversitywasonlyanalysed
and reproductive ramets of H. metallica in a rectangle of 3m intermsofclonaldiversityR.
width along each sample transect. We used the total number of Analysesofgeneticdiversitywereperformedatthegenetleveland
ramets to calculate population density (range: 0.23–4.73 ramets restrictedtopopulationsthatcontainedatleastfourgenets,because
m)2, TableS1b) and also calculated the proportion of reproduc- estimatorsofgeneticvariationstronglydependonsamplesize.We
tiverametsforeachsite(range:0–41%). calculatedNei’sgenediversity,H,andthepercentageofpolymor-
e
phicloci,P ,withaflp-surv1.0(Vekemansetal.2002),estimating
m
allelefrequencieswiththesquare-rootmethodassuminganinbreed-
AFLP GENOTYPING
ing coefficient of F=0.0. H and P were closely correlated
e m
PlantgenomicDNAwasextractedfrom10to15mgofdriedleaftis- (r=0.73) and genetic diversity among riverine populations was
sue or seed, respectively, using the DNeasy 96 Plant Kit (Qiagen, comparedintermsofpercentageofpolymorphicloci.Intheflood-
Crawley,UK).PriortoDNAextractionfromseeds,embryoswere plaininterior,geneticdiversitywasnotanalysed,because15of24
excisedtoremovethematernaltissueoftheseedcoat.TheDNAwas patchesconsistedoflessthanfourgenets.
eluted in 50lL of buffer AE obtaining concentrations of Inthefloodplaininterior,wetestedtheeffectoffloodingregimeon
16nglL)1±2.5 for leaves and 3.8nglL)1±2.9 for embryos. theclonaldiversityinthe24patcheswithanovaandpartitionedthe
AFLP(Vosetal.1995)analysiswasperformedwithamodifiedver- overallfloodingeffectintotwoorthogonalcontrastsoffloodedvs.
sionoftheAFLPplantmappingkitprotocolofAppliedBiosystems non-flooded,andfrequentlyvs.occasionallyfloodedpatches.More-
(Carlsbad,CA,USA)(ABI,fordetailsseeAppendixS1). over, we used simple regression analyses to investigate the effects
Wetested26primercombinationsfromwhichsixwereselected ofcanopyopenness,patchdensity,andproportionofreproductive
because of clear and scorable banding patterns: EcoRI-ACT- rametsonclonaldiversity.Atthelargespatialscale,weusedtheriver
FAM⁄MseI-CTA, ACG-JOE⁄CAG, ACC-TAMRA⁄CAG, ACT- distancesbetweenneighbouringsamplesitestodeterminetheposi-
FAM⁄CTC, AAG-JOE⁄M-CTC and AAC-TAMRA⁄CTG.Frag- tionofeachpopulationalongtheriver.Thepositionofthepopula-
mentanalysiswasperformedonanABIPrism310geneticanalyser tionatthealtitudinalrangemarginofH.metallicawassettozero.
withinternalsizestandardGeneScan-500-Rox.TheAFLPlociwere Wecarriedoutsimpleregressionanalysestotesttheeffectsofriver
scoredwiththe Genographer1.6.0 software(Benham etal. 1999), position,populationdensity,andproportionofreproductiveramets
usingfragmentsbetween50and500bp,andexportedasabinary onclonaldiversity.BecauseofthelowerabundanceofH.metallica
matrix.Outofatotalof625leafand79seedsamples,wescored180 ontheriverbanksofup-anddownriversites,weexpectedahump-
fragments,ofwhich103(57.2%)werepolymorphicandusedforthe shapedeffectofriverpositiononclonaldiversityandincorporatedan
analyses.Weperformedreplicateanalysesof16samplestodetermine additionalquadratictermofriverpositionintotheregressionmodel.
the reproducibility of AFLP genotyping and obtained an overall Furthermore,we analysed the effects of riverposition,population
errorrateof3.1%. density,andproportionofreproductiverametsongeneticdiversity.
Toadjustforthedifferentnumberofgenetsinthepopulations,we
additionally included the number of genets in multiple regression
IDENTIFICATION OF CLONES
models.
