F on ie s in Ma ine Science | www. on ie sin.o g 1 Janua y 2021 | Volume 7 | A icle 606088
ORIGINAL RESEARCH
published: 25 Janua y 2021
doi: 10.3389/ ma s.2020.606088
Edi ed by:
Xa ie Pochon,
Caw h on Ins i u e, NewZealand
Re iewed by:
Ch is ophe Somes,
GEOMAR Helmhol z Cen e o
Ocean Resea ch Kiel, Ge many
Eleono a Puccinelli,
Uni e si é de B e agne Occiden ale,
F ance
*Co espondence:
Rui Albuque que
[email p o ec ed]
†ORCID:
An onio Bode
o cid0000-0002-9535-254
Special y sec ion:
This a icle was submi ed o
Ma ine Ecosys em Ecology,
a sec ion o he jou nal
F on ie s in Ma ine Science
Recei ed: 14 Sep embe 2020
Accep ed: 15 Decembe 2020
Published: 25 Janua y 2021
Ci a ion:
Albuque que R, Bode A,
González-Go dillo JI, Dua e CM and
Quei oga H (2021) T ophic S uc u e
o Neus on Ac oss T opical and
Sub opical Oceanic P o inces
Assessed Wi h S able Iso opes.
F on . Ma . Sci. 7:606088.
doi: 10.3389/ ma s.2020.606088
T ophic S uc u e o Neus on Ac oss
T opical and Sub opical Oceanic
P o inces Assessed Wi h S able
Iso opes
RuiAlbuque que
1
*, An onioBode
2†, JuanIgnacioGonzález-Go dillo
3, Ca losM.Dua e
4 and
Hen iqueQuei oga
1
1 Depa amen o de Biologia, CESAM – Cen o de Es udos do Ambien e e do Ma , Uni e sidade de A ei o, A ei o, Po ugal,
2 Ins i u o Español de Oceanog a ía, Cen o Oceanog á ico de A Co uña, A Co uña, Spain, 3 Ins i u o Uni e si a io de
In es igación Ma ina (INMAR), Campus de Pue o Real, Uni e sidad de Cádiz, Cádiz, Spain, 4 Red Sea Resea ch Cen e
(RSRC) and Compu a ional Bioscience Resea ch Cen e (CBRC), King Abdullah Uni e si y o Science and Technology,
Thuwal, Saudi A abia
The ma ine neus on, o ganisms li ing in he icini y o he ocean su ace, is one o he
leas s udied zooplank on g oups. Neus on occupies a es ic ed ecological niche and is
a ec ed by a wide ange o endogenous and exogenous p ocesses while also being a
ood sou ce o zooplank on ish mig a ing om he deep laye s and seabi ds. In his s udy,
he neus onic communi ies we e cha ac e ized along he Malaspina global expedi ion
sampling opical and sub opical oceanic p o inces using s able ca bon and ni ogen
iso opes o explo e hei ophic s uc u e and ela ionships wi h en i onmen al a iables.
The di e ences in s able iso opes mi o ed he pa e ns in en i onmen al cha ac e is ics
o each p o ince. High δ13C alues we e associa ed wi h a mosphe ic ca bon inpu s, while
he p esence o dino lagella es, coccoli hopho ids, and upwelling in luence is ela ed o
low δ13C alues. Simila ly, p o inces p esen ing high δ15N alues we e associa ed wi h
deni i ica ion and ni a e di usi e luxes, whe eas he p esence o low δ15N is a ibu able
o ni ogen supplied h ough N2 ixa ion by diazo ophs. Neus on showed a la ge o e lap
among he iso opic niches o ou unc ional g oups, wi h chae ogna hs and de i i o es
gene ally exhibi ing a smalle deg ee o o e lap compa ed o ca ni o es and omni o es/
he bi o es. These esul s suppo he hypo hesis o a common ophic s uc u e in he
neus on communi y ac oss he ocean. Howe e , he size o he niche, small in coas al
a eas and hose in luenced by upwelling and la ge in oligo ophic egions, and hei o e lap,
low in mo e p oduc i e p o inces and high in oligo ophic p o inces, may beassocia ed
wi h ood a ailabili y. Small ophic niches a e associa ed wi h a dominance o specialized
o e -oppo unis ic eeding in p oduc i e en i onmen s.
Keywo ds: s able iso opes, ophic g oups, neus on, biogeochemical p o inces, niche size
Albuque que e al. T ophic S uc u e o Neus on
F on ie s in Ma ine Science | www. on ie sin.o g 2 Janua y 2021 | Volume 7 | A icle 606088
INTRODUCTION
The neus on, one o he less desc ibed and known aqua ic
ecological g oups, is pa adoxically he closes o ou sampling
pla o ms as i inhabi s he uppe cen ime e s o he ocean.
The e m neus on was coined in 1972 (Hempel and Weike ,
1972) o de ine he pelagic o ganisms ha occupy he icini y
o he su ace laye , albei o en in a empo ally- es ic ed and
a iable manne . Neus on occupies a delimi ed ecological niche
and is gene ally g ouped in o h ee ecological ca ego ies: (a)
euneus on: o ganisms wi h maximum abundance in he icini y
o he su ace on which hey eside day and nigh ; (b) acul a i e
neus on: o ganisms concen a ing a he su ace only du ing
ce ain hou s o he day, usually du ing da kness; and (c)
pseudoneus on: o ganisms wi h maximum concen a ions a
deepe laye s bu eaching he su ace laye a leas du ing
ce ain hou s (Ma shall and Bu cha d , 2005). The neus onic
communi y s uc u e is condi ioned by sunligh and an a ay
o endogenous (o ganic ma e , espi a o y, pho osyn he ic,
decomposi ional p ocesses) and exogenous (a mosphe ic
deposi ion, ino ganic ma e , winds, wa e ac ion, p ecipi a ion,
UV adia ion, oceanic cu en s, su ace empe a u e) a iables
and p ocesses a ec ing nu ien inpu s and ecycling (Ma shall
and Bu cha d , 2005; Rawlinson e al., 2005; Rezai e al., 2019).
