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SUPPLEMENTARY ONLINE MATERIAL
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Assessing trophic position from nitrogen isotope ratios: effective
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calibration against spatially varying baselines
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Paul Woodcock1, David P. Edwards1, Rob J. Newton2, Felicity A. Edwards1,
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Chey Vun Khen3, Simon H. Bottrell2, Keith C. Hamer1
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1
Institute of Integrative and Comparative Biology, University of Leeds, Leeds, LS2 9JT, UK
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2
School of Earth and Environment, University of Leeds, Leeds, LS2 9JT, UK.
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3
Sepilok Forest Research Centre, Sandakan, Sabah, Malaysia
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Corresponding author: Paul Woodcock; email: bgy3pw@leeds.ac.uk
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Table S1:
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Details of approaches to baseline correction in 34 recent stable isotope studies
conducted in terrestrial ecosystems.
Table S2:
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Mean and standard deviation for repeated isotope analyses of ant and plant
samples.
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Table S3:
Standard deviations of random effects in linear mixed models.
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Figure S1:
Variation in plant δ15N values between transects, with and without samples of
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the Fabaceae.
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Table S1: Approaches to baseline correction in recent stable isotope studies conducted in terrestrial ecosystems
Study
# distinct consumer # spatially
Max. distance
Baseline
# Samples
Question, study taxa and
sampling sites
distinct baselines between sampling
material
per baseline ecosystem
recognised
used to interpret locations assumed
data
to have a constant
baselinea
No Baseline
Duyck et al.
(2011)
Pisanu et al.
(2011)
4
0 and 2b
? - but total sampling
area is 920m2
N/A
N/A
2
0 and 1b
4km
N/A
N/A
Bihn et al.
(2010)
12
0
? – Not given
N/A
N/A
O’Grady et
al. (2010)
1
0
? – but sampling area is
4 x 2km
N/A
N/A
Prochazka et
al. (2010)
1
0
120m
N/A
N/A
Vidal and
Sabat (2010)
5
0 and 2b
100km
N/A
N/A
Smith et al.
(2008)
4
0
Several kmc
N/A
N/A
2
0
? – Not given
N/A
N/A
1
0 and 1b
? – Not given
N/A
N/A
Tillberg et al.
(2006)
Yi et al.
(2006)
Examine effects of cover crops on
arthropod food web structure
Determine trophic positions of
rats on a Subantarctic island
δ15N used as a trait to assess
changes in ant functional diversity
along a successional gradient
Trophic ecology of ants in
temperate grassland.
Compare trophic positions of 4
understorey bird species across a
forest-savannah ecotone
Divergence in resource use
amongst mainland and island
lizard populations
Investigate intra- and intercolonial variation in diet of
harvester ant castes
Inter and intra-colonial variation
in trophic position of ants
Describe food web in alpine
meadow
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a
No baseline means that the study effectively assumes that all sampling locations have a constant baseline of zero.
A combination of uncorrected and corrected data were presented/analysed
c
Principal conclusions on caste determination unaffected, because all castes are collected from all nests (thereby averaging out baseline variation), but cannot be confident
on whether intercolonial variation is genuine.
b
Single Baseline
Study
# distinct
consumer
sampling sites
recognised
# spatially
distinct baselines
used to interpret
data
Max. distance
between sampling
locations assumed
to have a constant
baselinea
Pisanu et al.
(2011)
2
0 and 1b
4km
Hyodo et al.
(2010)
1
1
?- Not given
6
1
? – Not given, but
sampling area is 2000m2
Leaf litter, grass
and top soil
Leaves, litter,
dead wood and
soil
Leaf litter and
roots
29
1
Several km
Soil, plants
382
5
1
? – Not given
Moss, herbs and
grass
18
1
1
< 50m
Termites
11
1
0 and 1b
? - Not given
Herbivores
?