Inordertoidentifyclones,i.e.geneticallyidenticalramets,weused Weexaminedthenormalityoftheresidualsofalllinearmodels
themethodofDouhovnikoff&Dodd(2003)todistinguishclones andangular-transformedclonaldiversityRtoimprovemodelresid-
fromcloselyrelatedsiblingsandtocalculateathresholdofsimilar- uals. Angular transformation of the proportion of reproductive
ityabovewhichrametswereconsideredtobelongtothesamegenet. ramets and of the proportion of polymorphic loci was not neces-
WedeterminedtheJaccardsimilaritybetweenthe79siblingsfrom sary. Analyses were carried out with R 2.10.1 (R Development
the21seedfamilies(120within-familycomparisons),andbetween92 CoreTeam2009).
rametsgrowinginthe6·36mplot(4186within-genetcomparisons)
because92ofthe96genotypedrametsinthisplotbelongedtothe
GENETIC STRUCTURE WITHIN AND AMONG
samegenet.Siblingsandclonesweredistinguishedbyathreshold
POPULATIONS
Jaccard similarity T defined by the following equation: T=
(SDclones·meansibs+SDsibs·meanclones)⁄(SDsibs+SDclones), Inordertodetectwhetherfloodplainpatchesandriverinepopula-
withmeansibs=0.728,SDsibs=0.058,meanclones=0.966,SDclones= tionsshowedasmall-scalegeneticstructure,weanalysedthespatial
(cid:2)2011TheAuthors.JournalofEcology(cid:2)2011BritishEcologicalSociety,JournalofEcology,99,373–382
Population genetics of anAmazonian herb 377
geneticautocorrelationoframetsandgenetswiththeprogramSPA- 1.00 (a) r2 = 0.40
GeDI(Hardy&Vekemans2002),poolingdatafromthetwospatial 0.95 P = 0.010
scales. We used the relationship coefficient for dominantmarkers, R
y 0.80
defined by Hardy (2003), as a measure of genetic relatedness. To sit
determinespatialcoordinates,weusedsamplingpositionsalongtran- ver 0.60
di 0.40
sectsorsamplinggridsfortherametsandcentroidcoordinatesfor al
n 0.20
thegenets. o
Cl
Toestimategeneticdifferentiationamongpopulations,wecalcu- 0.05
latedNei’sgeneticdistance(Nei1972)andF-statistics(Wright1951), 0.00
incorporatingonlypopulationsthatcontainedatleastfourgenets.
Both measures were calculated with aflp-surv 1.0 following the 60 (b) r = 0.49
approachofLynch&Milligan(1994),estimatingallelefrequencies %) 55 P = 0.046
withthesquare-rootmethodatF=0.0.Geneticpopulationstruc- (m 50
ture was further investigated by analyses of molecular variance Pm 45
(amova) with Arlequin 3.0 (Excoffier,Laval & Schneider 2005). In s
ordertotestforisolationbydistanceamongpopulations,weanaly- orphi 40
sed whether Nei’s genetic distances between pairs of floodplain ym 35
ol
patchesorriverinepopulationswererelatedtogeographicdistances P 30
ortoriverdistances,respectively,usingMantel-tests(10000itera-
25
tions,veganpackage1.17-2inR2.10.1,Oksanenetal.2010).Inaddi- 0 100 200 300 400 500 600
Relative river position (km)
tion, we applied a Bayesian approach to cluster individuals into
‘panmictic’ phylogroups using baps 5.2 (Corander, Sire´n & Arjas
Fig.3.EffectofthepositionofapopulationontheMadredeDios
2008).Weranbothanon-spatialandaspatialpopulationmixture
Riveron(a)clonaldiversityin21,and(b)percentagepolymorphism
analysiswiththemaximalnumberofgroups,K,rangingfrom1to20 in17populationsofH.metallica.Riverdistancesbetweenpopula-
with three replicates that identified the most likely partitioning tionswereusedtoestimatetheriverpositionofeachpopulation;pop-
accordingtomarginallog-likelihood. ulation1wassettozero.NoteangularscaleforclonaldiversityR.
