# Nutritional adaptation in the marine to freshwater establishment process - how do diet and genotype shape phenotype?

Cornelia Twining, Cameron Hudson, Jernej Bravničar, Antonella Carosi, Gaël

P.J. Denys, Philine Feulner, Žiga Fišer, Hanna Rosinger, Verena Saladin,

Linda Zanella, et al.

► To cite this version:

Cornelia Twining, Cameron Hudson, Jernej Bravničar, Antonella Carosi, Gaël P.J. Denys, et al. Nutritional adaptation in the marine to freshwater establishment process - how do diet and genotype shape phenotype?. Journal of Experimental Biology, In press, 10.1242/jeb.251462. hal-05532790

HAL Id: hal-05532790

https://hal.science/hal-05532790v1

Submitted on 2 Mar 2026

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# Nutritional adaptation in the marine to freshwater establishment process - how do diet and genotype shape phenotype?

Cornelia W. Twining<sup>1,2,*†</sup>, Cameron M. Hudson<sup>1,3,*</sup>, Jernej Bravničar<sup>4</sup>, Antonella Carosi<sup>5</sup>, Gael P. J. Denys<sup>6</sup>, Philine G.D. Feulner<sup>1,7</sup>, Žiga Fišer<sup>8</sup>, Hanna Rosinger<sup>1</sup>, Verena Saladin<sup>7</sup>, Linda Zanella<sup>9</sup>, Davor Zanella<sup>9</sup>, Catherine L. Peichel<sup>7</sup>, Blake Matthews<sup>1</sup>

## Abstract

Nutrients, including vital organic compounds, vary in availability across ecosystems, with the potential to act as a strong source of selection for traits that increase nutrient acquisition and biosynthesis. Compared to freshwaters, marine ecosystems are much richer in the omega-3 long-chain polyunsaturated fatty acid docosahexaenoic acid (DHA) and thus marine animals establishing new freshwater populations are faced with the challenge of acquiring DHA. However, the relative roles of DHA synthesis capacity and diet in the freshwater establishment process remain unresolved. We used common garden experiments to explore phenotypic responses to dietary nutrient content in threespine sticklebacks (*Gasterosteus aculeatus*) that

<footnote n="1"><sup>1</sup>Department of Fish Ecology and Evolution, Eawag, Kastanienbaum, Switzerland</footnote>

<footnote n="2"><sup>2</sup>Department of Environmental Systems Science, ETH Zürich, Zürich, Switzerland</footnote>

<footnote n="3"><sup>3</sup>Department of Aquatic Ecology, Eawag, Dübendorf, Switzerland</footnote>

<footnote n="4"><sup>4</sup>University of Ljubljana, Biotechnical Faculty, Department of Animal Science, Ljubljana, Slovenia</footnote>

<footnote n="5"><sup>5</sup>University of Perugia, Department of Chemistry, Biology, and Biotechnologies, Perugia, Italy</footnote>

<footnote n="6"><sup>6</sup>UAR Patrinat 2006, Museum of Natural History (OFB - MNHN - CNRS - IRD), Paris, France</footnote>

<footnote n="7"><sup>7</sup>Division of Evolutionary Ecology, Institute of Ecology and Evolution, University of Bern, Bern, Switzerland</footnote>

<footnote n="8"><sup>8</sup>University of Ljubljana, Biotechnical Faculty, Department of Biology, Ljubljana, Slovenia</footnote>

<footnote n="9"><sup>9</sup>Department of Biology, Faculty of Science, University of Zagreb, Zagreb, Croatia</footnote>

<footnote>*Co-first authors</footnote>

<footnote>†Author for correspondence: cornelia.twining@eawag.ch</footnote>
varied in their genetic capacity for DHA synthesis. We found that diet as well as presumed metabolic-
based adaptation to freshwater nutritional environments (population identity) had strong effects on
stickleback phenotypes and performance. Sticklebacks enriched with marine-derived fatty acids including
DHA grew more and were in better condition compared to controls. Those fed diets with more DHA also
accumulated more DHA in muscle tissue. Freshwater sticklebacks accumulated more DHA compared to
those from a marine population. However, populations with greater *fads2* copy numbers did not
consistently have higher performance or DHA accumulation. Together, these results suggest that DHA
availability during development can strongly influence phenotypes and performance, with the potential to
act as a source of selection. Our findings also suggest that mechanisms beyond the accumulation of *fads2*
copies, such as plasticity in expression or variation in other DHA synthesis pathway genes, could be
important adaptations to the nutritional constraints of freshwater.