5
1
? - Not given
Plants, litter and
soil
10
4
1
? – Not given, but
sampling area is 2000m2
Plants and
stubble
19
10
1d
>50 km
Plants
15-29
2
1
≈3km. Within-site
sampling area not givene
Leaves
37
Pollierer et
al. (2009)
Traugott et
al. (2008)
Sanders and
Platner
(2007)
Kupfer et al.
(2006)
Yi et al.
(2006)
Halaj et al.
(2005)
Schmidt et
al. (2004)
Tooker and
Hanks
(2004)
Blüthgen et
al. (2003)
Baseline
material
# Samples
per baseline
?
?
15
Question, study taxa and
ecosystem
Determine trophic positions of
rats on a Subantarctic island
Describe tropical forest food
web, including birds, mammals
and invertebrates
Describe soil food web in
temperate forest
Trophic positions of wireworm
species in European farmland
Intraguild interactions between
predatory arthropods in
grassland and meadow.
Describe tropical forest food
web.
Describe alpine meadow food
web.
Describe soil invertebrate food
web in coniferous forest and
examine effects of thinning
Soil food web in arable field
Investigate trophic position of
flower beetle on 2 plant species
in Illinois and Indiana prairies
Omnivory and food web
structure in arboreal rainforest
ants
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d
e
Study presents data separately for beetles on 2 different plant species, with each plant species used as a separate baseline
Models investigating interspecific variation in ant δ15N values included the δ15N value of the plant from which the ant was collected as a covariate.
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Multiple Baselines
Study
# distinct
consumer
sampling sites
recognised
# spatially
distinct baselines
used to interpret
data
Duyck et al.
(2011)
4
0 and 2b
Gibb and
Cunningham
(2011)
12
Hawke and
Clark (2010)
Baseline
material
# Samples
per baseline
Question, study taxa and
ecosystem
? - but each baseline
applies to an area of
460m2
Plants
?
Examine effects of cover crops
on arthropod food web structure
12
<6m
Leaf litter,
grass, top soil
3 homogenised
samples
Comparing ant community
across regenerating pastures
2
2
<5m
Soil
Homogenised
sample analysed
in duplicate
Describe arthropod food web in
penguin burrows
Menke et al.
(2010)
4
4
? – Not given, but may
be >100m
Plants
4-5 plant species
Smith and
Suarez (2010)
8
8
0m
Seeds
1 homogenised
sample
Vidal and Sabat
(2010)
5
0 and 2f
50km
Seeds &
fruits
?
2
2
30m
Plants, soil
and litter
25
10
2 and 10g
5m in some analyses, 601000m in other analyses
Plants
?
McGlynn et al.
(2009)
7
7
<10m?
Leaf litter
York and
Billings (2009)
5
5
? – Not given
Plants
El-Wakeil
(2009)
Kozhu et al.
(2009)
Max. distance
between sampling
locations assumed
to have a constant
baselinea
3 replicates of 3
homogenised
samples
1-7 per
sampling
location
Examine spatial variation in
trophic position of
chaparral/scrub ants
Trophic position of harvester ant
castes
Divergence in resource use
amongst mainland and island
lizard populations
Description of soil food web in
coniferous forest
Description of arthropod and
mammal food web on grasslands
Determining predictors of ant
d15n values
Compare trophic level within
and between fruit bat species
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f
g
2 mainland sites and 3 island sites examined, each separated by ≈50-100km. Correction carried out at mainland and island levels. Within site distances not given.
Data sometimes interpreted at the site level (= 2 distinct baselines) and sometimes at each of 5 sampling locations within each site (= 10 distinct baselines)
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Multiple Baselines (contd.)
Study
# distinct
consumer
sampling sites
recognised
# spatially
distinct baselines
used to interpret
data
Max. distance
between sampling
locations assumed
to have a constant
baselinea
Baseline
material
Hyodo et al.
(2008)
6
2
1kmh
Soil, leaf litter
& grass
4-5
Takimoto et al.
(2008)
36
36
<20m
Leaves
5-10
Daugherty and
Briggs (2007)
4
4
0m
Leaves
10-31
Tillberg et al.
