Telome e dynamics in pa asi ic g ea spo ed cuckoos and hei
magpie hos s
J. J. SOLER*, C. RUIZ CASTELLANO*, J. MARTI
´NEZ-DE LA PUENTE†,G.TOM

AS*,
M. RUIZ-RODRI
´GUEZ* & J. FIGUEROLA†
*Depa amen o de Ecolog
ıa Funcional y E olu i a, Es aci
on Expe imen al de Zonas

A idas (CSIC), Alme 
ıa, Spain
†Depa amen o de Ecolog
ıa de Humedales, Es aci
on Biol
ogica Do~
nana (CSIC), Se illa, Spain
Keywo ds:
b ood pa asi ism;
cell di ision;
incuba ion pe iod;
li e his o y;
nes ling g ow h;
oxida i e damage;
elome ase ac i i y.
Abs ac
Al hough li le is known on he impac o en i onmen on elome e leng h
dynamics, i has been sugges ed o be a ec ed by s ess, li es yle and/o li e-
his o y s a egies o animals. We he e compa ed elome e dynamics in e y-
h ocy es o ha chlings and ledglings o he b ood pa asi e g ea spo ed
cuckoos (Clama o glanda ius) and o magpies (Pica pica), hei main hos in
Eu ope. In magpie chicks, elome e leng h dec eased om ha ching o ledg-
ing, whe eas no signi ican change in elome e leng h o g ea spo ed
cuckoo chicks was ound. Mo eo e , we ound in e speci ic di e ences in
he associa ion be ween laying da e and elome e sho ening. In e speci ic
di e ences in elome e sho ening we e in e p e ed as a consequence o di -
e ences in li es yle and li e-his o y cha ac e is ics o magpies and g ea spo -
ed cuckoos. In compa ison wi h magpies, cuckoos expe ience educed
sibling compe i ion and highe access o esou ces and, consequen ly, lowe
s ess ul en i onmen al condi ions du ing he nes ling phase. These cha ac-
e is ics also explain he associa ions be ween elome e a i ion and en i-
onmen al condi ions (i.e. laying da e) o magpies and he absence o
associa ion o g ea spo ed cuckoos. These esul s he e o e i expec a ions
on elome e dynamics de i ed om in e speci ic di e ences in li es yle and
li e his o y o b ood pa asi es and hei bi d hos s.
In oduc ion
En i onmen al condi ions expe ienced du ing he i s
days o li e may ha e impo an consequences la e ,
du ing adul hood, in e ms o su i al p ospec s (Mon-
aghan, 2008). Recen ly, i has been sugges ed ha hese
delayed e ec s a e a leas pa ially d i en by he in lu-
ence o ea ly en i onmen al condi ions on elome e
dynamics (Bee y e al., 2012) and suppo ing e idences
ha e been accumula ed du ing he las ew yea s (Hall
e al., 2004; Heidinge e al., 2012; Boonekamp e al.,
2014; He bo n e al., 2014; Ne le e al., 2015). Telom-
e es a e specialized s uc u es a he end o he ch omo-
some, which consis on sho s epea s o he noncoding
DNA sequence TTAGGG ha , among o he unc ions,
p o ec wholeness o gene ic in o ma ion du ing cell
di ision (Blackbu n, 1991). Telome e a i ion has adi-
ionally been ela ed wi h ageing (Sahin & DePinho,
2010) and is closely ela ed o s ess e en s o di e en
na u e ha include hose associa ed wi h cell di ision
(i.e. g ow h) and wi h ele a ed concen a ions o eac-
i e oxygen species (Monaghan & Haussmann, 2006).
Ageing, li es yle, li espan and li e-his o y cha ac e is-
ics o animals a e closely ela ed in na u e (Monaghan
& Haussmann, 2006); o example, animals wi h ela-
i e sho li espan g ow as e and ep oduce ea lie a
he cos o inc easing me abolic and cellula degene a-
ion a es (Rickle s & Finch, 1995), which would a ec
elome e dynamics. Telome e leng h and dynamics a e
conside ed good indica o s o i ness in e ms o su -
i al p ospec o he adop ed li es yle and/o li e-his o y
s a egies (Monaghan & Haussmann, 2006; Ko schal
e al., 2007). Thus, de ec ing e idence linking li es yle
and/o li espan wi h elome e leng h and sho ening is
o p ime impo ance o unde s and he balance o cos s
Co espondence: Juan J. Sole , Depa amen o de Ecolog
ıa Funcional y
E olu i a, Es aci
on Expe imen al de Zonas

A idas (CSIC), C a.
