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Consumers feeding at the aquatic–terrestrial ecosystem interface may obtain a mixture of aquatic and terrestrial diet resources that vary in nutritional composition. However, in lake riparian spiders, the relative significance of aquatic versus terrestrial diet sources remains to be explored. We investigated the trophic transfer of lipids and polyunsaturated fatty acids (PUFA) from emergent aquatic and terrestrial insects to spiders at varying distances from the shoreline of a subalpine lake in Austria, using differences in fatty acid profiles and compound-specific stable carbon (δ¹³C) and hydrogen (δ²H) isotopes. The omega-3 PUFA content of emergent aquatic insects was higher than that of terrestrial insects. Emergent aquatic insects contained on average 6.6 times more eicosapentaenoic acid (EPA) and 1.2 times more α-linolenic acid (ALA) than terrestrial insects, whereas terrestrial insects contained on average 2.6 times more linoleic acid (LIN) than emergent aquatic insects. Spiders sampled directly on the lake and in upland habitats had similar EPA contents, but this EPA was derived from different diet sources, depending on the habitat. The δ¹³CEPA and δ²HEPA values of ‘lake spiders' revealed an aquatic diet pathway (i.e. EPA of aquatic origin). In contrast, EPA of spiders collected in terrestrial habitats was depleted in both ¹³C and ²H compared to any potential food sources, and their ALA isotopic values, suggesting that EPA was partly bioconverted from its dietary precursor ALA (i.e. internal pathway). The δ²H values of fatty acids clearly indicated that diet sources differed depending on the spider's habitat, which was less evident from the δ¹³C values of the fatty acids. Our data highlight that spiders can use two distinct pathways (trophic versus metabolic) to satisfy their physiological EPA demand, depending on habitat use and dietary availability.
© 2021 Nordic Society Oikos. Published by John Wiley & Sons Ltd
Subject Editor: Jerome Spitz
Editor-in-Chief: Dries Bonte
Accepted 31 August 2021
00: 1–12, 2021
doi: 10.1111/oik.08513
00 1–12
Consumers feeding at the aquatic–terrestrial ecosystem interface may obtain a mixture
of aquatic and terrestrial diet resources that vary in nutritional composition. However,
in lake riparian spiders, the relative significance of aquatic versus terrestrial diet sources
remains to be explored. We investigated the trophic transfer of lipids and polyunsatu-
rated fatty acids (PUFA) from emergent aquatic and terrestrial insects to spiders at
varying distances from the shoreline of a subalpine lake in Austria, using differences
in fatty acid profiles and compound-specific stable carbon (δ13C) and hydrogen (δ2H)
isotopes. e omega-3 PUFA content of emergent aquatic insects was higher than that
of terrestrial insects. Emergent aquatic insects contained on average 6.6 times more
eicosapentaenoic acid (EPA) and 1.2 times more α-linolenic acid (ALA) than terres-
trial insects, whereas terrestrial insects contained on average 2.6 times more linoleic
acid (LIN) than emergent aquatic insects. Spiders sampled directly on the lake and
in upland habitats had similar EPA contents, but this EPA was derived from different
diet sources, depending on the habitat. e δ13CEPA and δ2HEPA values of ‘lake spiders’
revealed an aquatic diet pathway (i.e. EPA of aquatic origin). In contrast, EPA of spi-
ders collected in terrestrial habitats was depleted in both 13C and 2H compared to any
potential food sources, and their ALA isotopic values, suggesting that EPA was partly
bioconverted from its dietary precursor ALA (i.e. internal pathway). e δ2H values of
fatty acids clearly indicated that diet sources differed depending on the spider’s habitat,
which was less evident from the δ13C values of the fatty acids. Our data highlight that
spiders can use two distinct pathways (trophic versus metabolic) to satisfy their physi-
ological EPA demand, depending on habitat use and dietary availability.
Keywords: bioconversion, carbon isotopes of fatty acids, eicosapentaenoic acid,
emerging aquatic insects, hydrogen isotopes of fatty acids, riparian consumers
Dietary availability determines metabolic conversion of long-
chain polyunsaturated fatty acids in spiders: a dual compound-
specific stable isotope approach
Margaux Mathieu-Resuge, Matthias Pilecky, Cornelia W. Twining, Dominik Martin-Creuzburg,
Tarn Preet Parmar, Simon Vitecek and Martin J. Kainz
M. Mathieu-Resuge ( (, M. Pilecky (,
S. Vitecek ( and M. J. Kainz (, WasserCluster Lunz – Biologische Station
GmbH, Lunz am See, Austria. SV also at: Univ. of Natural Resources and Life Sciences, Vienna, Inst. of Hydrobiology and Aquatic Ecosystem Management,
Vienna, Austria. MJK also at: Faculty of Medicine and Health, Danube Univ. Krems, Krems, Austria. – C. W. Twining (
8856), Max Planck Inst. of Animal Behavior, Radolfzell, Germany. – CWT, D. Martin-Creuzburg ( and T. P.
Parmar (, Limnological Inst., Univ. of Konstanz, Konstanz, Germany.
Nutrient and energy transfer across ecosystem boundaries is
a key process ensuring ecosystem functioning and food web
stability (Barnes et al. 2018). Insects that spend their juvenile
life stages in aquatic ecosystems before emerging (i.e. emergent
aquatic insects) into terrestrial ecosystems as adults are impor-
tant vectors of dietary nutrients, including omega-3 long-chain
polyunsaturated fatty acids (n-3 LC-PUFA; 20C). In aquatic
ecosystems, n-3 LC-PUFA are mostly synthesized by aquatic
primary producers and subsequently transferred across trophic
levels within the aquatic food web, while these dietary nutri-
ents are generally lacking in terrestrial food webs (Twining et al.
2016a, 2019). During their larval stages, emergent aquatic
insects acquire n-3 LC-PUFA, such as eicosapentaenoic acid
(EPA, 20:5n-3), from aquatic resources (Torres-Ruiz et al.
2007, Martin-Creuzburg et al. 2017, Scharnweber et al. 2019,
Twining et al. 2019). e n-3 LC-PUFA are considered to be
essential for animals because they are required to maintain vital
physiological processes but cannot be synthesized de novo by
many taxa (Cook and McMaster 2002, Twining et al. 2016a,
Fritz et al. 2017). However, many consumers are able to synthe-
size n-3 LC-PUFA from dietary C18 PUFA precursors, albeit
in quantities that are often insufficient to cover their needs
(Twining et al. 2018). Even when bioconversion from dietary
precursors is possible, it is likely more energetically demand-
ing than dietary acquisition of specific n-3 LC-PUFA (Parrish
2009). erefore, many consumers can benefit from obtaining
n-3 LC-PUFA through their diet to meet their physiological
demands. e differences in fatty acid profiles between aquatic
and terrestrial insects can help reveal aquatic–terrestrial linkages,
e.g. by comparing fatty acid profiles of consumers, like spiders,
with their potential prey (Fritz et al. 2017, Chari et al. 2020).
Spiders and other insectivorous riparian consumers prey
upon a wide range of insects and thus obtain a mixture of
resources differing in nutritional quality (Paetzold et al. 2005,
Fritz et al. 2017, Twining et al. 2019, Chari et al. 2020). Web-
constructing spiders are relatively stationary predators and
mostly insectivorous (Nyffeler 1999); they occur along broad
ecological gradients and can disperse by means of ballooning
(Bonte 2013). In spiders, random colonisation by ballooning
can be elicited by kin competition (Berger-Tal et al. 2016)
or poor resource availability (Mestre and Bonte 2012). is
strategy is risky and energetically costly (Bonte 2013), but
can provide access to high quality resources (e.g. emergent
aquatic insects rich in n-3 LC-PUFA). In contrast to spiders
that drift within terrestrial habitats, spiders that inciden-
tally drift to floating structures (e.g. docks, boats, emergence
traps) on aquatic habitats likely encounter food sources that
are richer in n-3 LC-PUFA, i.e. aquatic insects. e avail-
ability of n-3 LC-PUFA from aquatic resources can affect the
performance of riparian insectivores (Twining et al. 2016b,
2018, 2019), including spiders (Fritz et al. 2017). However,
how spiders process diets from aquatic or terrestrial origin has
not been studied yet.
Bulk carbon and nitrogen stable isotopes as well as fatty
acid profiles have been successfully used to reveal food sources
and the degree to which aquatic or terrestrial resources
are used by different consumers (Vander Zanden et al.
1999, Iverson et al. 2004, Perga et al. 2006, Iverson 2009,
Galloway et al. 2015), even though their application is con-
strained by the fact that several resources may share similar
stable isotope/fatty acid compositions (Cloern et al. 2002,
Guo et al. 2018, Twining et al. 2020, Ebm et al. 2021).
Compound-specific stable isotope analysis (CSIA) offers an
innovative and promising approach to these limitations. e
stable isotope values of fatty acids can indicate their dietary
or metabolic origin (Bec et al. 2011, Burian et al. 2020,
Kühmayer et al. 2020, Pilecky et al. 2021). Twining et al.
(2020) recently proposed that stable hydrogen isotopes of
fatty acids can be used at much finer-scale resolutions to
assess the origin of dietary resources and trophic interactions
than it would be possible from stable carbon isotopes, espe-
cially in systems in which the carbon stable isotope values of
potential resource fatty acids overlap (Pilecky et al. 2021).
Here, we applied these methods to determine whether spi-
ders obtain physiologically important n-3 LC-PUFA, such as
EPA, from aquatic or terrestrial sources.
