Submi ed 11 July 2016
Accep ed 7 Decembe 2016
Published 7 Feb ua y 2017
Co esponding au ho s
Ja ie A. Gui ián,
[email p o ec ed]
Ma ía Losada,
[email p o ec ed]
Academic edi o
Dezene Hube
Addi ional In o ma ion and
Decla a ions can be ound on
page 12
DOI 10.7717/pee j.2882
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2017 Gui ián e al.
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Di e ences in pollina ion success
be ween local and o eign lowe colo
pheno ypes: a ansloca ion expe imen
wi h Gen iana lu ea (Gen ianaceae)
Ja ie A. Gui ián1, Ma Sob al2, Tania Veiga1, Ma ía Losada2, Pablo Gui ián1
and José M. Gui ián2
1Depa amen o de Bo ánica/Facul ade de Bioloxía, Uni e sidade de San iago de Compos ela, San iago de
Compos ela, A Co uña, Spain
2Depa amen o de Bioloxía Celula e Ecoloxía/Facul ade de Bioloxía, Uni e sidade de San iago de Compos ela,
San iago de Compos ela, A Co uña, Spain
ABSTRACT
Backg ound. The adap i e main enance o lowe colo a ia ion is equen ly a -
ibu ed o pollina o s pa ly because hey p e e en ially isi ce ain lowe pheno ypes.
We es ed whe he Gen iana lu ea—which shows a lowe colo a ia ion ( om o ange
o yellow) in he Can ab ian Moun ains ange (no h o Spain)—is locally adap ed o
he pollina o communi y.
Me hods. We ansplan ed o ange- lowe ing indi iduals o a popula ion wi h yellow-
lowe ing indi iduals and ice e sa, in o de o assess whe he he e is a pollina ion
ad an age in he local mo ph by compa ing i s isi a ion a e wi h he o eign mo ph.
Resul s. Ou ecip ocal ansplan expe imen did no show clea local mo ph
ad an age in o e all isi a ion a e: local o ange lowe s ecei ed mo e isi s han
o eign yellow lowe s in he o ange popula ion, while bo h local and o eign lowe s
ecei ed he same isi s in he yellow popula ion; hus, he e is no e idence o local
adap a ion in Gen iana lu ea o he pollina o assemblage. Howe e , some lo al isi o
g oups (such as Bombus p a o um,B. so oensis anca icus and B. lapida ius decipiens)
consis en ly p e e ed he local mo ph o he o eign mo ph whe eas o he s (such as
Bombus e es is) consis en ly p e e ed he o eign mo ph.
Discussion. We concluded ha he e is no e idence o local adap a ion o he pollina o
communi y in each o he wo G. lu ea popula ions s udied. The consequences o
local adap a ion o pollina o on G. lu ea lowe colo would depend on he a ia ion
along he Can ab ian Moun ains ange in mo ph equency and pollina o communi y
composi ion.
Subjec s Conse a ion Biology, Ecology, En omology, E olu iona y S udies, Plan Science
Keywo ds Flowe colo a ia ion, Local adap a ion, Pollina o p e e ences, Recip ocal
ansplan s, Gen iana lu ea
INTRODUCTION
Flo al e olu ion is p ima ily d i en by pollina o s (B adshaw & Schemske, 2003;Whi all
& Hodges, 2007). The di e si y in lo al ai s ound in angiospe ms, such as lowe colo ,
could be ela ed wi h ansi ions in hese ai s (Raushe , 2008), as a esul o di e en
How o ci e his a icle Gui ián e al. (2017), Di e ences in pollina ion success be ween local and o eign lowe colo pheno ypes: a
ansloca ion expe imen wi h Gen iana lu ea (Gen ianaceae). Pee J 5:e2882; DOI 10.7717/pee j.2882
selec i e p essu es exe ed by pollina o s wi h s ong p e e ences o ce ain cha ac e is ics
(Schemske & B adshaw, 1999;S eis eld & Kohn, 2007) o by changes in pollina o
communi y composi ion (Ellis & Johnson, 2009). Thus, lo al di e si ica ion and specia ion
among closely ela ed plan popula ions may esul om he isola ion and/o local
adap a ion o he mos e icien pollina o s, which inc ease he numbe o isi s pe plan
depending on hei lowe p e e ences (Ha de & Johnson, 2009; and e e ences he ein).