Sac amen o s/n, La Ca~
nada de San U bano, Alme 
ıa, Spain. Tel.: +34
950281045; ax: +34 950277100; e-mail: [email p o ec ed]
ª2015 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY. J. EVOL. BIOL.
1
JOURNAL OF EVOLUTIONARY BIOLOGY ª2015 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY
doi: 10.1111/jeb.12680
and bene i s associa ed wi h species-speci ic li e his o-
ies (Salomons e al., 2009b; Monaghan, 2014). Pa icu-
la ly in e es ing is he s udy o elome e dynamics
du ing he i s ew days o li e o al icial bi ds o se -
e al easons. Na u al selec ion and elome e sho ening
is pa icula ly appa en a his s age due o e y high
cell di ision a e (Salomons e al., 2009b). Mo eo e ,
he ela i ely down- egula ion o elome ase ac i i y
(i.e. educed elome e epai ) du ing he exponen ial
g ow h s age (Fo sy h e al., 2002; bu see Haussmann
e al., 2007) makes easie he de ec ion o changes in
elome e leng h o explo e he p edic ed associa ions
wi h li e-his o y cha ac e is ics o en i onmen al condi-
ions.
In addi ion, en i onmen al condi ions expe ienced by
nes lings la gely a y in associa ion wi h laying da e,
wi h la e ha chlings ypically de eloping in wo se, mo e
s ess ul condi ions han ea ly ha chlings, due o poo
pa en al quali y o b eede s and/o de e io a ed
esou ce a ailabili y (De Ne e e al., 2004; Ve huls &
Nilsson, 2008). Fu he , he s eng h o he associa ion
be ween laying da e and esou ce a ailabili y o de el-
oping nes lings depends on species-speci ic li e-his o y
ai s (S ensson, 1995) and, hus, we should expec
elome e dynamics o a y wi h laying da e (Foo e
e al., 2011), a leas o some species. As a as we
know, his p edic ed associa ion has ne e been es ed
in he wild.
Finally, sibling compe i ion o pa en al ca e is one
impo an cause o physiological s ess (Ma 
ınez-Padilla
e al., 2004; Blanco e al., 2006) likely a ec ing elome e
dynamics du ing g ow h. Consequen ly, p e ious s ud-
ies ha e ound ha he s eng h o in ab ood compe i-
ion o ood in jackdaw (Co us monedula) (Salomons
e al., 2009a) and Eu opean s a ling (S u nus ulga is)
(Ne le e al., 2013) nes lings was posi i ely ela ed wi h
elome e a i ion du ing hei i s ew days o li e.
Thus, because he s eng h o sibling compe i ion
widely a ies wi hin and among bi d species (Mock &
Pa ke , 1997), i can be expec ed ha nes lings o spe-
cies wi h in ense sibling i al y had be e mechanisms
o cope wi h s ess and o educe elome e sho ening,
han hose o species wi h low sibling compe i ion. A
he in aspeci ic le el, elome e leng h and body size o
ba n swallow (Hi undo us ica) nes lings posi i ely
co a ied (Cap ioli e al., 2013), and elome e a i ion
due o sibling compe i ion was nega i ely ela ed wi h
nes ling hie a chy in Eu opean s a lings (Ne le e al.,
2013, 2015); bo h esul s sugges ing he possibili y o
physiological adjus men depending on nes ling hie a -
chy (i.e. body mass) ha could e en be media ed by
ma e nal e ec s. Mo eo e , expe imen ally inc eased
b ood size also esul ed in la ge elome e a i ion in
zeb a inch (Taeniopygia gu a a) pa en s (Reiche e al.,
2014). Thus, species di e ing in he s eng h o in a-
b ood compe i ion o ood may also di e in elome e
dynamics du ing g ow h.
Wi h he aim o con ibu ing o unde s and ac o s
explaining elome e dynamics o de eloping animals,
we he e explo ed he associa ion be ween elome e
leng h o ha chlings and ledglings o magpies (Pica
pica), and o hei b ood pa asi ic g ea spo ed cuckoos
(Clama o glanda ius), in ela ion o laying da e. Explo -
ing hese associa ions in his b ood pa asi e–hos sys em
has se e al ad an ages. Fi s , magpies a e he main
Eu opean hos o g ea spo ed cuckoos, and he e o e,
nes lings o bo h species a e ea ed by magpie adul s
(Sole e al., 1999; Sole & Sole , 2000). Second, en i-
onmen al condi ions expe ienced by de eloping g ea
spo ed cuckoo and magpie nes lings usually di e , he
la e de eloping in a mo e compe i i e en i onmen
han he o me . G ea spo ed cuckoo nes lings usually
ou compe e os e siblings soon a e ha ching. Mo e-
o e , pos -ha ching g ow h a e o g ea spo ed cuck-
oos is much highe han ha o magpie hos s (Sole &
Sole , 1991). Consequen ly, i elome e dynamics
depend on cell di ision a es, ha o g ea spo ed cuck-
oos and magpies should di e . Finally, because g ea
spo ed cuckoo nes lings usually sha e he nes s wi h
ewe chicks han magpies do, i is likely ha he
expec ed e ec s o en i onmen al condi ions ela ed o
esou ce a ailabili y (i.e. laying da e (De Ne e e al.,
2004)) on elome e dynamics a e weake o nes lings
o he b ood pa asi ic species.
