In eg a ing Gamma us insensibilis in bio loc sys ems: A sus ainable app oach
o nu ien en ichmen and was e alo isa ion in aquacul u e
Ma a Cas illa-Ga il´
an
a,b,*
, Jos´
e Manuel Gue a-Ga cía
c
, Ismael Hache o-C uzado
b,d
a
Ins i u des Subs ances e O ganismes de la Me , ISOme , UR 2160, Nan es Uni e si ´
e, 44000 Nan es, F ance
b
IFAPA Cen o El To u˜
no, Camino Ti o Pich´
on s/n, El Pue o de San a Ma ía, 11500 C´
adiz, Spain
c
Labo a o io de Biología Ma ina, Depa amen o de Zoología, Facul ad de Biología, Uni e sidad de Se illa, A da. Reina Me cedes 6, 41012 Se illa, Spain
d
“C ecimien o Azul”, Cen o IFAPA el To u˜
no, Unidad Asociada al CSIC, El Pue o de San a Ma ía, 11500 C´
adiz, Spain
ARTICLE INFO
Keywo ds:
Sus ainable aquacul u e
Nu i ion
Low- ophic aquacul u e
Was e alo isa ion
Ci cula economy
ABSTRACT
This esea ch add esses he cul i a ion o Gamma us insensibilis in bio loc sys ems. The aim o he wo k was o
alo ise and o bio emedia e aquacul u e e luen s while enhancing he nu i ional alue o bio locs h ough he
in oduc ion o amphipods, which can be g own in close aquacul u e sys ems. Two expe imen al die s (aqua-
cul u e was e de i us and comme cial ish eed) we e es ed o amphipods in iplica ed bio loc sys ems, agains
h ee con ol anks wi hou bio loc. The expe imen was conduc ed o e a six-week pe iod a a empe a u e o
20 ◦C in he absence o ligh . The wa e quali y, he su i al, lipid p o ile and a y acid composi ion o he
amphipods and he bio locs p oduced we e s udied. Signi ican di e ences we e obse ed in he wa e quali y
be ween he ea men g oups and he con ol, since ni i ying bac e ia we e p esen in he bio locs o med in he
ea men anks. Indeed, 100 % mo ali y was eached in he con ol anks om he second week o he
expe imen . The ish pelle s die and he pelle -based bio loc esul ed in a signi ican ly highe o al lipid con en
han he de i us die and bio loc. Howe e , no di e ences we e ound in he essen ial long-chain poly-
unsa u a ed a y acids (LC-PUFAs) con en (ARA, EPA and DHA) be ween he comme cial pelle s and he
de i us-based bio loc, hus jus i ying he en ichmen o he bio loc wi h amphipods. Indeed, he wild amphipods
displayed highe le els o sa u a ed a y acids and omega-6 PUFAs compa ed o he expe imen al g oups, which
demons a ed highe omega-3 PUFAs con en , pa icula ly DHA. The s udy highligh s he nu i ional ad an ages
o pelle -based die s o amphipods, including imp o ed su i al and lipid con en , while also emphasising he
enhanced nu i ional p o ile o he en iched de i us-based bio loc. We sugges ha he in eg a ion o aquacul-
u e de i us wi h supplemen al aqua eed in BFT sys ems will suppo bo h he bio loccula ion p ocess and he
nu i ional needs o he amphipods, he eby c ea ing a sus ainable and e icien cycle o was e alo isa ion and
li e ood p oduc ion. Fu he esea ch is equi ed in o de o s udy he comple e li e cycle o amphipods and hei
ep oduc i e capaci y in hese sys ems. A lowe wa e empe a u e and he inocula ion o mic oalgae should be
conside ed in o de o achie e highe su i al a es and PUFAs con en .
1. In oduc ion
Bio loc echnology (BFT) was o igina ed in he 1970s (Eme enciano
e al., 2021) and has been widely adop ed due o i s en i onmen al and
economic bene i s (Zimme mann e al., 2023). This echnology is based
on ze o-wa e exchange sys ems wi h he o ma ion o bio locs in he
cul u e medium whe e unea en ood, excess ino ganic nu ien s and
aeces agg ega e along wi h mic oo ganisms (bac e ia, mic oalgae such
as dia oms, p o ozoa…) (Khanjani e al., 2024b). This agg ega ion is
enabled by a ma ix o ex acellula polyme ic subs ances sec e ed by
he mic oo ganisms (Ha g ea es, 2013). By main aining a high ca bon/
ni ogen a io (Ha g ea es, 2013) h ough he addi ion o a ca bon
sou ce (molasses, glyce ol, lou s; Zhao e al. (2016) o he cul u e
medium, he colonies o chemoau o ophic bac e ia p esen in he bio-
locs assimila e he ammonium exc e ed by he cul u ed species du ing
he ni i ica ion p ocess, ans o ming i in o ni a e (Khanjani e al.,
2022). He e o ophic bac e ia can also di ec ly assimila e he ammo-
nium in o bac e ial biomass. Addi ionally, mic oalgae con ibu e o ni-
ogen abso p ion du ing pho osyn hesis (Ebeling e al., 2006). BFT
sys ems ha e been iden i ied as a p omising echnology o sus ainable
* Co esponding au ho a : Ins i u des Subs ances e O ganismes de la Me , ISOme , UR 2160, Nan es Uni e si ´
e, 44000 Nan es, F ance.
E-mail add ess: [email p o ec ed] (M. Cas illa-Ga il´
an).
Con en s lis s a ailable a ScienceDi ec
Aquacul u e
jou nal homepage: www.else ie .com/loca e/aquacul u e
h ps://doi.o g/10.1016/j.aquacul u e.2024.741922
Recei ed 6 Sep embe 2024; Recei ed in e ised o m 18 No embe 2024; Accep ed 18 No embe 2024
Aquacul u e 597 (2025) 741922
A ailable online 19 No embe 2024
0044-8486/© 2024 The Au ho (s). Published by Else ie B.V. This is an open access a icle unde he CC BY license (
h p://c ea i ecommons.o g/licenses/by/4.0/ ).
ood p oduc ion, p omo ing low- ophic-le el species cul i a ion, poly-
cul u e, and he de elopmen o a ci cula economy model (Khanjani
e al., 2024c). The eeding e iciency and was e managemen a e
enhanced in hese sys ems, as hey ha e he capaci y o inc ease nu ien
u ilisa ion and minimize en i onmen al impac , achie ed h ough
educed wa e and e ilize use (Knowle e al., 2020; Zimme mann
e al., 2023). Thus, he need o wa e exchange is minimized, which in
u n educes pumping cos s, p e en s luc ua ions in empe a u e and
educes he ene gy equi emen o hea ing (McCuske e al., 2023). In
BFT sys ems, ood equi emen s a e also educed, and an inc ease in he
su i al and g ow h a es o cul i a ed species has been obse ed
(Khanjani and Sha i inia, 2024). Bio locs p esen a a ou able nu i-
ional p o ile (C ab e al., 2012) and hei consump ion ha e been
demons a ed o enhance g ow h and o i y he immune sys em o
cul i a ed species by inc easing he ac i i y o hei diges i e enzymes,
esul ing in an inc ease in he eed con e sion a e (Xu and Pan, 2013).
BFT is p ima ily employed in he cul i a ion o penaeids (C ab e al.,
2010b) bu nume ous s udies ha e applied his echnology o he
cul i a ion o il e eede s, de i i o es and e en some he bi o ous o
omni o ous in ish, which a e able o eed di ec ly on pa icula e
o ganic ma e (C ab e al., 2012; Das and Mandal, 2021; Dinda e al.,
2020). These species a e ole an o changes in he concen a ions o
dissol ed oxygen in he wa e and ni ogenous compounds and can
wi hs and high cul u e densi ies and high concen a ions o suspended
solids (El-Sayed, 2021; Eme enciano e al., 2021).