Fu he mo e, he neus on p o ides a ood sou ce o he
zooplank on mig a ing om deepe laye s o he su ace (Hempel
and Weike , 1972), as well as o seabi ds oaming o e he
oceans (Cheng e  al., 2010). Fo hese easons, he neus onic
communi y is belie ed o play a c i ical ole on he s uc u e
and unc ion o ma ine ood webs. Ye , esea ch on neus on
communi ies o da e ocused p edominan ly on geog aphically-
limi ed egions o he ocean (Zai se , 1971; Hempel and Weike ,
1972; Holdway and Maddock, 1983; Ebbe s and Wing, 1997;
Rezai e  al., 2019) o coas al a eas (B odeu , 1989; Le Fe e
and Bou ge , 1991; Padma a i and Goswami, 1996). Consequen ly,
neus on complexi y is s ill poo ly unde s ood as s udies on
he communi y s uc u e and he axonomical composi ion o
o ganisms inhabi ing his ecological niche emain ew
(Rezai e  al., 2019), and global scale analyses a e ye lacking.
The neus onic animals o m a subse o he zooplank on
communi y, which plays a pi o al ole in he unc ioning o
ma ine ecosys ems. Zooplank on a e pa ially esponsible o he
ac i e ene gy lux be ween supe icial and deep laye s o he
ocean (Tu ne , 2002; Jónasdó i e  al., 2015; He nández-León
e  al., 2020). Zooplank on species composi ion, biomass, and
seconda y p oduc ion in luence a wide ange o ophic le els
in ma ine communi ies, as hey cons i u e a link be ween p ima y
p oduc ion and seconda y consume s (Li chman e  al., 2013;
Benede i e  al., 2016; de Oli ei a Sod é and Bozelli, 2019).
Copepods cons i u e he mos abundan zooplank on axon in
e ms o biomass and di e si y wo ldwide (Kiø boe, 2011;
Neumann-Lei ão e al., 2018); he e o e, changes in hei communi y
composi ion can hus impac he biogeochemical cycles (Bianchi
and Mislan, 2016) and migh be indica i e o clima e a iabili y
impac s on ecosys em unc ioning (Hoo and Pe e son, 2006).
His o ically, zooplank on assemblages esea ch has ocused
mainly on axonomic s udies and hose ela ed o communi y
s uc u e (Pome leau e  al., 2015). Howe e , ecen ly, esea ch
has ee ed owa d an al e na i e ai -based app oach (Pome leau
e  al., 2015; Benede i e  al., 2016; Campos e  al., 2017),
p o iding a pe spec i e mo e ocused on g oups o species
wi h analogous unc ional ai s. This allows indi iduals o
be classi ied in o ypes cha ac e ized by he p esence/absence
o ce ain alleles o a gene, in o size classes, ecological guilds,
o unc ional g oups (FGs; Tuomis o, 2010). Func ional ai s
a e pheno ypes a ec ing o ganism i ness, g ow h, su i al, and
ep oduc i e abili y (Violle e  al., 2007; de Oli ei a Sod é and
Bozelli, 2019). These a e egula ed by he exp ession o genes
wi hin species, and he exp ession o ai s egula e, in u n,
he species i ness unde con as ing bio ic and abio ic
ci cums ances (Ba on e al., 2013). Mo eo e , a speci ic unc ional
ai can also de elop om he in e ac ions be ween o he
ai s and en i onmen al condi ions (Kiø boe, 2011), leading
o a gi en ai g ouping being a o ed unde ce ain condi ions.
Zooplank on ai s can beclassi ied in acco dance o ecological
unc ions – eeding, g ow h, ep oduc ion, su i al, and o he
cha ac e is ics such as mo phology, physiology, beha io , o
li e his o y (Li chman e  al., 2013; Hun e  al., 2015; B un
e al., 2016). Pa icula ly, eeding s a egies and ophic g oups
a e ele an o asce ain eeding e iciency and associa ed
p eda ion isk (B un e  al., 2017). Addi ionally, hey acili a e
he unde s anding o ecosys em se ices associa ed wi h
zooplank on, such as he dis ibu ion o ishe ies o
biogeochemical cycling (P owe e al., 2019) while also allowing
he posi ioning o zooplank on axa in he ood web
(Benede i e  al., 2016, 2018).
S able iso ope analysis (SIA) has been widely used o explo e
he ood web s uc u e; o iden i y an o ganism’s ophic posi ion;
o quan i y ca bon, ni ogen, and ene gy luxes; and o
cha ac e ize ophic niches (F y, 2006; Bouillon e  al., 2011;
Middelbu g, 2014). Gene ally, s able iso opes unde go a
p edic able ophic en ichmen be ween p ey and consume
(Minagawa and Wada, 1984) and e lec he o ganism’s die
o e a conside able pe iod o ime (Vande Zanden e  al.,
2015). Ra ios o ca bon iso opes (δ13C) gene ally ha e a low
ophic en ichmen and a e commonly used o iden i y ca bon
sou ces (DeNi o and Eps ein, 1978; Vande Zanden and
Rasmussen, 2001; Pos , 2002a). Ni ogen iso ope a ios (δ15N)
show p og essi e en ichmen be ween p ey and consume s
(DeNi o and Eps ein, 1978; Minagawa and Wada, 1984; Pos ,
2002b; McCu chan e  al., 2003) and we e hus employed o
es ima e ophic posi ions (F y and She , 1984; Minagawa and
Wada, 1984; Pe e son and F y, 1987; Owens, 1988; Pos , 2002b).
Thus, by measu ing he a ios o δ13C and δ15N, i is possible
o in e he ophic s uc u e o ma ine ood webs (F y, 2006).