Results
In riverine populations, mean clonal diversity R was 0.47
(range:0–0.90),Simpson’sindexD0.74(range:0–0.98),and
CLONAL AND GENETIC DIVERSITY
Evenness E 0.56 (range: 0–1, TableS1b). The effect of river
ClonaldiversityRdifferedstronglyamongpatchesintheinte- position on clonal diversity R was best described by a qua-
riorofthesinglefloodplainforeststudied(mean:0.16,range: dratic regression (r2=0.40, P=0.010, Fig.3a), indicating
0–0.46,TableS1a).Consistentwithourfirsthypothesis,Rwas thatclonaldiversityRwashighestinpopulationsatthemiddle
much higher in flooded than in non-flooded patches rivercourseandlowerinup-anddownriverpopulations.Clo-
(F1,22=20.9,P<0.001,Fig.2),butfrequentlyfloodedand nal diversity R was not related to population density
occasionally flooded patches did not differ (F1,14=0.07, (r=0.31,P=0.174)ortheproportionoffloweringramets
P=0.791).Incontrasttothesecondhypothesis,Rdidnot inapopulation(r=0.02,P=0.936).
increase with canopy openness (r=)0.12, P=0.563). TheaverageofNei’sgenediversityH forriverinepopula-
e
Moreover,clonaldiversitywasnotinfluencedbypatchdensity tions was 0.22 (range: 0.20–0.26) and of percentage of poly-
(r=)0.15, P=0.496) or the proportion of reproductive morphismP 42.3%(range:30.1–54.4%,TableS1b).Inline
m
ramets(r=)0.13,P=0.532). withourthirdhypothesis,percentagepolymorphismmonoto-
nously increased downriver (r=0.49, P=0.046, Fig.3b).
0.30 a We found an increase in genetic diversity even when we
a
adjustedforthenumberofgenetsinapopulation(b=0.40,
R 0.20 t=2.94,P=0.011),andwhenweremovedtwooutlierpop-
y ulationsthatstronglyaffectedthepartialregressioncoefficient
sit 0.12
er (b=0.18, t=2.28, P=0.042). Percentage polymorphism
v
di P was not related to population density (r=0.19, P=
al 0.05 m
n b 0.459) or theproportion of flowering ramets of H. metallica
o
Cl (r=)0.40,P=0.115);incorporatingthenumberofgenets
0.01
inamultiplemodeldidnotchangetheresultsqualitatively.
0.00
Frequently Occasionally
Non-flooded
Flooded GENETIC STRUCTURE WITHIN AND AMONG
POPULATIONS
Fig.2.Effectsofdifferentfloodingregimesontheclonaldiversityin
Theanalysisofspatialgeneticautocorrelationoverallsamples
24patchesofH.metallicainafloodplainforest.Differentlettersindi-
catesignificantdifferencesbetweenfloodingregimesinaposthoctest revealedasignificantspatialstructureatboththerametand
(Tukey,P<0.01);noteangularscaleforclonaldiversityR. genetlevel.Relationshipcoefficientsofgenetsdecreasedwithin
(cid:2)2011TheAuthors.JournalofEcology(cid:2)2011BritishEcologicalSociety,JournalofEcology,99,373–382
378 M.Schleuninget al.
(a) bothpools.Pools3and4werealmostexclusivelyrestrictedto
nt 1.0 Ramets several (11) or a few (2) floodplain patches, respectively. In
e
effici 0.8 Genets cwoanstrfoasutntdointhtehenospna-stpiaaltlyiailnafonramlyesdis,clounsltyeraansianlgylseisg(elongempoaor-l
co 0.6
p ginallikelihood-7001),underliningthelowgeneticdivergence
shi 0.4 amongpopulations.