(2007)
6
6
? – Not given
Plants
6-21
Gratton and
Denno (2006)
36
4
? – Not given, but >10m
Plants, leaf
litter & soil
core
5-10
6
6
0m
Pine needles
5
4
4
5m
Leaf litter and
bark
8
3
3
? – Not given
Leaves
?
Mooney and
Tillberg
(2005)
Schneider et
al. (2004)
Davidson et al.
(2003)
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29
30
31
# Samples
per
baseline
Question, study taxa and
ecosystem
Effect of humification on termite
& earthworm δ15N values in
forest & savannah
Effect of disturbance on food
chain length, focusing on lizards
& spiders
Trophic structure of arthropod
community in pear orchards
Spatial variation in trophic
position of invasive ant species
in woodland/pasture
Effect on arthropod food web of
removing an invasive plant from
salt marshes
Spatial variation in ant omnivory
in pine forest
Niche differentiation in orbatid
mites in temperate forest.
Herbivory and food web
structure in arboreal ant food
web of rainforests
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Within vegetation types, δ15N values differ by ≈1.5‰
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Blüthgen, N, Gebauer G, Fiedler K (2003) Disentangling a rainforest food web using stable isotopes: dietary diversity in a species-rich ant
community. Oecologia 137:426-435
Daugherty MP, Briggs CJ (2007) Multiple sources of isotopic variation in a terrestrial arthropod community: challenges for disentangling food
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El-Wakeil KF (2009) Trophic structure of macro- and meso-invertebrates in Japanese coniferous forest: carbon and nitrogen stable isotope
analyses. Biochem Syst Ecol 37:317-324
Gibb H, Cunningham SA (2011) Habitat contrasts reveal a shift in the trophic position of ant assemblages. J Anim Ecol 80:119-127
Gratton C, Denno RF (2006) Arthropod food web restoration following removal of an invasive wetland plant. Ecol Appl 16:622-631
Halaj J, Peck RW, Niwa CG (2005) Trophic structure of a macroarthropod litter food web in managed coniferous forest stands: a stable isotope
analysis with δ15N and δ13C. Pedobiologia 49:109-118
Hawke DJ, Clark JM (2010) Isotopic signatures (13C/12C; 15N/14N) of blue penguin burrow soil invertebrates: carbon sources and trophic
relationships. New Zeal J Zool 37:317-321
Hyodo F, Tayasu I, Konate S, Tondoh JE, Lavelle P, Wada E (2008) Gradual enrichment of 15N with humification of diets in a below-ground
food web: relationship between 15N and diet age determined using 14C. Funct Ecol 22:516-522
Hyodo F, Matsumoto T, Takematsu Y, Kamoi T, Fukuda D, Nakagawa M, Itioka T (2010) The structure of a food web in a tropical rain forest in
Malaysia based on carbon and nitrogen stable isotope ratios. J Trop Ecol 26:205-214
Kohzu A, Iwata T, Kato M, Nishikawa J, Wada E, Amartuvshin N, Namkhaidorj B, Fujita N (2009) Food webs in Mongolian grasslands: the
analysis of 13C and 15N natural abundances. Isot Environ Healt S 45:209-219
Kupfer A, Langel R, Scheu S, Himstedt, W, Maraun M (2006) Trophic ecology of a tropical aquatic and terrestrial food web: insights from
stable isotopes (15N). J Trop Ecol 22:469-476
McGlynn TP, Choi HK, Mattingly ST, Upshaw A, Poirson EK, Betzelberger J (2009) Spurious and functional correlates of the isotopic
composition of a generalist across a tropical rainforest landscape. BMC Ecology 9:23
Menke SB, Suarez AV, Tillberg CV Chou CT, Holway DA (2010) Trophic ecology of the invasive argentine ant: spatio-temporal variation in
resource assimilation and isotopic enrichment. Oecologia 164:763-771
Mooney KA, Tillberg CV (2005) Temporal and spatial variation in ant omnivory in pine forests. Ecology 86:1225-1235
O’ Grady A, Schmidt O, Breen, J (2010) Trophic relationships of grassland ants based on stable isotopes. Pedobiologia 53:221-225
Pisanu B, Caut S, Gutjah S, Vernon P, Chapuis J-L (2011) Introduced black rats Rattus rattus on Ile de la Possession (Iles Crozet, Subantarctic):
diet and trophic position in food webs. Polar Biol 34:169-180