Keywords: threespine sticklebacks, fatty acids, fads2, marine, freshwater, diet, synthesis

**Introduction**

Fish have established new freshwater populations and lineages from marine ancestral populations
numerous times (e.g., Seehausen and Wagner 2014). Such establishments have required them to adapt to
novel abiotic conditions, notably lower salinity (Lee and Bell 1999; Velotta et al. 2022), as well as novel
biotic conditions, including resources that vary in nutritional composition compared to those in marine
ecosystems. In particular, the physiologically-important omega-3 long chain polyunsaturated fatty acids
(n-3 LC-PUFA) eicosapentaenoic acid (EPA; 20:5n-3) and docosahexaenoic acid (DHA; 22:6n-3) are
both abundant throughout marine food webs and relatively scarcer, especially in the case of DHA, in
freshwater food webs (Twining et al., 2021). EPA and DHA are critically important for a host of
physiological functions in animals, including neural, immune, and cardiac function, and reproduction
(Bell & Tocher, 2009; Brenna et al., 2009; Brenna & Carlson, 2014; Das, 2006) and serve as limiting
nutrients for many fish (Sargent et al., 1995; Tocher, 2003). As limiting nutrients, n-3 LC-PUFA
availability may serve as a source of selection for fish and other marine taxa seeking to exploit the novel
nutritional conditions of freshwaters, and thus promote the evolution of nutritional adaptations.
Consequently, as marine fish establish new freshwater populations, they can provide a unique window
into the role of nutrients in the process of adaptation to freshwater environments.

One way that consumers can escape n-3 LC-PUFA limitation is through increased synthesis from
dietary precursors like alpha linolenic acid (ALA; 18:3n-3; Twining et al. 2021). Some animals can
source all of the DHA that they require by adding double bonds and carbon atoms to these precursors via
the Sprecher or Δ4 pathway and then using a combination of fatty acid desaturase (*fads*) and elongase
(elovl) enzymes (Oboh et al., 2017), encoded by *fads* and *elovl* genes respectively (Castro et al., 2012, 2016; Monroig & Kabeya, 2018). Freshwater fishes appear to have adapted to freshwater nutritional constraints through changes in the function, efficiency, and expression of these genes (Zhang et al., 2023). For instance, freshwater flatfish species have increased DHA synthesis capacity through changes in the efficiency and function of the *fads2* and *elovl5* genes (Matsushita et al., 2020) while freshwater threespine sticklebacks have increased their copy numbers of *fads2* (Ishikawa et al., 2019).

In this study, we focus on threespine stickleback (*Gasterosteus aculeatus*), which have repeatedly established freshwater populations from ancestral marine populations, across a range of different freshwater nutritional environments (i.e., in lakes as well as streams) across the Northern Hemisphere following deglaciation (Fang et al., 2018, 2020; McKinnon & Rundle, 2002; Hudson et al., 2024; Ishikawa et al., 2021). Ishikawa et al. (2019) found that multiple freshwater lineages of threespine stickleback had more copies of the *fads2* gene than contemporary marine stickleback populations, presumably allowing freshwater lineages to more readily synthesize DHA. In addition, stream fish in the Pacific Northwest with more DHA-poor benthic diets had higher *fads2* copy numbers than lake fish that consumed more DHA-rich pelagic prey (Ishikawa et al., 2021). Across lakes in Greenland, stickleback in lakes with a higher proportion of DHA-rich copepods had fewer *fads2* copies compared to those from nearby lakes with fewer copepods, suggesting that local dietary DHA availability can select for nutritional adaptation (Hudson et al., 2024).