n
o
ati 0.2
el
R 0.0 Discussion
3 10 30 100 300 1000
Spatial distance (m)
ENVIRONMENTAL EFFECTS ON CLONAL AND GENETIC
(b) DIVERSITY
0.10
ance 0.08 rM =a n0t.e3l4-P < 0.01 Flooding,butnotcanopyopennessstronglyaffectedtheclonal
st diversityofH.metallica.Clonaldiversitywasmuchhigherat
c di 0.06 floodedthanatnon-floodedsites,becauseseedlingrecruitment
neti 0.04 ofH.metallicaislargelylimitedtofloodedsites(Schleuning,
e
s g 0.02 Huama´n&Matthies2009).Ourstudysupportstheconclusion
ei' froma demographicstudythatnon-floodedpopulations are
N 0.00
maintainedonlybymeansofclonalpropagation(Schleuning,
0 100 200 300 400 500
Distance along the river (km) Huama´n&Matthies2008).Thesepopulationscouldberem-
nant populations that have survived for several decades or
Fig.4.Isolation by distance within and between populations of evencenturiesaftertheirsiteswerenolongerflooded(Schleu-
H.metallica.(a)Spatialautocorrelogramofrelationshipcoefficients
ning,Huama´n&Matthies2008)andmayhavelosttheirclonal
oframetsandgenetsoverallpopulations.Meanrelationshipcoeffi-
diversity over time. Alternatively, non-flooded populations
cientsandtheir95%confidenceintervalsaregiven;notelog-scalefor
spatialdistance.(b)Nei’sgeneticdistancesbetweenpairsof17river- couldhavebeenestablishedbyasinglecolonizationeventdur-
inepopulationsinrelationtodistancealongtheriver. inganextremefloodandexperiencedstrongfoundereffects.
Althoughnotonlyflooding,butalsotheformationofcanopy
gaps, increases seedling recruitment (Schleuning, Huama´n &
patches(atdistances<10m),suggestingthatgeneflowisspa- Matthies2009),gapsdidnotaffectclonaldiversity.Thismight
tiallyvery restricted. Spatialautocorrelationdeclinedsteeper beduetotheephemeralcharacterofsmallcanopygapsthat
forrametsthanforgenets(Fig.4a).Atdistances<10m,rela- increaselightavailabilityintheunderstoreyoftropicalforests
tionshipcoefficientsoframetswereveryhigh,butdroppedto onlyforabout2years(Smith,Hogan&Idol1992;Schleuning,
levelssimilartothoseofgenetsatdistances>50m.Accord- Huama´n&Matthies2008).
ingly,agenetneveroccurredinmorethanonepopulation,and In temperate understorey herbs, environmental conditions
rametsbelongingtothesamegenetextendedoveramaximum stronglyaffecttheclonaldiversityofpopulations,buteffects
distanceof55m. dependontheenvironmentaldriverandtheplantspeciescon-
F-statistics indicated that 16% of the genetic variation sidered.Highsoilmoistureincreasesseedlingrecruitmentand
residedamongpatchesinthefloodplainforest(F =0.158, clonal diversity of Paris quadrifolia (Jacquemyn et al. 2005),
ST
SE=0.075,P<0.001),butonly6%amongriverinepopu- whilehumantramplingdecreasesclonaldiversityofAnemone
lations at the large spatial scale (F =0.062, SE=0.231, nemorosa (Rusterholz, Kissling & Baur 2009). Canopy gaps
ST
P<0.001).Nei’sgeneticdistancesbetweenpairsofpopula- affectclonaldiversitybothpositively(Kudohetal.1999)and
tions were higher between patches in the floodplain forest negatively(Vandepitteetal.2010),dependingontheresponses
(0.051±95% CI 0.011) than between riverine populations ofsexualandclonaloffspringtotheincreaseinlightavailabil-
(0.021±95%CI0.003).Thehigherdifferentiationwithinthe ity (Matlaga & Horvitz 2009; Vandepitte et al. 2010). The
floodplain than along the river is consistent with our fourth results from our study suggest that flooding influences the
hypothesis suggesting that gene flow by ornithochory is less genetic structure of plant populations in Amazonian flood-
effective than by hydrochory. Genetic distances were not plainforestsmorestronglythanephemeralcanopygaps.
related to geographic distances in the floodplain interior Thepositionalongtheriverinfluencedtheclonaldiversity
(r=0.035, Mantel-P>0.40), but increased with distance ofriverinepopulationsofH.metallica.Incontrasttopopula-
alongtheriveramongriverinepopulations(r=0.338,Man- tionsalongthemiddlecourse,upriveranddownriverpopula-
tel-P<0.01,Fig.4b). tions only consisted of a single or a few different genets.