Recen ly, SIA has been inc easingly employed o he
cha ac e iza ion o ophic niche (Layman e al., 2007a,b, 2012;
Hun e  al., 2015). The ophic niche o a single species,
communi y, o ecosys em is he agg ega e o he in e ac ions
be ween i s cons i uen s and he ecosys em (El on, 1927), hence
ep esen a i e o he cha ac e is ics o i s habi a and ophic
posi ion (Leibold, 1995). The ophic niche can be in e ed
om he iso opic niche (e.g., he δ13C–δ15N bi-plo space) o
unco e ele an aspec s o ophic s uc u e. Fo ins ance,
Albuque que e al. T ophic S uc u e o Neus on
F on ie s in Ma ine Science | www. on ie sin.o g 3 Janua y 2021 | Volume 7 | A icle 606088
niche-based quan i a i e me ics can be used o es ecological
heo y and ophic esponses o an h opogenic impac s (Layman
e  al., 2007a; Schmid e  al., 2007), including he deg ee o
o e lap be ween dis inc ophic niches (Jackson e  al., 2011).
To ou knowledge, he use o iso opic niche me ics on
he s udy o he neus onic zooplank on communi y has ne e
been a emp ed be o e. He e, we b idge his gap by explo ing
he a ia ions in he size o he ophic niche o neus onic
zooplank on ac oss sub opical and opical oceanic p o inces.
Ou main hypo hesis is ha he ophic s uc u e o he
neus onic communi y is p ese ed in spi e o di e ences in
nu ien sou ces and p oduc i i y along oceanic p o inces. Mo e
speci ically, in his s udy we (1) cha ac e ize a ios o C and
N o neus on ac oss sub opical and opical oceanic p o inces,
(2) de e mine ophic s uc u e simila i ies ac oss oceanic
p o inces, and (3) analyze he ela ionships be ween selec ed
en i onmen al a iables and he ophic s uc u e among oceanic
p o inces. Wedo so based on he samples collec ed along he
Malaspina Ci cumna iga ion Expedi ion, which ci cumna iga ed
he sub opical and opical ocean in 2010–2011 (Dua e, 2015).
MATERIALS AND METHODS
Sample Collec ion
The neus on samples we e collec ed along he Malaspina 2010
Expedi ion, which ci cumna iga ed he globe and was ca ied
ou be ween Decembe 2010 and July 2011 ac oss opical,
sub opical, and empe a e egions o he A lan ic, Indian, and
Paci ic Oceans be ween 35° N and 40° S (Dua e, 2015).
Sampling s a ions we e dis ibu ed o cha ac e ize pelagic
communi ies ac oss egions o he open ocean in he no he n
and sou he n hemisphe e (Dua e, 2015). In o de o allow
o he in e compa abili y o he obse a ions and a oid ad e se
wea he du ing sampling, he c uise was scheduled o isi
mos egions du ing hei sp ing-summe mon hs. The sampling
loca ions we e assigned o Longhu s Biogeochemical p o inces
(Longhu s , 2007). The e, ou biomes (Pola , Wes e lies, T ades
and Coas al) and a o al o 56 p o inces we e iden i ied, based
on he cha ac e iza ion o p ima y p oduc ion, mixed dep h
laye , nu ien s a ailabili y, pho ic dep h, algal biomass, B un -
Väisälä equency, and he Rossby adius o in e nal de o ma ion.
Speci ically, he neus on samples epo ed he e included 10
oceanic Longhu s p o inces (Longhu s , 2007). Fou p o inces
in he A lan ic [No heas A lan ic sub opical gy al (NASE;
n=16); No h A lan ic opical gy al (NATR; n=14); Wes e n
opical A lan ic (WTRA; n=15); Sou h A lan ic gy al (SATL;
n=12)], ou in he Paci ic [Sou h Paci ic gy e (SPSG; n=18);
Paci ic equa o ial di e gence (PEQD; n = 12); No h Paci ic
opical gy e (NPTG; n=14); No h Paci ic equa o ial coun e
cu en (PNEC; n=15)], and wo in he Indian Ocean [Indian
Sou h sub opical gy e (ISSG; n = 18); Sou h sub opical
con e gence (SSTC; n=16); Figu e1; Supplemen a y Table S1].
Be ween h ee and ou s a ions we e sampled wi hin each
p o ince. Samples we e collec ed wice a day a 12 pm and
4 am, by owing a neus on sample wi h a mou h opening
o 80 × 30 cm and a mesh size o 200 μm, a 2–3 kno s o
10–15 min, which sampled he i s 15 cm o he wa e
a a dis ance o 5 m om he s a boa d o he essel
(González-Go dillo e  al., 2012). The con en o each sample
FIGURE1 | Loca ion o he s a ions sampled in his s udy, indica ed by g een ci cles. S a ions we e g ouped in dis inc biogeochemical p o inces acco ding o
Longhu s (2007) wi h ac onyms depic ed. Shape ile adap ed om Flande s Ma ine Ins i u e (2009).
Albuque que e al. T ophic S uc u e o Neus on
F on ie s in Ma ine Science | www. on ie sin.o g 4 Janua y 2021 | Volume 7 | A icle 606088
was s o ed in 4% o maldehyde un il analysis. En i onmen al
da a we e acqui ed using CTD cas s deployed a each s a ion
om he su ace o 4,000 m o 100 m abo e he sea loo
when his was shallowe han 4,000 m.
Sample P ocessing, Taxonomical
Iden i ica ion, and Func ional G ouping
Samples we e s ained wi h Bengal Rose in o de o acili a e
he iden i ica ion o he o ganisms o axonomic le el o a
amily which, whene e possible, was de e mined unde a
s e eomic oscope (Olympus SZX16) using app op ia e guides
(Bol o skoy, 1999; Cas ellani and Edwa ds, 2017). The mos
abundan and ep esen a i e axa selec ed o SIA we e copepods
o he amilies Aca iidae, Calanidae, Co ycaeidae, Oncaeidae,
and Pon ellidae, and Phylum Chae ogna ha. The la e we e
included as a ep esen a i e o op plank onic p eda o s. Each
o hese axa ha e speci ic ophic oles, acco ding o he
li e a u e. Fo ins ance, copepod amilies we e assigned o FGs
acco ding o Benede i e  al. (2016, 2018). Fou FGs we e
conside ed and axa we e assigned as ollows: FG1=de i i o es
(Oncaeidae); FG2 = he bi o es/omni o es (Aca iidae and
Calanidae); FG3 = ca ni o es (Co ycaeidae and Pon ellidae);
and FG4 = p eda o s (Chae ogna ha). All specimens selec ed
o analysis we e adul s acco ding o hei mo phological aspec
unde he mic oscope.