Despi e all hese ad an ages, bio locs ha e been obse ed o p esen
low p opo ions o long-chain polyunsa u a ed a y acids (LC-PUFAs)
(C ab e al., 2010a), maybe linked wi h he absence o PUFAs in bac e ia
(Zhuko a and Kha lamenko, 1999), and some essen ial amino acids
(EAA) such as me hionine (Abbaszadeh e al., 2022). In his sense,
se e al s udies ha e demons a ed ha he inco po a ion o li e ood,
such as dia oms, o i e s, A emia o copepods, in o BFT can enhance he
nu i ional alue o he bio locs, as well as he g ow h and su i al a es
o he cul i a ed species (Abbaszadeh e al., 2022; B i o e al., 2016; de
And ade e al., 2021). Bio loc en ichmen wi h amphipods has been
add essed in only one p e ious s udy (P om hale e al., 2021). These
au ho s epo ed a high su i al a e o amphipods in he BFT condi ions
and a high po en ial o he en iched bio locs o eplace ishmeal. Am-
phipods cul i a ion in e es has inc eased in he las yea s due o hei
high-quali y nu i ional p o ile (Baeza-Rojano e al., 2014; Gue a-
Ga cía e al., 2016; Jim´
enez-P ada e al., 2018) adequa e o eeding
high ma ke alue ishes o cephalopods (Baeza-Rojano e al., 2010;
Baeza-Rojano e al., 2013; Suon ama e al., 2007). Indeed, he Spanish
Ins i u e o Oceanog aphy (IEO) holds a pa en o he me hodology o
cul i a ion o he common oc opus pa ala ae (Tu e al., 2020). This
pa en delinea es pa icula zoo echnical condi ions and he u ilisa ion
o amphipods as an al e na i e li e p ey o c us acean zoeae, and is
cu en ly being u ilised by he company “Nue a Pescano a” o he
cul i a ion o Oc opus ulga is. Howe e , he upscaling o he sys ems
equi es inc easing amoun s o li e ood, being a bo leneck o he
de elopmen o hese species aquacul u e (Ga cía-Fe n´
andez, 2022).
The ma ine amphipod Gamma us insensibilis S ock, 1966 has been
highligh ed as a p omising species o being in ensi ely cul u ed due o
i s adequa e nu i ional p o ile, la ge body size and high na u al den-
si ies (Jim´
enez-P ada e al., 2020). Mo eo e , as a de i i o ous species,
G. insensibilis can eed on aquacul u e sludge and was es (Cas illa-Ga -
il´
an e al., 2023).
Wi h his backg ound, we sugges ha G. insensibilis has he po en ial
o u ilisa ion as a li e ood sou ce and could be cul i a ed in ensi ely in
BFT sys ems, he eby en iching bio locs nu i ional alue. Mo eo e ,
hei abili y o eed on de i us would pe mi he alo isa ion o aqua-
cul u e was es and he bio emedia ion o he e luen s (Cas illa-Ga il´
an
e al., 2023). In he p esen s udy, G. insensibilis, ed wi h comme cial
aqua eed o de i us om aquacul u e e luen s, has been cul i a ed in
BFT sys ems. The wa e quali y, he su i al a e and he nu i ional
p o ile o he amphipods as well as he bio loc p oduced we e analysed.
2. Ma e ials and me hods
Specimens o G. insensibilis we e sampled in Ma ch 2023 om a
na u al ma sh pond a he IFAPA Cen e “El To u˜
no” (El Pue o de San a
Ma ia, Cadiz Bay, Spain). Amphipods on hei o iginal sub ac (Ul a sp.
halli, whe e hey mainly inhabi ) we e anspo ed o IFAPA acili ies
and ans e ed in o a 400 L ank o allow hei acclima ion du ing a
week p io expe imen a ion. This ank was connec ed o a eci cula ion
sys em (RAS) equipped wi h a cooling, mechanical il e , p o ein
skimme , ul a iole ligh s and bio il e (Cas illa-Ga il´
an e al., 2023).
They we e illumina ed (1.5–1.7
μ
mol pho ons m
−2
s
−1
) wi h luo es-
cen ubes in a 8:16 h (ligh :da kness) o acclima e amphipods o he
da kness, acco ding o P om hale e al. (2021). The seawa e was
main ained a he same condi ions han in he na u al en i onmen ,
17 ◦C and 38 g L
−1
salini y, and was enewed daily in con inuous wa e
low.
P io o he expe imen , 2700 adul s o G. insensibilis we e sampled
om he ank, de oid o algae and placed in 9 cul u e uni s (300 spec-
imens pe cul u e uni ) illed wi h 25 L o ae a ed and mixed seawa e .
Th ee hund ed addi ional “wild” specimens we e sampled and s o ed a
−80 ◦C o lipid p o ile analysis and compa ison wi h he specimens
used in he expe imen . The cul u e uni s we e main ained in da kness
wi hou any wa e exchange, ollowing P om hale e al. (2021). An
a i icial subs a e o amphipods was p o ided in all cul u e uni s
(plas ic mesh, ollowing Cas illa-Ga il´
an e al., 2023). Amphipods we e
s a ed o 24 h o s anda dize hei nu i ional condi ions. Two
expe imen al ea men s we e es ed agains a con ol, a 20 ◦C
(P om hale e al., 2021), in iplica e cul u e uni s. In he i s ea men ,
G. insensibilis specimens we e ed wi h g ounded sea b eam comme cial
pelle s (0.2 g day
−1
). In he second ea men , he eed was was e
de i us (1.2 g day
−1
) ob ained h ough he cleaning o he il e s o a
Senegalese sole RAS uni and consis ed p ima ily o ish aeces and
unea en ish eed pelle s. The quan i y o ood was de e mined by
conside ing he o ganic ma e pe cen age o each ype, wi h de i us
con aining six imes less o ganic ma e han pelle s. Th ee samples o
each ood ype we e also s o ed a −80 ◦C o lipid p o ile analyses.
Be ween h ee and ou imes a week (20 sampling days in o al),
h ee wa e samples we e aken om he anks and il e ed on GF/C
ibe glass il e s ha we e ea ed ollowing Cas illa-Ga il´
an e al.
(2023), in o de o de e mine o al suspended solids in he wa e and he
o ganic ma e con en o hese solids. Wa e was hen analysed: dis-
sol ed ni a e and ni i e concen a ions we e sequen ially quan i ied by
he educ ion o NO
3
−
o NO
2
−
wi h anadium (VCl
3
) ollowing he
me hod o Ga cía-Robledo e al. (2014). Ammonium concen a ion was
de e mined by he indophenol-blue me hod (Amino e al., 1997), and
ca bon concen a ion was hen adjus ed o a 16:1C:N a io h ough he
addi ion o co n lou (P om hale e al., 2021). The con ol ea men
was also ed wi h de i us, and wa e was also sampled daily o moni-
o ing pu poses and ammonium concen a ion analyses, bu no ca bon
sou ce was added in o de o a oid he bio- loccula ion p ocess. Wa e
pa ame e s (mean dissol ed oxygen, empe a u e, salini y and pH) we e
checked daily in all anks and we e ound o be 6.1 ±0.04 mg L
−1
DO,
21.1 ±0.33 ◦C, 40.1 ±4.14 g L
−1
salini y ( ypical o ma sh en i on-
men s), and 8.1 ±0.09 pH. The expe imen las ed six weeks excep o
he con ol anks, since 100 % mo ali y was obse ed om he second
week. A he end o he expe imen , su i ing amphipods and he sludge
(bio locs) p oduced in he 6 expe imen al cul u e uni s we e sampled,
amphipods we e coun ed and all samples we e s o ed a −80 ◦C p io o
biochemical analysis.
All he eeze samples we e eeze-d ied and g ounded in o a ine
powde . Ashes, mois u e, o al lipids and a y acids p o ile we e ana-
lysed. To al lipids (TL) and a y acids (FA) p o ile me hods we e hose
p e iously desc ibed by Hache o-C uzado e al. (2014). B ie ly, TL we e
ex ac ed om powde ollowing he me hodology o Folch e al. (1957)
wi h sligh modi ica ion by Ch is ie and Han (2012). To ex ac FA, o al
lipid ex ac s we e subjec ed o acid-ca alysed ansme hyla ion. Fa y
M. Cas illa-Ga il´
an e al.
Aquacul u e 597 (2025) 741922
2
acids me hyl es e s (FAME) we e sepa a ed and quan i ied by gas
ch oma og aphy and he iden i ied FA we e compa ed o he au hen ic
s anda ds (FAME Mix C37) and well-cha ac e ized ish oil (Menhaden
Oil, SUPELCO, USA) Ch is ie and Han (2012).
Mean and s anda d de ia ion (SD) we e calcula ed o all da a. S a-
is ical analyses and plo s we e pe o med using RS udio so wa e.
Su i al was analysed h ough a one-way ANOVA es be ween am-
phipods in bo h ea men s and he con ol. Di e ences in he ino ganic
ni ogen concen a ion o he di e en ea men s we e es ed wi h wo-
way ANOVAs ollowing an expe imen al design wi h o hogonal ac o s:
ea men , a ixed ac o wi h h ee le els (con ol s pelle s s de i us)
and ime, a ixed ac o wi h wen y le els (sampling days). Di e ences
in TL be ween amphipods in he di e en ea men s and be ween
p oduced bio locs and die s we e also s udied h ough one-way
ANOVAs. Whe e ANOVAs indica ed signi ican di e ences (p <0.05),
he sou ce o he di e ences was iden i ied using a Tukey es . P io o
ANOVAs, he no mali y was es ed wi h Shapi o-Wilk es (p >0.05) and
he homogenei y o a iances was es ed wi h Le ene es (p >0.05). I
a iances emained he e ogeneous e en a e da a ans o ma ion, un-
ans o med da a we e s ill analysed, as ANOVA is a obus s a is ical
es and is ela i ely una ec ed by he he e ogenei y o a iances
(Unde wood, 1997). In such cases, he le el o signi icance was educed
o <0.01 o a oid ype I e o . Addi ionally, p incipal componen ana-
lyses (PCA) we e conduc ed o FA ma ixes o he o dina ion o he
samples.