Previous data suggest that nutritional habitat-based differences can lead to increased *fads2* copy numbers and may consequently improve stickleback performance in DHA-poor environments as part of adaptation to freshwater. However, this has not been tested experimentally along a gradient of marine to freshwater populations varying in their *fads2* copy numbers and history of freshwater establishment. Importantly, while experimental previous studies have compared the phenotypes and performance either marine (i.e., Ishikawa et al., 2019) or freshwater (i.e., Hudson et al., 2022) population pairs, we lack comparative data on marine and freshwater stickleback performance and phenotypes under common garden conditions. Moreover, previous studies comparing the fatty acid content of two marine or freshwater populations did not manipulate diet, leaving the importance of *fads2* copy numbers versus diet quality in shaping freshwater stickleback phenotypes unclear.

Here, we explored adaptation to the nutritional constraints of freshwater environments in threespine sticklebacks, through common garden experiments with both marine and freshwater fish. We included freshwater populations that had contrasting *fads2* copy numbers from lineages of variable estimated ages since freshwater establishment. We also manipulated dietary n-3 LC-PUFA availability to understand how n-3 LC-PUFA synthesis and n-3 LC-PUFA retention from diet affect fatty acid phenotypes and performance, including survival. This design allowed us to examine both the general
freshwater nutritional adaptation process (i.e., marine-freshwater differences) as well as the specific importance of *fads2* (i.e., variation across freshwater populations) in this process.

# Materials and Methods

## *Study design*

To understand the effects of both population and diet on fatty acid phenotypes as well as performance, we conducted a common garden experiment, where we manipulated nutritional quality directly for multiple populations of sticklebacks that varied in their *fads2* genotypes (Tables 1, S1; Figure 1). Five European freshwater populations from across Baltic, northern/central European, and southern European Mediterranean lineages, plus one North American marine population (Puget Sound, USA) were raised under common conditions on chironomid-based diets that varied in DHA content, mimicking either marine or freshwater prey. We examined the effects of diet and population-identity (representing genetically divergent lineages) on survival, growth, body condition, and fatty acid phenotypes of these populations.

We chose populations from European lineages that varied in their time of freshwater establishment (Fang et al., 2018; Hudson, Lucek, et al., 2021); Tables 1, S1). Stickleback populations from the Baltic Sea lineage, which have been introduced around the Bodensee (i.e., Lake Constance; our Chriesbach, CH), were estimated to have established freshwater populations within the last 10-5 kya during the post-Pleistocene (Fang et al., 2020; Hudson, Lucek, et al., 2021; Liu et al., 2016; Mäkinen & Merilä, 2008; Marques et al., 2019). Populations from the Swiss midlands (i.e., Binnenkanal, CH), western Switzerland (i.e., Chessel, CH) and the Seine drainage around Paris, North of France (i.e., Réveillon, FR) were assumed to be of mixed European lineages. These admixed populations originate from lineages that established during the Pleistocene in southern Europe in the lower Rhône and other Mediterranean drainages up to 40 kya as well as those that established after the Pleistocene (i.e., ~11.7-5 kya) in northern and central Europe in the Seine, Rhein, and other Atlantic or North Sea drainages (Fang et al., 2020; Hudson et al., 2023; Lucek et al., 2010). We also included a central Italian population (i.e., Clitunno River, IT) estimated to be from an older (≥ 25 kya) pre-glacial southern Mediterranean lineage (Coll-Costa et al., 2024; Fang et al., 2018). All freshwater populations came from streams because macroinvertebrate prey from streams typically have lower DHA availability compared to those in lakes, making them the most challenging freshwater nutritional environment for establishment by animals of marine origin (Twining et al. 2021).
Breeding and husbandry

In spring of 2022, we first visually assessed the egg ripeness of females and only sacrificed those that looked most ready to breed. We also sacrificed adult males that were in breeding color. After sacrificing and rinsing tricaine methanesulfonate (i.e., MS-222) off with clean stream water, we gently released clutches of eggs from females into petri dishes. We then dissected testes out of males, mashed these between two glass slides, and washed this mixture over eggs with stream water. After letting the egg clutches and testes sit for ~5-10 minutes to allow fertilization to occur, we washed fertilized clutches into clean plastic test tubes, leaving head space for gas exchange, and brought tubes back to the lab in coolers of cool stream water.