S able Iso ope Analysis
Analysis o he na u al abundance o ca bon and ni ogen
iso opes we e pe o med on p e iously d ied (50°C, 48 h)
neus on samples. E en when specimens we e classi ied a genus
o species le el, we pooled indi iduals om he same sample
s a ion, including day and nigh samples, a amily (Aca iidae,
Calanidae, Co ycaeidae, Oncaeidae, and Pon ellidae) o phylum
(Chae ogna ha) le el in o de o ob ain ca. 1 mg d y weigh
o iso opic de e mina ion and o achie e a minimum o h ee
da a poin s o each unc ional g oup and minimize he
cons ain s o small sample size (Jackson e  al., 2011). D ied
samples we e packed in o in capsules and measu ed in an
elemen al analyze (Ca lo E ba CHNSO 1108) coupled o an
iso ope- a io mass spec ome e (Finnigan Ma Del a Plus).
Iso opic analyses we e pe o med by he Se icio de Análisis
Ins umen al o he Uni e sidade da Co uña (Spain). Samples
we e no acidi ied in o de o emo e ca bona es since he
selec ed axa we e only sligh ly calci ied, and i has been
demons a ed ha his p ocedu e could impac ni ogen
measu emen s (Ma eo e  al., 2008). No co ec ions we e made
o he possible e ec o o maldehyde on he s able iso ope
composi ion as mos s udies speci ically made on ma ine
zooplank on samples poin ou o no signi ican e ec s in δ15N
and gene ally a dec ease o less han 2‰ in δ13C a e se e al
yea s o s o age (e.g., Mullin e  al., 1984; Bicknell e al., 2011;
de Lecea e al., 2011). Taking in o conside a ion ha all samples
ha e been p ese ed simila ly and ha he ime be ween
collec ion and SIA la gely exceeded 1 yea , we assumed ha
di e ences in s able iso ope alues e lec genuine a ia ions
in he p o inces and selec ed FGs.
Values o na u al abundance o s able iso opes we e exp essed
as δ13C and δ15N (‰) ela i e o Vienna Pee Dee Belemni e
and a mosphe ic ni ogen, espec i ely (Coplen, 2011). Ce i ied
iso ope s anda ds (USGS40 and L-alanine) we e analyzed along
wi h in e nal ace anilide and sample s anda ds wi h s anda d
de ia ion (SD) be ween ce i ied and measu ed alues <0.1‰.
The p ecision [s anda d e o (SE)] o eplica e de e mina ions
o s anda ds and samples was <0.05‰ o bo h iso opes (n=4).
As he C:N mass a io o mos samples exceeded 3.5 ( hus
sugges ing a signi ican and a iable lipid con en ), δ13C alues
we e no malized using an empi ical linea eg ession wi h he
sample C:N alue de e mined o aqua ic o ganisms (Pos e al.,
2007). This p ocedu e aimed a emo ing he e ec o he low
δ13C associa ed o lipids and was p e e ed due o low biomass
cons ain s ha hinde ed he use o lipid emo al me hodologies
such as dichlo ome hane and an accele a ed sol en ex ac ion
sys em (Bodin e  al., 2009).
En i onmen al Va iables
En i onmen al a ibu es o each o he sampled s a ions we e
cha ac e ized by a numbe o a iables collec ed in si u o
sa elli e-de i ed. (Supplemen a y Table S2). These a iables
we e al eady employed in a p e ious analysis o Malaspina
2010 c uise, whe e me hodological de ails can be ound
(Mompeán e al., 2013, 2016b; Fe nández-Cas o e al., 2015).
In b ie , he s a i ica ion o he wa e column was ep esen ed
by he dep h o he mixing laye (MLD, m), he mean squa ed
B un –Väisälä equency (N2, s−2), and he dep h o he
chlo ophyll maximum (DCM, m), all es ima ed om e ical
p o iles o a CTD equipped wi h a luo escence senso
(Fe nández-Cas o e al., 2015). Simila ly, nu ien inpu s om
deep laye s we e es ima ed by di usi i y due o u bulence
(KT, m2 s−1), de e mined om e ical cas s o a mic os uc u e
u bulence p o ile (Fe nández-Cas o e  al., 2015).
Phy oplank on biomass was ep esen ed by su ace and pho ic-
zone in eg a ed chlo ophyll-a (Chlas and Chlai, mg m−3 and
mg m−2, espec i ely), de e mined om ace onic ex ac s o
phy oplank on (Es ada e al., 2016). Mic oplank on (40–200μm)
cha ac e is ics we e indica ed by an abundance o he ni ogen-
ixe T ichodesmium (T icho, cells ml−1), ca bon biomass (C40–200,
mg C m−3), and na u al abundance o ni ogen iso opes (δ15N40–200,
‰) measu ed in samples collec ed by e ical ows o a plank on
ne be ween he su ace and 200 m dep h (Mompeán e  al.,
2013, 2016b). Sa elli e de i ed a iables we e conside ed o
ep esen condi ions p e ailing o e la ge spa ial and empo al
scales han hose conside ed du ing he speci ic sampling o
each s a ion and included annual a e ages o p ima y p oduc ion
(PP) o 2010 (mg C m−2 d−1) and mean mon hly a mosphe ic
dus deposi ion (MDU, g m−2mon h−1). The o me ep esen ed
egional p oduc i i y and was de i ed om he da a p o ided
by he Ocean P oduc i i y websi e (h p://www.science.
o egons a e.edu/ocean.p oduc i i y/index.php) in a g id o
0.17°×0.17° including each s a ion posi ion. Dus deposi ion,
a p oxy o a mosphe ic inpu s o key nu ien s as Fe o P,
was es ima ed om Aqua-MODIS Ae osol Op ical Dep h a
550 nm and Ae osol Small Mode F ac ion da a p o ided by
he Gio anni online da a sys em (NASA Godda d Ea h Sciences)
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om a g id o 1°×1° nea each s a ion (Mompeán e al., 2016b).