3. Resul s
3.1. Su i al a e
Su i al was signi ican ly highe in he anks ed wi h pelle s han in
hose ed wi h de i us (one-way ANOVA; d =2; SS =0.28; MS =0.14; F
alue =7.95; p alue =0.0206; Fig. 1). No amphipods we e obse ed in
he con ol anks om he nin h day.
3.2. Wa e quali y
Rega ding ino ganic ni ogen concen a ions, s a is ics a e summa-
ized in he Table 1. Ammonium (N-NH
4
+
) le els we e signi ican ly
highe in he con ol anks han in he o he ea men s (Fig. 2), wi h no
di e ences obse ed be ween he la e (Table 1). Ni i e (N-NO
2
−
) and
ni a e (N-NO
3
−
) concen a ions we e in gene al low (Fig. 3), and no
di e ences we e ound be ween anks ed wi h pelle s o de i us
(Table 1).
3.3. Nu i ional p o ile
The Fig. 4 illus a es he o ganic ma e (OM) and he o al lipids (TL)
con en o he samples. Signi ican di e ences we e obse ed in TL
con en be ween he expe imen al die s and he bio locs (one-way
ANOVA; d =4, SS =228.68, MS =57.17, F alue =42.14, p alue
<0.001). The pelle s die exhibi ed he signi ican ly highes TL con en
(Tukey es ; p alues <0.001) and he bio loc om he pelle s ea men
p esen ed a signi ican ly highe con en han he bio loc om he
de i us ea men (Tukey es ; p alue =0.047). Conce ning amphipods,
no di e ences we e ound be ween hem (one-way ANOVA; d =2, SS =
14.38, MS =7.19, F alue =1.65, p alue =0.268). Di e ences in he
OM con en we e only ound be ween he die s and he bio locs (one-
way ANOVA; d =3; SS =6447; MS =2148.9; F alue =210.1; p alue
<0.001). The pelle s p esen ed he highes con en (Tukey es ; p alues
<0.001), ollowed by he pelle -based bio loc, which p esen ed a
signi ican ly highe con en han he wo o he ea men s (Tukey es ; p
alues <0.001). No di e ences we e ound be ween de i us-based die
and bio loc (Tukey es ; p alue =0.6509).
The a y acid p o ile (in pe cen age) o he die s and bio locs ana-
lysed and he one-way ANOVAs a e summa ized in he Table 2. The
isoC
16
a y acid, which is cha ac e is ic o bac e ia (Tanaka e al.,
2014), was analysed among he SFA. While he pelle s p esen ed no
e idence o his FA, he de i us die and he di e en bio locs did
(Table 2), he eby co obo a ing he p esence o bio loc- o ming bac-
e ia. Majo FA we e he SFA 16:0 (PA), he MUFAs 18:1n7 (cis-VA) and
18:1n9 (OA), he omega-6 PUFA 18:2n6 (LA) and he omega-3 PUFA
22:6n3 (DHA). The pelle -based die p esen ed he signi ican ly highes
le el o OA (Tukey es ; p alue <0.001). Howe e , he pelle -based
bio loc did no show he same p opo ion o OA, and displayed a
signi ican ly lowe con en han he de i us-based bio loc (Tukey es ; p
alue =0.008) o no di e ences wi h he de i us-based die (Tukey es ;
p alue =0.392). The pelle -based bio loc had he signi ican ly highes
le el o cis-VA (Tukey es ; p alue ≤0.01), while he pelle s p esen ed
he lowe con en (Table 2). These indings suppo he p esence o
bio loc- o ming bac e ia, as cis-VA is a p oduc o many he e o ophic
bac e ial species (Gillan and Sands om, 1985).
In gene al, he de i us-based die p esen ed he signi ican ly highes
con en o SFA and he lowes MUFAs con en . The pelle -based die
exhibi ed he signi ican ly highes con en o omega-3 and omega-6
Fig. 1. Mean amphipods su i al a es (%) in he di e en ea men s. Ba s
ep esen he maximum and minimum su i al a es.
Table 1
Resul s o he wo-way ANOVAs o ammonium, ni i e and ni a e
concen a ions.
Sou ce o a ia ion d SS MS F alue P alue
Ammonium
T ea men (T) 2 11.48 5.74 15.33 0.000
Time ( ) 19 18.40 0.97 2.59 0.002
T: 21 1.80 0.09 0.23 0.999
Residuals 86 32.20 0.37
Le ene Tes 42 4.10 0.343
Tukey Tes
con ol >de i us 0.000
con ol >pelle s 0.000
de i us =pelle s 0.997
Ni i e
T ea men 1 0.00 0.00 0.06 0.805
ime 4 0.00 0.00 5.07 0.006
T: 4 0.00 0.00 1.04 0.410
Residuals 20 0.00 0.00
Le ene Tes 9 1.58 0.189
Ni a e
T ea men 1 0.01 0.01 3.12 0.093
ime 4 0.14 0.04 9.73 0.000
T: 4 0.04 0.01 2.38 0.086
Residuals 20 0.07 0.00
Le ene Tes 9 1.37 0.266
M. Cas illa-Ga il´
an e al.
Aquacul u e 597 (2025) 741922
3
PUFAs (Table 2). Howe e , he bio loc gene a ed in his ea men
p esen ed he lowes p opo ion o PUFAs, showing no signi ican di -
e ences wi h he de i us-based die (Tukey es ; p alue =0.990) and a
signi ican ly lowe con en han he de i us-based bio loc (Tukey es ; p
alue =0.010).
The esul s o he PCA based on die s and bio locs we e also in
ag eemen wi h he p e ious esul s and iden i ied OA, cis-VA, LA and
PA as majo con ibu o s o he o al a iance. The i s p incipal
componen explained 57 % o he o al a iance and he second p incipal
componen explained 34.5 %. PC1 co ela ed posi i ely wi h he a y
acids OA ( =0.950, n =20, p <0.005) and LA ( =0.925, n =20, p <
0.005), and nega i ely wi h cis-VA ( = − 0.756, n =20, p <0.005). PC2
Fig. 2. Ammonium concen a ion (mean ±SD) in he di e en cul u e uni s du ing he expe imen .
Fig. 3. Ni a e and ni i e concen a ions (mean ±SD) in he di e en cul u e uni s du ing he expe imen .
M. Cas illa-Ga il´
an e al.
Aquacul u e 597 (2025) 741922
4
co ela ed posi i ely wi h PA ( =0.902, n =20, p <0.005) and
nega i ely wi h cis-VA ( = − 0.603, n =20, p <0.005). The axes clea ly
sepa a e he die s om he bio locs, and di e en ia e he bio locs based
on hei PA and cis-VA con en (Fig. 5).
The a y acid composi ion o he wild amphipods was compa ed
wi h ha o he amphipods used in he expe imen ( ed wi h pelle s o
de i us). Table 3 shows esul s (in %) and one-way ANOVAs. The majo
a y acids we e: he sa u a ed a y acids (SFA) 16:0 (palmi ic acid; PA)
and 18:0 (s ea ic acid; SA); he monounsa u a ed a y acids (MUFAs)
16:1n7 (palmi oleic acid, POA), 18:1n9 (oleic acid, OA) and 18:1n7 (cis-
accenic acid; cis-VA); and he polyunsa u a ed a y acids (PUFAs)
18:2n6 (linoleic acid, LA), 20:4n6 (a achidonic acid, ARA), 20:5n3
(eicosapen aenoic acid, EPA) and 22:6n3 (docosahexaenoic acid, DHA).
Only o al SFA composi ion showed signi ican di e ences be ween he
amphipods, wi h he wild animals displaying a highe concen a ion
han hose used in he expe imen (Tukey es ; p alue <0.001). This was
mainly a ibu ed o a highe indi idual PA con en (Table 2). Con-
ce ning MUFAs, wild amphipods p esen ed a signi ican ly highe con-
en o POA while amphipods ed wi h pelle s showed a highe con en o
OA (Table 2). Wild amphipods also showed he highes con en o
omega-6 PUFAs due o he highe LA and ARA le els, while he am-
phipods used in he expe imen p esen ed highe con en s o omega-3
DHA. The p incipal di e en ia ing ac o s be ween he wo eeding e-
gimes we e he signi ican ly highe le els o PA (Tukey es ; p alue =
0.012) and OA (Tukey es ; p alue <0.001) obse ed in he pelle s
g oup.