Ma e ials and me hods
S udy si es and ieldwo k
The s udy was pe o med in he Guadix (37°180N,
3°110W, sou he n Spain) magpie popula ion, si ua ed in
a high-al i ude pla eau (app ox. 1000 m a.s.l.). The
ege a ion o he a ea is spa se, including uncul i a ed
a eas and many g o es o almond (P unus dulcis) and
pine (Pinus halepensis and Pinus pinas e ) ees and oaks
(Que cus o undi olia) in which magpies p e e o build
hei nes s. B ood pa asi ism by g ea spo ed cuckoos is
qui e common in he a ea (Sole e al., 2001, 2013)
and, al hough g ea spo ed cuckoos usually ou com-
pe e magpie os e siblings (Sole & De Ne e, 2013),
pa asi ic nes lings do no e ic hos nes lings a e
ha ching and, he e o e, bo h species may g ow up
oge he in he same nes (Ma 
ın-G
al ez e al., 2011).
Fieldwo k was conduc ed du ing he 2013 b eeding
season. A he beginning o sp ing in Ma ch–Ap il, we
in ensely sea ched o nes s o magpies ha we e e is-
i ed egula ly o de e mine hei laying da e. Expec ed
ha ching da e o magpies and g ea spo ed cuckoos was
es ima ed as 25 and 19 days, espec i ely, a e laying
da e o he i s magpie egg. Magpie nes s we e hen
isi ed du ing hese and subsequen days up o sam-
pling magpie ha chlings (i.e. 0–1 day old) and, in he
case o pa asi ized nes s, g ea spo ed cuckoo ha ch-
lings. Because ma e nal and gene ic ac o s a e known
ª2015 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY. J. EVOL. BIOL. doi: 10.1111/jeb.12680
JOURNAL OF EVOLUTIONARY BIOLOGY ª2015 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY
2JUAN J. SOLER ET AL.
o de e mine ea ly li e elome e leng h (Asgha e al.,
2015b), we collec ed a d op o blood om he b achial
ein o wo andomly selec ed magpie ha chlings and
om all cuckoo ha chlings in a magpie nes . All mag-
pies and g ea spo ed cuckoo nes lings in he nes s 18
and 16 days a e ha ching, espec i ely, we e again
blood-sampled.
To inc ease he numbe o pa asi ized nes s wi h
magpies and g ea spo ed cuckoo ledglings g owing up
oge he , we expe imen ally pa asi ized 11 magpie nes s
wi h a single cuckoo egg close o ha ching ime om
mul ipa asi ized magpie nes s. In h ee o hese nes s,
only he cuckoo ledged [cuckoos ha ched 3.33
(SE =0.67) days be o e magpies]; in one, only magpies
( ou magpie ledglings ha ha ched 2 days in ad ance
o cuckoos); and in se en nes s, bo h he g ea spo ed
cuckoo and magpies (a e age numbe o ledglings pe
nes =2.6, SD =0.98) ledged [cuckoos ha ched 2.14
(SE =0.86) days be o e magpies]. In addi ion, we sam-
pled 27 na u al magpie nes s. F om eigh o hese nes s,
only cuckoos ledged (mean =1.13, SD =0.35), om
18 only magpies ledged (mean =4.5, SD =1.04), and
om he las nes , one g ea spo ed cuckoo and one
magpie success ully eached he ledging s age. How-
e e , duplica ed es ima es o elome e leng h o some
samples we e no consis en ly simila (i.e. CV >5%,
see below) and we e no conside ed in he analyses.