Few analyses of spiders’ responses to variations in diet
composition and quality at small spatial scales have been
conducted so far, and such studies have largely focused on
river ecosystems (Paetzold et al. 2005, Chari et al. 2020,
Siebers et al. 2021). In addition, in spite of studies suggest-
ing that aquatic-derived n-3 LC-PUFA can be beneficial for
riparian spiders (Fritz et al. 2017), previous studies have not
resolved whether lake riparian spiders acquire n-3 LC-PUFA
from aquatic or terrestrial sources. In this context, the aim of
our study was to characterize the food resources and origin
of n-3 LC-PUFA in lake riparian consumers as well as their
metabolic fate. We used carbon and hydrogen stable isotopes
of fatty acids to investigate the origin of fatty acids and dem-
onstrate their potential as dietary tracers in field studies. To
understand how habitat changes affect resource use by spi-
ders, we assessed PUFA sources of (long-jawed) orb-weaving
and sheet-weaving spiders (Tetragnathidae, Araneidae and
Linyphiidae) within the riparian zone of a subalpine lake. We
collected potential insect prey emerging from the lake and
the surrounding terrestrial habitat as well as Tetragnathid,
Araneid and Linyphiid spiders. Based on total lipid content,
fatty acid and stable isotope profiles, as well as CSIA of fatty
acids, we tested the following hypotheses: 1) total lipid and
n-3 LC-PUFA contents of emerging aquatic insects are higher
than those of terrestrial insects; 2) spider fatty acid and bulk
stable isotope profiles reflect those of their insect prey; 3) the
origin of spider fatty acids differs between spiders collected
directly on the lake and those collected around the lake.
Material and methods
Study sites and sampling
Insects and spiders were collected twice a week, from mid-
June to the end of September 2019, at the subalpine Lake
Lunz, Austria and in its adjacent terrestrial habitat (47°85N,
15°05E). Six floating emergence traps were deployed on
Lake Lunz (Fig. 1A–B), to collect emerging aquatic insects.
Araneidae, Tetragnathidae and Linyphiidae spiders coloniz-
ing emergence traps (‘lake spiders’ hereafter) were collected
by hand. Nine window/malaise hybrid traps were placed
along three different terrestrial transects, each along a distance
gradient from the lake shore (1, 70 and 150 m; Fig. 1A, C).
Two transects of three window traps each were deployed in
an orchard (roughly corresponding to CLC 2.4.2 Complex
Cultivation Patterns), while the third transect was installed in a
coniferous forest (CLC 3.1.2), all along the southern margin of
Lake Lunz; again, Araneidae, Tetragnathidae and Linyphiidae
spiders colonizing window traps were collected by hand.
Traps collected a broad range of taxa that were grouped as
‘aquatic insects’ and ‘terrestrial insects’. ‘Aquatic insects’ com-
prised Chironomidae and Trichoptera (Phryganeidae and
Limnephilidae) from emergence traps on Lake Lunz, while
Cicadidae and a set of terrestrial Diptera (mostly Muscidae,
Phoridae, Psychodidae, Sciaridae and Dolichopodidae)
from window traps were considered ‘terrestrial insects’. Each
month, Araneidae, Tetragnathidae and Linyphiidae spiders
(total n = 5 per habitat) from the outside of traps were col-
lected in each habitat. Correspondingly, spiders collected on
the lake traps are considered ‘lake spiders’, while spiders col-
lected on terrestrial traps are considered ‘terrestrial spiders’.
Each emergence trap placed on Lake Lunz consisted of
four floatable tubes covering a surface area of 0.36 m2 form-
ing a pyramid-shaped construction covered with extra fine
mosquito net (mesh size ~500 μm) (Martin-Creuzburg et al.
2017, Fig. 1B). Window traps (0.36 m2 area) were cov-
ered with same nets as the lake traps. Terrestrial traps were
suspended from trees and equipped with collecting vials
(Fig. 1C).
All collected insects and spiders were transported to the
laboratory within 1 hour, frozen at 80°C, then freeze-dried
for 24 h and identified to order or family level. Both spiders
and insects were counted and put in pre-weighed tin cups,
weighed and stored at 20°C until further analyses.
Fatty acid analysis
After freeze-drying, a minimum of 2 mg of insect (n = 10 per
order and per habitat) and spider (n = 5 per habitat) samples
were homogenized and lipids were extracted according to the
method described by Guo et al. (2016). Extracted lipids were
transmethylated to obtain fatty acid methyl esters (FAME)
that were subsequently analyzed on a gas chromatograph
(Trace GC; ermo Scientific; FID 250°C, carrier gas: He:
1 ml min1, detector gases: H2: 35 ml min1, make-up gas
flow 30 ml min1, air: 350 ml min1, temperature ramp of
the oven: 140°C at 20°C min1 for 5 min, to 170°C at 4°C
min1 and to 240°C at 2°C min1 for 8 min), equipped with
a flame-ionization detector (FID, set at 250°C). FAME were
separated by a Supelco SP-2560 column (100 m, 0.25 mm
i.d., 0.2 mm film thickness), identified by comparison of their
retention times with standards (37-component FAME Mix,
Supelco 47885-U; Bacterial Acid Methyl Ester Mix, Supelco
47080-U) and quantified with reference to seven-point cali-
bration curves based on known standard dilution raw con-
centrations. All fatty acids were measured and reported as
FAME, and their contents are expressed in mass fractions (i.e.
mg FAME g dw1), and in percentages (%) of total fatty acids.
Bulk and compound-specific stable isotope analyses
Freeze-dried and homogenized samples (ca 0.3 mg) of insects
(n = 10 per order and per habitat) and spiders (n = 5 per
habitat) were put into tin capsules. eir bulk stable isotope
Figure 1. Location of the sampling habitats around Lake Lunz (A; 47°85N, 15°05E; Austria). Lake traps are represented by dark blue
squares, and terrestrial traps by red squares. Floating traps on the lake (B) and window trap on terrestrial habitat (C).
(δ13C and δ15N) values were quantified using an A flash HT
Plus CNSOH elemental analyzer interfaced with a Conflo
IV device to a continuous flow stable isotope ratio mass spec-
trometer (Delta V Advantage IRMS). Values were normalized
against reference gas injections of N2 and CO2 and standard-
ized using international standards using international stan-
dards IAEA-N-1, and IAEA-N-2 for nitrogen, and USGS24,
and IAEA-CH-7 for carbon.
Compound-specific stable isotope analyses (CSIA) were
performed to assess the isotopic composition of selected FA.
We selected insect samples (n = 5 per order, i.e. n = 10 per
habitat) collected in both habitats in August, and spiders
(n = 5 per habitat) of each habitat from June to September.
FAME were separated using a gas chromatograph linked to
the Delta V Advantage IRMS via Isolink 2 and Conflo IV.
A Split/Splitless Liner with Single Taper (4 × 6.3 × 78.5
mm, vat. no. 453A1355) was used, the injector temperature
was kept at 250°C and all samples were injected in splitless
mode. For δ13C, FAME were separated on a VF-WAXms 60
m/0.25 mm i.d./0.25 µm film thickness column (Agilent
Technologies) at a flow rate of 1.2 ml min1, followed by oxi-
dation to CO2 in a combustion reactor, filled with Ni, Pt
and Cu wires, at a temperature of 1000°C. For δ2H, FAME
were separated on a VF-WAXms 30 m/0.32 mm i.d./1 µm
film thickness column) at a flow rate of 1.0 ml min1, fol-
lowed by reduction to H2 by passing through a high thermal
conversion reactor (empty ceramic tube) kept at 1420°C.
e temperature gradient for δ13C analysis started at 80°C,
which was kept for 2 min, after which the temperature was
raised by 30°C min1 to 175°C, by 5°C min1 to 200°C and
finally by 2.4°C min1 to 250°C, which was maintained for
30 min. e temperature gradient for δ2H analysis started at
80°C, which was kept for 2 min, after which the temperature
was raised by 30°C min1 to 175°C, and then by 5°C min1
to 240°C, and held for 35 min. FAME were identified as
for GC-FID using 37-component FAME Mix. Results are
expressed in delta (δ) units with respect to international stan-
dards (Vienna Standard Mean Ocean Water for δ2H, Vienna
Pee Dee Belemnite for δ13C and atmospheric nitrogen for
δ15N), following the equation: δ2H, δ13C or δ15N = [(Rsample/
Rstandard) 1] × 103 (expressed in ‰), where R is 2H/1H,
13C/12C or 15N/14N. e 16:1n-7 and 16:1n-9, as 18:1n-7
and 18:1n-9 coelute on the column, therefore they are simul-
taneously analyzed and reported as Σ16:1 and Σ18:1.
Statistical analysis
Normality distribution and homoscedasticity of data were
analyzed using Shapiro–Wilks and Bartlett’s tests; both pre-
requisites were not met and thus data were analyzed using
non-parametric tests. First, we tested for differences in spi-
ders collected along a distance gradient (i.e. aquatic, ripar-
ian and more distant terrestrial). As there were no significant
differences in δ13C and δ2H values of fatty acids among these
three sites (due to low number of replicates in different sites),
we combined riparian and distant terrestrial spiders and com-
pared ‘lake’ versus terrestrial spiders.
To assess the difference of nutritional values of insects
depending on their habitats of origin, Kruskal–Wallis (KW)
tests followed by Conover–Iman multiple comparisons with
Bonferroni adjustment method (post hoc tests) allowed us
to compare the mean seasonal lipid content; as well as LIN,
ALA, EPA, total PUFA contents and for each other single
fatty acid found in insects of each habitat (in %, Table 1).
e diet of the spiders was characterized according to
their habitats. For that, we first compared the diets’ lipid and
PUFA contents using KW tests. To represent lipid contents,
we combined the ‘violin’ and ‘box’ plots to show the density
distribution of our data together with the median, and quar-
tiles of it, where dots represent outliers. Violin plots show
the probability density of the total lipid contents (Hintze
and Nelson 1998; Fig. 2). Particular fatty acid contents (mg
FAME g1) of interest are depicted in histograms (Fig. 3).
e KW tests also allowed to test for differences in the bulk
stable isotope values (carbon and nitrogen) between habitats
of spiders and between their potential preys, for each month
independently. Bulk stable isotope values (δ13C and δ15N)
of insects and spiders (i.e. including only Tetragnathidae) in
August were then used to assess differences in dietary sources
and consumers between both habitats (95% confident ellipses
for spiders were estimated using the stat_ellipse function of
ggplot2. Fig. 4).