The adap i e main enance o lowe colo a ia ion is equen ly a ibu ed o pollina o s,
in pa because hey end o p omo e asso a i e ma ing, i.e., p e e en ially isi ing ce ain
pheno ypes (Wase & P ice, 1981;S an on, 1987;Jones & Rei hel, 2001). Howe e , many
o he easons could explain he e olu ion o lo al di e si ica ion, such as he po en ial
uncoupling o he e olu ion o lo al ai s om pollina o -media ed selec ion (S auss
& Whi all, 2006). Fo example, pleio opic e ec s on he biosyn he ic pa hways o lo al
pigmen a ion may d i e he e olu ion o al e na i e lowe colo s (A mb us e , 1993;
Schemske & Bie zychudek, 2007;Cooley, Ca allo & Willis, 2008).
Many s udies suppo ha animal pollina o p e e ences may cause selec i e p essu es
on lowe colo , which may luc ua e depending on he s uc u e and/o composi ion o he
pollina o communi y (Melendez-Acke man & Campbell, 1998;Gómez & Zamo a, 2000). In
his con ex , ecip ocal ansplan expe imen s be ween popula ions can be a powe ul ool
o assess how changes in lowe colo p e e ences o he pollina o assemblage con ibu e o
an adap i e ad an age o he local mo ph o e he o eign mo phs (S eis eld & Kohn, 2007).
Gen iana lu ea L. shows lowe colo a ia ion along he Can ab ian Moun ains
(No he n Spain). Gen iana lu ea lowe s a e ypically yellow (G. lu ea a . lu ea), hough
no hwes Ibe ian popula ions ha e o ange- lowe ing indi iduals, which cons i u es a
di e en a ie y o he species (G. lu ea a . au an iaca; Renobales, 2012). The pollina o
assemblages a e made up mos ly o bumblebee species (Veiga e al., 2015;Sob al e al.,
2015). P e ious s udies conclude ha : (i) Gen iana lu ea lowe colo a ia ion does
no esul om adap a ion o en i onmen al ac o s, such as ele a ion, empe a u e,
adia ion, and p ecipi a ion (Veiga e al., 2016); (ii) Gen iana lu ea is s ongly dependen on
pollina o s o ep oduc ion, and he plan has a posi i e ela ionship be ween he numbe
o pollina o isi s and he numbe o seeds i p oduces (Losada e al., 2015); (iii) he e
is a pa ial hyb idiza ion ba ie among G. lu ea colo mo phs (Losada e al., 2015); (i )
Gen iana lu ea lowe colo in luences pollina o isi s and i s a ia ion is ela ed o changes
in pollina o communi y composi ion ac oss popula ions, since pa o his a ia ion is
explained by di e en pollina o selec i e p essu es exe ed on lowe colo among G. lu ea
popula ions (Sob al e al., 2015); and ( ) he mos abundan pollina o s o ou s udy
species, Bombus e es is and B. p a o um (Veiga e al., 2015), possess pho o ecep o s wi h
low sensi i i y o ed colo s; bu o he pollina o s in e ac ing wi h G. lu ea plan s, such
as Bombus lapida ius, has high sensi i i y o ed colo s despi e he lack o pho o ecep o s
equi ed (Pei sch e al., 1992).
In o de o de e mine whe he G. lu ea popula ions a e locally adap ed o he pollina o
communi y, we e alua ed he e ec o local and o eign lowe colo mo phs on pollina o
isi a ion a e by means o ecip ocal ansplan s in wo Gen iana popula ions. Mo e
speci ically, we es ed whe he he e is an adap i e ad an age in he local lowe colo
Gui ián e al. (2017), Pee J, DOI 10.7717/pee j.2882 2/15
Figu e 1 Colo mo phs o Gen iana lu ea L. a No he n Spain. (A) a . lu ea, he yellow mo ph in
Pon ón loca ion; and (B) a . au an iaca, he o ange mo ph in Anca es loca ion.
mo ph ela i e o he o eign mo ph in pollina o isi s by compa ing he (i) o e all
pollina o assemblage, and (ii) di e en isi o g oups sepa a ely. We p edic ed ha he
local lowe colo mo ph would ecei e mo e pollina o isi s han he o eign mo ph, and
ha he wo di e en colo mo phs would ecei e isi s by di e en pollina o g oups in
hei local popula ions.
