Supplementary material
B.B. Bressendorff & S.Toft:
Dome-shaped functional response induced by nutrient imbalance of the prey
Materials and methods
The wolf spider Pardosa amentata has an annual life cycle in Denmark, with reproduction in early
spring (May) and hibernating in the subadult instar. For the experiment with adult females (see
below) they were collected as subadults in April and raised in the laboratory to the adult stage. For
the experiment with subadults, the animals were collected in September.
Experimental procedures
Experiment with adult females: In the laboratory the spiders were fed fruit flies raised on dogfoodenriched medium (1) until their maturation moult. They were then kept at 10 °C until all the
spiders had moulted. Before the experiment started they were fed fruit flies for two days and
starved for one day, so that all individuals were in the same life-history stage and nutritional
condition. For the experiments, fruit flies of two nutritional compositions were used (2): highprotein flies raised on a medium of 40% Drosophila medium and 60% casein; high-lipid flies raised
on a medium of 80% Drosophila medium and 20% sucrose. The lipid-rich flies contained a
lipid:protein ratio of 0.89, while the protein-rich flies contained a lipid:protein ratio of 0.10 (3). By
using the same prey raised on different media instead of two different prey types we avoided a
potentially confounding factor.
The experiment was run at a fixed temperature of 21 °C and 14 h light/10 h dark
photoperiod. The spiders were weighed and placed individually in 14 cm petri-dishes with moist
filter paper. The animals were divided randomly in two groups that received different pretreatments, lipid-rich flies or protein-rich flies, ad libitum. This intended to create nutritional
imbalance in two opposite directions. After four days of pre-treatment the spiders were starved
for one day before the functional response part of the experiment. The animals from each of the
pre-treatment groups were divided in eight sub-groups. These were split between four prey
densities (2, 5, 10 or 15 flies per petri-dish) and two treatments (lipid-rich flies, protein-rich flies)
with 6 replicates for each combination. The functional response part of the experiment was
repeated during seven days with daily prey replacement: the number of flies eaten was recorded,
non-eaten flies and prey remnants were removed, and new flies were added. Samples of the lipidrich and protein-rich flies used as food were also collected each day. For logistical reasons the
experiment was completed in two blocks, separated by one week, with 3 replicates of all
combinations in each block. Sample flies and food remnants from each day were dried in a
vacuum oven at 60 °C for at least 24 hours and weighed on a Sartorius balance (precision 0.001
mg). This allowed calculation of the daily individual food consumption of the spiders.
Experiment with subadults: This experiment was run as the adult experiment with a few minor
differences. After collection the spiders were kept at 10 °C until the pre-treatments started, i.e.
without any standardization. Due to the subadult spiders’ smaller size, the prey densities were 1,
3, 5, and 8 flies per petri dish per day. The experiment was also completed in two blocks,
separated by a month, and run in a basement room with near-constant temperature of 20 °C and
a 12h light/12h dark photoperiod. Prey remnants were not collected, so only the number of prey
killed was analyzed.
Statistical analysis: The data from each experimental series were first analyzed with a repeated
measures ANOVA, using the number of flies killed or biomass of flies eaten as the dependent
variable, and days as the repeated factor. Prey density, pre-treatment fly type, functional response
fly type, and block were factors, and spider body mass was entered as covariate; all possible
interaction terms of the three first-mentioned factors were also included in the initial models.
Subsequently, the results for each day were analyzed by 3-way ANCOVA with prey density, pretreatment fly type and functional response fly type as factors and spider body mass as covariate. A
significant prey-density*fly-type interaction (shown by asterisks after day # in each panel of Fig. 1)
will here indicate a possible dome-shaped response. Similar 3-way ANCOVAs were also made on
the results for the two highest prey densities alone, since it is in this range that a difference
between a simple type II and a dome-shaped type II response would be visible (significance
indicated by asterisks in the space between data points for the two highest densities in each panel
of Fig. 1). Finally, a comparison between fly type treatments at the highest prey density was made
with the t-test (or a Welch t-test if variances were unequal) (significant differences shown by
asterisks to the right of the curves in each panel of Fig. 1). Reduced models were obtained by
backwards deletion of non-significant interaction terms. A total of 89 adults and 92 subadults were
used in the two experiments, respectively.