In dep h in o ma ion on sampling and analy ical me hodology
employed h oughou he Malaspina 2010 c uise can be ound
in Mo eno-Os os (2012).
S a is ical Analysis
Di e ences in δ13C and δ15N a ios o Oncaeidae among oceanic
p o inces we e es ed wi h ANOVA and Bon e oni pos hoc
es on S a is ica 12 (S a So , Inc., Tulsa, OK, USA). Analysis
o he di e ences in s able iso ope alues among oceanic
p o inces was assessed using a pe mu a ional mul i a ia e
analysis o a iance (PERMANOVA) and pai wise es s
on PRIMER 6.0 (Cla ke and Go ley, 2006) and he
add-on package PERMANOVA+ (Ande son e  al., 2008;
Supplemen a y Table S3). Fo subsequen analysis, a
ep esen a i e p ima y consume was used o no malize s able
iso ope alues acco ding o Cla k and F i z (1997) and S asko
e  al. (2018) o accoun o spa ial he e ogenei y in ca bon
and ni ogen a ios. To his end, consume δ13C alues we e
no malized ela i e o a pelagic baseline (Δ13Cpel) as:
∆13 3
1000
1000
110Cpel w
c
=+
+−









×
d
d, we e δC is he consume δ13C
alue and δW is he p o ince-speci ic mean δ13C alue o he
copepod amily Oncaeidae, conside ed as he e e ence baseline.
Consume δ15N alues we e no malized by sub ac ing he
p o ince-speci ic mean δ15N alue o he copepod amily
Oncaeidae om he consume δ15N, ep esen ed he ea e as
δ15Nn. The use o Oncaeidae as baseline is no in ended o
e lec he base o he ood web bu a he o se a homogeneous
s a ing poin along he δ13C and δ15N con inuum. All s a is ical
analyses we e om his poin on pe o med on Oncaeidae-
no malized alues.
Analysis o he iso opic niche size we e made in he iso opic
space de ined by Δ13Cpel–δ15Nn o each p o ince/ unc ional g oup
combina ion wi h he package SIBER (Jackson e  al., 2011).
Bayesian es ima es o he s anda d ellipse a ea (SEAb) we e
calcula ed o each combina ion o cha ac e ize he ull a iabili y
in o aging habi s and used esou ces (Layman e  al., 2007a,
2012). These es ima es accoun ed o unce ain y due o he
numbe o samples. A minimum o h ee samples was equi ed
o hese es ima ions. SEAb s a is ics we e compu ed om 104
simula ions pe p o ince/ unc ional g oup a angemen ,
al hough only he maximum likelihood ellipses we e used o
g aphical ep esen a ions. Pai wise compa isons o SEAb
(Supplemen a y Table S4) we e pe o med by calcula ing he
p opo ion o ellipse size ha di e ed be ween wo gi en
combina ions being in e p e ed as a di ec p oxy o he
p obabili y ha one combina ion is di e en om he o he
(Jackson e  al., 2011). Because o he limi a ions imposed by
he a ailable sample biomass o iso opic analyses, da a o some
FGs could no be ob ained o all p o inces.
P incipal componen analysis (PCA) was used o
in es iga e he ela ionships be ween en i onmen al a iables
and he niche size (as a p oxy o ophic s uc u e) o
he FGs among oceanic p o inces using PRIMER 6.0
(Cla ke and Go ley, 2006). Fi s , no malized en i onmen al
a iables measu ed on each sampling si e we e employed o
compu e he eigen alues, eigen ec o s, and p incipal componen
(PC) coo dina es. Then, Pea son’s co ela ion coe icien be ween
SEAb a e age and 95% c edible in e als (CI) ange o each
unc ional g oup/p o ince combina ion and he en i onmen al
PC coo dina es we e compu ed using he co () unc ion on
R 3.6.3 (R Co e Team, 2020). Finally, he co ela ion coe icien s
be ween he niche me ics and he PCA axes we e plo ed
on he a iable space o he PCA i s wo p incipal componen
coo dina es axis.
RESULTS
δ13C and δ15N Ra ios
The mean δ13C o Oncaeidae a ied signi ican ly among
p o inces (Figu e2A). Bon e oni pos hoc es e ealed h ee
dis inc g oups (a, b, and c) o dec easing mean alues (−21.2,
−21.8, and −22.3‰, espec i ely). The highes alues we e
ound in NATR and SPSG p o inces, while he lowes alues
occu ed in NASE. Simila ly, signi ican di e ences in mean
alues o δ15N be ween p o inces we e also ound (Figu e2B).
All δ15N alues we e posi i e, wi h maximum alues in SSTC
p o ince, ollowed by hose in PNEC and NPTG, and minimum
alues (<2‰) in NATR. Bon e oni pos hoc es s also e ealed
h ee g oups (d, e, and ) o dec easing mean δ15N (10.1, 5.4,
and 3.5 ‰, espec i ely).
A
B
FIGURE2 | Mean and SE alues o δ13C (A) and δ15N (B) o Oncaeidae by
Longhu s P o inces. G oups o signi ican ly di e en means (ANOVA and Bon e oni
pos hoc es , p<0.05) a e indica ed by di e en le e s (a, b, c, d, e, and ).

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Iso opic Niche Size o Func ional G oups
The analysis o he iso opic niches e ealed di e ences be ween
p o inces and FGs in all oceans. The maximum likelihood
ellipses we e dis inc om each o he in all p o inces and
FGs bu showed la ge o e laps be ween hem in some p o inces/
unc ional g oup a angemen s.