TheBayesianclusteranalysisatthegenetlevelrevealedfour Accordingly,theoverallabundanceofH.metallicawaslower
genepools(logmarginallikelihood-6853,Fig.S1inSupport- ontheriverbanksintheAndeanfoothillsandintheEastern
ing Information), that where rather closely related with lowlands. Differences in both clonal diversity and overall
pairwise Nei’s distances ranging from 0.029 to 0.143. Gene abundance could be due to differences in flooding regimes
pools1and2wherepresentbothalongtheentireriverandin along the river because the flow velocity strongly decreases
thefloodplainforestwithmostpopulationsbeingamixtureof fromtheAndeanfoothillstotheEasternlowlands(Goulding
(cid:2)2011TheAuthors.JournalofEcology(cid:2)2011BritishEcologicalSociety,JournalofEcology,99,373–382
Population genetics of anAmazonian herb 379
etal.2003),providingsuitableintensitiesoffloodingforseed- utedandcolonizationofnewsiteswasexclusivelyduetothe
ling recruitment mainly at mid-altitudes (see Schleuning, recruitment of seedlings and not to the dispersal of clonal
Huama´n & Matthies 2009). Alternatively, changes in soil propagules.
propertiesorlongerdroughtperiodsintheEasternlowlands Intheonefloodplainforeststudied,thegeneticdifferentia-
could limit the distribution of H. metallica (see Schleuning, tion among patches of H. metallica was pronounced
Huama´n & Matthies 2008). A lower clonal diversity at the (F =0.16).Patchesclusteredintofourdifferentgenepools,
ST
distributional margins is consistent with previous studies althoughallweresituated withina small area of 2km2ofa
reporting that sexual reproduction and seedling recruitment floodplainforest.Stronggeneticdifferentiationhasalsobeen
decrease at the range margins of plant species (Eckert 2001; found in studies of clonal herbs of temperate forests (Auge
Jump&Woodward2003;Beattyetal.2008). et al. 2001; Jacquemyn et al. 2006a). In tropical floodplain
The genetic diversity of H. metallica increased downriver. forests,geneflowbypollenorseedsappearstobelow.Recruit-
Tosomeextent,thiscouldbetheconsequenceofrareseedling ment of seedlings of H. metallica in the floodplain mostly
recruitmentinupriverpopulations,resultinginlowfrequencies occursadjacenttoreproductiverametsindicatingthatornith-
ofmultilocusgenotypesinthesepopulations.However,inour ochorous seed dispersal of H. metallica is spatially restricted
analyses,weonlyincludedpopulationswithatleastfourgenets (Schleuning,Huama´n&Matthies2009).However,inH.met-
andstatisticallyaccountedfordifferentnumbersofgenetsper allica, pollen flow between patches is likely to occur and
population.Moreover,differencesinseedlingrecruitmentcan- flowersofsolitaryrametsreceivepollenofhighqualityfrom
notexplainthehigh-geneticdiversityindownriverpopulations distantsources(Schleuningetal.2010).Thisisnotsurprising
where clonal diversity was comparatively low. The monoto- becausehermithummingbirds,thepredominantpollinatorsof
nous increase of genetic diversity in riverine populations of H.metallica,foragealongregularlyfollowedroutes(i.e.trap
H. metallica is rather consistent with the hypothesis that lines)ofupto1kmlength(Stiles&Wolf1979).Nevertheless,
unidirectional geneflow leads to an accumulation of genetic pollenflowbetweenpatchesofH.metallicaisapparentlynot
diversity in downstream populations (Barrett, Eckert & sufficienttopreventtheformationofastrongspatialgenetic
Husband1993).IntheMadredeDiosriversystem,geneflow structureinthefloodplain.