References
1. 1. Mayntz,D. & Toft, S. 2001 Nutrient composition of the prey’s diet affects growth and
survivorship of a generalist predator. Oecologia 127, 207-213.
2. Mayntz, D., Raubenheimer, D., Salomon, M., Toft, S. & Simpson, S.J. 2005 Nutrient-specific
foraging in invertebrate predators. Science 307, 111-113.
3. Jensen, K., Mayntz,D., Toft, S., Raubenheimer,D. & Simpson, S.J. 2011 Prey nutrient
composition has different effects on Pardosa wolf spiders with dissimilar life histories.
Oecologia (DOI 10.1007/s00442-010-1811-1)
Table S1. Statistical analysis (repeated measures ANOVA – reduced models) of the number of flies
killed or biomass of flies consumed by the wolf spider Pardosa amentata during functional
response experiments run over 7 days. All models originally included the factors fly type, pretreatment fly type, prey density, and all their interactions, as well as block and spider mass. Pretreatment fly type was consistently non-significant. Analysis of each day’s results, see Table S2.
A.
Exp.: dep. var.
Source
F
df
P
Adults: No. flies killed
Spider mass
5.01
1,84
0.0278
Prey density
62.6
1,84 <0.0001
Days*fly type
2.0
Days*prey density
B.
C.
Adults: consumption
Subadults: No. flies killed
10.4
6,79
0.0145
6,79 <0.0001
Days*fly type*prey density
2.8
6,79
0.0172
Spider mass
4.4
1,83
0.0380
Prey density
61.9
Days*block
2.6
6,78
0.0251
Days*fly type
2.6
6,78
0.0238
Days*prey density
9.4
6,78 <0.0001
Days*fly type*prey density
2.8
6,78
0.0154
Prey density
15.0
1,89
0.0002
Days*fly type
4.7
6,84
0.0004
Days*prey density
11.0
1,83 <0.0001
6,84 <0.0001
Table S2. Statistical analyses of each day’s results from functional response experiments (overall
analyses in Table S1). Full models included the factors pre-treatment fly type, experimental fly
type, prey density and their interactions, and spider mass as covariate. Models were reduced by
backwards deletion of non-significant interactions.
Source
Adult exp.
P
Exp fly-type*prey
density (two highest
densities)
Exp. fly type (highest
density only)
Day 1
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
0,6967
0,0621
146,9009
4,4754
0,4062
0,8037
<,0001
0,0373
F1,41=0.70
P=0.41
t=0.41
P=0.69
Day 2
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
0,4808
1,4105
31,6015
3,6450
0,4900
0,2383
<,0001
0,0597
F1,41=0.74
P=0.39
t=0.16
P=0.87
Day 3
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
Exp fly type * Prey density
0,9485
1,2734
19,3743
2,4000
4,0643
0,3329
0,2624
<,0001
0,1251
0,0470
F1,41=6.11
P=0.0177
t=2.45
P=0.0270
Day 4
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
Pre-tr fly type * Exp fly type
Exp fly type * Prey density
3,3799
4,2143
16,6037
2,3798
5,0528
8,2375
0,0696
0,0433
0,0001
0,1268
0,0273
0,0052
F1,41=4.61
P=0.0378
t=3.22
P=0.0045
Day 5
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
Exp fly type * Prey density
1,4728
3,5967
14,6270
1,4671
5,9558
0,2283
0,0614
0,0003
0,2292
0,0168
F1,41=1.97
P=0.17
t=2.73
P=0.0154
Day 6
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
2,4725
1,9221
8,4790
0,0688
0,1196
0,1693
0,0046
0,7937
F1,41=0.20
P=0.66
t=0.62
P=0.54
Day 7
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
0,1444
1,2558
13,8102
2,5518
0,7049
0,2656
0,0004
0,1139
F1,41=0.85
P=0.36
t=1.62
P=0.12
Day 1-7
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
0,8158
0,4144
60,5780
4,3556
0,3690
0,5215
<,0001
0,0399
F1,41=4.46
P=0.0407
t=2.18
P=0.0440
Day 1
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
0,8617
6,4476
125,0008
3,2476
0,3559
0,0130
<,0001
0,0751
F1,41=0.46
P=0.50
t=0.71
P=0.49
Day 2
Pre-treatment fly type
Experimental fly type
Prey density
0,6200
6,0190
31,7061
0,4333
0,0162
<,0001
F1,41=0.66
P=0.42
t=0.79
P=0.44
No. flies killed
(df=4,84)
Adult exp.