Finally, the mean δ13C and δ2H values of ‘terrestrial’ and
‘aquatic’ spider fatty acids were compared using the same
method of KW tests (Supporting information), yielding dif-
ferences of PUFA origins between habitats. en, to trace the
origin of EPA in spiders, δ13C and δ2H values of EPA and
ALA were compared between spiders (i.e. consumers includ-
ing only Tetragnathidae) and insects (i.e. prey) from August
in each habitat, and represented by boxplots (fig. 5).
All statistical analyses were performed and visualized with
R ver. 3.6.1 (<>), using vegan, stat and
PMCMR, lm4 packages.
Differences in the nutritional value of emergent aquatic
versus terrestrial insects
e total lipid content of emergent aquatic insects was on
average 1.2 times higher than in terrestrial insects (KW test,
H25 = 6.00, p < 0.05, Fig. 2). e total PUFA content of
insects did not differ significantly between aquatic and ter-
restrial habitats (KW test, H25 = 3.40, p = 0.06; Fig. 3D).
On average, terrestrial insects contained 2.6 times more
LIN than emergent aquatic insects (KW test, H25 = 21.06,
p < 0.001; Fig. 3A), whereas emergent aquatic insects con-
tained 1.2 times more ALA than terrestrial insects (KW
test, H25 = 9.10, p < 0.01; Fig. 3B). In particular, the EPA
content of emergent aquatic insect was on average 6.6 times
higher than that of terrestrial insects (KW test, H25 = 63.40,
p < 0.001; Fig. 3C). However, emergent aquatic insects
had higher n-3 PUFA, but lower n-6 PUFA than terrestrial
insects, indeed the n-3/n-6 ratios of emergent aquatic insects
were on average 5.4 times higher than those of terrestrial
insects (Table 1).
Diet of spiders after dispersal in different habitats
Unlike insects, spiders collected on the lake (‘lake spiders’)
had on average 1.5 times lower lipid contents than spi-
ders from the terrestrial habitat (KW test, H25 = 10.40, p
< 0.01, Fig. 2). Total PUFA (KW test, H25 = 13.10, p <
0.001), LIN (KW test, H25 = 14.30, p < 0.001) and ALA
(KW test, H25 = 11.40, p < 0.001) contents were on aver-
age 1.7, 2.2 and 2.3 times higher in terrestrial than in ‘lake
spiders’, respectively (Fig. 3). e EPA contents of spiders
did not differ significantly between the two habitats (KW
test, H25 = 0.03, p = 0.87; Fig. 3C). e n-3/n-6 ratios of
spiders from both habitats were < 1, with ‘lake spiders’ show-
ing a significantly higher ratio (0.8) compared to terrestrial
spiders (0.5) (KW test, H25 = 7.90, p < 0.001). e EPA
content of emergent insects did not differ compared to ‘lake
spiders’ (KW test, H25 = 1.86, p = 0.17; Table 1), while
the terrestrial spiders have higher EPA contents than ter-
restrial insects (KW test, H25 = 21.86, p < 0.001; Table 1).
Terrestrial spiders accumulated higher contents of n-3 fatty
acids (included n-3 HUFA) compared to terrestrial insects
(KW test, H25 = 5.77, p < 0.05; Table 1). In comparison,
emergent insects had higher n-3 HUFA than ‘lake spiders
(KW test, H25 = 10.22, p < 0.01; Table 1).
Emergent aquatic insects from Lake Lunz were 13C depleted
compared to insects from terrestrial habitats throughout the
study period (KW tests, p < 0.05; Table 2), and compared to
spiders from both habitats (KW tests, p < 0.05). Despite the
differences between insects, bulk carbon stable isotope values
did not differ significantly between ‘lake spiders’ and spiders
collected from terrestrial traps (Table 2) with the exception of
July, when ‘lake spiders’ were more 13C depleted than riparian
spiders (KW tests, H25 = 5.80, p < 0.05; Table 2). e bulk
nitrogen stable isotope values of spiders from both habitats
were also similar over the season, as well as when comparing
insects with spiders (Table 2, Fig. 4). For example, in August,
spiders from both habitats did not differ in their bulk stable
isotope values (Fig. 4), but inter-individual variation within
‘lake spiders’ was larger compared to inter-individual varia-
tion within terrestrial spiders.
Stable carbon and hydrogen values of PUFA in
e δ13CALA values in emergent aquatic insects were lower
compared to those in terrestrial insects (Supporting infor-
mation), but the δ13CEPA values did not significantly differ
between habitats (Supporting information). e δ2HEPA val-
ues were lower in emergent aquatic insects than in terrestrial
insects (Δδ2H = 121.9‰; Supporting information). e
δ13CEPA values in ‘lake spiders’ were higher compared to those
of terrestrial spiders (Δδ13C = 3.8‰; KW test, H25 = 4.56,
p < 0.05; Supporting information), while the δ13C values of
other PUFA were not significantly different between habi-
tats (Supporting information). e δ2HALA values were higher
in ‘lake spiders’ than in terrestrial spiders (Δδ2H = 60.6‰;
Supporting information) and the δ2HEPA values not sig-
nificantly different between habitats, but ‘lake spiders’ were
isotopically slightly depleted compared to terrestrial spiders
(Supporting information).
To assess the origin of EPA in spiders, the δ13C and δ2H
values of ALA and EPA were compared between insects and
spiders in each habitat (Fig. 5). e δ13CALA values of ‘lake
spiders’ and emergent aquatic insects did not differ signifi-
cantly (KW test, H25 = 5.40, p = 0.07; Fig. 5A). e δ2HALA
values of ‘lake spiders’ were significantly higher than those of
Trichoptera and similar to those of Chironomidae (KW test,
H25 = 7.30, p < 0.05; Fig. 5B). e δ13CEPA values of ‘lake
spiders’ were similar to those of emergent Chironomidae,
and significantly higher than those of Trichoptera (KW
test, H25 = 9.12, p < 0.05; Fig. 5C), while the δ2HEPA val-
ues of ‘lake spiders’ were significantly higher than those of
Trichoptera and similar to those of Chironomidae (KW
test, H25 =5.90, p < 0.05; Fig. 5D). Terrestrial spiders had
Table 1. Total lipids (TL mean ± SE; mg g1) and fatty acids contents
(mean ± SE; mass percentage of total FA, %) of insects and spiders
from both aquatic and terrestrial habitats. Only the fatty acids
accounting for > 1% in at least one sample are shown. Different
letters indicate significant difference between insect and spider (KW
tests and Conover–Iman multiple comparisons, significant level p <
0.05), for each environment.
Lake Lunz Terrestrial
Insects Spiders Insects Spiders
ΣBFA 6.6 ± 0.6 8.1 ± 0.8 2.1 ± 0.3a4.3 ± 0.7b
14:0 2.5 ± 0.3a1.8 ± 0.4b1.9 ± 0.4a2.6 ± 0.4b
16:0 17.2 ± 0.5a13.4 ± 0.8b15.8 ± 0.7 15.1 ± 0.6
18:0 6.5 ± 0.2a12.3 ± 0.6b6.6 ± 0.6a8.1 ± 0.2b
ΣSFA 28.2 ± 0.7 30.0 ± 1.0 25.8 ± 1.1 27.2 ± 0.9
16:1n-7 9.8 ± 0.8a6.8 ± 0.8b9.3 ± 1.4 7.8 ± 0.7
18:1n-9 13.3 ± 1.1a16.2 ± 1.5b23.4 ± 1.3 24.3 ± 1.3
18:1n-7 3.8 ± 0.4a6.1 ± 0.8b0.8 ± 0.2a3.5 ± 0.6b
ΣMUFA 28.1 ± 1.1 30.0 ± 2.0 34.7 ± 1.9 36.7 ± 1.5
18:2n-6 (LIN) 10.2 ± 0.7a18.0 ± 1.1b25.8 ± 2.1 21.2 ± 1.3
18:3n-3 (ALA) 9.7 ± 1.1a3.9 ± 0.5b7.3 ± 1.5a5.1 ± 0.6b
18:4n-3 1.2 ± 0.2a0.2 ± 0.0b0.0 ± 0.0a0.2 ± 0.1b
20:4n-6 2.5 ± 0.2 3.8 ± 0.6 2.1 ± 0.4 2.3 ± 0.2
20:5n-3 (EPA) 14.9 ± 1.4 12.2 ± 1.3 2.2 ± 0.5a6.5 ± 0.8b
ΣPUFA 41.9 ± 1.2 39.6 ± 2.6 38.6 ± 1.8 35.9 ± 1.5
Σn-3 26.1 ± 1.7a17.1 ± 1.6b9.7 ± 1.4a11.9 ± 0.9b
n-3 HUFA 15.3 ± 1.4 13.0 ± 1.5 2.3 ± 0.5a6.6 ± 0.9 b
Σn-6 13.6 ± 0.8a23.0 ± 1.6b28.6 ± 1.9 24 ± 1.2
n-3/n-6 2.7a0.8b0.5 0.5
TL 182.7 ± 7.8 165.8 ± 13.1 155.49 ± 8.7a 246.6 ± 18.2b
SFA: saturated fatty acids; MUFA: monosaturated fatty acids; PUFA:
polyunsaturated fatty acids; BFA: Bacterial fatty acids (sum of 15:0,
iso15:0, anteiso15:0, iso16:0, 17:0, iso17:0 and anteiso17:0, 18:1n-
7, 18:1n-6); Σn-3: sum of n-3 fatty acids; Σn-6: sum of n-6 fatty
acids; n-3 HUFA: n-3 highly unsaturated fatty acids (sum of 20:3n-3,
20:4n-3, 20:5n-3, 22:3n-3, 22:5n-3, 22:6n-3); n-3/n-6: sum of n-3
fatty acids/sum of n-6 fatty acids.
significantly lower δ13CALA & EPA, δ2HALA & EPA values than ter-
restrial Diptera (KW tests, H25 = 7.30, 6.20 and 4.2, p <
0.05, respectively; Fig. 5).