A lan ic P o inces
The e was a clea sepa a ion o he ellipses o op p eda o s
(FG4) and de i i o es (FG1) in he p o inces whe e hese
g oups we e analyzed (Figu e 3). In con as , ellipses o
he bi o es/omni o es (FG2) and ca ni o es (FG3) showed la ge
o e laps. The a iabili y o SEAb (Bayesian s anda d ellipse
a ea) was highes o FG3 and lowes o FG2 (Table 1).
Pai wise compa isons o he iso opic niche a eas using SEAb
e ealed ha only FG3 a ied signi ican ly om small alues
in NASE, in e media e in NATR and high alues in SATL,
while no signi ican di e ences esul ed when compa ing o he
FG among A lan ic p o inces (Supplemen a y Table S4).
Indian P o inces
Dis inc maximum likelihood ellipses we e p esen in bo h
p o ince/FGs a ays; none heless, signi ican o e laps a e
disce nible h oughou he isospace (Figu e4; Table 1). SEAb
was consis en ly la ge o all FGs in he ISSG p o ince and
he la ges ophic a iabili y was ound o FG2. Mo eo e ,
SEAb we e he smalles o bo h FG1 and FG3 in SSTC.
The e we e no signi ican di e ences in SEAb o equi alen
FG among Indian p o inces (Supplemen a y Table S4).
Paci ic P o inces
The isospace a ea o he Paci ic p o inces ollows he end
desc ibed o A lan ic and Indian p o inces, wi h pe cep ible
di e ences in he dis inc p o inces/FGs a angemen s, bu he
la ge o e lap was ound in many cases, especially in PNEC
and SPSG (Figu e 5). The la ges alues o SEAb we e ound
o FG4 in SPSG and he minimum alues o FG2 in PEQD
(Table1). NPTG and PEQD p esen ed na owe SEAb compa ed
o SPSG and PNEC. Pai wise compa isons o SEAb indica ed
ha he alues o FG1 we e signi ican ly smalle o PNEC
compa ed o hose o SPSG and ha hose o FG2in NPTG
and PEQD we e smalle han hose in PNEC and SPSG, while
hose o FG3 in PNEC and SPSG we e la ge han hei
equi alen a eas in NPTG and PEQD. Finally, SEAb o FG4in
PNEC and SPSG we e la ge han hei compa able g oup in
NPTG (Supplemen a y Table S4).
En i onmen al Va iables and T ophic
S uc u e
The PCA on he en i onmen al a iables e ealed dis inc i e
condi ions among p o inces, as s a ions a e clea ly clus e ed
wi hin each p o ince (Figu e 6). The i s wo p incipal
componen s o he PCA accoun ed o 66.4% o he o al
a iance (Table 2). Su ace chlo ophyll had he la ges posi i e
FIGURE3 | Isospace o Δ13Cpel and δ15Nn including indi idual sample no malized alues o neus onic zooplank on o biogeochemical p o inces in he A lan ic
Ocean: NASE, NATR, WTRA, and SATL. The maximum likelihood es ima es o he s anda d ellipse a eas ( hick lines) o each unc ional g oup (FG) a e shown.
FG 1=de i i o es; FG 2=he bi o es/omni o es; FG 3=ca ni o es; FG 4=p eda o s. Dec eases in Δ13Cpel e lec inc eases in aw δ13C alues due o no maliza ion.
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loadings wi h he i s componen (PC1), ollowed by he
mic oplank onic ca bon biomass and u bulence di usi i y
(Figu e7). Con e sely, PC1 was nega i ely co ela ed wi h he
dep h o chlo ophyll maximum. The second componen (PC2)
was posi i ely co ela ed wi h p ima y p oduc ion, δ15N40–200,
and he mixed laye dep h, while showing nega i e co ela ions
wi h a mosphe ic dus deposi ion, T ichodesmium abundance,
and pho ic-zone in eg a ed Chl-a. The emaining en i onmen al
a iables ha e a mino ele ance in explaining he epo ed
di e ences (Figu e 7).
The size o he iso opic niche (indica ed by median SEAb
alues and 95% CI) o FG4 was posi i ely co ela ed wi h
PC1, while hose o FG3 showed nega i e co ela ions wi h
his componen . In u n, SEAb o FG1 and FG2 we e mainly
nega i ely co ela ed wi h PC2 (Figu e 7).
DISCUSSION
In he cu en s udy, we p o ide he i s compa ison o he
ophic s uc u e o he neus onic communi y in opical and
sub opical p o inces o he A lan ic, Indian, and Paci ic Oceans
(Longhu s , 2007; Dua e, 2015). The analysis o s able ca bon
and ni ogen iso opes o neus on FGs allowed o he
cha ac e iza ion o ca bon and ni ogen a ios, he quan i ica ion
o he size o he iso opic niche as a p oxy o ophic s uc u e,
and i s ela ionships wi h en i onmen al a iables ac oss dis inc
oceanic p o inces. No wi hs anding he limi a ions imposed
by he analy ical equi emen and, hus, he small numbe o
samples employed, as well as he samples p ese a ion me hod,
he esul s o his s udy allow us o es ablish an app oxima ion
o he ophic in e ac ions in he neus on and hei ole in
pelagic ood webs a a global scale.
δ13C and δ15N Ra ios
The di e ences ound in ca bon and ni ogen appea o e lec
a ia ions in he en i onmen al egimes o he dis inc p o inces.
P o ided ha di e en phy oplank on axa a o he use o
dis inc ca bon sou ces o p ima y p oduc ion, i is plausible
ha i could lead o a ia ions in δ13C ha can be passed
upon he ood web. Fo ins ance, hea ie δ13C alues ha e
TABLE1 | Iso opic niche a ea (‰2) es ima es o each unc ional g oup and
p o ince.