We hus collec ed in o ma ion wi h adequa e es ima es
o elome e a i ion o 36 magpie nes s. In eigh o
hese nes s, magpies and g ea spo ed cuckoos g ew
oge he , bu in o ma ion o magpies and cuckoo nes -
lings was only a ailable o ou o hem. Fo he o he
ou nes s wi h bo h species, only in o ma ion o mag-
pie nes lings was a ailable o he analyses. In addi ion,
we collec ed in o ma ion on elome e a i ion o 18
and 10 nes s, espec i ely, whe e only chicks o one
species, ha is ei he magpies o g ea spo ed cuckoos,
g ew up. Fo some addi ional nes s, in o ma ion o
elome e leng h was only a ailable o ha chling o o
ledglings. Thus, o cuckoo nes lings, inal sample size
was 18 o elome e leng h o ha chling and 14 o
elome e a i ion. Fo magpie nes lings, inal sample
size used in he analyses was 26 and 27 o elome e
leng h o ha chlings and ledglings, espec i ely, and 26
o elome e a i ion.
Labo a o y wo k
DNA was ex ac ed om blood samples using a s an-
da d chlo o o m–isoamyl alcohol-based p o ocol (see
Fe agu i e al., 2013). DNA concen a ion was adjus ed
o 20 ng lL
1
using dis illed wa e and conse ed o-
zen un il u he analyses. Rela i e elome e leng h
(he ea e elome e leng h) was es ima ed by q-PCR ol-
lowing he p o ocol and p ime s o C iscuolo e al.
(2009). As con ol single-copy gene we used he glyce -
aldehyde-3-phospha e dehyd ogenase (GAPDH).
GAPDH was used o no malize he quan i y o elome e
sequence o he amoun o DNA in he q-PCR. The
inal PCR olume was 20 lL con aining 10 lLo
Ligh Cycle 480 SYBR G een I Mas e (Roche Diagnos-
ics GmbH, Mannheim, Ge many) and 1 lL o DNA a
20 ng lL
1
o DNA. The eac ions o elome es o
GAPDH we e done in di e en pla es due o he di e -
en ial PCR condi ions. Telome e PCR condi ions we e
10 min a 95 °C ollowed by 30 cycles o 1 min a
56 °C and 1 min a 95 °C. GAPDH PCR s a ed wi h
10 min a 95 °C ollowed by 40 cycles o 1 min a
60 °C and 1 min a 95 °C, bo h pe o med in a
Ligh Cycle 480 RT-PCR Sys em (Roche). Each sample
was un in duplica e, and samples wi h a coe icien o
a ia ion highe o 5% we e emo ed om he analy-
ses. Each 96-well pla e included se ial dilu ions o DNA
(40 ng, 10 ng, 2.5 ng, 0.66 ng o DNA pe well) om a
e e ence pool ( he in e nal con ol) un in iplica e,
which we e used o gene a e he s anda d cu es, and
a blank con ol wi h no DNA. Quan i ica ion cycle al-
ues (C ) we e ans o med in o no malized ela i e
quan i ies (NRQs) ollowing Hellemans e al. (2007)
p ocedu e, which con ols o he ampli ying e iciency
o each qPCR. Ampli ica ion e iciency o elome e
p oduc s anged be ween 1.85 and 2.01 and o he
GAPDH p oduc be ween 1.88 and 2.14. The slope o
he calib a ion cu e anged be ween -3.792 and -3.277
o he elome e p oduc and -3.647 and -3.031 o he
GAPDH p oduc . The mel ing cu es o he con ol gene
cycles con i med no e idence o p ime dime o non-
speci ic ampli ica ion. Di e en echniques a e a ailable
o measu ing elome e leng h in wildli e ( e iewed in
Nussey e al., 2014). Each o hese echniques has
ad an ages and limi a ions ha may a ec he eliabil-
i y o he esul s. In he case o q-PCR, an impo an
limi a ion is ha in e s i ial elome ic sequences may
occu on he genome and consequen ly may bias es i-
ma es o ela i e elome e leng h. Consequen ly, q-PCR
es ima ed elome e leng hs may be no adequa e o
compa e elome e leng h among species (Nussey e al.,
2014). Howe e , i is adequa e o compa e pa e ns o
a ia ion ac oss species based on epea ed measu es o
he same indi iduals ac oss ime, as we ha e done in
his s udy ( o a simila app oach, see Asgha e al.,
2015a). Consequen ly, we will no deal wi h in e speci-
ic di e ences in elome e leng h bu compa e he em-
po al and de elopmen al pa e ns o a ia ion in
elome e leng h be ween magpies and cuckoos.