Emerging aquatic insects were richer in n-3 LC-PUFA, espe-
cially EPA, than terrestrial insects that contained more of
the n-6 PUFA LIN, indicating the higher nutritional value
(i.e. n-3 LC-PUFA) of emergent aquatic insects. Our data
revealed that riparian spiders used dietary sources from both
ecosystems and obtained their n-3 LC-PUFA via two distinct
pathways. ‘Lake spiders’ collected from Lake Lunz acquired
their EPA directly via the consumption of emergent aquatic
insects, most likely from Chironomidae, while terrestrial spi-
ders biosynthesized their EPA from dietary precursors, i.e.
ALA and/or stearidonic acid (18:4n-3). e combined use
of stable hydrogen and carbon isotopes of fatty acids empha-
sized the significance of these two distinct pathways for spi-
ders in the acquisition of EPA, which was not possible to
reveal from fatty acid profiles or bulk stable isotopes.
PUFA of terrestrial and emergent aquatic insects as
food sources
e higher n-3 LC-PUFA contents, in particular EPA, in
emergent aquatic insects compared to terrestrial insects, are in
line with previous findings (Twining et al. 2019). ese differ-
ences in the nutritional quality of insects appear to be a result
of ecosystem-based differences in the feeding history of insect
larvae (Guo et al. 2018, Scharnweber et al. 2019). Emergent
aquatic insects likely obtain their ALA and EPA from dietary
algae rich in PUFA, especially EPA (Hixson et al. 2015,
Twining et al. 2016a). In contrast, terrestrial insects mainly
derive their LIN and ALA from the base of the terrestrial food
web (Gladyshev et al. 2013, Twining et al. 2019), which pro-
vides mainly ALA, but only traces of EPA (Gladyshev et al.
2009, Hixson et al. 2015, Taipale et al. 2015). Because n-3
LC-PUFA are important for development, somatic growth
and reproduction of animals (Twining et al. 2016b, 2019),
their higher contents in emergent aquatic insects compared
to terrestrial insects results in differences in dietary quality
supplied to riparian insectivores. erefore, emergent aquatic
insects may generally be of higher nutritional value, in terms
of n-3 PUFA content, than terrestrial insects for insectivo-
rous riparian consumers, such as spiders.
Interestingly, the high ALA contents in both emerging
aquatic and terrestrial insects indicate ALA to be a poor diet
source biomarker in this study. is fatty acid can be syn-
thetized by both aquatic and terrestrial primary producers
(Twining et al. 2016a). ALA is a physiologically important
fatty acid (e.g. as cell membrane component), and serves as
a potential precursor for n-3 LC-PUFA (Brenna et al. 2009,
Hixson et al. 2015). e capacity to convert C18 to C20
PUFA (e.g. ALA to EPA) differs among consumers (Bell
and Tocher 2009). In addition, this conversion is metaboli-
cally cost-intensive (Parrish 2009). Hence, riparian spiders
that have direct access to dietary EPA may have a metabolic
Figure 2. Total lipid contents (mg g dw1) of (A) insects and (B) spiders from aquatic and terrestrial habitat. Significant differences (KW
tests and Conover–Iman multiple comparisons) between origin of insects (i.e. aquatic versus terrestrial) are indicated with lowercase letters,
while differences between origin of spiders (i.e. aquatic versus terrestrial) are indicated with capital letters. e violin plots present the prob-
ability density distribution of the total lipids (therefore the ‘distribution’ of the data). In the boxplots, the median is represented by the thick
horizontal line; the box limits are the 25% (lower part) and the 75% (upper part) quartiles of the dataset; the vertical bars represent 1.5
times the interquartile range (IQR (i.e. the difference between the first and third quartile) above the upper quartile and below the lower
quartile; and dots represent outliers which are therefore the observations that are above q0.75 + 1.5 × IQR or below q0.25 1.5 × IQR.
advantage over spiders that need to biosynthesize EPA from
dietary ALA.
Diet of spiders after random colonization of
different habitats
Dietary sources of spiders were difficult to discern based on
bulk carbon and nitrogen stable isotope values. Bulk δ13C
and δ15N values of ‘lake spiders’ and terrestrial spiders did not
differ and thus no dietary distinction was possible between
spiders from aquatic and terrestrial habitats based on bulk
stable isotope data. e turnover of the stable isotopes in
spiders ranged between one and three weeks (Belivanov and
Hambäck 2015). erefore, it is most likely that ‘lake spi-
ders’ have recently dispersed to the aquatic environment, as
reflected by the stable isotope values closer from the terres-
trial habitat, where they have been feeding before dispersal.
In a recent study, bulk stable carbon isotope values of stream
riparian spiders did not match the ones of aquatic source,
suggesting that they did not obtain carbon from dietary
Figure 3. Contents in (A) LIN, (B) ALA, (C) EPA and (D) total PUFA of insects and spiders from aquatic and terrestrial habitats (mean ±
SE; mg FAME g1). Significant differences (KW tests and Conover–Iman multiple comparisons) between origin (i.e. aquatic versus terres-
trial) of insects are indicated with lowercase letters, while differences between origin (i.e. aquatic versus terrestrial) of spiders are indicated
with capital letters.
aquatic resources (Siebers et al. 2021). Chitin and proteins
represent up to 90% of the material found in spider cuticles
(Sewell 1955, Nentwig 2012), therefore their nitrogen iso-
topic composition should reflect the one of the habitat in
which they were hunting before moulting. However, these
limitations can be overcome by combining elemental analysis
(e.g. bulk δ13C) with molecular (e.g. fatty acids) biomarkers
(Perga et al. 2006, Jardine et al. 2015), and/or compound-
specific stable isotope analyses to trace dietary sources and
specific compounds within food chains (Kohl et al. 2015,
Taipale et al. 2015, Twining et al. 2020).
Spiders collected on the lake traps had lower total lipid,
PUFA, LIN and ALA contents than terrestrial spiders, but the
equal EPA contents suggest preferential retention of this fatty
acid. e particular retention of EPA (%) and n-3 HUFA by
terrestrial spiders compared to terrestrial insects highlights
the importance of long-chain PUFA in riparian consumers.
However, the PUFA composition of terrestrial spiders was
characterized by higher C18 PUFA contents, i.e. LIN and
ALA, compared to ‘lake spiders’. ese differences in PUFA
contents suggest that terrestrial spiders are part of a distinct
terrestrial food chain rich in LIN and ALA, but poor in EPA
(Budge and Parrish 1998, Twining et al. 2016a). e lower
lipid content observed in ‘lake spiders’ may have resulted
from a lower food availability on the floating lake traps, and/
or before their dispersal. In contrast, terrestrial spiders may
have been able to maintain higher lipid contents because of
a higher prey density in the riparian zone and the absence of
dispersal costs. It is also possible that ‘lake spiders’ may have
used C18 PUFA preferentially for gaining energy, but retained
EPA in their cell membranes, suggesting a hierarchy in the use
of biochemical components when facing changing resources.
Moreover, the high EPA content in emergent aquatic insects
supports the hypothesis that spiders that drifted to aquatic
habitats via ballooning likely have higher dietary access to this
n-3 LC-PUFA. Our results corroborate that EPA is an excel-
lent biomarker of aquatic-derived subsidies in riparian con-
sumers (Chari et al. 2020), and highlight the importance of
n-3 LC-PUFA for both riparian spiders . In particular, these
data reveal two different trophic trajectories through which
spiders, or riparian insectivores in general, obtain PUFA.
Origin of PUFA in spiders
e CSIA data suggest that spiders acquired their EPA from dif-
ferent sources. ‘Lake spiders’ likely obtained their EPA directly
Table 2. Bulk carbon and nitrogen isotope composition (mean ± SD; δ13C and δ15N, ‰) of insects and spiders from aquatic (lake) and ter-
restrial habitats. Different letters indicate significant differences (KW tests and Conover–Iman multiple comparisons) between habitats;
Δδ13C = difference in δ13C values (‰) between aquatic and terrestrial habitats.
