1
Supplementary Methods
2
Comparison of Metabolism Estimates to Alternative Reference Lake
3
We produced metabolism estimates using methods described in the main text for an
4
alternative reference lake, Paul Lake, which has been used as a reference lake in many whole-
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ecosystem experiments over 30+ years (e.g. Carpenter et al. 1987). We evaluated the treatment
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effect on all metabolic variables by comparing differences in paired sampling events pre- and
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post-manipulation in the treatment basin and Paul Lake using the Welch t test (Stewart-Oaten et
8
al. 1992). The treatment basin significantly increased compared to the alternative reference lake
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(Paul Lake) in both GPP and R and significantly decreased in NEP (Welch t test for GPP n pre =
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105, n post = 192, t = -3.61, df =292.49, p < 0.001; Welch t test for R n pre = 105, n post = 192, t
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= -9.05, df = 287.85, p < 0.001; Welch t test for NEP n pre = 105, n post = 192, t = 5.28, df =
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230.24, p < 0.001). This indicates that the increase in GPP and R and decrease in NEP is
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significant in our treatment basin and is not due to just a negative DOC treatment in the reference
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basin. For all subsequent analyses, we use our reference lake, West Long, for interpretation of
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the results as supplementary data (e.g. allochthonous DOC loading) was not available for Paul
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Lake.
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Supplementary Table
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Table S1. Mean and standard error (in parenthesis) of additional physiochemical variables not
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discussed in the main text. Data are organized into the treatment and reference basins both pre-
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and post-manipulation. Filtered water color (Color) was measured spectrophotometrically using
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a 10 cm cuvette and expressed as an absorption coefficient for 440 nm (g440 = 2.303 x
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(absorbance at 440nm / 0.1 m) (Houser 2006). Chlorophyll a (Chl a) was measured using
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methanol extraction and a fluorometric method (Welschmeyer 1994). pH was measured weekly
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at 0.5 m intervals using a YSI multiprobe profiler (YSI Professional Plus; YSI Incorporated), and
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we average all measurements in the surface mixed layer for each sampling date. Particulate
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organic carbon (POC) was analyzed by filtering approximately 200 mL of water from the
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surfaced mixed layer through 0.45 µm GF/F filters. POC samples were run on a Finnigan Delta
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Plus isotope ratio mass-spectrometer (Thermo Scientific, Waltham, Massachusetts, USA).
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Soluble reactive phosphorus (SRP) was measured on filtered water using a colorimetric assay
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(Menzel and Corwin 1965). Total nitrogen (TN) was analyzed using a spectrophotometric
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method following persulfate digestion of unfiltered water (Olsen 2008). Iron (Fe) concentrations
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were measured by inductively coupled argon plasma spectrophotometry (ICP) on a Thermo
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Elemental Iris Intrepid ICP-OES using Standard Method 3120 B (APHA 1995; EPA 1994) with
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a method detection limit of 0.05 mg L-1. Fe measurements were done in the Grand Rapids
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Forestry Sciences Lab of the USDA Forest Service. Color, TN, Fe, and Chl a significantly
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increased in the treatment basin while pH significantly decreased (Welch t test for Color n pre =
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27, n post = 29, t = -15.59, df =51.94, p < 0.001; for TN n pre = 25, n post = 16, t = -2.19, df
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=16.02, p = 0.044; for Fe n pre = 7, n post = 7, t = -4.38, df = 7.86, p = 0.002; for Chl a n pre =
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25, n post = 30, t = -2.23, df = 43.83, p = 0.031; for pH n pre = 22, n post = 11, t = 2.70, df =
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13.49, p = 0.018). Particulate organic carbon and SRP did not significantly change due to the
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experimental manipulation (Welch t test for POC n pre = 27, n post = 26, t = -1.37, df =43.01, p
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= 0.18; for SRP n pre = 24, n post = 24, t = -0.71, df = 45.81, p = 0.48). Observations of the pre-
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(n pre) and post-manipulation (n post) periods are from paired sampling time points.
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Basin (Pre / Post
Color
Chl a
manipulation)
(g440)
(µg L-1)
Treatment (Pre)
3.99 (0.15)
6.48 (0.55)
Reference (Pre)
3.52 (0.19)
Treatment (Post)
Reference (Post)
pH
POC
SRP
TN
Fe
(mg L-1)
(µg L-1)
(µg L-1)
(mg L-1)
6.07 (0.11)
0.72 (0.08)
5.17 (0.29)
446.58 (12.36)
0.04 (0.01)
6.22 (0.48)
6.50 (0.13)
0.65 (0.05)
5.23 (0.82)
433.43 (9.74)
0.03 (0.01)
7.02 (0.23)
7.93 (1.14)
5.94 (0.18)
0.86 (0.08)
6.63 (0.79)
498.67 (60.59)
0.12 (0.04)
3.16 (0.11)
4.95 (0.60)
6.91 (0.10)
0.67 (0.05)
5.85 (0.84)
348.10 (21.20)
0.06 (0.05)
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46
References
47
APHA. 1995. Standards methods for the examination of water and wastewater, 19th ed.,
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American Public Health Association / American Waters Works Association / Water
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Environment Federation, Washington, DC.
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