MATERIAL AND METHODS
Plan species
Gen iana lu ea L. is an he baceous pe ennial geophy e (Fig. 1). Du ing summe , he hizome
de elops a lowe ing s alk o ca. 80 cm high, wi h a basal ose e o med by 4–8 oblong
lea es. Flowe s g ow in wo opposed cymose g oups (each one wi h 15 lowe s) a di e en
s alk le els, abo e wo ellip ic lea es; and one lowe c owns he op o he s alk. Flowe s
ea u e a o a e co olla wi h 5–7 lobules, 5–7 ee s amens, and a ixed o a y wi h wo
nec a ies on he base. The ui is an o oid capsule, holding ellip ic and winged seeds o 3–4
mm (Renobales, 2012). This species needs pollina o s—mainly bumblebees— o p oduce
seed (see Veiga e al., 2015;Sob al e al., 2015).
Gen iana lu ea lowe colo a ies be ween o ange and yellow (see Fig.1) along he
Can ab ian Moun ains (No he n Spain; Fig. 2). The wes e n popula ions (wes o Somiedo
Na u al Pa k) ha e o ange- lowe ing indi iduals, whe eas he eas e n popula ions (eas o
Pue o de Ven ana) ha e yellow- lowe ing indi iduals; and he in e media e popula ions
show indi iduals wi h lowe s o a g ada ion be ween hese wo colo mo phs.
S udy si e
The s udy was conduc ed in July 2012 a he no hwes o he Ibe ian Peninsula in wo
popula ions, one loca ed in Sie a de Anca es (o ange- lowe ed popula ion) and o he
in Pue o del Pon ón (yellow- lowe ed popula ion), sepa a ed by app oxima ely 160 km
(Fig. 2). Popula ion size was es ima ed by he o al numbe o G. lu ea indi iduals (>3,000
in bo h popula ions).
Flowe colo o 75 haphaza dly selec ed lowe s ( h ee lowe s pe plan , 25 plan s) was
measu ed he p e ious yea in bo h popula ions by means o a spec ome e (USB2000+;
Gui ián e al. (2017), Pee J, DOI 10.7717/pee j.2882 3/15
Figu e 2 Geog aphic loca ion o Gen iana lu ea popula ions a No he n Spain in which ecip ocal
ansplan s we e conduc ed. Black do , Anca es, he o ange mo ph popula ion; whi e do , Pon ón, he
yellow mo ph popula ion.
Ocean Op ics, Inc., Dunedin, FL, USA) and pe al colo spec a we e p ocessed using he
Spec aSui e R
so wa e (Ocean Op ics, Inc., Dunedin, FL, USA). The CIELab Colo ime ic
Sys em (CIE P., 2004) was chosen o desc ibe lowe colo a ia ion in he isible ange o
he elec omagne ic spec um, h ough h ee a iables educed by p incipal componen
analysis ( o addi ional de ails, see Veiga e al., 2015): L(b igh ness o colo , om black
o whi e), a( ed colo a ia ion, om g een o ed) and b(yellow colo a ia ion, om
blue o yellow). Indi iduals om he Sie a de Anca es popula ion showed o ange-colo ed
lowe s (L=16.42; a=11.77; b=23.61), and om he Pon ón popula ion, yellow ones
(L=19.64; a=3.84; b=27.84).
Al hough ce ain pollina o species may de ec UV ligh , a p e ious s udy ound no
di e ences among G. lu ea popula ions and be ween indi iduals wi h di e en colo
mo phs (o ange o yellow, disce nible by human eye) in he UV ligh ange (Veiga e al.,
2015). Thus, we p esume ha UV ligh does no d i e local adap a ion among popula ions,
and conside lowe colo in bo h, he UV and isible ligh anges in his s udy.
Expe imen al design
In July 2012, a ecip ocal ansplan expe imen was conduc ed o analyze local adap a ion
o colo mo phs o pollina o assemblage o bo h popula ions ( hanks o he ield
Gui ián e al. (2017), Pee J, DOI 10.7717/pee j.2882 4/15
pe mission issued by he En i onmen al Te i o ial Se ice ins i u ion om León, Regional
Go e nmen o Cas illa and León, Te i o ial Delega ion o Go e nmen o Spain—
Iden i ie : 12_LE_325_RNA_PuebladeLilio_INV—Re e ence: 06.01.013.016/ROT/abp—
File numbe : AEN/LE/103/12). Gi en he di icul y o ansplan ing plan s in po s due
o hei ex ensi e hizomes, lowe ing s ems used o he expe imen we e cu a he
heigh o hizome, placed in plas ic con aine s wi h wa e (0.5 L), sealed wi h adhesi e
ape, and bu ied a g ound le el o p e en e apo a ion and ensu e longe lowe li e.