S a is ical analyses
F equency dis ibu ion o NRQ alues es ima ed o
nes lings and ledglings o cuckoos and magpies did no
di e om no mal dis ibu ions (Kolmogo o –Smi no
es o con inuous a iables, P>0.05). Mo eo e ,
be ween-nes a ia ion in elome e leng h o magpie
ledglings was signi ican ly la ge han wi hin-nes a i-
ª2015 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY. J. EVOL. BIOL. doi: 10.1111/jeb.12680
JOURNAL OF EVOLUTIONARY BIOLOGY ª2015 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY
B ood pa asi ism and elome e dynamics 3
a ion (R=0.67, F
26,77
=4.99, P<0.0001); hus, we
used wi hin-nes mean alues o subsequen analyses.
Telome e a i ion was hus es ima ed as he di e ence
be ween a e age alues a he ime o ha ching and a
he ime o ledging. Laying da e did no di e om
no mal dis ibu ion (Kolmogo o –Smi no es o con-
inuous a iables, P>0.05), and hus, we used pa a-
me ic s a is ical es s o compa ing elome e leng h
and a i ion o g ea spo ed cuckoos and magpies and
i s ela ionship wi h hei espec i e laying da e.
Whe he o no cuckoo o magpie chicks g ew up
oge he wi h nes lings o he o he species did no a ec
elome e leng h and elome e a i ion o ledgling cuck-
oos [sample size: N(only cuckoos) =10, N(cuckoos and
magpies) =4, <1.02, P>0.33] o o magpie nes lings
[sample size: N(only magpies) =19, N(cuckoos and
magpies) =8, <0.35, P>0.73]. Thus, his ac o was
no conside ed in u he analyses. Some nes lings o
bo h species died la e du ing he nes ling phase be o e
he second blood sampling, and hus, sample sizes o
g ea spo ed cuckoos and magpies in pa asi ized nes s
ea ing nes lings o bo h species do no coincide.
Telome e dynamics o g ea spo ed cuckoos and mag-
pies we e i s explo ed sepa a ely o each species by
means o epea ed-measu es ANOVAs wi h es ima es om
he same nes a di e en imes as wi hin ac o and b ood
size and laying da e as co a iables. In e speci ic di e -
ences in he associa ion be ween laying da e and ei he
elome e leng h a di e en nes s age (i.e. ha ching and
ledging) o elome e sho ening we e explo ed wi h AN-
COVAs wi h species iden i y as he ca ego ical independen
ac o , laying da e o he i s magpie egg as he co a iable
and he in e ac ion be ween laying da e and species iden-
i y as he ac o in o ming on he in e speci ic di e -
ences. Rela ionships be ween laying da e and ei he
elome e leng h o elome e sho ening we e explo ed
using Pea son co ela ions.
All s a is ical es s we e pe o med in S a is ica 10.0
(S a so Inc., 2011).
Resul s
As magpie nes lings expe ienced a ela i ely d as ic
elome e sho ening du ing he nes ing pe iod
( epea ed-measu es ANOVA,F
1,25
=45.65, P<0.0001,
Fig. 1), his was no he case o cuckoo nes lings
( epea ed-measu es ANOVA,F
1,13
=0.19, P=0.67, Fig. 1;
in e speci ic compa isons o elome e sho ening, in e -
ac ion be ween de eloping ime and species iden i y,
F
1,38
=16.68, P=0.0002, Fig 1). O e all, elome e
leng h o magpies declined by 64.3% om ha ching o
ledging. Including he nonsigni ican e ec s o laying
da e and b ood size (P>0.28) in he s a is ical models
explo ing in e speci ic di e ences did no change any
conclusions.
The associa ion be ween elome e leng h and laying
da e di e ed o g ea spo ed cuckoo and magpie
ha chlings (ANCOVA, in e ac ion be ween species iden i y
and laying da e, F
1,40
=7.48, P=0.009, Fig. 2). Telom-
e e leng h o magpie ha chlings showed a dec ease in
ela ion o laying da e (R=0.44, =2.37, N=26,
P=0.026, Fig. 2), whe eas elome e leng h o cuckoo
ha chlings was posi i ely ela ed o laying da e
(R=0.49, =2.28, N=18, P=0.037, Fig. 2).
A he ime o ledging, he associa ion be ween elom-
e e leng h and laying da e o g ea spo ed cuckoos and
magpies also di e ed signi ican ly (ANCOVA, in e ac ion
be ween species iden i y and laying da e, F
1,41
=6.40,
P=0.015, Fig. 3). Telome e leng h o ledgling magpies
inc eased as he season p og essed (R=0.49, =2.83,
N=27, P=0.009, Fig. 3), e en a e con olling o he
e ec o elome e leng h a ha ching (laying da e: pa ial
Fig. 1 Mean con idence in e als (95%) o elome e leng h
(NRQ alues) o g ea spo ed cuckoo and magpie ha chlings and
ledglings. Lines connec ing alues o he same species indica e
elome e dynamics du ing he nes ing phase.