P incipal componen s analyses (PCA) we e consis en wi h he e-
sul s shown abo e. The i s p incipal componen explained 57 % o he
o al a iance and he second p incipal componen explained 23 %.
P incipal componen 1 (PC1) co ela ed posi i ely wi h he a y acids
DHA ( =0.854, n =9, p <0.005) and cis-VA ( =0.705, n =9, p <
0.025), and nega i ely wi h PA ( = − 0.958, n =9, p <0.005), EPA ( =
−0.695, n =9, p <0.025), ARA ( = − 0.934, n =9, p <0.025) and LA (
= − 0.718, n =9, p <0.025). PC2 co ela ed posi i ely wi h OA ( =
0.821, n =9, p <0.005). Axes clea ly sepa a ed samples by ea men s
(wild, pelle s o de i us) (Fig. 6).
4. Discussion
4.1. Su i al a e
The e a e limi ed epo s on he amphipods cul u e. Ne e heless,
he su i al a e obse ed in his s udy aligns wi h he indings o
Jim´
enez-P ada e al. (2020), who epo ed alues o app oxima ely 40 %
o e a 21-days expe imen . Simila ly, Ribes-Na a o e al. (2022)
obse ed compa able su i al a es o Gamma us locus a when wa e a
20 ◦C and/o die s ich in sho -chain PUFAs and SFAs we e u ilised in
hei expe imen . Howe e , hei esul s indica ed ha lowe empe a-
u es (5–15 ◦C) and die s ich in LC-PUFAs we e associa ed wi h
enhanced su i al a es.
4.2. Wa e quali y
No s a is ically signi ican di e ences we e ound in wa e quali y
pa ame e s among he ea men s in his s udy. The esul s on nu ien s
concen a ion in he ea men s anks, when compa ed o he con ol,
con i med he e ec o he ca bon sou ce on he colonisa ion o he -
e o ophic bac e ia and he loccula ion p ocess (C ab, 2010; Ha -
g ea es, 2013; Khanjani e al., 2024a): he concen a ion o ammonium
was signi ican ly lowe , while ni a e cumula es in he ea men anks,
hus indica ing ha ni i ying bac e ia we e p esen in he bio loc and
being consis en wi h p e ious wo ks (Abbaszadeh e al., 2022; B i o
e al., 2016).
4.3. Nu i ional p o ile
The loccula ion success was also co obo a ed by he lipid p o ile o
he bio locs, which exhibi ed majo a y acids including PA, cis-VA, OA
and LA, as al eady obse ed in p e ious wo k (Anand e al., 2014).
Essen ial LC-PUFAs we e no among he majo FA. Howe e , he p esen
s udy e ealed clea ly highe amoun s o ARA (0.8 %), EPA (3.1 %) and
DHA (5.5 %) han Anand e al. (2014), who ound only 0.4 % ARA and
did no de ec DHA o EPA in hei bio loc. These alues we e simila o
he FA p o ile o he comme cial aqua eed used in he p esen s udy
(pelle s), which is o mula ed o mee he nu i ional needs o ish (see
Table 2: no di e ences we e ound in he ARA, EPA and DHA con en
be ween he pelle s and he de i us-based bio loc). This jus i ies he
en ichmen o he bio loc wi h amphipods. Indeed, amphipods showed a
p o ile ich in EPA, DHA, LA and ARA, as also demons a ed by Jim´
enez-
P ada e al. (2018). The gamma ids om bo h ea men s showed he
same amoun o TL as he wild specimens, ye demons a ed a educ ion
in ARA and EPA, which was coun e balanced by a highe amoun o
DHA. In e es ingly, Ribes-Na a o e al. (2021) ha e demons a ed ha
gamma id amphipods a e de oid o desa u ase genes ha a e comple-
men a y o he elongases needed o he biosyn hesis o LC-PUFAs.
Consequen ly, hey a e unable o syn hesise hese compounds de
no o. Simila ly o he s udy conduc ed by P om hale e al. (2021), ou
expe imen was pe o med in da kness. Thus, i ’s unlikely ha hese
compounds we e acqui ed h ough he die (phy oplank on). I has been
p oposed ha he high pe cen ages o DHA de ec ed in gamma ids, e en
i ed wi h die s lacking LC-PUFAs, a e he esul o an accumula ion o
p e ious die s (Ribes-Na a o e al., 2022), which could explain he
esul s obse ed in he p esen s udy. These selec i e e en ion mecha-
nisms would enable hem o su i e du ing pe iods o limi ed o no
bioa ailabili y o hese nu ien s (Ribes-Na a o e al., 2022).
Fig. 4. O ganic ma e (OM) and o al lipids (TL) con en (%) in he wo die s
es ed (pelle s and de i us), in he bio locs sampled om he di e en ea -
men s a he end o he expe imen , and in he amphipods in bo h ea men s
and hose sampled om he wild (mean ±SD).
M. Cas illa-Ga il´
an e al.
Aquacul u e 597 (2025) 741922
5
The de i us-based bio loc p esen ed a highe TL con en and a mo e
nu i ious FA p o ile han he de i us-based die , p omo ed by he
p esence o he mic oo ganism assemblage and he amphipods. Mo e-
o e , his bio loc showed highe pe cen ages o PA, OA and LA han he
pelle -based bio loc. This is p obably due o he highe mo ali y a e in
his ea men , wi h ewe indi iduals being sampled o biochemical
analyses and u he en iching he bio loc. I is impo an o no e ha
hese a e ela i e esul s and ha absolu e alues o hese FA we e
simila in bo h ea men s (da a no shown), since he pelle -based
bio loc showed a signi ican ly highe amoun o TL. Howe e , he
Table 2
Fa y acids composi ion (mean % ±SD) o he die s used in he expe imen (i.e. pelle s, de i us) and he bio locs sampled om hese wo ea men s a he end o he
expe imen .
Fa y acid pelle s de i us Bio loc pelle s Bio loc de i us P alue Tukey
SFA
14:0 1.44 ±0.00 3.81 ±0.13 1.57 ±0.44 2.64 ±0.16 0.000** d >all; bd >bp; p =bp
15:0 0.24 ±0.01 0.78 ±0.01 0.66 ±0.19 0.60 ±0.05 0.017* d >p; p =bp =bd; d =bp =bd
16:0 16.43 ±0.03 38.21 ±2.12 22.06 ±0.92 26.78 ±1.37 0.000*** d >bd >bp >p
iso-C
16
0.00 ±0.00 0.84 ±0.05 1.48 ±0.32 0.79 ±0.15 0.000** bp >bd >d >p; d =bd
17:0 0.33 ±0.00 1.09 ±0.16 1.31 ±0.26 0.48 ±0.31 0.000** d =bp >p; p =bd; bp >bd
18:0 4.87 ±0.09 8.49 ±0.06 8.22 ±1.09 10.37 ±1.03 0.000** bd >bp >p; d =bd
20:0 0.36 ±0.00 0.64 ±0.06 0.82 ±0.09 0.70 ±0.03 0.000*** bp >bd =d >p
22:0 0.29 ±0.02 0.65 ±0.07 1.47 ±0.36 1.19 ±0.22 0.000*** p <bp; p =d =bd; bp =d =bd
24:0 0.19 ±0.00 0.76 ±0.08 1.08 ±0.27 1.16 ±0.16 0.000*** bd =bp =d >p
∑SFA 24.14 ±0.15 55.28 ±2.46 38.68 ±1.01 44.71 ±2.05 0.000*** d >bd >bp >p
MUFAs
16:1n5 0.03 ±0.03 0.02 ±0.02 0.36 ±0.19 0.39 ±0.23 0.044
16:1n7 3.68 ±0.04 3.21 ±0.13 8.16 ±2.14 5.83 ±1.65 0.001** bp >p =d; bp =bd
18:1n5 0.16 ±0.00 0.06 ±0.01 0.58 ±0.06 0.41 ±0.23 0.000*** bp =bd >d; d =p; bp >p; bd =p
18:1n7 3.31 ±0.09 4.73 ±0.09 18.99 ±5.55 8.01 ±4.27 0.000*** bp >p =d =bd
18:1n9 30.67 ±0.03 9.59 ±0.71 8.30 ±0.87 10.67 ±1.64 0.000*** p >all; bd >bp: d =bp; d =bd
20:1n9 2.30 ±0.01 2.38 ±0.12 0.73 ±0.31 2.42 ±0.33 0.000*** bp <p =d =bd
22:1n11 1.22 ±0.01 2.30 ±0.06 0.87 ±0.41 3.06 ±0.45 0.003** bd >bp; bd =p =d; bp =p =d
24:1 0.51 ±0.01 1.88 ±0.06 1.02 ±0.23 2.39 ±0.28 0.000*** p =bp; d =bd; d >p; bd >bp
∑MUFAs 41.89 ±0.09 24.16 ±0.84 39.01 ±2.72 33.19 ±3.98 0.000*** p =bp >bd >d
n-6 PUFAs
18:2n6 14.93 ±0.01 4.15 ±0.21 3.96 ±0.33 5.04 ±0.23 0.000*** p >bd >bp >d
20:4n6 0.85 ±0.00 0.41 ±0.12 0.68 ±0.33 0.80 ±0.24 0.440
∑n-6 PUFAs 15.78 ±0.02 4.56 ±0.34 4.64 ±0.24 5.84 ±0.17 0.000*** p >all; d =bp; bd >d; bd >bp
n-3 PUFAs
18:3n3 2.36 ±0.01 0.48 ±0.02 0.32 ±0.25 0.37 ±0.12 0.000*** p >d =bp =bd
20:5n3 3.68 ±0.00 2.28 ±0.15 1.87 ±0.59 3.11 ±0.90 0.002** p >bp; p =d =bd; bp =d =bd
22:5n3 0.94 ±0.00 0.46 ±0.01 0.21 ±0.03 0.48 ±0.08 0.000** p >all; bd >bp; bd =d
22:6n3 6.73 ±0.03 3.76 ±0.18 3.92 ±1.12 5.54 ±1.66 0.010* p >d =bp; d =bp =bd; p =bd
∑n-3 PUFAs 13.72 ±0.04 6.99 ±0.37 6.31 ±1.73 9.50 ±2.63 0.000*** p >all; bd >bp; d =bd; d =bp
∑PUFAs 29.49 ±0.03 11.56 ±0.70 10.95 ±1.87 15.34 ±2.77 0.000*** p >all; bd >bp;d =bd; d =bp
p =pelle -based die ; d =de i us-based die ; bp =pelle -based bio loc; bd =de i us-based bio loc; SFA =sa u a ed a y acids; MUFA =monounsa u a ed a y acids;
PUFA =polyunsa u a ed a y acid.