byhydrochoricseeddispersalfromlowlandtributaries(Colo- The lack of replication of the small-scale study of spatial
rado,InambariandTambopataRiver,seeFig.1)couldfur- geneticstructureofH.metallicacouldaffectthegeneralityof
ther contribute to the accumulation of genetic diversity in our findings.Detailed sampling was only feasible in a single
downriverpopulations.Increasinggeneticdiversitydownriver floodplainforestwheretrailsprovidedeasyaccesstotheforest
wasfoundinseveralplantspeciesoftemperateriversystems interiorandwherefloodingfrequenciesweremonitoredovera
(Lundqvist&Andersson2001;Liu,Wang&Huang2006;Fe´r 3-yearperiod.However,thestudysitemightberepresentative
&Hroudova´ 2008;Chenetal.2009;Polluxetal.2009),but for many other sites along the middle course of Amazonian
wasnotevidentinmanyothers(Lundqvist&Andersson2001; rivers where spatially extended floodplain forests have been
Imbert & Lefe`vre 2003; Tero et al. 2003; Hu et al. 2010). formedalongrivermeanders(Saloetal.1986;Foster1990).
According to these studies, the relative contribution of dis- Although each floodplain may be unique in its flooding his-
persalsystemsotherthanhydrochoryandtheimportanceof tory,inwaterandsoilchemistryandintherelativeproportions
geneflowbypollendeterminethedistributionofgeneticdiver- ofcurrentlyfloodedandnon-floodedareas(Saloetal.1986),
sityalongriversystems.InH.metallica,geneflowbyornith- theseforestsshareaheterogeneousmicro-topography,dueto
ochorous pollen and seed dispersal seems to be far less variationinelevationabovetheriver atasmallspatialscale
important than water-mediated seed dispersal downriver. (Foster1990;King2003).Moreover,theyharboursimilarpol-
Therefore,weassumethathydrochoryisanimportantdriver linatorandfrugivorebirdcommunitiesintheforestundersto-
of the population genetic structure of plant species growing rey (Robinson & Terborgh 1990). Thus, we assume that the
alongriverswithstrongdynamics,suchastheAmazonandits spatialgeneticstructureofH.metallicaissimilarlystrongin
tributaries. otherAmazonianfloodplainforests.
Genetic differentiation among riverine populations of
H. metallica (F =0.06) was much lower than among the
ST
SPATIAL GENETIC STRUCTURE AT DIFFERENT
patchesintheonefloodplainforest(F =0.16).Differences
ST
SPATIAL SCALES
inpopulationagecancontributetothesedifferencesbecause
WefoundthattheextensionofgenetsofH.metallicawasup recentlyfoundedpopulationsofclonalplantsarelessdifferen-
to50mandthatthosegenetsweremuchlargerthanthegenets tiatedfromeachotherthanoldpopulations(Jacquemynetal.
ofunderstoreyherbsoftemperateforests(Kudohetal.1999; 2006b).InH.metallica,floodplainpopulationsarepresumably
Jacquemynetal.2005;Vandepitteetal.2010).GenetsofH. older than riverine populations that might have rather
metallicacouldbemanydecadesorevencenturiesoldbecause recently established on the riverbanks.More importantly, in
rhizomegrowth,inparticularatclosed-canopysites,israther plantsthataredispersedbywater,likeH.metallica,geneflow
slow (maximum of 65cmyear)1; Schleuning, Huama´n & islikelytobehigherinhabitatscloseoradjacenttothewater
Matthies 2008). Nevertheless, relationship coefficients of flow (Kudoh & Whigham 2001; Hu et al. 2010). Consistent
ramets of H. metallica dropped to background levels at dis- withthisconclusion,inH.metallicaalmostallriverinepopula-
tances >50m, indicating that genets were patchily distrib- tionsclusteredintotwocloselyrelatedgenepools,indicating
(cid:2)2011TheAuthors.JournalofEcology(cid:2)2011BritishEcologicalSociety,JournalofEcology,99,373–382
380 M.Schleuninget al.
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the estimates of clonal and genetic diversity of H. metallica in 24 Technicalsupportissuesarisingfromsupportinginformation(other
patchesinafloodplainforest,andin21riverinepopulations. thanmissingfiles)shouldbeaddressedtotheauthors.
Asaservicetoourauthorsandreaders,thisjournalprovidessupport-
inginformationsuppliedbytheauthors.Suchmaterialsmaybere-or-
(cid:2)2011TheAuthors.JournalofEcology(cid:2)2011BritishEcologicalSociety,JournalofEcology,99,373–382