Fig. 2 Rela ionships be ween elome e leng h a ha ching ime
and laying da e o magpies (open ci cles, con inuous lines) and
g ea spo ed cuckoos ( ull ci cles, do ed line). Values a e mean
nes alues and lines a e eg ession lines.
ª2015 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY. J. EVOL. BIOL. doi: 10.1111/jeb.12680
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4JUAN J. SOLER ET AL.
R=0.50, P=0.021; elome e leng h a ha ching: pa ial
R=0.02, P=0. 93). This end was no de ec ed in cuck-
oos o which la e b oods had nonsigni ican smalle
elome es han hose o ea ly b oods (R=0.30,
=1.25, N=18, P=0.23, Fig. 3) e en a e con olling
o he e ec o elome e leng h a ha ching (laying da e:
pa ial R=0.01, P=0.96; elome e leng h a ha ching:
pa ial R=0.27, P=0.36).
The abo e epo ed associa ions be ween laying da e
and elome e leng h o ha chlings and ledglings o
g ea spo ed cuckoos and magpies sugges ha he
associa ions be ween elome e a i ion and laying da e
should also di e o magpie and g ea spo ed cuckoo
nes lings (ANCOVA, in e ac ion be ween species iden i y
and laying da e, F
1,36
=10.15, P=0.003, Fig. 4).
Telome e a i ion du ing he nes ing phase was nega-
i ely associa ed wi h laying da e o magpie nes lings
(R=0.53, =3.10, N=26, P=0.005, Fig. 4) e en
a e con olling o he e ec o elome e leng h a
ha ching (laying da e: pa ial R=0.46, P=0.021;
elome e leng h a ha ching: pa ial R=0.95,
P<0.0001). The posi i e associa ion de ec ed o g ea
spo ed cuckoos (R=0.44, =1.69, N=14, P=0.12,
Fig. 4) was a om s a is ical signi icance a e con ol-
ling o elome e leng h a ha ching (laying da e: pa ial
R=0.01, P=0.96; elome e leng h a ha ching: pa -
ial R=0.69, P=0.008).
Discussion
Ou main inding is ha elome e leng h o magpie
nes lings is d as ically educed du ing de elopmen a
he nes , whe eas no changes occu in cuckoo nes lings.
This sugges s ha magpies a e su e ing highe le els o
s ess han cuckoos o ha cuckoos can deal wi h such
s ess be e han magpies. Mo eo e , we ound in e -
speci ic di e ences in he associa ion be ween laying
da e and elome e leng h and sho ening. The s onge
associa ions we e de ec ed in magpies o which la e
ha chlings had he sho e elome es, whe eas elome e
a i ion was s onge o magpie nes lings o ea ly
b oods. Below we discuss hese esul s in he con ex s
o in e speci ic di e ences in de elopmen al s a egies
o g ea spo ed cuckoos and magpies ha may a ec
elome e leng h and en i onmen al in luences on he
dynamics o elome e change.
In e speci ic di e ences in elome e leng h along
he nes ling pe iod
When g ea spo ed cuckoos success ully pa asi ize mag-
pie nes s be o e he onse o incuba ion, he accele a ed
emb yonic de elopmen al a es o pa asi es ela i e o
hose o hei hos s con e pa asi ic o sp ing wi h huge
ad an ages when compe ing o ood wi h hos nes -
lings (Sole & Sole , 1991). In hese pa asi ized nes s,
hos nes lings ha ch i e o six days a e cuckoos
ha ch, a a ime when pa asi ic os e siblings weigh
mo e han 60 g, which is en imes mo e han he
weigh o magpie ha chlings (J.J. Sole , unpublished
in o ma ion). Because magpie adul s p e e o eed he
la ges nes ling in he nes (Sole e al., 1995), magpie
o sp ing has no op ion o su i e in hese nes s whe e
success ul pa asi ism occu s be o e incuba ion s a ed
(Sole e al., 1998). Thus, i is likely ha a e paying
he cos s o accele a ed a e o cell di ision du ing hei
sho emb yonic s age, cuckoos will enjoy a peace ul
nes ling pe iod wi h no appa en esou ce limi a ion o
g owing (Sole & Sole , 2000). In con as , magpie nes -
lings should always compe e o ood wi h siblings and
migh he e o e su e inc eased cos s in e ms o elom-
e e a i ion (see In oduc ion). In ag eemen wi h his
Fig. 3 Rela ionships be ween elome e leng h a ledging ime and
laying da e o magpies (open ci cles, con inuous lines) and g ea
spo ed cuckoos ( ull ci cles, do ed line). Values a e mean nes
alues and lines a e eg ession lines.