Fig. 5. P incipal componen analysis (PCA) plo based on FA composi ion (in % o FA) o expe imen al die s (i.e. pelle s o de i us) and bio loc sampled om he
expe imen al anks. Only majo FA a e shown. 16:0 =PA, 18:1n7 =cis-VA, 18:1n9 =OA, 18:2n6 =LA. D =de i us; P =pelle s; BD =de i us-based bio loc; BP =
pelle -based bio loc.
M. Cas illa-Ga il´
an e al.
Aquacul u e 597 (2025) 741922
6
pelle -based bio loc did no p esen a be e p o ile han he pelle -based
die , highligh ing he nu i ional alue o he die , bu also poin ing he
in e es o alo ising aquacul u e was es in BFT sys ems (Das e al.,
2023; Kala e al., 2023). Fu he mo e, he pelle -based bio loc exhibi ed
a signi ican ly highe pe cen age o isoC
16
and cis-VA, commonly
associa ed wi h bac e ia (Ron ani e al., 2003). This is a ibu ed o he
signi ican ly highe con en o OM in he pelle s and pelle -based bio loc,
which p o ides essen ial nu ien s and acili a es he es ablishmen o
he e o ophic bac e ia and he con e sion o he OM in o bac e ial
biomass (Zhuko a and Kha lamenko, 1999). This demons a es he
ad an age o he pelle s ea men in e ms o bio loccula ion capaci y
(Faus , 2014), in addi ion o he imp o ed su i al a es. I has been
shown ha he FA p o ile o amphipods as well as hei su i al a e a e
closely linked o he die (Ribes-Na a o e al., 2022). Thus, he in e-
g a ion in BFT sys ems o aquacul u e was e de i us wi h a supple-
men a ion o comme cial aqua eed could po en ially enhance he o e all
nu i ional p o ile while enhancing he bio loccula ion p ocess o he
bio emedia ion o he aquacul u e e luen s (Cas illa-Ga il´
an e al.,
2023). Mo eo e , i will be sui able o ca y ou he expe imen unde
ligh condi ions and wi h he addi ion o dia oms o chlo ophy es, which
ha e been highligh ed by o he au ho s o imp o e he LC-PUFAs and
EAA p o iles (Kha oon e al., 2009; Ma ins e al., 2016), and wa e
quali y (B i o e al., 2016; de And ade e al., 2021).
The in e es in in eg a ed BFT sys ems (a combina ion o in eg a ed
mul i- ophic aquacul u e, IMTA, and BFT) has inc eased in ecen yea s
(Bo ges e al., 2020; Ca alho e al., 2023; Lega da e al., 2021; Lima
e al., 2021). In hese sys ems, he esidues o one ed species se e as a
sou ce o OM o bio locs, which can be en iched wi h a second species,
which in u n is ed o a hi d species. This app oach aims o minimize
he p oduc ion o was e and he use o aqua eed. The p esen s udy
shows ha he e luen o a ish RAS can be alo ised o eed he medium
o a BFT sys em en iched wi h amphipods. Bo h bio locs and amphipods
could be u he used as ood, as hei a y acids p o ile would sa is y
he equi emen s o a ious species o in e es in aquacul u e as a e
salmonids (Ca e al., 2023), ma ine c us aceans (Gonz´
alez-F´
elix e al.,
2002), sea b eam (Ibeas e al., 1996) o u bo (Cas ell e al., 1994). The
use o bio locs o eed ish has been demons a ed o be an e ec i e
app oach, as e idenced by he indings o se e al s udies (Bo ges e al.,
2020; Holanda e al., 2023; Lega da e al., 2021; Pinho e al., 2021;
Poe sch e al., 2021; Sil a e al., 2022). Fu he mo e, amphipods ha e
also been success ully employed as a die a y sou ce o ish (Mo en e al.,
2006; Suon ama e al., 2007) and cephalopods (Baeza-Rojano e al.,
2010; Baeza-Rojano e al., 2013). Finally, he p esen s udy suppo s he
g owing in e es in he use o amphipods in aquacul u e, wi h a
pa icula ocus on BFT sys ems. I aims o p omo e u he esea ch in
his opic, gi en ha o da e, BFT sys ems ha e been p edominan ly
s udied in mono-speci ic o mul i-speci ic decapod c us aceans aqua-
cul u e (Baj acha ya e al., 2024; Ekasa i e al., 2014; Eme enciano
e al., 2021; Galasso e al., 2024; Huang e al., 2023; Lima e al., 2021;
Nguyen e al., 2024; P´
e ez de Jesús e al., 2024; P´
e ez-Velasco e al.,
2023; Pimen el e al., 2024; Qiu e al., 2023; Reis e al., 2023). Few
s udies ha e been conduc ed on echinode ms (Chen e al., 2018a,
2018b) and only a single s udy has been ca ied ou on amphipod
c us aceans (P om hale e al., 2021).
5. Conclusion
This s udy demons a es he po en ial o using G. insensibilis in BFT
sys ems, ocusing on i s su i al and nu i ional p o ile, when ed wi h
comme cial aqua eed o de i us om aquacul u e e luen s. The ind-
ings e eal ha G. insensibilis can be e ec i ely cul i a ed in BFT sys-
ems, le e aging i s abili y o eed on de i us, which p omo es
bio loccula ion and enhances he nu ien p o ile o he bio loc. The
p esence o bio locs en iched wi h amphipods signi ican ly imp o es he
o e all a y acid composi ion, pa icula ly in essen ial LC-PUFAs such
as EPA, DHA, and ARA. This en ichmen add esses he inhe en nu i-
ional de iciencies o bio locs, making hem mo e sui able as ood o
high- alue aquacul u e species. The s udy highligh s he dual bene i s o
his app oach: enhancing he nu i ional alue o bio locs while
con ibu ing o he bio emedia ion o aquacul u e e luen s. Fu he -
mo e, he use o close aquacul u e sys ems educes cos s and main ain
Table 3
Fa y acids composi ion (mean % ±SD) o G. insensibilis om sal ma shes and
hose used in he expe imen , ed wi h ish pelle s o de i us.