Insects Spiders
δ13C (‰) Δδ13C
δ15N (‰) δ13C (‰) Δδ13C
δ15N (‰)
Lake Lunz Terrestrial Lake Lunz Terrestrial Lake Lunz Terrestrial Lake Lunz Terrestrial
June 34.2 ± 1.6b27.7 ± 2.0a6.5 3.0 ± 1.4 2.4 ± 2.3 28.7 ± 1.9 28.5 ± 1.3 0.2 2.0 ± 1.4 1.7 ± 0.8
July 31.0 ± 3.1b27.9 ± 1.6a3.1 1.8 ± 1.8 2.0 ± 2.5 29.3 ± 1.1b27.3 ± 0.7a2.0 2.4 ± 1.0 1.5 ± 1.6
August 30.5 ± 1.8b26.8 ± 1.9a3.7 1.3 ± 2.2 1.8 ± 3.0 27.9 ± 2 29.0 ± 0.8 1.1 1.7 ± 1.6 1.3 ± 1.4
September 30.2 ± 2.0b27.5 ± 2.2a2.7 0.6 ± 3.6b2.1 ± 2.2a27.7 ± 1.4 28.5 ± 0.6 0.8 1.4 ± 2.5 3.2 ± 0.9
Figure 4. Bulk stable isotopes carbon and nitrogen ratios (δ13C and δ15N, ‰) of insects and spiders from aquatic and terrestrial habitats in
August. e 95% normal ellipse from aquatic spiders is represented in blue, and from terrestrial spiders in dark green.
from their aquatic diet, while terrestrial spiders likely obtained
EPA from both diet and bioconversion (i.e. they converted
dietary precursors to EPA intrinsically). Based on δ13CEPA val-
ues, ‘lake spiders’ obtained their EPA from Chironomidae,
one of the most abundant emergent aquatic insect taxa found
in lakes (Armitage 1995, Martin-Creuzburg et al. 2017,
Selene et al. 2020, Mathieu-Resuge et al. 2021b), mak-
ing them a likely food source for ‘lake spiders’. Because the
δ13CEPA values of emergent aquatic insects overlapped with
those from terrestrial insects, it was not possible to trace the
origin of spider EPA using δ13CEPA values alone. However, the
δ2HEPA values of ‘lake spiders’ were close to those of emer-
gent aquatic insects, which were clearly distinct from those of
terrestrial insects (Δδ2H = 121.9‰). us, the δ2HEPA values
provided isotopic evidence for the aquatic origin of EPA in
‘lake spiders’, highlighting the potential of using dual com-
pound-specific stable isotope approaches in tracing the origin
of fatty acids in natural ecosystems.
Spiders from terrestrial habitats appear more likely to have
obtained their EPA partially from diet sources and partially
Figure 5. Stable carbon and hydrogen isotope values (δ13C and δ2H, ‰) of the n-3 polyunsaturated fatty acids ALA (A and B, respectively)
and EPA (C and D, respectively) of insects and spiders collected from aquatic (Lake Lunz) and terrestrial habitats in August. e letters
indicate significant differences (KW tests and Conover–Iman multiple comparisons) between stable isotope values of n-3 PUFA between
taxa from aquatic and terrestrial habitats.
through bioconversion from dietary ALA. As it was the case
for ‘lake spiders’, δ13CEPA & ALA values of aquatic and terres-
trial insects overlapped, precluding us from drawing strong
inferences about fatty acid origin for terrestrial habitat spi-
ders based upon δ13CEPA & ALA. Terrestrial spiders had δ2HEPA
& ALA values between aquatic and terrestrial insects, suggesting
that EPA can come from both aquatic (i.e. direct assimila-
tion and accumulation) and terrestrial (i.e. bioconversion
from precursors) sources. However, in contrast to the δ2HEPA
values of ‘lake spiders’, the δ2HEPA values of terrestrial spi-
ders were substantially higher than those of emergent aquatic
insects (Δδ2H = 79‰), making it highly unlikely that they
only obtained their EPA directly from feeding on emergent
aquatic insects. As terrestrial insects contained very little
EPA, compared to terrestrial spiders and because the δ2HEPA
values of terrestrial spiders were lower than those of terres-
trial insects (Δδ2H = 43‰) it is also unlikely that spiders
obtained their EPA directly through diet from terrestrial
insects. Instead, terrestrial spiders had isotopically lighter val-
ues of δ13CEPA & ALA and δ2HE PA & ALA compared to terrestrial
insects, pointing towards innate PUFA conversion, as sug-
gested for other taxa (Bec et al. 2011, Burian et al. 2020,
Twining et al. 2020). Moreover, the δ2HE PA values were
lower than the δ2HALAδ2H = 13‰), supporting the idea
of internal bioconversion from ALA to EPA. Indeed ALA
(%) was high in insects from both environments, but was
also specifically accumulated by terrestrial spiders. Because
isotopically lighter ALA should be faster converted to EPA
(Bec et al. 2011, Twining et al. 2020), the remaining pool
of ALA remains isotopically more enriched compare to the
EPA bioconverted. Lower dietary access to emerging aquatic
resources may select for increased bioconversion in spider
populations in terrestrial habitats, or increased bioconversion
may be the result of plasticity in response to the relative avail-
ability of EPA and its precursors. Future laboratory feeding
studies will be necessary to distinguish between these pos-
sibilities, by characterising fatty acid fractionations depend-
ing on dietary and/or metabolic acquisition. erefore, these
results imply that terrestrial spiders do not necessarily rely on
dietary EPA from aquatic sources, but given that bioconver-
sion is costly this suggests that riparian spiders preferentially
use dietary EPA whenever it is available.
Compound-specific stable isotope analysis on fatty acids
is a powerful tool to better understand trophic transfer of
dietary energy in and across ecosystems (Pilecky et al. 2021).
e simultaneous use of stable hydrogen and carbon isotopes
of fatty acids allows to better discriminate the origin of fatty
acids. Even if both hydrogen and carbon stable isotopes allow
discrimination among food sources at the compound-specific
level (Burian et al. 2020, Twining et al. 2020), the isotopic
discrimination between source and consumer is greater for
hydrogen than carbon stable isotopes (Pilecky et al. 2021). In
our study, the distinction between terrestrial and ‘lake spiders’
and their food sources were much more evident for δ2HEPA
δ2H = 42.9‰ and 121.9‰, respectively) than for δ13CEPA
values (Δδ13C = 3.80‰ and 2.50‰, respectively). However,
this study lacks information on how H stable isotopes are
processed during lipid metabolism (i.e. elongation, desatu-
ration and retro-conversion) and, more precisely, what are
the specific values of hydrogen isotopic fractionations during
incorporation and internal biosynthesis of fatty acids. One
way to solve these uncertainties and fully characterise the
fractionation processes is by conducting laboratory feeding
experiments (e.g. knowing the food source fatty acids’ stable
isotope composition). Such controlled experiments can effec-
tively characterise precisely the changes in isotopic fraction-
ations of fatty acids between dietary and metabolic pathways.
Our study therefore illustrates the added value that can be
gained from this new method in trophic ecology, allowing
us to explore how the fatty acid requirements of consumers
change with habitat and how bioconversion is triggered by
dietary availability.
is field study underlines the importance of aquatic
resources in terrestrial consumer diets (e.g. n-3 PUFA) and
thus the interconnectivity between aquatic and terrestrial
habitats. Our results highlight the intrinsic hierarchy of bio-
chemical compounds in spiders, with a preferential retention
of EPA, compared to LIN and ALA. e dual use of stable
hydrogen and carbon isotopes of fatty acids is a novel and
promising approach for trophic ecology to characterize vari-
ous diet sources and pathways to consumers. Results of this
field study suggest that spiders acquire EPA from both ter-
restrial and aquatic resources using two pathways: 1) direct
dietary acquisition (trophic pathway), and/or, 2) bioconver-
sion (intrinsic pathway). Specifically, spiders collected from
Lake Lunz acquired their EPA directly from emergent aquatic
insects – most likely from Chironomidae – while terrestrial
spiders seemed to have obtained their EPA partially from
diet sources (i.e. from aquatic insects) and partially through
bioconversion from dietary precursors (e.g. ALA and/or stea-
ridonic acid). us, spiders appear to be capable of biosynthe-
sizing their n-3 LC-PUFA from dietary precursors depending
on dietary PUFA availability. Finally, the combined use of
stable hydrogen and carbon isotopes of fatty acids provides
better understanding of dietary energy transfer in aquatic and
terrestrial food webs than it would be possible from using
only fatty acid profiles or bulk stable isotope values, but it
will be necessary to better calibrate these methods with labo-
ratory feeding experiments to better understand fractionation
along dietary and metabolic pathways.
Acknowledgements – We thank H. H. Hager, K. Winter, S.-K.
Kämmer, E. Wassenaar, L. Perez and S. Damodaran for their field
assistance, and lipid analyses.
Funding – is study was funded by the Austrian Science Fund
(FWF; I 3855-B25) and the German Research Foundation (DFG;
MA 5005/8-1) within the framework of the DACH collaboration
(project ‘AquaTerr’).
Conflict of interest – e authors declare no conflict of interest.
Author contributions
Margaux Mathieu-Resuge: Conceptualization (equal);
Formal analysis (equal); Investigation (lead); Methodology
(equal); Validation (equal); Visualization (lead); Writing
– original draft (lead). Matthias Pilecky: Formal analy-
sis (equal); Methodology (equal); Validation (equal);
Visualization (equal); Writing – review and editing (equal).
Cornelia W. Twining: Investigation (equal); Methodology
(equal); Validation (equal); Visualization (equal); Writing
review and editing (equal). Dominik Martin-Creuzburg:
Conceptualization (equal); Funding acquisition (lead);
Investigation (equal); Methodology (equal); Project admin-
istration (lead); Supervision (equal); Validation (equal);
Visualization (equal); Writing – review and editing (equal).
Tarn-Preet Parmar: Methodology (equal); Validation (equal);
Visualization (equal); Writing – review and editing (equal).
Simon Vitecek: Methodology (equal); Validation (equal);
Visualization (equal); Writing – review and editing (equal).
Martin J. Kainz: Conceptualization (equal); Funding acqui-
sition (lead); Investigation (lead); Methodology (equal);
Project administration (lead); Resources (lead); Supervision
(lead); Validation (equal); Visualization (equal); Writing
original draft (equal).