Fig. 4 Rela ionships be ween elome e a i ion du ing
de elopmen in magpies (open ci cles, con inuous lines) and g ea
spo ed cuckoos ( ull ci cles, do ed line) and laying da e. Values
a e mean nes alues and lines a e eg ession lines.
ª2015 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY. J. EVOL. BIOL. doi: 10.1111/jeb.12680
JOURNAL OF EVOLUTIONARY BIOLOGY ª2015 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY
B ood pa asi ism and elome e dynamics 5

scena io, we ound ha elome es o magpies, bu no
o g ea spo ed cuckoos, dec eased signi ican ly o e
he nes ing pe iod, which is in acco dance wi h di e -
en ial cos s associa ed wi h in ense in ab ood compe i-
ion in e ms o elome e a i ion de ec ed in some
o he species (Salomons e al., 2009b; Ne le e al.,
2013).
Pa icula ly in iguing is he lack o e idence o con-
sis en elome e a i ion o cuckoo nes lings o e he
nes ing pe iod. E en mo e when conside ing ha o
one- hi d o he s udied nes s wi h g ea spo ed cuck-
oos, elome e leng h inc eased a he han dec eased
(Fig. 4), al hough some o hese cases may be explained
by measu emen e o s. Telome e enla gemen o e he
nes ling pe iod has been p e iously de ec ed in s a -
lings, bu because elome ase ac i i y is conside ed o
be down- egula ed in ju eniles as a umou -p o ec ing
mechanism (W igh & Shay, 2001), and i was only
de ec ed in wo indi iduals, hese we e conside ed as
ou lie s due o measu emen e o s (Ne le e al., 2013).
Howe e , con a y o he idea ha elome ase ac i i y
is mainly supp essed in mos soma ic issues o de elop-
ing animals, Haussmann e al. (2007) de ec ed ha i is
main ained h oughou he li espan o bi ds, including
he ha chling age class. Thus, i is possible ha elom-
e ase ac i i y was especially high in g ea spo ed
cuckoo nes lings, coun e ac ing degene a i e e ec s o
he apid g ow h expe ienced du ing he nes ling s age,
possibly e en allowing some eco e y o elome e
leng h a e hei ex emely speedy g ow h du ing he
emb yonic phase (and he p esumably e y high elom-
e e a i ion a e du ing his phase, see abo e). How-
e e , i is also possible ha he in e speci ic di e ences
in elome e ajec o ies we e due o di e ences in he
capaci y o con ol oxida i e s ess (i.e. highe access o
an ioxidan s gi en a highe access o ood o mo e
in ense an ioxidan enzyma ic ac i i y) (Bad
as e al.,
2015).
Finally, because ledgling cuckoos we e sampled
when hey we e wo days younge han magpies, di -
e ence in age migh explain he in e speci ic di e -
ences de ec ed in elome e a i ion. This las possibili y
is, howe e , unlikely as cuckoos de elop as e han
magpies and nes lings o bo h species a e a simila
de elopmen al s age a he age o sampling (Sole &
Sole , 1991).
Telome e leng h and sho ening, and laying da e
Laying da e is one o he mos employed a iables
e lec ing e i o y quali y and a ailabili y o esou ces
and/o pa en al quali y (B inkho e al., 1993; Mo eno,
1998; De Ne e e al., 2004; Ve huls & Nilsson, 2008).