The e ec i eness o his me hod was p e iously de e mined by checking he du a ion
o lowe ing and pollina ion in a con ol popula ion and e i ying ha he lowe s we e
well-p ese ed in he i s ou days.
Fo he ecip ocal ansplan expe imen , 15 lowe ing s ems we e cu in he o ange
popula ion (Anca es) and ans e ed o he yellow popula ion (Pon ón) immedia ely
a e collec ion, while simul aneously, 15 yellow s ems we e cu in Pon ón and ans e ed
o he Anca es popula ion ( o eign ansplan ea men ). Addi ionally, 15 plan s o each
popula ion (o ange in Anca es and yellow in Pon ón) we e subjec ed o he same p ocedu es
(cu , ansplan ed, and bu ied) o he local ansplan ea men .
The same day, plan con aine s we e placed in each popula ion wi hin 1 m2squa es, se
in andom layou by picking he g id poin using a andom numbe able. Addi ionally,
15 plan s we e haphaza dly ma ked o se e as a con ol in bo h popula ions. Thus,
each popula ion con ained 30 expe imen al plan s and 15 con ol plan s (45 plan s pe
popula ion, 90 plan s o e all). No e ha he s em heigh , he numbe o lo al who ls pe
in lo escence, and he numbe o lowe s we e measu ed in all plan indi iduals o co ec
o possible plan mo phological e ec s in he expe imen al esul s.
Pollina o censuses
We conduc ed 15 2-minu e pollina o censuses on each o he 45 plan s in bo h lowe colo
mo ph popula ions o e 3 days. Acco ding o p e ious s udies, he obse a ion sampling
e o (30 min pe plan , 1,350 min in each popula ion) is app op ia e o ob ain an adequa e
ep esen a ion o he pollina o spec a o his species and in his geog aphic a ea (see
Ma e ials and Me hods sec ion in Veiga e al., 2015;Sob al e al., 2015). Fo each census,
he species ha accessed he lowe s and he numbe o lowe s hey isi ed we e eco ded.
E e y day and be o e he census, he numbe o open lowe s on each plan was coun ed
o co ec o he e ec o lo al display in pollina o a ac ion. Plan censuses we e
andomly conduc ed unde sunny condi ions be ween 0800 and 1800 h. The numbe o
isi s pe plan ( isi a ion a e pe 30 min) was used as a measu e o pollina ion success.
A he end o each census, a sample o pollina o axa obse ed was collec ed and sen o
u he iden i ica ion by en omological specialis s om he Uni e si y o O iedo (Spain).
Co ec ing any disc epancies be ween ou isual iden i ica ions o bumblebee species
and he lab de e mina ions, species we e g ouped o analyses in o eigh mo phological
ca ego ies based on p oboscis leng h (Obeso, 1992). The eigh mo phological g oups
we e: g oup 1 (Bombus e es is, B. luco um); g oup 2 (B. ho o um, B. jonellus); g oup 3
(B. p a o um, B. so oensis anca icus, B. lapida ius decipiens); g oup 4 (Bombus wu lenii);
Gui ián e al. (2017), Pee J, DOI 10.7717/pee j.2882 5/15

g oup 5 (B. mesomelas); g oup 6 (B. pascuo um); g oup 7 (B. (Psi hy us) upes is); g oup 8
(B. hypno um).
S a is ical analyses
Se e al gene alized linea models (GLM) we e pe o med, whe e he numbe o isi s
pe plan was he dependen a iable. The locali y (Anca es and Pon ón), ea men
(con ol, local ansplan , and o eign ansplan ), pollina o g oup (desc ibed ea lie ),
and he in e ac ions among hese ac o s we e included as ixed e ec s. The models we e
pe o med o all pollina o species pooled oge he and o each o he eigh pollina o
g oups independen ly. Residuals o he dependen a iable we e i ed in all models wi h a
Poisson dis ibu ion and a loga i hmic link unc ion.