In the Réveillon River at Villecresnes (France) we caught two males, which we mated with one ripe female, resulting in one split clutch (i.e., half of the clutch had one male parent and the other half had the other male parent). In the Clitunno River (Italy) we produced two successful clutches from two males and two females. In Grand Canal Chessel, Binnenkanal, and Chriesbach (Switzerland), we produced eight, five, and five successful clutches respectively from male and female pairs. After breeding in the field, we took fin clips from each individual and stored these in absolute ethanol for genetic analysis (see below). In addition, we acquired nine clutches of Puget Sound (USA) marine sticklebacks that were already in culture at the University of Bern.

All clutches were initially placed in plastic cups with window screen mesh bottoms that were suspended in aquaria to allow water circulation and gas exchange. Air bubblers were placed on the bottoms of the aquaria directly under cups in order to ensure oxygenation and frequent water movement. We visually inspected all clutches daily and manually removed eggs with fungi with the aid of a dissecting microscope. Eggs took between 8-14 days to hatch with entire clutches hatching within a few hours of each other. When eggs hatched, we removed bubblers from the bottom of tanks and reduced air flow until fish began swimming freely and feeding. We initially fed juveniles with Salt Lake *Artemia spp.*, nauplii that were hatched in salt water, rinsed and filtered into fresh water. When fish were approximately 28-35 days old, we began introducing them to finely chopped frozen chironomid larvae, gradually acclimating them to a full, non-chopped chironomid diet. We used aquarium heaters to keep all aquaria at approximately 18 °C throughout the experiment and we added NaCl (100 mg/L) to all aquaria to help prevent fungal infections while maintaining salinity below that of typical marine conditions. To minimize tank-specific effects, we moved fish to new tanks halfway through experiments. Tank water was changed weekly or more frequently if fish showed signs of disease or stress.

In late summer to fall of 2022, we placed fish into experimental treatments when they were approximately 75-90 days old (variation due to differences in clutch hatching dates within the same population). Clutch number and egg survival to initial larval stage varied across populations, resulting in
different numbers of clutches per population so we mixed clutches within populations and did not include clutches as a random effect in our models (see below). Fish from multiple mixed clutches within populations were placed in each 100 L aquaria and then randomly assigned to diet treatments with either *ad libitum* thawed chironomids or *ad libitum* thawed chironomids soaked in a marine oil enriching mixture (Selco Easy DHA artemia enricher, INVE Aquaculture; Figure 1a, S1). Due to variation in breeding success and clutch survival among populations, we created five replicated tanks of each diet treatment for the marine population as well as the three Swiss populations (i.e., Grand Canal Chessel, Binnenkanal, and Chriesbach), three replicate tanks per diet treatment for the Italian population, and one tank per treatment for the French population. We took samples of both diet treatments monthly for analyses of fatty acid content.

We examined the effects of diet on growth and body condition by weighing and photographing each fish three times over the course of the experiment at approximately 158 ± 5 days (10 weeks in experiment), 201 ± 6.7 days (16 weeks in experiment), and 242 ± 8 days (22 weeks in experiment) old. We used ImageJ (Ferreira & Rasband, 2011) to calculate fish standard lengths from photos taken of fish next to a ruler and calculated average tank body condition as tank average fish mass divided by tank average fish length. We concluded experiments when fish were approximately 246-270 days old and close to reproductive age. At this point, we photographed each individual in a cuvette with color blocks and length bars, then weighed it, and sacrificed it *via* MS-222 overdose. After sacrifice, we dissected each fish to confirm its sex based on its gonads and took skinned tail muscle samples for fatty acid analysis.

Marine populations experienced high mortality within the first two months of our experiments. One individual from a marine oil-enriched diet treatment died within the first eight days of the experiment. Two additional marine tanks on control chironomid diets experienced 80% mortality (i.e., four fish from each tank of five died) within the first two months of the experiment. We replaced all of these individuals with fresh individuals from stock tanks of the same age and mix of parents (from breeding described above) that had been fed control chironomids. All individuals in one of these tanks subsequently died before the next measurement. We included fish from the one surviving replacement tank in fatty acid content analyses, but not in subsequent analyses of mortality or performance.