F1
Amounts eaten
(df=4,84)
Spider mass
0,0867
Day 3
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
0,4457
0,0000
16,4777
1,8664
0,5062
0,9951
0,0001
0,1755
F1,41=6.03
P=0.0184
t=1.84
P=0.084
Day 4
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
Pre-tr fly type * Prey density
Exp fly type * Prey density
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
Exp fly type * Prey density
2,7788
0,4420
13,5228
2,3543
4,5104
7,6541
0,0993
0,5080
0,0004
0,1288
0,0367
0,0070
F1,41=4.44
P=0.0412
t=2.73
P=0.0127
1,0510
0,2674
13,0448
1,4241
4,9675
0,3083
0,6065
0,0005
0,2361
0,0285
F1,41=1.77
P=0.19
t=1.96
P=0.067
Day 5
Subadult
experiment
3,0055
Day 6
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
1,9168
6,3557
9,2967
0,0450
0,1699
0,0136
0,0031
0,8326
F1,41=0.30
P=0.59
t=1.01
P=0.32
Day 7
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
0,0201
0,0013
13,2135
2,2666
0,8875
0,9710
0,0005
0,1359
F1,41=0.96
P=0.33
t=1.02
P=0.32
Day 1-7
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
0,4100
2,2775
61,1106
4,0986
0,5237
0,1350
<,0001
0,0461
F1,41=4.70
P=0.0360
t=0.95
P=0.35
Day 1
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
0,1041
0,6902
41,5253
0,1013
0,7477
0,4084
<,0001
0,7511
F1,42=1.02
P=0.32
t=0.94
P=0.36
Day 2
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
0,3860
3,6944
1,2708
2,1111
0,5360
0,0579
0,2627
0,1498
F1,42=3.63
P=0.064
t=2.61
P=0.0169
Day 3
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
0,0876
5,5958
0,0002
0,2738
0,7680
0,0202
0,9878
0,6021
F1,42=0.60
P=0.44
t=1.82
P=0.086
Day 4
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
0,0200
13,0048
0,0184
0,0012
0,8878
0,0005
0,8925
0,9720
F1,42=0.15
P=0.70
Welch t=2.55
P=0.0227
Day 5
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
0,3770
2,0804
0,0001
0,5862
0,5408
0,1528
0,9911
0,4460
F1,42=0.94
P=0.34
t=1,.50
P=0.15
Day 6
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
0,5508
0,1903
7,1029
0,0000
0,4600
0,6638
0,0092
0,9984
F1,42=0.87
P=0.36
t=0.30
P=0.77
No. flies killed
(df=4,87)
Day 7
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
2,1124
1,8706
4,3991
0,0406
0,1497
0,1749
0,0389
0,8408
F1,42=0.23
P=0.64
Welch t=1.29
P=0.22
Day 1-7
Pre-treatment fly type
Experimental fly type
Prey density
Spider mass
0,1289
1,7235
14,1760
0,4072
0,7204
0,1927
0,0003
0,5251
F1,42=1.18
P=0.28
t=1.13
P=0.27