Data availability statement
Data are available from the Dryad Digital Repository:
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... Because all of the contributions considered at least one explicit variable to quantify the nutritional quality of the resource, none of them falls within the cluster of 'undefined food quality' (in green). Five of the articles of the Special issue (Chouvelon et al. 2022, Lowman et al. 2022, Mathieu-Resuge et al. 2022, Sentis et al. 2022, van Deurs et al. 2022) fall within the cluster aquatic ecosystem/ stoichiometry-lipids/trophodynamics-fitness, a proportion that mirrors well the actual predominance of those themes within the overall research (Fig. 2c). Two more articles of this Special issue (Le Gall et al. 2022, Zaguri et al. 2022 belong to the cluster linking the nutritional geometry to physiological aspects, which is current ranked second in term of contribution to the overall research. ...
... The articles published in this special issue are well representative of these trends since one third of the published articles question resources quality through the consideration of resources elemental content (Chouvelon et al. 2022, Lowman et al. 2022, Sentis et al. 2022, while a second third focuses on resources macro-nutrient contents, alone (Le Gall et al. 2022, Zaguri et al. 2022 or in association with fatty acids (van Deurs et al. 2022). Finally, three articles specifically focus on fatty acids/lipids (Hudson et al. 2022, Leal et al. 2022, Mathieu-Resuge et al. 2022. ...
... Terrestrial and aquatic ecosystems are almost equally represented in our analysis of existing scientific literature, which contrasts with the articles published in the special issue, largely dominated by aquatic studies (only two articles 6 considering terrestrial consumers: Le Gall et al. 2022, Mathieu-Resuge et al. 2022. Interestingly, parameters used for describing resources quality significantly differ between ecosystems. ...
Our understanding of ecosystem functioning is strongly linked to the study of predator–prey relationships and food web structures. However, trophic ecology has often focused on identifying taxonomic relationships and quantifying the biomass or energy ingested by consumers, but has often failed to integrate the importance of the nutritional quality of resources in ecological dynamics. Underlying this gap is the multi‐dimensional nature of resource quality which has hampered any consensus on the definition of resource nutritional quality. In this special issue, we aimed at gathering a subset of articles exemplifying the diversity of variables by which resources quality is quantified, the diversity of research topics that can be tackled in ecology – from physiological or evolutionary aspects to ecosystem processes – and propose some perspectives on the integration of nutritional quality within broader ecological concepts. Using a semi‐automated literature analysis, we map the current landscape of the ‘resources nutritional quality' research of the last 30 years. We depict how it has been quantified through physical, biological or chemical indicators, the use of these parameters being largely dependent on the type of ecosystem studied and on the investigated ecological process. We then position the articles published in this special issue of Oikos within this landscape, showing they cover a small but relatively well representative subset of the domains of resources quality‐related issues. Articles in this special issue browse a range of individual and population‐level approaches (embracing evolutionary questions) to community related questions, include methodological issues and ecosystem‐wide approaches using trophic quality indicators as tracers of resources origin. Based on these studies and on the literature review, we identify a non‐exhaustive list of challenges and perspectives of research that we consider of highest priority in the large topic of trophic ecology.
... In contrast, vascular terrestrial plants typically contain only C18-PUFA, such as linoleic acid (LIN; 18:2n-6) and α-linolenic acid (ALA; 18:3n-3), and lack the ability to synthesize LC-PUFA (Uttaro, 2006;Hixson et al., 2015). This ecosystem-based (i.e., aquatic compared to terrestrial) difference in primary producers' FA profiles is transferred to higher trophic levels and is also observed between the FA profiles of aquatic and terrestrial insects (Guo et al., 2018;Kowarik et al., 2021;Mathieu-Resuge et al., 2021b;Twining et al., 2021b). ...
... This significant LIN comparison may be a product of the additional sampling from June to October, including Orthoptera as well as a higher proportion of Dermaptera and Neuroptera to the analysis, all of which having significantly higher LIN content. Although Mathieu-Resuge et al. (2021b) found no differences in the total PUFA content between aquatic and terrestrial insects around Lake Lunz (Austria), similar to the current study, ALA and EPA were higher in aquatic insects while LIN was higher in terrestrial insects. ...
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Emergent insects represent a key vector through which aquatic nutrients are transferred to adjacent terrestrial food webs. Aquatic fluxes of polyunsaturated fatty acids (PUFA) from emergent insects are particularly important subsidies for terrestrial ecosystems due to high PUFA contents in several aquatic insect taxa and their physiological importance for riparian predators. While recent meta-analyses have shown the general dichotomy in fatty acid profiles between aquatic and terrestrial ecosystems, differences in fatty acid profiles between aquatic and terrestrial insects have been insufficiently explored. We examined the differences in fatty acid profiles between aquatic and terrestrial insects at a single aquatic-terrestrial interface over an entire growing season to assess the strength and temporal consistency of the dichotomy in fatty acid profiles. Non-metric multidimensional scaling clearly separated aquatic and terrestrial insects based on their fatty acid profiles regardless of season. Aquatic insects were characterized by high proportions of long-chain PUFA, such as eicosapentaenoic acid (20:5n-3), arachidonic acid (20:4n-6), and α-linolenic acid (18:3n-3); whereas terrestrial insects were characterized by high proportions of linoleic acid (18:2n-6). Our results provide detailed information on fatty acid profiles of a diversity of aquatic and terrestrial insect taxa and demonstrate that the fundamental differences in fatty acid content between aquatic and terrestrial insects persist throughout the growing season. However, the higher fatty acid dissimilarity between aquatic and terrestrial insects in spring and early summer emphasizes the importance of aquatic emergence as essential subsidies for riparian predators especially during the breading season.
... [30][31][32] As a result, terrestrial insects have n-3 LCPUFA contents only 3%-25% those of aquatic insects of the same mass. 33 Thus, the availability of desirable fatty acids for insectivores is likely proportional to the relative availability of aquatic versus terrestrial insects ( Figure 1). ...
... The phenology of aquatic and terrestrial insect communities drives the seasonal availability of n-3 LCPUFA, which are important nutrients for a diversity of consumers reliant on insects, 33,54,55 including avian aerial insectivores. 11,14,56 We find that the peak biomass of aquatic insect emergence tends to occur earlier in the season and then declines in total output, whereas terrestrial insect biomass progressively increases throughout our observed season. ...
Climate change can decouple resource supply from consumer demand, with the potential to create phenological mismatches driving negative consequences on fitness. However, the underlying ecological mechanisms of phenological mismatches between consumers and their resources have not been fully explored. Here, we use long-term records of aquatic and terrestrial insect biomass and egg-hatching times of several co-occurring insectivorous species to investigate temporal mismatches between the availability of and demand for nutrients that are essential for offspring development. We found that insects with aquatic larvae reach peak biomass earlier in the season than those with terrestrial larvae and that the relative availability of omega-3 long-chain polyunsaturated fatty acids (n-3 LCPUFAs) to consumers is almost entirely dependent on the phenology of aquatic insect emergence. This is due to the 4- to 34-fold greater n-3 LCPUFA concentration difference in insects emerging from aquatic as opposed to terrestrial habitats. From a long-sampled site (25 years) undergoing minimal land use conversion, we found that both aquatic and terrestrial insect phenologies have advanced substantially faster than those of insectivorous birds, shifting the timing of peak availability of n-3 LCPUFAs for birds during reproduction. For species that require n-3 LCPUFAs directly from diet, highly nutritious aquatic insects cannot simply be replaced by terrestrial insects, creating nutritional phenological mismatches. Our research findings reveal and highlight the increasing necessity of specifically investigating how nutritional phenology, rather than only overall resource availability, is changing for consumers in response to climate change.
... Although they also had access to freshwater lakes and a terrestrial environment comprising mainly boreal forest, the composition of their red blood cells nevertheless indicates that they foraged primarily in the marine environment. In contrast, urban gulls nesting at Long Pond and Spaniard's Bay relied more heavily on terrestrial and anthropogenic food sources, as evidenced by their overall low levels of EPA and DHA and high levels of ARA and LA in their red blood cells (Gladyshev and Sushchik, 2019;Mathieu-Resuge et al., 2021). Their isotopic signatures were also similar to those of terrestrial and anthropogenic food sources and to those of consumers of such foods, which further suggests a primarily terrestrial and anthropogenic diet (Davis et al., 2017;de Faria et al., 2021;Garthe et al., 2016). ...
Species and populations with greater cognitive performance are more successful at adapting to changing habitats. Accordingly, urban species and populations often outperform their rural counterparts on problem-solving tests. Paradoxically, urban foraging also might be detrimental to the development and integrity of animals' brains because anthropogenic foods often lack essential nutrients such as the long-chain omega-3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), which are important for cognitive performance in mammals and possibly birds. We tested whether urbanization or consumption of EPA and DHA are associated with problem-solving abilities in ring-billed gulls, a seabird that historically exploited marine environments rich in omega-3 fatty acids but now also thrives in urban centres. Using incubating adults nesting across a range of rural to urban colonies with equal access to the ocean, we tested whether urban gulls preferentially consumed anthropogenic food while rural nesters relied on marine organisms. As we expected individual variation in foraging habits within nesting location, we characterized each captured gulls' diet using stable isotope and fatty acid analyses of their red blood cells. To test their problem-solving abilities, we presented the sampled birds with a horizontal rendition of the string-pull test, a foraging puzzle often used in animal cognitive studies. The isotopic and fatty acid profiles of urban nesters indicated a diet comprising primarily anthropogenic food, whereas the profiles of rural nesters indicated a high reliance on marine organisms. Despite the gulls' degree of access to urban foraging habitat not predicting solving success, birds with biochemical profiles reflecting anthropogenic food (less DHA and a higher carbon-13 ratio in their red blood cells) had a greater probability of solving the string-pull test. These results suggest that experience foraging on anthropogenic food is the main explanatory factor leading to successful problem-solving, while regular consumption of omega-3s during incubation appears inconsequential.