Fa y acid wild pelle s de i us P alue Tukey
SFA
14:0 1.46 ±
0.12
0.75 ±
0.15
0.63 ±
0.17 0.000*** w >p =d
15:0 0.32 ±
0.27
0.41 ±
0.02
0.47 ±
0.02 0.538
16:0 20.40 ±
0.14
16.44 ±
0.37
15.47 ±
0.26 0.000*** w >p >d
16:0i 0.18 ±
0.03
0.90 ±
0.03
0.79 ±
0.39 0.057
17:0 0.12 ±
0.01
1.03 ±
0.04
1.22 ±
0.05 0.000*** w <p <d
18:0 3.46 ±
0.12
3.83 ±
0.15
4.39 ±
0.15 0.000*** w <p <d
22:0 0.30 ±
0.26
0.37 ±
0.16
0.70 ±
0.06 0.073
∑SFA 26.23 ±
0.51
23.74 ±
0.42
23.67 ±
0.29 0.000 w >p =d
MUFAs
16:1n7 5.97 ±
0.28
3.67 ±
0.29
4.16 ±
1.42 0.036* w >p =d
18:1n9 15.89 ±
0.27
22.17 ±
0.85
16.14 ±
0.95 0.000*** p >w =d
18:1n7 4.30 ±
0.16
6.48 ±
1.25
8.35 ±
3.17 0.115
18:1n5 0.19 ±
0.01
0.65 ±
0.28
0.77 ±
0.39 0.092
20:1n9 0.80 ±
0.03
1.22 ±
0.03
0.79 ±
0.06 0.000*** p >w =d
20:1n7 0.33 ±
0.04
0.52 ±
0.11
0.57 ±
0.26 0.243
∑MUFAs 27.47 ±
0.63
34.70 ±
2.55
30.77 ±
4.40 0.064
n-6 PUFAs
18:2n6 7.43 ±
0.09
6.77 ±
0.63
5.69 ±
0.69 0.021* w >d; w =p;
p =d
20:2n6 0.94 ±
0.02
0.85 ±
0.17
0.84 ±
0.18 0.636
20:4n6 7.59 ±
0.12
4.90 ±
0.52
5.49 ±
1.23 0.013* w >p =d
22:5n6 0.55 ±
0.01
0.77 ±
0.15
0.64 ±
0.08 0.084
∑n-6
PUFAs
15.51 ±
0.22
13.29 ±
1.44
12.66 ±
2.13 0.040* w >d; w =p;
p =d
n-3 PUFAs
18:3n3 0.75 ±
0.04
0.37 ±
0.32
0.19 ±
0.34 0.108
20:5n3 15.25 ±
0.28
11.37 ±
0.87
13.43 ±
3.34 0.136
22:5n3 0.97 ±
0.03
0.56 ±
0.09
0.76 ±
0.15 0.008** w >p; w =d;
p =d
22:6n3 4.82 ±
0.00
9.47 ±
0.41
11.81 ±
2.17 0.002** w <p =d
∑n-3
PUFAs
21.79 ±
0.26
21.77 ±
1.21
26.19 ±
4.83 0.176
16:2n4 0.44 ±
0.48
0.05 ±
0.05
0.39 ±
0.07 0.261
16:4n1 1.08 ±
0.47
0.87 ±
0.60
1.33 ±
0.30 0.526
∑PUFAs 39.82 ±
1.12
35.98 ±
2.66
40.57 ±
6.36 0.386
w =wild; p =pelle s; d =de i us; SFA =sa u a ed a y acids; MUFA =
monounsa u a ed a y acids; PUFA =polyunsa u a ed a y acid.
M. Cas illa-Ga il´
an e al.
Aquacul u e 597 (2025) 741922
7
isola ion om na u al en i onmen . The in eg a ion o aquacul u e
de i us wi h supplemen al aqua eed in BFT sys ems will suppo bo h
he bio loccula ion p ocess and he nu i ional needs o he amphipods,
he eby c ea ing a sus ainable and e icien cycle o was e alo isa ion
and ood p oduc ion.
The use o G. insensibilis in BFT sys ems ep esen s a p omising
s a egy o imp o ing he sus ainabili y and p oduc i i y o aquacul u e
ope a ions. By op imizing he use o o ganic was e and p oducing
nu i ionally ich bio locs, his app oach can signi ican ly con ibu e o
he de elopmen o a ci cula economy in aquacul u e, educing en i-
onmen al impac s and enhancing he g ow h and heal h o cul i a ed
species. Fu he esea ch is needed in o de o s udy he comple e li e
cycle o amphipods and hei ep oduc i e capaci y in hese sys ems. A
lowe wa e empe a u e and he inocula ion o mic oalgae should be
conside ed in o de o achie e highe su i al a es and PUFAs con en .
CRediT au ho ship con ibu ion s a emen
Ma a Cas illa-Ga il´
an: W i ing – e iew & edi ing, W i ing –
o iginal d a , Valida ion, Me hodology, In es iga ion, Fo mal analysis,
Da a cu a ion, Concep ualiza ion. Jos´
e Manuel Gue a-Ga cía: Vali-
da ion, Supe ision, Me hodology, Da a cu a ion. Ismael Hache o-
C uzado: Valida ion, Supe ision, Me hodology, Da a cu a ion,
Concep ualiza ion.
Decla a ion o compe ing in e es
The au ho s decla e ha hey ha e no known compe ing inancial
in e es s o pe sonal ela ionships ha could ha e appea ed o in luence
he wo k epo ed in his pape .
Da a a ailabili y
Da a will be made a ailable on eques .
Acknowledgemen s
Au ho s wish o hank he Eu opean Ma i ime and Fishe ies Fund
(EMFF) o he unding du ing his esea ch included in he p ojec
“Acuicul u a y ci cula idad: alo izaci´
on de esiduos de la acuicul u a
median e o ganismos de bajo ni el ´
o ico (ACUICIRC)” unde ag ee-
men EI.FEM.PPA202200.003. Du ing his s udy MCG was suppo ed by
a “Ma ga i a Salas” pos -doc o al g an om he Spanish Minis y o
Science, Inno a ion and Uni e si ies unded by he Eu opean Union –
Nex Gene a ion EU p og am. Thank you o Melissa San ’ Anna da Sil a
o he con ibu ion du ing he mas e ’s in e nship.
Re e ences
Abbaszadeh, A., Mozanzadeh, M.T., Qasemi, A., Ouji a d, A., Na isi Bahabadi, M., 2022.
E ec s o he addi ion o Calanopia ellip ica, A emia anciscana, and B achionus
o undi o mis in a nu se y bio loc sys em on wa e quali y, g ow h, gu mo phology,
heal h indices, and ansc ip ional esponse o immune and an ioxidan - ela ed
genes in Penaeus annamei. Aquac. In . 30, 653–676. h ps://doi.o g/10.1007/
s10499-021-00823-1.
Amino , A., Ki kwood, D.S., K´
e ouel, R., 1997. De e mina ion o ammonia in seawa e by
he indophenol-blue me hod: e alua ion o he ICES NUTS I/C 5 ques ionnai e. Ma .
Chem. 56, 59–75. h ps://doi.o g/10.1016/S0304-4203(96)00080-1.
Anand, P.S.S., Kohli, M.P.S., Kuma , S., Sunda ay, J.K., Roy, S.D., Venka eshwa lu, G.,
Sinha, A., Pailan, G.H., 2014. E ec o die a y supplemen a ion o bio loc on g ow h
pe o mance and diges i e enzyme ac i i ies in Penaeus monodon. Aquacul u e
418–419, 108–115. h ps://doi.o g/10.1016/j.aquacul u e.2013.09.051.
Baeza-Rojano, E., Ga cía, S., Ga ido, D., Gue a-Ga cía, J.M., Domingues, P., 2010. Use
o amphipods as al e na i e p ey o cul u e cu le ish (Sepia o icinalis) ha chlings.
Aquacul u e 300, 243–246.
Baeza-Rojano, E., Domingues, P., Gue a-Ga cía, J.M., Capella, S., No e˜
na-Ba oso, E.,
Caamal-Mons eal, C., Rosas, C., 2013. Ma ine gamma ids (C us acea: Amphipoda): a
new li e p ey o cul u e Oc opus maya ha chlings. Aquacul . Res. 44, 1602–1612.
Baeza-Rojano, E., Hache o-C uzado, I., Gue a-Ga cía, J.M., 2014. Nu i ional analysis o
eshwa e and ma ine amphipods om he s ai o Gib al a and po en ial
aquacul u e applica ions. J. Sea Res. 85, 29–36.
Baj acha ya, S., Fisk, J.C., Fleckens ein, L.J., Ray, A.J., 2024. Sal ype, suga addi ion,
and sys em ype in in ensi e RAS o Paci ic whi e sh imp (Li openaeus annamei)
p oduc ion. Aquacul u e 586, 740755.
