1
APPENDIX S1: Source of genotypes
2
Table A1 shows the source lake for each genotype. Clone lines were started from a
3
single parthenogenetic female from each lake, ensuring that individuals in each line
4
are genetically identical.
5
6
Table A1 Source lake characteristics for the four genotypes of Daphnia pulicaria.
Location
Surface
Lake Clone
(closest city)
Lindsay 2C
Total P (μg/L)
0.1250a
13.7a
8.0b
44°55’ N 76°40’ W
0.155b
26.0b
Mississippi Station, ON
45°08’ N 78°30’ W
Glen GLEN
0.1630d
15.0d
Haliburton, ON
46°28’ N 80°56’ W
Ramsey RAM
7.9520d
20.5d
Sudbury, ON
a
Provided by Nelson Laboratory, Queen’s University, Kingston, ON
Long
7
44°32’ N 76°23’ W
Lake Opinicon, ON
Area
Maximum Depth (m)
(km2)
4B
8
b
From Reavie & Smol, 2001.
9
c
From Reavie et al, 2006.
10
d
Provided by Dorset Environmental Science Center (DESC), Dorset, ON
11
12
4.1c
10.3d
11.5d
13
APPENDIX S2: Maternal effects
14
Since we were interested in studying life-history traits across a range of resources that
15
went from near starvation to moderate abundance, there were some resource levels
16
where reproduction did not occur (see Results). As such, it was not possible to control
17
for maternal effects for all resource levels. To address the issue of maternal effects,
18
we conducted two rounds of experiments. In the first round, all offspring used in the
19
treatments came from mothers raised at the highest resource level and thus all life-
20
history responses include effects due to the difference between maternal and offspring
21
resource environment. In the second round, we took offspring from mothers raised at
22
resource levels where reproduction occurred and raised these individuals in the same
23
environment as their mother. Thus, we were able to compare the influence of maternal
24
environment on life-history trait correlations for the four highest resource levels. Note
25
that the two experiments were conducted concurrently, and are only distinguished for
26
ease of communicating the analyses and results.
27
To evaluate whether maternal effects had an influence on correlation
28
coefficient, we estimated correlation coefficients for those individual with maternal
29
effects, and those without maternal effects (see Methods), which yielded two
30
estimates of correlation coefficients for each genotype at the four highest resource
31
levels (0.5, 0.42, 0.34, 0.26 mgC/day). Reproduction was insufficient at lower
32
resource levels to raise enough individuals with controlled maternal effects. A two-
33
factor ANOVA with maternal effects and treatment indicated no significant impact of
34
maternal effects on correlation coefficient for any pair of traits (1- correlation:
35
F=2.9823, df=1,24, p=0.0970; 1- correlation: F=1.2463, df=1,24, p=0.2753, -
36
correlation: F=0.8018, df=1,24, p=0.3794).
37
38
2
39
APPENDIX S3: Life-history characterization
40
Figure S1 Example growth trajectories (circles) and corresponding fit von Bertalanffy
41
growth model (lines). Fit maximum growth rate (1) is shown beside each trajectory.
0.407
2.5
0.3882
0.36
0.3287
Length (mm)
2.0
0.2676
1.5
1.0
0
10
20
30
40
50
60
70
Age (days)
42
43
44
Figure C2 Correlation between maximum growth rate (1) and the common metric of
45
growth at fixed age (Vanni & Lampert 1992, Boersma & Vijverberg 1994), assuming
46
instar duration of 2.5 days (Taylor & Gabriel 1993).
3
0.45
Max Growth Rate
0.40
0.35
0.30
0.25
1.2
1.4
1.6
1.8
2.0
2.2
Length at age 9
47
48
49
50
51
52
4
53
APPENDIX S4: Shapiro-Wilk test for normality of life-history traits
54
Table D1 Results (p-value) of a Shapiro-Wilk test for normality of each life-history
55
trait (1=growth, =reproduction, =age at death). Values less than 0.05 indicate non-
56
normality.
Genotype
GLEN
4B
Ramsey
2C
Trait
F=0.04
F=0.08
F=0.16
F=0.26
F=0.34
F=0.42
F=0.5
1
0.472
0.447
0.597
0.903
0.223
0.103
0.023

-
-
0.048
0.029
0.100
0.218
0.157

0.000
0.036
0.520
0.011
0.000
0.159
0.027
1
0.092
0.048
0.105
0.034
0.453
0.489
0.373

-
-
0.000
0.009
0.017
0.445
0.033

0.000
0.000
0.111
0.174
0.185
0.750
0.010
1
0.053
0.137
0.248
0.163
0.690
0.294
0.553