... A recent study suggested that in lake ecosystems where phytoplankton produce less long-chain PUFAs, the biosynthesis of DHA from ALA in perch is intensified (Scharnweber et al., 2021). Terrestrial sources tend to be depleted especially with long-chain omega-3 (n3) FAs and enriched with omega-6 (n6) FAs, especially LIN (Hixson et al., 2015;Taipale et al., 2015;Mathieu-Resuge et al., 2021). Growth experiments have shown that invertebrates fed with allochthonous matter containing LIN will result high ARA content in their tissues (Goedkoop et al., 2007;Taipale et al., 2015). ...
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Environmental change, including joint effects of increasing dissolved organic carbon (DOC) and total phosphorus (TP) in boreal northern lakes could potentially affects food web energy sources and the biochemical composition of organisms. These environmental stressors are enhanced by anthropogenic land-use and can decrease the quality of polyunsaturated fatty acids (PUFAs) in seston and zooplankton, and therefore, possibly cascading up to fish. In contrast, the content of mercury in fish increases with lake browning potentially amplified by intensive forestry practises. However, there is little evidence on how these environmental stressors simultaneously impact beneficial omega-3 fatty acid (n3-FA) and total mercury (THg) content of fish muscle for human consumption. A space-for-time substitution study was conducted to assess whether environmental stressors affect Eurasian perch (Perca fluviatilis) allochthony and muscle nutritional quality [PUFA, THg, and their derivative, the hazard quotient (HQ)]. Perch samples were collected from 31 Finnish lakes along pronounced lake size (0.03–107.5 km²), DOC (5.0–24.3 mg L⁻¹), TP (5–118 μg/ L) and land-use gradients (forest: 50.7–96.4%, agriculture: 0–32.6%). These environmental gradients were combined using principal component analysis (PCA). Allochthony for individual perch was modelled using source and consumer δ²H values. Perch allochthony increased with decreasing lake pH and increasing forest coverage (PC1), but no correlation between lake DOC and perch allochthony was found. Perch muscle THg and omega-6 fatty acid (n6-FA) content increased with PC1 parallel with allochthony. Perch muscle DHA (22:6n3) content decreased, and ALA (18:3n3) increased towards shallower murkier lakes (PC2). Perch allochthony was positively correlated with muscle THg and n6-FA content, but did not correlate with n3-FA content. Hence, the quality of perch muscle for human consumption decreases (increase in HQ) with increasing forest coverage and decreasing pH, potentially mediated by increasing fish allochthony.
Omega-3 long-chain polyunsaturated fatty acids (n-3 LC-PUFA) are essential micronutrients for aquatic consumers. Synthesized by aquatic primary producers, n-3 LC-PUFA are transferred across trophic levels and may eventually end up accumulating in fish. However, if short in dietary supply, fish may also biosynthesize n-3 LC-PUFA from dietary precursors (i.e., n-3 C18-PUFA). We applied compound-specific hydrogen stable isotope analysis (CSIA) of fatty acids to investigate sources and metabolic processes of n-3 LC-PUFA, and in particular of docosahexaenoic acid (22:6n-3, DHA), in Common Carp (Cyprinus carpio) raised in semi-intensive aquaculture ponds. Carp were feeding on natural pond zooplankton and benthic macroinvertebrates rich in n-3 LC-PUFA and cereal-based pellet feeds rich in C18-PUFA. Results provide isotopic evidence that carp obtained a significant amount of dietary lipids and nitrogen from added cereal-based feeds, while n-3 LC-PUFA were generally acquired by feeding on benthic macroinvertebrates and zooplankton. However, DHA retained in carp was also generated endogenously via bioconversion from dietary PUFA precursors, such as EPA. DHA was isotopically lighter than EPA and likely not supplied in sufficient quantities to meet the physiological requirements for DHA in carp. Our data show that depending on the natural abundance of dietary DHA in these eutrophic ponds, farmed carp can obtain DHA by two different pathways; i.e., directly via dietary uptake and indirectly via bioconversion. This field study highlights the importance of dietary LC-PUFA supply in eutrophic aquatic ecosystems and the ability of carp to biosynthesize highly valuable LC-PUFA, eventually also benefiting human health.
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Omega-3 long-chain polyunsaturated fatty acids (n-3 LC-PUFA) are key structural lipids and their dietary intake is essential for brain development of virtually all vertebrates. The importance of n-3 LC-PUFA has been demonstrated in clinical and laboratory studies, but little is known about how differences in the availability of n-3 LC-PUFA in natural prey influence brain development of wild consumers. Consumers foraging at the interface of aquatic and terrestrial food webs can differ substantially in their intake of n-3 LC-PUFA, which may lead to differences in brain development, yet this hypothesis remains to be tested. Here we use the previously demonstrated shift towards higher reliance on n-3 LC-PUFA deprived terrestrial prey of native brown trout Salmo trutta living in sympatry with invasive brook trout Salvelinus fontinalis to explore this hypothesis. We found that the content of n-3 LC-PUFA in muscle tissues of brown trout decreased with increasing consumption of n-3 LC-PUFA deprived terrestrial prey. Brain volume was positively related to the content of the n-3 LC-PUFA, docosahexae-noic acid, in muscle tissues of brown trout. Our study thus suggests that increased reliance on diets low in n-3 LC-PUFA, such as terrestrial subsidies, can have a significant negative impact on brain development of wild trout. Our findings provide the first evidence of how brains of wild vertebrate consumers response to scarcity of n-3 LC-PUFA content in natural prey.
Not all insects are created equal and those emerging from wetlands are nutritionally superior to those from uplands. Insectivorous birds have timed reproduction to coincide with insect pulses, but new work shows how climate change has disconnected this synchrony, creating reductions in insect quality with profound implications for conservation.
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• The ecological role of emergent aquatic insects from lakes in exporting dietary polyunsaturated fatty acids (PUFA) across the freshwater-land interface is still poorly understood. • In this field study, we explored the seasonal biomass export of emergent insects from three subalpine lakes and investigated how lipids of emergent insects were related to lake bathymetry, lipids of organic matter in lake sediments (i.e., basal resources), and the taxonomic composition of insects. • The total lipid and PUFA fluxes of emergent insects were strongly related to taxonomy and lake bathymetry, but weakly associated with the PUFA content of the uppermost lake sediment layers. PUFA flux estimates of the dominant taxon, Chironomidae, from the shallowest lake (3 m depth; 125 g PUFA m⁻² season⁻¹) were considerably higher than those from the deepest lake (33 m depth; 56 g PUFA m⁻² season⁻¹), due to the higher per area biomass of emergent insects from this shallow lake. Insect taxonomy also affected the composition of PUFA transfer to land: Chironomidae were richer in ω-6 PUFA, such as linoleic acid (18:2n-6) and arachidonic acid (20:4n-6), whereas Ephemeroptera and Trichoptera contained more ω-3 PUFA, especially α-linolenic acid (18:3n-3) and eicosapentaenoic acid (20:5n-3). • Our findings suggest that taxon-specific differences in PUFA content and lake bathymetry jointly shape PUFA fluxes and thus the provisioning of emergent insects as dietary sources of physiologically important PUFA for riparian consumers.
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Rationale Compound-specific stable isotope analysis (CSIA) is a powerful tool to better understand trophic transfer of dietary molecules in and across ecosystems. Hydrogen isotope values (δ2H) in consumer tissues have potential to more clearly distinguish dietary sources than 13C or 15N within and among habitats but have not been used at the fatty acids level for ecological purposes. Methods Here we demonstrate a new online high-capacity gas chromatography-isotope ratio-mass spectrometry technique (2H-CSIA) that offers accurate and reproducible determinations of δ2H values on a range of fatty acids from organisms of aquatic food webs. Results We show that δ2H of lipid extracts obtained from aquatic organisms, such as biofilms, leaves, invertebrates, or fish muscle tissue have distinctive δ2H that can be used to assess sources and trophic interactions, as well as dietary allocation and origin of fatty acids within consumer tissue. Conclusions The new 2H-CSIA method can be applied to evaluate sources and trophic dynamics of fatty acids in organisms ranging from food web ecology, migratory connectivity.
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The River Continuum Concept implies that consumers in headwater streams have greater dietary access to terrestrial basal resources, but recent studies have highlighted the dietary importance of high-quality algae. Algae provide consumers with physiologically important omega-3 (n-3) polyunsaturated fatty acids (PUFA), particularly eicosapen-taenoic acid (EPA). However, terrestrial plants and most benthic stream algae lack the long-chain (LC) n-3 PUFA docosahexaenoic acid (DHA, 22:6n-3), which is essential for neural development in fish and other vertebrates. We sampled subalpine streams to investigate how the PUFA composition of neural (brain and eyes), muscle, and liver tissues of freshwater fish is related to their potential diets (macroin-vertebrates, epilithon, fresh and conditioned terrestrial leaves). The PUFA composition of consumers was more similar to epilithon than to terrestrial leaves. Storage lipids of eyes most closely resembled dietary PUFA (aquatic invertebrates and algae). However, DHA and arachidonic acid (ARA, 20:4n-6) were not directly available in the diet but abundant in organs. This implies that algal PUFA were selectively retained or were produced internally via enzymatic PUFA conversion by aquatic consumers. This field study demonstrates the nutritional importance of algal PUFA for neural organs in aquatic consumers of headwater regions.