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Pollierer MM, Langel R, Scheu S, Maraun M (2009) Compartmentalisation of soil animal food web as indicated by dual analysis of stable
isotope ratios (15N/14N and 13C/12C). Soil Biol Biochem 41:1221-1226
Prochazka P, Reif J, Horak D, Klvana P, Lee RW, Yohannes E (2010) Using stable isotopes to trace resource acquisition and trophic position in
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Sanders D, Platner C. (2007) Intraguild interactions between spiders and ants and top-down control in a grassland food web. Oecologia 150:611624
Schmidt O, Curry JP, Dyckmans J, Rota E, Scrimgeour CM (2004) Dual stable isotope analysis (δ13C and δ15N) of soil invertebrates and their
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Schneider K, Migge, S, Norton, RA, Scheu S, Langel R, Reineking A, Maraun M (2004) Trophic niche differentiation in soil microarthropods
(Orbatida, Acari): evidence from stable isotope ratios (15N/14N). Soil Biol & Biochem 36:1769-1774
Smith CR, Anderson KE, Tillberg CV, Gadau, J, Suarez AV (2008) Caste determination in a polymorphic social insect: nutritional, social and
genetic factors. Am Nat 172:497-507
Smith CR, Suarez AV (2010) The trophic ecology of castes in harvester ant colonies. Funct Ecol 24:122-130
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Table S2: Mean and standard deviation (SD) of δ15Nair values for repeated isotope analyses
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of ants collected from within single sampling points: Lophomyrmex bedoti (Ant1),
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Pachycondyla obscurans (Ant2 and Ant3), Paratrechina sp3 (Ant4) and Pseudolasius sp1
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(Ant5) and sp2 (Ant6). The standard deviation for repeats of a homogeneous leaf sample
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(EUP) run in several columns is also shown. Numbers of repeats analysed per sample are
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shown in brackets.
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Ants
Plant
Sample1 Sample2 Sample3 Sample4 Sample5 Sample6 EUP
(5)
(3)
(3)
(3)
(3)
(3)
(38)
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Mean
5.77
(‰)
6.89
7.54
6.03
5.64
5.46
0.00
SD
(‰)
0.20
0.21
0.07
0.36
0.36
0.25
0.18
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Table S3: Standard deviation of random effects in linear mixed models of corrected ant δ15N
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values versus local baseline for overall ant community, ant subfamily, and ant species
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analyses within unlogged and logged forests. The greater the standard deviation, the more
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variation is attributable to the random effect.
Overall
Unlogged
Logged
Subfamily
Unlogged Logged
Species
Unlogged
Logged
subfamily
1.85
1.58
-
-
0.20
0.32
species (within subfamily)
species
0.96
-
0.86
-
1.01
0.87
-
-
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Figure S1: Mean plant δ15N values (‰) + for the entire dataset (n=160 plants; ALL) and for
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each transect (n = 20 plants per transect) in (a) unlogged forest and (b) logged forest. Shaded
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columns have Fabaceae samples removed for comparison. Transects are presented in rank
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order, from highest plant δ15N value to lowest.
(a)
3.5
All leaf samples
3
Mean plant δ15N
Excluding Fabaceae samples
2.5
2
1.5
1
0.5
0
ALL
-0.5
1
2
3
4
5
6
7
8
Sampling location in rank order
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(b)
Sampling location in rank order
ALL
1
2
3
4
5
6
7
8
Mean plant δ15N
0
-0.5
-1
-1.5
-2
All leaf samples
Excluding Fabaceae samples
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-2.5
10