Bo ges, B.A.A., Rocha, J.L., Pin o, P.H.O., Zacheu, T., Chede, A.C., Magno i, C.C.F.,
Ce quei a, V.R., A ana, L.A.V., 2020. In eg a ed cul u e o whi e sh imp Li openaeus
Fig. 6. P incipal componen analysis (PCA) plo based on FA composi ion (in % o FA) o amphipods om he wild o ed wi h expe imen al die s (i.e. pelle s o
de i us). Only majo FA a e shown. 16:0 =PA, 20:4n6 =ARA, 20:5n3 =EPA, 18:1n9 =OA, 22:6n3 =DHA, 18:1n7 =cis-VA. D =de i us; P =pelle s; W =wild.
Th ee eplica es we e analysed, bo h in he wild and in each ea men (3 anks pe ea men ). Each eplica e consis ed on a pool o amphipods.
M. Cas illa-Ga il´
an e al.
Aquacul u e 597 (2025) 741922
8
annamei and mulle Mugil liza on bio loc echnology: Zoo echnical pe o mance,
sludge gene a ion, and Vib io spp. educ ion. Aquacul u e 524, 735234. h ps://doi.
o g/10.1016/j.aquacul u e.2020.735234.
B i o, L.O., dos San os, I.G.S., de Ab eu, J.L., de A aújo, M.T., Se e i, W., G`
al ez, A.O.,
2016. E ec o he addi ion o dia oms (Na icula spp.) and o i e s (B achionus
plica ilis) on wa e quali y and g ow h o he Li openaeus annamei pos la ae ea ed
in a bio loc sys em. Aquacul . Res. 47, 3990–3997.
Ca , I., Glenc oss, B., San igosa, E., 2023. The impo ance o essen ial a y acids and
hei a ios in aqua eeds o enhance salmonid p oduc ion, wel a e, and human
heal h. F on . Anim. Sci. 4, 1147081.
Ca alho, A., Cos a, L.C.O., Holanda, M., Poe sch, L.H., Tu an, G., 2023. In luence o
To al suspended solids on he g ow h o he sea le uce Ul a lac uca in eg a ed wi h
he Paci ic whi e sh imp Li openaeus annamei in a bio loc sys em. Fishes 8, 163.
h ps://doi.o g/10.3390/ ishes8030163.
Cas ell, J.D., Bell, J.G., Toche , D.R., Sa gen , J.R., 1994. E ec s o pu i ied die s
con aining di e en combina ions o a achidonic and docosahexaenoic acid on
su i al, g ow h and a y acid composi ion o ju enile u bo (Scoph halmus
maximus). Aquacul u e 128, 315–333.
Cas illa-Ga il´
an, M., Gue a-Ga cía, J., Mo eno-Oli a, J., Hache o-C uzado, I., 2023.
How much was e can he amphipod Gamma us insensibilis emo e om aquacul u e
e luen s? A i s s ep owa d IMTA. Aquacul u e 573, 739552.
Chen, J., Ren, Y., Li, Y., Xia, B., 2018a. Regula ion o g ow h, in es inal mic obio a, non-
speci ic immune esponse and disease esis ance o sea cucumbe Apos ichopus
japonicus (Selenka) in bio loc sys ems. Fish Shell ish Immunol. 77, 175–186. h ps://
doi.o g/10.1016/j. si.2018.03.053.
Chen, J., Ren, Y., Wang, G., Xia, B., Li, Y., 2018b. Die a y supplemen a ion o bio loc
in luences g ow h pe o mance, physiological s ess, an ioxidan s a us and immune
esponse o ju enile sea cucumbe Apos ichopus japonicus (Selenka). Fish Shell ish
Immunol. 72, 143–152.
Ch is ie, W.W., Han, X., 2012. Chap e 4 - analysis o simple lipid classes. In: Ch is ie, W.
W., Han, X. (Eds.), Lipid Analysis, Fou h edi ion. Oily P ess Lipid Lib a y Se ies.
Woodhead Publishing, pp. 69–90. h ps://doi.o g/10.1533/9780857097866.69.
C ab, R., 2010. Bio locs Technology: An In eg a ed Sys em o he Remo al o Nu ien s
and Simul aneous P oduc ion o Feed in Aquacul u e. Ghen Uni e si y.
C ab, R., Chielens, B., Wille, M., Bossie , P., Ve s ae e, W., 2010a. The e ec o di e en
ca bon sou ces on he nu i ional alue o bio locs, a eed o Mac ob achium
osenbe gii pos la ae. Aquacul . Res. 41, 559–567.
C ab, R., Lambe , A., De oi d , T., Bossie , P., Ve s ae e, W., 2010b. The applica ion o
bio locs echnology o p o ec b ine sh imp (A emia anciscana) om pa hogenic
Vib io ha eyi. J. Appl. Mic obiol. 109, 1643–1649.
C ab, R., De oi d , T., Bossie , P., Ve s ae e, W., 2012. Bio loc echnology in
aquacul u e: bene icial e ec s and u u e challenges. Aquacul u e 356, 351–356.
Das, S.K., Mandal, A., 2021. Supplemen a ion o bio loc in ca p (Cyp inus ca pio a .
Communis) cul u e as a po en ial ool o esou ce managemen in aquacul u e.
Aqua . Li ing Resou . 34, 20. h ps://doi.o g/10.1051/al /2021019.
Das, S.K., Mondal, B., Sa ka , U.K., Das, B.K., Bo ah, S., 2023. Unde s anding and
app oaches owa ds ci cula bio-economy o was ewa e euse in ishe ies and
aquacul u e in India: an o e iew. Re . Aquac. 15, 1100–1114. h ps://doi.o g/
10.1111/ aq.12758.
de And ade, R.J.V., dos San os, E.P., de Almeida Cos a, G.K., da Sil a Campos, C.V.F., da
Sil a, S.M.B.C., G´
al ez, A.O., B i o, L.O., 2021. E ec o di e en equencies o he
addi ion o B achionus plica ilis on he pe o mance o Li openaeus annamei in a
nu se y bio loc sys em wi h ice b an (anae obic and ae obic) as an o ganic ca bon
sou ce. Aquacul u e 540, 736669.
Dinda, R., Mandal, A., Das, S.K., 2020. Neem (Azadi ach a indica a. Juss) supplemen ed
bio loc medium as al e na i e eed in common ca p (Cyp inus ca pio a . communis
Linnaeus) cul u e. J. Appl. Aquac. 32, 361–379. h ps://doi.o g/10.1080/
10454438.2019.1645076.
Ebeling, J.M., Timmons, M.B., Bisogni, J., 2006. Enginee ing analysis o he
s oichiome y o pho oau o ophic, au o ophic, and he e o ophic emo al o
ammonia–ni ogen in aquacul u e sys ems. Aquacul u e 257, 346–358.
Ekasa i, J., Angela, D., Waluyo, S.H., Bach ia , T., Su awidjaja, E.H., Bossie , P., De
Sch y e , P., 2014. The size o bio loc de e mines he nu i ional composi ion and
he ni ogen eco e y by aquacul u e animals. Aquacul u e 426, 105–111.
El-Sayed, A.M., 2021. Use o bio loc echnology in sh imp aquacul u e: a comp ehensi e
e iew, wi h emphasis on he las decade. Re . Aquac. 13, 676–705.
Eme enciano, M.G., Mi anda-Baeza, A., Ma ínez-Po chas, M., Poli, M.A., Viei a, F.,
2021. Bio loc echnology (BFT) in sh imp a ming: pas and p esen shaping he
u u e. F on . Ma . Sci. 8, 813091.
Faus , L., 2014. Bio loccula ion o Was ewa e O ganic Ma e a Sho Re en ion Times.
Wageningen Uni e si y and Resea ch.
Folch, J., Lees, M., Sloane S anley, G.H., 1957. A simple me hod o he isola ion and
pu i ica ion o o al lipids om animal issues. J. Biol. Chem. 226, 497–509.
Galasso, H.L., Owa a i, M.S., A ana, L.A.V., Lapa, K.R., 2024. E ec s o scaled-down
dissol ed ai lo a ion sys em on suspended solids emo al om Penaeus annamei
cul u e unde bio loc condi ions. Aquac. Eng. 104, 102396.
Ga cía-Fe n´
andez, P., 2022. Es udio de la ase la a ia planc ´
onica del pulpo común,
oc opus ulga is cu ie , 1797 en e a dis in as condiciones de cul i o median e el
empleo de ecnologías ´
omicas e iden i icaci´
on de genes bioma cado es.
Ga cía-Robledo, E., Co zo, A., Papaspy ou, S., 2014. A as and di ec spec opho ome ic
me hod o he sequen ial de e mina ion o ni a e and ni i e a low concen a ions
in small olumes. Ma . Chem. 162, 30–36. h ps://doi.o g/10.1016/j.
ma chem.2014.03.002.