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Introduction of exotic species is one of the major human impacts for decline in autochthonous biodiversity. In freshwater lentic environments, alien fish introduction heavily shapes macrobenthic invertebrate structure, with special regard for Chironomidae (Diptera) community. These effects could be more evident in alpine lakes whose environments are considered as remote and undisturbed, but extremely susceptible and vulnerable to both natural and anthropogenic impacts. In this context, modern and subfossil chironomid assemblages were studied in a high-altitude lake (Balma Lake, Piedmont, Italy) which was not previously investigated and subject to brook trout introduction for recreational fishing at the end of the twentieth century (after 1970). Seasonal samplings (summer, autumn) were performed in five littoral and three deep sites, while a core sample was extracted from the deepest point of the lake. The analysis highlighted significant differences in subfossil chironomid communities before and after fish introduction and between subfossil and modern communities, with notable decrease in recent diversities. Dissimilarities were mainly related to Corynocera oliveri, Zavrelimyia, Micropsectra, Metriocnemus, and Heterotrissocladius marcidus type. Therefore, this study highlights the importance and effects of anthropic environmental events, such as fish introduction, in climate reconstructions and their interpretation, especially regarding the last 100/200-year period, when human environmental issues have become more significant.
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According to the River Continuum Concept, headwater streams are richer in allochthonous (e.g. terrestrial leaves) than autochthonous (e.g. algae) sources of organic matter for consumers. However, compared to algae, leaf litter is of lower food quality, particularly ω‐3 polyunsaturated fatty acids (n‐3 PUFA), and would constrain the somatic growth, maintenance, and reproduction of stream invertebrates. It may be thus assumed that shredders, such as Gammarus, receive lower quality diets than grazers, e.g. Ecdyonurus, that typically feed on algae. The objective of this study was to assess the provision of dietary PUFA from leaf litter and algae to the shredder Gammarus and the grazer Ecdyonurus. Three different diets (algae, terrestrial leaves, and an algae–leaf litter mix) were supplied to these macroinvertebrates in a flume experiment for 2 weeks. To differentiate how diet sources were retained in these consumers, algae were isotopically labelled with 13C. Both consumers became enriched with 13C in all treatments, demonstrating that both assimilated algae. For Gammarus, n‐3 PUFA increased, whereas n‐6 PUFA stayed constant. By contrast, the n‐3 PUFA content of Ecdyonurus decreased as a consequence of declining algal supply. Results from compound‐specific stable isotope analysis provided evidence that the long‐chain n‐3 PUFA eicosapentaenoic acid (EPA) in both consumers was more enriched in 13C than the short‐chain n‐3 PUFA α‐linolenic acid, suggesting that EPA was taken up directly from algae and not from heterotrophic biofilms on leaf litter. Both consumers depended on algae as their carbon and EPA source and retained their EPA from high‐quality algae.
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• Aquatic and terrestrial ecosystems differ fundamentally in the abundance of long‐chained polyunsaturated fatty acids (PUFAs), such as eicosapentaenoic acid (EPA; 20:5n‐3), which are produced by aquatic algae, but only in low quantities by terrestrial plants. Aquatic insects, such as Chironomidae (non‐biting midges) feed on algae during their larval stage, making them rich in EPA and therefore high‐quality prey for insectivores after emergence. However, the magnitude of EPA subsidies from aquatic insects may be different among water bodies in response to abiotic (e.g. nutrient load) as well as biotic factors (e.g. food web structure). • To test the predation effects of crucian carp (Carassius carassius), nutrient concentrations, and Chironomidae community composition on the fatty acid export from aquatic ecosystems, we conducted a 25‐day experiment across 20 1,500‐L mesocosms covering a total phosphorus (TP) gradient of 20–1,000 µg/L. • Twice a week, we collected adult emerging Chironomidae and found differences in fatty acid composition in the two most abundant chironomid species emerging from the mesocosms. Two PUFAs, α‐linolenic acid and EPA, contributed to most of the variation in Chironomidae fatty acid content across the nutrient gradient. Whereas the proportions of α‐linolenic acid were positively correlated to the mesocosm TP concentration, we found a negative correlation for the proportions of Chironomidae EPA and mesocosm TP concentration. However, despite lower biomass‐specific EPA content at higher TP, higher biomass of emerging Chironomidae at intermediate TP concentrations resulted in higher total export of PUFAs from water to land. • Predation pressure from carp decreased the biomass of emerging Chironomidae on average 8‐fold. Chironomidae biomass showed a hump‐shaped relationship along the TP‐gradient and was strongly influenced by periphyton biomass. • Export rates of EPA and fatty acids in general responded in a quadratic manner along the nutrient gradient, reaching a maximum value at a TP of 400 µg/L and decreasing thereafter. • These findings highlight that the export of fatty acids from aquatic systems via adult Chironomidae is highly dependent on fish predation pressure, but also the nutrient concentrations of the system.
Drying in alpine streams might decrease aquatic-terrestrial trophic linkages by reducing terrestrial predation on aquatic prey. We tested this hypothesis by investigating whether a common riparian predator (hunting spiders) in alpine environments assimilated a lower proportion of aquatic prey with increasing stream intermittency. We used high temporal-resolution data from electrical resistance sensors to map patterns of naturally-occurring flow intermittency across 30 headwater streams of Val Roseg, a glacierized catchment in the Swiss Alps. We collected riparian hunting spiders, as well as potential terrestrial and aquatic macroinvertebrate prey, from streams and their associated riparian zones across two seasons (Alpine spring and summer). We estimated aquatic contributions to spider diets (pA) using (i) a gradient approach with aquatic invertebrate and spider carbon stable isotope ratio values (δ13C), and (ii) Bayesian carbon and nitrogen (δ15N) isotope mixing models. Spider pA from the gradient method were not statistically different from zero in spring (0.08 ± 0.10) and low in summer (0.16 ± 0.04). Mixing models also estimated low dependence on aquatic prey in both seasons, although with potentially higher contributions in summer. Spider diet did not vary with increasing flow intermittency in either season. Our results suggested that alpine hunting spiders obtain most of their carbon from terrestrial prey. The slight increase in spider pA during summer may correlate with peak emergence periods for aquatic insects, indicating opportunistic feeding by this riparian predator.
Stream and riparian food webs can be strongly linked by inputs of aquatic emergent insect prey to terrestrial predators. However, quantifying these linkages and understanding how they vary in time and space is challenging. We investigated the dynamic width of a riverine trophic subsidy zone by determining the relationship between perpendicular distance from a river and dietary contributions of aquatic insect prey to web-building spiders' diets. To assess this relationship, riparian web-building spiders at two river sites were sampled during four seasons and analysed for the fatty acids 16:0, 16:1ω7 and 20:5ω3, their total ω3-fatty acid content and their ω3:ω6 ratio to evaluate trophic subsidies reaching them from an adjacent river. River-derived fatty acids generally declined with increased distance from the river, indicating a diffusion of aquatically derived subsidies into the riparian zone. While the river was only 16 m wide at its broadest, river-derived trophic subsidies were detected up to four times that distance from the river edge. Spiders at a downstream section of the river, characterised by generally higher emergence rates of aquatic insects, contained higher proportions of aquatic indicator fatty acids compared with spiders located upstream, where emergence rates were lower. Similarly, proportions of aquatic indicator fatty acids in spiders were lowest during winter when aquatic insect emergence rates were lowest. The fatty acid 20:5ω3 (eicosapentaenoic acid; EPA) held the best promise as a biomarker of aquatic-derived tropic subsidies and could be developed as a useful tool for riparian research and management.
Compound-specific isotope analyses (CSIA) of fatty acids (FA) constitute a promising tool for tracing energy flows in food-webs. However, past applications of FA-specific carbon isotope analyses have been restricted to a relatively coarse food-source separation and mainly quantified dietary contributions from different habitats. Our aim was to evaluate the potential of FA-CSIA to provide high-resolution data on within-system energy flows using algae and zooplankton as model organisms. First, we investigated the power of FA-CSIA to distinguish among four different algae groups, namely cyanobacteria, chlorophytes, haptophytes and diatoms. We found substantial within-group variation but also demonstrated that δ ¹³ C of several FA (e.g. 18:3 ω 3 or 18:4 ω 3) differed among taxa, resulting in group-specific isotopic fingerprints. Second, we assessed changes in FA isotope ratios with trophic transfer. Isotope fractionation was highly variable in daphnids and rotifers exposed to different food sources. Only δ ¹³ C of nutritionally valuable poly-unsaturated FA remained relatively constant, highlighting their potential as dietary tracers. The variability in fractionation was partly driven by the identity of food sources. Such systematic effects likely reflect the impact of dietary quality on consumers' metabolism and suggest that FA isotopes could be useful nutritional indicators in the field. Overall, our results reveal that the variability of FA isotope ratios provides a substantial challenge, but that FA-CSIA nevertheless have several promising applications in food-web ecology. This article is part of the theme issue ‘The next horizons for lipids as ‘trophic biomarkers’: evidence and significance of consumer modification of dietary fatty acids’.
To understand consumer dietary requirements and resource use across ecosystems, researchers have employed a variety of methods, including bulk stable isotope and fatty acid composition analyses. Compound-specific stable isotope analysis (CSIA) of fatty acids combines both of these tools into an even more powerful method with the capacity to broaden our understanding of food web ecology and nutritional dynamics. Here, we provide an overview of the potential that CSIA studies hold and their constraints. We first review the use of fatty acid CSIA in ecology at the natural abundance level as well as enriched physiological tracers, and highlight the unique insights that CSIA of fatty acids can provide. Next, we evaluate methodological best practices when generating and interpreting CSIA data. We then introduce three cutting-edge methods: hydrogen CSIA of fatty acids, and fatty acid isotopomer and isotopologue analyses, which are not yet widely used in ecological studies, but hold the potential to address some of the limitations of current techniques. Finally, we address future priorities in the field of CSIA including: generating more data across a wider range of taxa; lowering costs and increasing laboratory availability; working across disciplinary and methodological boundaries; and combining approaches to answer macroevolutionary questions. This article is part of the theme issue ‘The next horizons for lipids as ‘trophic biomarkers’: evidence and significance of consumer modification of dietary fatty acids’.