-
-
0.000
0.077
0.026
0.138
0.328

0.000
0.001
0.628
0.082
0.024
0.002
0.000
1
0.017
0.495
0.004
0.046
0.661
0.307
0.212

-
-
0.000
0.000
0.006
0.099
0.377

0.000
0.000
0.004
0.005
0.028
0.011
0.003
57
58
59
5
60
APPENDIX S5: Growth variation comparison
61
We observed growth variation among genetically identical individuals raised in
62
carefully controlled environments, which is common in zooplankton experiments (see
63
analysis in Ananthasubramaniam et al. 2011). To ensure this was not an artifact of our
64
transfer protocol, we compared the growth variance in our experiment to that found in
65
the literature. The highest resolved data is found in Lynch (1988), which shows an
66
increase in growth variance with incresing age that peaks around 10 days old (Figure
67
E1). Converting to a common basis of carbon per day, two of our food levels (0.5 and
68
0.42 mgC/day) correspond to the lowest food level studied in Lynch (1988).
69
Comparing these two studies, the dynamics of growth variance in our experiments
70
follow a very similar age pattern and is quantitatively similar in terms of both peak
71
and average variance values (Figure E2).
72
73
74
75
76
77
78
79
80
81
82
83
84
6
85
FIGURE E1. Dynamics of growth variance among genetically identical zooplankton
86
individuals from Lynch (1988) across nine food levels.
0.05
0.04
Variance
0.03
0.02
0.01
0.00
0
10
20
30
40
Age (days)
87
88
89
90
91
92
93
94
7
95
FIGURE E2. Dynamics of the growth variance among genetically identicals
96
observed in our experiments. a) Variance dynamics at 0.5 mgC/day for all four
97
genotypes (2C: blue, 4B: green, GLEN: yellow, RAMSEY: red) compared to the 0.1
98
mg C/day food level from Lynch (1988; dotted). b) Variance dynamics at 0.42
99
mgC/day for all four genotypes (2C: blue, 4B: green, GLEN: yellow, RAMSEY: red)
100
compared to the 0.8 mg C/day food level from Lynch (1988; dotted).
0.05
a
0.05
0.03
0.03
Variance
0.04
Variance
0.04
b
0.02
0.02
0.01
0.01
0.00
0.00
0
10
20
Age (days)
30
40
0
10
20
30
Age (days)
101
102
103
104
105
106
107
108
109
8
40
110
APPENDIX S6: Proportion reproducing
111
Figure F1. Proportion of adults that reproduced before death. Genotypes are shown in
112
different colors (red: RAMSEY, yellow: GLEN green: 4B, blue: 2C)
1.0
Proportion Reproducing
0.8
0.6
0.4
0.2
0.0
0.04
0.08
0.16
0.26
0.34
0.42
0.50
Treatment - mgC/vial/day
113
114
115
116
117
118
9
119
APPENDIX S6: Correlation coefficient analysis
120
Table G1 Statistical tests of whether the correlation coefficients differ from zero
121
(1=growth, 𝜔=reproduction, δ=age at death). Values in the top part of the table are p-
122
values from t-tests at each resource level. The next two blocks show if the null
123
hypothesis of no difference from zero is rejected (●) or not (◦). The first of these is
124
without adjusting the type I error rate, and the second is adjusting using Sidak’s
125
correction. Corresponding alpha values shown in parentheses in the first column.
Trait
Correlations
F=0.04
F=0.08
F=0.16
F=0.26
F=0.34
F=0.42
F=0.5
1-
-
-
0.0084
0.0004
0.0077
0.0028
0.0377
1-
0.0122
0.0940
0.357
0.0031
0.0117
0.0017
0.0005
-
-
-
0.391
0.257
0.106
0.0221
0.0392
Trait
Correlations
Significance without adjusting type I error rate
1- (α=0.05)
-
-
●
●
●
●
●
1- (α=0.05)
●
◦
◦
●
●
●
●
- (α=0.05)
-
-
◦
◦
◦
●
●
Trait
Correlations
Significance with adjusting type I error rate
1-
(α=0.0102)
-
-
●
●
●
●
◦
1-
(α=0.0073)
◦
◦
◦
●
◦
●
●
-
(α=0.0102)
-
-
◦
◦
◦
◦
◦
126
127
128
129
10
130
APPENDIX REFERENCES
131
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133
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135
Daphnia species and their hybrid in a eutrophic lake: evidence for food limitation.
136
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137
Brookfield, J.F.Y. (1984) Measurement of the intraspecific variation in population
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Genetica, 63, 161-174.
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12