Inc eased nes ling compe i ion educes esou ce a ail-
abili y o g owing nes lings, and de e io a ion o qual-
i y o pa en al ca e is pos -na al s esso s ypically
associa ed wi h laying da e (Pe ins, 1970; B inkho
e al., 1993). The e o e, an inc ease in endogenous glu-
coco icoids is p edic ed in nes lings om la e b eeding
a emp s (Bane jee e al., 2012), which consequen ly
should a ec elome e leng h dynamics (Monaghan,
2014). In acco dance wi h his scena io, we ound ha
la e magpie ha chlings had sho e elome e leng h han
ea ly ha chlings. Howe e , some o ou esul s we e no
in acco dance wi h his hypo hesis as elome e leng h o
g ea spo ed cuckoo ha chlings was posi i ely associa ed
wi h laying da e, and elome e a i ion expe ienced by
magpie nes lings dec eased o e he b eeding season; a
endency no de ec ed in cuckoos. The de ec ed associa-
ions be ween laying da e and ei he elome e leng h o
elome e a i ion did no depend on elome e leng h a
ha ching, and he e o e, hese associa ions canno be
explained by he commonly desc ibed highe elome e
a i ion in indi iduals wi h ini ially longe elome es
(Ma cand e al., 1999; Ka lsede e al., 2002; Hall e al.,
2004). Thus, in e speci ic di e ences in he associa ion
be ween elome e leng h and laying da e s ongly sug-
ges ha ela ed en i onmen al cha ac e is ics do in lu-
ence elome e dynamics o magpie and o g ea spo ed
cuckoo nes lings di e en ly. I is di icul , howe e , o
explain con adic o y esul s o he p oposed scena io,
and we can only specula e abou possible easons. I is,
o ins ance, possible ha he nega i e associa ion
be ween elome e leng h o magpie ha chlings and lay-
ing da e was due o a ia ion in esou ce a ailabili y o
laying emales (Pe ins, 1970; D en , 2006). La e magpie
nes lings ha e poo e immune sys em (So ci e al.,
1997) and lowe de elopmen al a e han ea ly nes -
lings (De Ne e e al., 2004), which may hus explain he
de ec ed nega i e co ela ion be ween laying da e and
elome e a i ion o magpies du ing de elopmen . How-
e e , his possibili y p edic s a posi i e ela ionship
be ween elome e a i ion and laying da e ha we did
no ind. Ano he possibili y explaining he de ec ed el-
a i ely la ge elome e a i ion a es o ea ly magpie
ha chlings is ha la e ha chlings expe ience a ela i ely
low wi hin-b ood compe i ion. Howe e , esou ce a ail-
abili y dec eases as he season p og esses in magpie
nes s (De Ne e e al., 2004), and i is likely nega i ely
ela ed wi h le el o wi hin-b ood compe i ion. Addi-
ionally, elome e leng h has a ma e nal and a he edi-
a y componen in bi ds (Monaghan, 2014; Asgha
e al., 2015b), and consequen ly, ela ionships be ween
ha chling elome e leng h and laying da e may e lec
an unde lying ela ionship be ween he gene ic basis o
elome e leng h o pa en s (mainly emales) and laying
da e.
Taken oge he , all hese esul s o he i s ime
poin ou a ela ionship be ween en i onmen al condi-
ions (laying da e) and elome e leng h and sho ening
ha a y be ween species expe iencing simila ecologi-
cal condi ions du ing g ow h. As elome e leng h and
elome e sho ening a e a iables e lec ing su i al
p ospec and cos s associa ed wi h s ess esponses
ª2015 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY. J. EVOL. BIOL. doi: 10.1111/jeb.12680
JOURNAL OF EVOLUTIONARY BIOLOGY ª2015 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY
6JUAN J. SOLER ET AL.
(Asgha e al., 2015a; Ne le e al., 2015), hese esul s
may sugges ha species a ying in de elopmen al
s a egies (i.e. nes ling g ow h and/o emb yonic g ow h
a es) also a y in su e ed cos s in e ms o elome e
sho ening and/o dynamics. The e o e, ou esul s i
expec a ions acco ding o in e speci ic di e ences in li e-
s yle and li e his o y. Explo ing possible causes o he
de ec ed in e speci ic di e ences and associa ions may
help o unde s and he e olu iona y ela ionship
be ween elome e dynamics and li e-his o y ai s o ani-
mals.
Acknowledgmen s
We hank Olga Co ona Fo e o, F ancisco Mi anda and
Olaya Ga c
ıa o echnical assis ance wi h DNA ex ac-
ion om blood samples and ampli ica ion o elome e
leng h es ima ion. Funding was p o ided by he Span-
ish Minis e io de Economia y compe i i idad and Eu o-
pean Funds (FEDER) (CGL2013-48193-C3-1-P,
CGL2012-30759) and Jun a de Andaluc
ıa (RNM 340,
RNM7038). JMP is suppo ed by a Juan de la Cie a
con ac . Blood sampling and nes isi ing we e done
ollowing e hical s anda ds and unde he pe mission
o Jun a de Andaluc
ıa, En i onmen al Managemen
Agency, which au ho ized ield p o ocol (Re .: SGMN/
GyB/JMIF).
S a emen o au ho ship
JJS and JF designed he s udy wi h conside able assis-
ance om JMP and CRC. CRC pe o med all molecula
analyses wi h conside able assis ance JMP. JJS, MRR
and GT pe o med mos o he ield wo k wi h assis-
ance by CRC. JJS pe o med all he s a is ical analyses
and w o e he manusc ip wi h subs an ial con ibu ion
om all au ho s
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ª2015 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY. J. EVOL. BIOL. doi: 10.1111/jeb.12680
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8JUAN J. SOLER ET AL.