Ocean P o ince FG SEAb 95% CI
A lan ic
NASE
1 0.446 0.130 o 2.128
2 0.339 0.088 o 1.144
3 0.377 0.115 o 0.999
4 0.212 0.040 o 0.929
NATR 2 0.481 0.173 o 1.403
3 1.259 0.526 o 3.921
WTRA
1 0.866 0.198 o 4.276
2 0.832 0.211 o 3.497
3 0.597 0.250 o 1.540
4 0.524 0.140 o 2.592
SATL 2 0.596 0.194 o 2.094
3 4.508 1.695 o 13.427
Indian
ISSG
2 1.411 0.598 o 3.888
3 1.001 0.500 o 2.510
4 0.468 0.134 o 2.273
SSTC
1 0.177 0.047 o 0.821
2 0.826 0.348 o 1.973
3 0.054 0.015 o 0.238
4 0.221 0.061 o 1.047
Paci ic
NPTG
2 0.338 0.119 o 1.253
3 0.554 0.167 o 1.541
4 0.250 0.063 o 1.284
PEQD
2 0.088 0.024 o 0.393
3 0.543 0.142 o 1.709
4 0.998 0.220 o 4.597
PNEC
1 0.397 0.107 o 2.010
2 1.258 0.367 o 4.325
3 2.913 1.027 o 8.495
4 2.232 0.487 o 10.505
SPSG
1 2.162 0.444 o 10.884
2 1.897 0.802 o 4.899
3 3.177 0.938 o 7.988
4 3.361 0.598 o 14.937
Es ima es a e p esen ed as he mode o he Bayesian s anda d ellipse a ea (SEAb).
Lowe and uppe 95% CI indica e he unce ain y in he SEAb es ima es.
FIGURE4 | Isospace o Δ13Cpel and δ15Nn including indi idual sample no malized alues o neus onic zooplank on o biogeochemical p o inces in he Indian
Ocean: ISSG and SSTC. The maximum likelihood es ima es o he s anda d ellipse a eas ( hick lines) o each FG is shown. FG 1=de i i o es; FG 2=he bi o es/
omni o es; FG 3=ca ni o es; FG 4=p eda o s. Dec eases in Δ13Cpel e lec inc eases in aw δ13C alues due o no maliza ion.
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been associa ed wi h highe phy oplank on ac iona ion o
ino ganic ca bon, and subsequen sinking o o ganic ma e
ela i ely deple ed in 13C om he su ace wa e s led o high
alues in he uppe ocean a sub opical and opical wa e s
(G ube e al., 1999; Schmi ne e al., 2013). Mo eo e , up ake
o a mosphe ic CO2 ia ai -sea exchange and consequen
ac iona ion in he con e sion p ocess o HCO−3 can esul
in high δ13C-dissol ed ino ganic ca bon (Zhang e  al., 1995).
Such p ocesses could explain he en ichmen in 13C (i.e., highe
δ13C) ound o he iso opic baseline (Oncaeidae) in he SPSG
and NATR. Addi ionally, he en ichmen in δ13C measu ed in
SPSG could also be ela ed o he p esence o dia oms, as
epo ed in he egion du ing he same c uise (Es ada e  al.,
2016). Al hough hey we e no he dominan phy oplank on
axa (Es ada e  al., 2016), hey may ha e disp opo iona ely
con ibu ed o he me azoan ood web, while he dominan
picocyanobac e ia would con ibu e o he mic obial ood web.
While cyanobac e ia up ake o ino ganic ca bon occu s ia
di ec HCO−3 anspo , dia oms a e also capable o anspo ing
CO2 de i ed om he ca alyzed dehyd a ion o HCO−3 (To ell
and Mo el, 2002). This implies a la ge iso opic ac iona ion
(i.e., less nega i e alues o δ13C) in dia oms when compa ed
wi h cyanobac e ia o o he mic oalgae (F y and Wain igh ,
1991). Ano he hypo hesis o he δ13C en ichmen measu ed
in NATR p o ince is ela ed o he abundance o Sa gassum
spp. on he wes e nmos s a ions (Gou êa e  al., 2020).
FIGURE5 | Isospace o Δ13Cpel and δ15Nn including indi idual sample no malized alues o neus onic zooplank on o biogeochemical p o inces in he Paci ic
Ocean: NPTG, PEQD, PNEC, and SPSG. The maximum likelihood es ima es o he s anda d ellipse a eas ( hick lines) o each FG a e shown. FG 1=de i i o es;
FG 2=he bi o es/omni o es; FG 3=ca ni o es; FG 4=p eda o s. Dec eases in Δ13Cpel e lec inc eases in aw δ13C alues due o no maliza ion.
FIGURE6 | P ojec ion o samples on he space o he i s (PC1) and second
(PC2) componen s o he p incipal componen analysis (PCA) o en i onmen al
a iables (DCM: dep h o he chlo ophyll maximum (m), δ15N40–200: na u al
abundance o mic oplank on ni ogen iso opes (‰), PP: annual mean p ima y
p oduc ion (mg C m−2 d−1), MLD: mixed laye dep h (m), C40–200: mic oplank on
(40–200μm) ca bon biomass (mg C m−3), KT: di usi i y due o u bulence (m2
s−1), Chlas: su ace chlo ophyll-a (mg m−3), N2: mean squa ed B un -Väisälä
equency (s−2), Chlai: pho ic-zone in eg a ed chlo ophyll-a (mg m−2), T icho:
abundance o T ichodesmium (cells ml−1), MDU: mean mon hy a mosphe ic
dus deposi ion (g m−2 mon h−1)) de e mined o neus on sampling s a ions.
P o ince ac onyms as in Supplemen a y Table S1.
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This mac oalgae has a δ13C anging om −16‰ o −18‰,
and i is concei able ha i could ha e in luenced he δ13C
baseline in NATR (Cabanillas-Te an e  al., 2019). Howe e ,
di e ences in δ13C could be he esul o many ac o s associa ed
wi h changes in phy oplank on CO2 ixa ion, which is known
o a y in ela ion o empe a u e, concen a ion o aqueous
CO2, phy oplank on composi ion, and he a ailabili y o dissol ed
ino ganic ca bon and nu ien s (Wong and Sacke , 1978;
Descolas-G os and Fon ugne, 1990; F ancois e  al., 1993;
Bu kha d e  al., 1999; Popp e  al., 1999).