Addi ionally, pai swise con as s we e pe o med o analyze he di e ences on he
isi a ion a e (numbe o isi s pe plan pe 30 min) by he eigh bumblebee g oups
be ween and wi hin he ansplan ea men s and con ol g oup a e aged o bo h
des ina ion locali ies. To es o local adap a ion, he mean di e ences on isi a ion a e
o g oup 1 and 3 bumblebees ( he mos abundan g oups) we e compa ed be ween each
pai o ea men s applied o he ansloca ion expe imen ( o eign ansplan s, local
ansplan s, con ols), conside ing bo h locali ies (Anca es and Pon ón). All analyses we e
pe o med using SPSS S a is ics 20 (IBM Co p., Some s, NY).
RESULTS
Pollina o censuses
In o al 951 insec s (99% bumblebees belonging o he genus Bombus, and he emaining
1% belongs o he genus Apis o he amily Vespidae) isi ed he plan s (538 in Anca es,
and 413 in Pon ón), which ep esen ed 3,268 o al cumula i e lowe isi s (see Da a S1).
Bombus e es is (g oup 1) and Bombus p a o um (g oup 3) we e he main lowe isi o
o Gen iana lu ea plan s in Pon ón, wi h 447 and 372 o al cumula i e isi s espec i ely o
each g oup (excluding con ol ea men , see Fig. 3). Bombus so oensis anca icus +Bombus
lapida ius decipiens (g oup 3) and Bombus (Psi hy us) upes is (g oup 7) we e he main
lowe isi o s in Anca es (338 and 224 o al cumula i e isi s espec i ely o each g oup,
only o ansplan ea men s), being he la es g oup absen in Pon ón, and Bombus
hypno um (g oup 8) was only p esen in his loca ion (Fig. 3).
Recip ocal ansplan s
In Anca es, a popula ion o o ange mo phs, mo e isi s pe 30 min we e obse ed o
o ange— han o yellow— lowe ing plan s (32.5% e sus 24.7% plan s isi ed, espec i ely
o each colo mo ph); whe eas in Pon ón, a popula ion consis ing o yellow lowe s, he e
was no di e ence in isi a ion a e be ween he wo mo phs (39.2% e sus 41% plan s
isi ed, espec i ely o yellow and o ange mo phs; see Da a S2). Numbe o isi s pe
plan depended on he lo al isi o g oup (P<0.001; Table 1). Addi ionally, he numbe
o isi s pe plan by lo al isi o g oups we e a ec ed di e en ly pe ea men : con ol,
local ansplan , and o eign ansplan (as sugges ed by he signi ican ea men * lo al
isi o s in e ac ion; P<0.05; Table 1), and also by locali y (as sugges ed by he signi ican
ea men * lo al isi o s*locali y in e ac ion; P<0.05; Table 1).
Gui ián e al. (2017), Pee J, DOI 10.7717/pee j.2882 6/15
Figu e 3 To al cumula i e numbe o isi s o he Gen iana lu ea ansplan ed plan s (black-colo ed
ba s, o eign; g ey-colo ed ba s, local) by each g oup o bumblebees in he wo s udy si es a No h-
e n Spain (Anca es wi h he o ange mo ph and Pon ón wi h he yellow mo ph). Abb e ia ions: T ans-
o eign, o eign ansplan ; T ans-local, local ansplan . Flo al isi o g oups we e he ollowing: g oup 1,
Bombus e es is,B. luco um; g oup 2, B. ho o um,B. jonellus; g oup 3, B. p a o um,B. so oensis anca i-
cus,B. lapida ius decipiens; g oup 4, B. wu lenii; g oup 5, B. mesomelas; g oup 6, B. pascuo um; g oup 7, B.
(Psi hy us) upes is; g oup 8, B. hypno um.
Resul s o he GLM o analyze he locali y and ea men e ec s on he numbe o isi s
pe plan o each pollina o g oup showed di e en lowe colo p e e ences, bu only
in he g oups 1 (B. e es is) and 3 (B. p a o um,B. so oensis anca icus +B. lapida ius
decipiens; signi ican ea men ac o ; g oup 1: P<0.05; g oup 3: P<0.01; Table 2).
G oup 1 and g oup 3 pollina o s isi ed con ol and local plan s simila ly in bo h colo
mo ph popula ions (Table 3 and Fig. 4).