Gillan, F.T., Sands om, M.W., 1985. Mic obial lipids om a nea sho e sedimen om
Bowling G een Bay, No h Queensland: he a y acid composi ion o in ac lipid
ac ions. O g. Geochem. 8, 321–328.
Gonz´
alez-F´
elix, M.L., Ga lin III, D.M., Law ence, A.L., Pe ez-Velazquez, M., 2002. E ec
o die a y phospholipid on essen ial a y acid equi emen s and issue lipid
composi ion o Li openaeus annamei ju eniles. Aquacul u e 207, 151–167.
Gue a-Ga cía, J.M., Hache o-C uzado, I., Gonz´
alez-Rome o, P., Jim´
enez-P ada, P.,
Cassell, C., Ros, M., 2016. Towa ds in eg a ed mul i- ophic aquacul u e: lessons
om cap ellids (C us acea: Amphipoda). PloS One 11, e0154776.
Hache o-C uzado, I., Rod íguez-Rua, A., Rom´
an-Padilla, J., Ponce, M., Fe n´
andez-
Díaz, C., Manchado, M., 2014. Cha ac e iza ion o he genomic esponses in ea ly
Senegalese sole la ae ed die s wi h di e en die a y iacylglyce ol and o al lipids
le els. Comp. Biochem. Physiol. Pa D Genomics P o eomics 12, 61–73.
Ha g ea es, J.A., 2013. Bio loc P oduc ion Sys ems o Aquacul u e. Sou he n Regional
Aquacul u e Cen e S one ille, MS.
Holanda, M., Ra agnan, E., La a, G., San ana, G., Fu ado, P., Ca dozo, A.,
Wasielesky J ., W., Poe sch, L.H., 2023. In eg a ed mul i ophic cul u e o sh imp
Li openaeus annamei and ilapia O eoch omis nilo icus in bio loc sys em: a pilo scale
s udy. F on . Ma . Sci. 10, 1060846.
Huang, H.-H., Li, C.-Y., Lei, Y.-J., Zhou, B.-L., Kuang, W.-Q., Zou, W.-S., Yang, P.-H.,
2023. E ec s o Bacillus s ain added as ini ial indigenous species in o he bio loc
sys em ea ing Li openaeus annamei ju eniles on bio loc p e o ma ion, wa e quali y
and sh imp g ow h. Aquacul u e 569, 739375.
Ibeas, C., Cejas, J., Gomez, T., Je ez, S., Lo enzo, A., 1996. In luence o die a y n−3
highly unsa u a ed a y acids le els on ju enile gil head seab eam (Spa us au a a)
g ow h and issue a y acid composi ion. Aquacul u e 142, 221–235.
Jim´
enez-P ada, P., Hache o-C uzado, I., Gi ´
aldez, I., Fe n´
andez-Diaz, C., Vilas, C.,
Ca˜
na a e, J.P., Gue a-Ga cía, J.M., 2018. C us acean amphipods om ma sh ponds:
a nu i ious eed esou ce wi h po en ial o applica ion in in eg a ed mul i- ophic
aquacul u e. Pee J 6, e4194.
Jim´
enez-P ada, P., Hache o-C uzado, I., Gue a-Ga cía, J., 2020. Aquacul u e was e as
ood o amphipods: he case o Gamma us insensibilis in ma sh ponds om sou he n
Spain. Aquac. In . 29, 139–153.
Kala, A., K ishnan, S., Rani, A.M., 2023. Valo isa ion o aquacul u e sludge in o
mic obial p o ein using bio eac o wi h an op imised nu ien s. Biochem. Eng. J.
198, 109014. h ps://doi.o g/10.1016/j.bej.2023.109014.
Khanjani, M.H., Sha i inia, M., 2024. Feeding Nile ilapia wi h a ying le els o bio loc:
e ec on g ow h pe o mance, su i al a e, diges i e and li e enzyme ac i i ies,
and mucus immuni y. Aquac. In . 32, 8171–8194. h ps://doi.o g/10.1007/s10499-
024-01561-w.
Khanjani, M.H., Mohammadi, A., Eme enciano, M.G.C., 2022. Mic oo ganisms in bio loc
aquacul u e sys em. Aquac. Rep. 26, 101300. h ps://doi.o g/10.1016/j.
aq ep.2022.101300.
Khanjani, M.H., Mohammadi, A., Eme enciano, M.G.C., 2024a. Wa e quali y in bio loc
echnology (BFT): an applied e iew o an e ol ing aquacul u e. Aquac. In .
h ps://doi.o g/10.1007/s10499-024-01618-w.
Khanjani, M.H., Mozanzadeh, M.T., Sha i inia, M., Eme enciano, M.G.C., 2024b.
B oods ock and seed p oduc ion in bio loc echnology (BFT): an upda ed e iew
ocused on ish and penaeid sh imp. Aquacul u e 579, 740278. h ps://doi.o g/
10.1016/j.aquacul u e.2023.740278.
Khanjani, M.H., Sha i inia, M., Eme enciano, M.G.C., 2024c. Bio loc echnology (BFT) in
aquacul u e: wha goes igh , wha goes w ong? A scien i ic-based snapsho . Aquac.
Nu . e7496572. h ps://doi.o g/10.1155/2024/7496572.
Kha oon, H., Bane jee, S., Yuso , F.M., Sha i , M., 2009. E alua ion o indigenous
ma ine pe iphy ic Ampho a, Na icula and Cymbella g own on subs a e as eed
supplemen in Penaeus monodon pos la al ha che y sys em. Aquacul . Nu . 15,
186–193. h ps://doi.o g/10.1111/j.1365-2095.2008.00582.x.
Knowle , D., Chopin, T., Ma ínez-Espi˜
nei a, R., Neo i, A., Nob e, A., Noce, A., Reid, G.,
2020. The economics o in eg a ed mul i- ophic aquacul u e: whe e a e we now and
whe e do we need o go? Re . Aquac. 12, 1579–1594. h ps://doi.o g/10.1111/
aq.12399.
Lega da, E.C., da Sil a, D., Mi anda, C.S., Pe ei a, P.K.M., Ma ins, M.A., Machado, C., de
Lo enzo, M.A., Hayashi, L., Do Nascimen o Viei a, F., 2021. Sea le uce in eg a ed
wi h Paci ic whi e sh imp and mulle cul i a ion in bio loc impac sys em
pe o mance and he sea le uce nu i ional composi ion. Aquacul u e 534, 736265.
h ps://doi.o g/10.1016/j.aquacul u e.2020.736265.
Lima, P.C.M., Sil a, A.E.M., Sil a, D.A., Sil a, S.M.B.C., B i o, L.O., G´
al ez, A.O., 2021.
E ec o s ocking densi y o C assos ea sp. in a mul i ophic bio loc sys em wi h
Li openaeus annamei in nu se y. Aquacul u e 530, 735913. h ps://doi.o g/
10.1016/j.aquacul u e.2020.735913.
Ma ins, T.G., Odeb ech , C., Jensen, L.V., D’Oca, M.G., Wasielesky J ., W., 2016. The
con ibu ion o dia oms o bio locs lipid con en and he pe o mance o ju enile
Li openaeus annamei (Boone, 1931) in a BFT cul u e sys em. Aquacul . Res. 47,
1315–1326.
McCuske , S., Wa be g, M.B., Da ies, S.J., Valen e, C.S., Johnson, M.P., Cooney, R.,
Wan, A.H.L., 2023. Bio loc echnology as pa o a sus ainable aquacul u e sys em: a
e iew on he s a us and inno a ions o i s expansion. Aquac. Fish Fish. 3, 331–352.
h ps://doi.o g/10.1002/a 2.108.
Mo en, M., Suon ama, J., Hem e, G.-I., Ka lsen, Ø., Olsen, R., Mundheim, H.,
Julshamn, K., 2006. Elemen concen a ions in meals om k ill and amphipods,—
possible al e na i e p o ein sou ces in comple e die s o a med ish. Aquacul u e
261, 174–181.
Nguyen, T.T.T., Foysal, M.J., Gup a, S.K., Tay, A., Fo eda , R., Gagnon, M.M., 2024.
E ec s o ca bon sou ce addi ion in ea ing wa e on sedimen cha ac e is ics,
g ow h and heal h o cul u ed ma on (Che ax cainii). Sci. Rep. 14, 1349.
P´
e ez de Jesús, D., He n´
andez-Ve ga a, M.P., P´
e ez-Ros o, C.I., F ías-Quin ana, C.A.,
2024. E ec o as ing on compensa o y g ow h and diges i e enzyma ic ac i i y o
eshwa e p awn pos la ae (Mac ob achium osenbe gii) du ing i s cul u e in
bio loc. Aquac. In . 32, 119–135.
M. Cas illa-Ga il´
an e al.
Aquacul u e 597 (2025) 741922
9