Bo h con inen al o ganic ma e and a mosphe ic CO2 a e
cha ac e ized by ha ing lowe alues o δ13C han hei equi alen s
in oceanic (Pe y e  al., 1999) o upwelled wa e s (G ube
e  al., 1999). Ou esul s ag ee wi h he epo o highe δ13C
alues in zooplank on om oligo ophic egions o he sub opical
No h A lan ic du ing he same c uise (Mompeán e al., 2013).
In con as , mo e nega i e δ13C alues o he Oncaeidae baseline,
such as hose measu ed in NPTG and NASE, can beexplained
by he dominance o dino lagella es and coccoli hopho es in
hese p o inces (Es ada e  al., 2016) and by he in luence o
upwelling in NASE (G ube e al., 1999; Mompeán e al., 2013).
Ra ios o ni ogen also a ied be ween p o inces, acco ding
o he di e ences ound in δ15N alues o Oncaeidae. Low
δ15N alues could bedue o diazo ophy since he a mosphe ic
ni ogen p esen s δ15N = 0‰, and i can be aced h ough
he ood web (McClelland e al., 2003; Mompeán e al., 2016a;
Bode and He nández-León, 2018). This p ocess is media ed
by specialized p oka yo es – diazo ophs – which h i e in
usually ni a e-poo , wa m and s a i ied wa e s such as he
ones o he sub opical and opical gy es (Falkowski, 1997;
Zeh e  al., 2003; Capone e  al., 2005; Luo e  al., 2012) and
in oduce in o he ocean bioa ailable ni ogen deple ed in δ15N
(Somes e al., 2010). Addi ionally, i is also known ha iso opic
ac iona ion occu s du ing phy oplank on up ake, and his
may cause low δ15N alues (<5‰) in plank on when dissol ed
ni ogen concen a ions a e high, as epo ed a he ini ial
phases o blooms (Wase e al., 2000). In u n, high concen a ions
o 15N-en iched ni a e o ma ine o igin a e simila ly e lec ed
in he δ15N alues o zooplank on (Owens, 1988; Mon oya
e al., 2002), as ound in he SSTC samples. Ou esul s s ongly
sugges ha he low δ15N alues measu ed o Oncaeidae e lec
he ole o N2 ixa ion by diazo ophs in supplying N in some
p o inces (mainly in NATR bu also in NASE, WTRA, SATL,
SPSG, and PEQD), while he high δ15N alues measu ed in
o he p o inces indica e inpu s o ni ogen de i ed om
deni i ica ion p ocesses (e.g., PNEC, NPTG, and SSTC). These
assump ions a e suppo ed by he consis ency wi h es ima ions
o he abundance o he N- ixe T ichodesmium (Es ada e al.,
2016; Mompeán e  al., 2016b) and N ixa ion a es and NO3−
di usi e luxes ob ained o he Malaspina c uise (Fe nández-
Cas o e  al., 2015), as well as ecen es ima es o ma ine
ni ogen ixa ion and deni i ica ion a he scale o he global
ocean (Knapp e  al., 2016; Bonne e  al., 2017; G ube , 2019;
Wang e  al., 2019).
Mo eo e , he δ15N alues epo ed he e o Oncaeidae
a e consis en wi h hose o mic oplank on collec ed
simul aneously du ing he Malaspina c uise (δ15N40–200) in
he uppe 200 m o he wa e column, hus suppo ing ou
assump ion o he ole o Oncaeidae as p ima y consume s
and c i ical nodes in he ood web. Fu he mo e, he gene al
cohe ence be ween he dis ibu ion pa e n o bo h Oncaeidae
and mic oplank on δ15N ac oss p o inces (e.g., Mompeán
e  al., 2016b; Supplemen a y Figu e S1) sugges s simila
u no e imes o he s able iso opes (and hence g ow h
a es) in epipelagic mic oplank on and, in ou case,
mesoneus on (>200 μm).
Spa ial Di e ences in Iso opic Niche
The small laye o su ace wa e he neus onic communi y
inhabi s would imply a sca ce a ailabili y o ood sou ces,
pa icula ly in mo e oligo ophic egions, such as mos o
p o inces in his s udy. This sugges s a s ong compe i ion o
TABLE2 | Eigen alues, pe cen o o al a iance (%Va iance), and pe cen o
cumula i e a ia ion (Cum. %Va ia ion) explained o he i s i e p incipal
componen s (PC) o he p incipal componen analysis (PCA) on en i onmen al
a iables.
Componen Eigen alue %Va iance Cum. %Va ia ion
PC1 4.49 40.8 40.8
PC2 2.82 25.6 66.4
PC3 1.13 10.3 76.7
PC4 0.86 7.8 84.5
PC5 0.72 6.5 91.1
FIGURE7 | Loadings ( ec o s) o he en i onmen al a iables on he space
o he i s (PC1) and second (PC2) componen s o he PCA o en i onmen al
a iables de e mined o neus on sampling s a ions and p ojec ions (do s) o
median alues o niche size (SEAb) and 95% c edible in e als (CI) o he
di e en FGs (indica ed by numbe s 1–4). DCM: dep h o he chlo ophyll
maximum (m), δ15N40–200: na u al abundance o mic oplank on ni ogen
iso opes (‰), PP: annual mean p ima y p oduc ion (mg C m−2 d−1), MLD:
mixed laye dep h (m), C40–200: mic oplank on (40–200μm) ca bon biomass
(mg C m−3), KT: di usi i y due o u bulence (m2 s−1), Chlas: su ace
chlo ophyll-a (mgm−3), N2: mean squa ed B un -Väisälä equency (s−2), Chlai:
pho ic-zone in eg a ed chlo ophyll-a (mg m−2), T icho: abundance o
T ichodesmium (cells ml−1), MDU: mean mon hly a mosphe ic dus deposi ion
(gm−2 mon h−1).