G oup 1 isi ed o eign ansplan indi iduals mo e o en han local ansplan
indi iduals, conside ing bo h loca ions (signi ican mean di e ences be ween o eign and
local ansplan s; P<0.05; Table 3). G oup 1 pollina o s isi ed con ol and local plan s
simila ly in bo h colo mo ph popula ions (Table 3 and Fig. 4A). Howe e , he absolu e
numbe o isi s by hese pollina o s was highe in Pon ón (yellow mo ph popula ion)
han in Anca es (o ange mo ph popula ion).
G oup 3 isi ed local ansplan s mo e o en han o eign ansplan s in bo h loca ions
(no signi ican ea men *locali y in e ac ion; Table 2). The numbe o isi s by g oup 3 o
con ol and o eign plan s was equal conside ing bo h popula ions (Table 3 and Fig. 4B).
Once again, he absolu e numbe o isi s by hese pollina o s was highe in Pon ón
(yellow- lowe ed popula ion) han in Anca es (o ange- lowe ed popula ion).
Gui ián e al. (2017), Pee J, DOI 10.7717/pee j.2882 7/15
Table 1 Resul s o he gene alized linea model o analyze he e ec o colo mo phs o Gen iana lu ea
on he isi a ion a e (numbe o isi s pe plan pe 30 min) by eigh bumblebee g oups in wo a eas
o No he n Spain ( he o ange colo mo ph was p esen a he Anca es si e, whe eas he yellow mo ph
was p esen a he Pon ón si e). We ma ked in bold he s a is ically signi ican ac o s (P<0.05). Fac o s:
locali y (Anca es and Pon ón, o ange and yellow mo ph popula ion espec i ely); ea men (con ol, lo-
cal ansplan s, and o eign ansplan s); and lo al isi o g oups (g oup 1, Bombus e es is,B. luco um;
g oup 2, B. ho o um,B. jonellus; g oup 3, B. p a o um,B. so oensis anca icus,B. lapida ius decipiens; g oup
4, B. wu lenii; g oup 5, B. mesomelas; g oup 6, B. pascuo um; g oup 7, B. (Psi hy us) upes is; g oup 8, B.
hypno um).
Fac o s Wald Chi-Squa e d. .P
Locali y 0.702 1 0.402
T ea men 0.110 2 0.946
T ea men * Locali y 4.336 2 0.114
Flo al isi o s 66.548 3 <0.001
T ea men * Flo al isi o s 16.703 6 0.010
T ea men * Flo al isi o s * Locali y 19.573 9 0.021
Table 2 Resul s o he gene alized linea model o analyze he e ec o locali y, ea men , and he co -
esponding in e ac ion be ween bo h, on he isi a ion a e (numbe o isi s pe plan pe 30 min) by
wo bumblebee g oups (g oup 1 and 3) in wo a eas o No he n Spain (Anca es and Pon ón, he o -
ange and yellow mo ph popula ion espec i ely). We ma ked in bold he s a is ically signi ican ac o s
(P<0.05). Fac o s: locali y (Anca es and Pon ón, o ange and yellow mo ph popula ion espec i ely);
ea men (con ol, local ansplan s, and o eign ansplan s); and lo al isi o g oups (g oup 1, Bom-
bus e es is,B. luco um; g oup 3, B. p a o um,B. so oensis anca icus,B. lapida ius decipiens). No e ha we
only included esul s o he isi o g oups ha showed signi ican model e ec s (G oup 1, G oup 3).
Flo al isi o s Fac o s Wald Chi-Squa e d. .P
Locali y 32.222 1 <0.001
T ea men 6.181 2 0.045
G oup 1
T ea men * Locali y 0.692 2 0.707
Locali y 1.867 1 0.172
T ea men 16.331 2 <0.001
G oup 3
T ea men * Locali y 0.114 2 0.945
DISCUSSION
Ou esul s show ha Gen iana lu ea plan s om bo h loca ions ace di e en se s o lowe
isi o s wi h di e en colo p e e ences—indica ed by di e ences in isi a ion a e—
sugges ing ha hese di e ences may play a ole in lo al colo di e gence pa e ns. We
ound ha he o ange- lowe ing plan s may be locally adap ed o he pollina o assemblage
since hey ecei ed mo e isi s han he yellow- lowe ing ansplan s in Anca es, he
popula ion o o ange mo phs. Howe e , we ound no e idence o local adap a ion o
yellow- lowe ing plan s o he o iginal popula ion since he numbe o bumblebee isi s
was simila o he o ange- lowe ing ansplan s in Pon ón. This esul di e s om p e ious
esea ch, which sugges ed ha lowe colo a ia ion among G. lu ea popula ions is ela ed
o selec i e p essu es exe ed by di e en pollina o assemblages (Sob al e al., 2015).
Se e al possible explana ions may help o cla i y ou indings. The di e en esponses
ob ained in he Anca es and Pon ón popula ions a e likely due o a ia ion in he pollina o
Gui ián e al. (2017), Pee J, DOI 10.7717/pee j.2882 8/15
Table 3 Resul s o pai wise con as s o analyze he mean di e ences on he isi a ion a e (numbe
o isi s pe plan pe 30 min) by bumblebee g oups 1 and 3, a e aged o wo a eas o No he n Spain
(Anca es and Pon ón, he o ange and yellow mo ph popula ion espec i ely) and pe ea men (con-
ol, local ansplan , o eign ansplan ). We ma ked in bold he s a is ically signi ican mean di e ences
be ween he pai o ea men s (P<0.05). Abb e ia ions: T ans-local, local ansplan s; T ans- o eign,
o eign ansplan s; and con ol, unmanipula ed g oups. No e ha we only included esul s om pai wise
con as s ha showed signi ican mean di e ences be ween pai o ea men s (i.e., pollina o g oups 1
and 3).
Flo al isi o s Pai o ea men s Mean-di e ence S.E. d. .P
Con ol T ans- o eign −5.090 2.801 1 0.069
Con ol T ans-local 1.390 2.331 1 0.551
G oup 1
T ans- o eign T ans-local 6.480 2.733 1 0.018
Con ol T ans- o eign 18.720 4.369 1 <0.001
Con ol T ans-local 8.210 4.992 1 0.100
G oup 3
T ans- o eign T ans-local −10.510 3.736 1 0.005
assemblage and/o lo al isi o p e e ences be ween he wo a eas (as expe imen ally
demons a ed, Jones & Rei hel, 2001). Anca es and Pon ón ha e di e en pollina o
assemblage and he species om he Anca es popula ion may p e e local o ange- lowe ing
plan s, while he species om Pon ón popula ion clea ly show no p e e ence o one
o he wo colo mo phs. Geog aphic a ia ion in lo al isi o p e e ences can esul
om changes in pollina o composi ion, abundance, and di e si y (P ice e al., 2005),
e en di e en plan popula ions ha sha e he same pollina o g oup ecei e di e en
p opo ions o lowe isi s om each species o unc ional g oup (Fens e e al., 2004;
Tas a d e al., 2014). Thus, pollina o -dependen o aging p e e ences may cause shi s in
he op imal lo al pheno ypes (see Gómez e al., 2009; and e e ences he ein).
Wi hin he pool o G. lu ea lowe isi o s in he Anca es popula ion, he mos abundan
species (Bombus e es is) has low sensi i i y o ed colo de ec ion (Pei sch e al., 1992;
B iscoe & Chi ka, 2001), which p obably explains he educ ion in his species isi a ion
a e when inc easing o ange- lowe ing indi iduals (Fig. 4A). We suspec ha o he
pollina o species wi h p obably highe sensi i i y o ed colo may inc ease i ness o
he o ange- lowe ing plan s. The e o e, he ac ha hey pe cei e ed colo does no imply
ha hey necessa ily would p e e his colo o e o he s. In ac , B. lapida ius, p esen
in o he o ange G. lu ea (JA Gui ián, pe s. obs., 2012), shows sensi i i y o ed colo s;
al hough i lacks ed colo ecep o s (Kugle , 1943;Pei sch e al., 1992). I pollina o species
wi h a highe sensi i i y o inna e p e e ences o ed colo s a e p esen , o ange- lowe ed
plan s would inc ease hei i ness h ough he ising isi a ion a e by hese pollina o s.
The e o e, a ia ion o he pollina o communi y composi ion migh a ec he selec i e
p essu es exe ed on G. lu ea lowe colo , and ul ima ely a ec lowe colo a ia ion
among popula ions ia local adap a ion.
A second explana ion is ha he p e e ence o ce ain pollina o g oups could be
dependen on he equency o each mo ph in he popula ion (i.e., he p e e ences o
o ange plan s in Anca es a e dilu ed in a con ex o yellow plan s in Pon ón). Empi ical
s udies ha examine plan species polymo phisms in lowe colo , mo phology, o sex
Gui ián e al. (2017), Pee J, DOI 10.7717/pee j.2882 9/15