A widespread *Vibrio* infection occurred throughout our aquaria facility midway through our rearing study causing 56%, 30%, 22%, 16%, 7%, and 4% mortality in marine, French, Binnenkanal, Chriesbach, Clitunno, and Chessel populations respectively. Fish in stock tanks began showing symptoms in late November and *Vibrio* infection was confirmed by mid-December by which time fish in experiments (~200 days old) began getting sick as well. Diagnosis of sick fish and assessment of potential treatments was conducted by the Institute for Fish and Wildlife Health at the University of Bern. In mid-December, we began treating fish showing signs of disease (fast breathing, lying at the bottom of tanks,
not eating) with Enrox® 10% (100 mg/mL Enrofloxacin). Tanks with fish showing signs of disease were given an initial 1 mL dose of Enrox, followed by a 50% percent water change and then subsequent dosing three and five days later.

## Genetic analyses

We used genomic quantitative PCR (g-qPCR) to investigate *fads2* copy number variation across our five freshwater populations (Figure 1; Table S1). In addition to the parents from all freshwater sites, we also included an additional juvenile female caught from the Reveillon River (France); an additional 16 females and 10 males from Clittuno River (Italy); an additional 16 females and 12 males from Binnenkanal (Switzerland); an additional 13 females and 17 males from Chessel (Switzerland); and an additional 17 females and 14 males from Chriesbach (Switzerland).

First, we extracted DNA from fin clips preserved in absolute ethanol using Quiagen ® DNeasy Blood & Tissue Kits and used the resulting extract for genomic-qPCR (g-qPCR). After extraction, we quantified DNA concentrations with a Nanodrop 1 (Thermo). As in Hudson et al. (2024) we modified our g-qPCR method from Ishikawa et al., (2019), using the same forward and reverse primers, and a TaqMan probe on a region conserved among *fads2* haplotypes. These primers amplify both *fads2* on stickleback chromosome 12 as well as the transposed copy (and any duplications of this) on chromosome 19 (i.e., the X chromosome). Following Ishikawa et al. (2019), we used thyroid stimulating hormone β2 (*tshβ2*), to calculate the relative copy number of *fads2* (henceforth copy number) for each individual. *tshβ2* is a single copy gene without known duplications in threespine stickleback, meaning that the two copies present in a diploid stickleback are normalized to a copy number of one. g-qPCR reactions were performed on a LightCycler ® 480 System (Roche) with Real-Time PCR Master Mix (Roche); we ran each reaction in duplicate (i.e., two reactions with *fads2* primers and two with *tshβ2* primers). We then calculated relative *fads2* copy numbers using the delta-delta Ct method (Livak & Schmittgen, 2001) to compare the mean Ct values for *fads2* and *tshβ2* across replicates. We excluded samples from further analyses and/or reanalyzed them when *fads2* or *tshβ2* copy number differences between replicates were greater than 0.5. As we normalized copy number relative to the single copy gene *tshβ2*, a relative copy number of one for *fads2* reflects a total genomic copy number of two.

## Fatty acid content analyses

We quantified fatty acid content of 90 sticklebacks and their chironomid diets from our experiments (Table S1). We either extracted lipids three times by immersing tissues in a 1:2 methanol:dichloromethanol (DCM) mixture, followed by placing ashed glass vials in an ultrasonic ice water bath for 10 minutes, transferring extract into new ashed and pre-weighed test tubes with glass
Pasteur pipettes, and drying down the extract with nitrogen gas (N2). We then added 50 µL of C23:0 at a 483.6 µg/mL concentration to each extracted lipid sample via Hamilton syringe as a recovery standard and then added 1 mL of Boron trifluoride (BF3) to each tube with a glass pipette, following vortexing and five minutes of water bath sonication. We then placed tubes in an oven at 100 °C for two hours. After removing methylated samples from the oven and allowing them to cool, we added 2 mL milliQ purified water to each, followed by approximately 1 mL hexane. We then vortexed samples, and used glass pipettes to transfer the hexane fraction to an HPLC vial three times. After transferring hexane to vials, we again dried them down with N2, and resuspended fatty acid methyl esters (FAME) in 500 µL EtOAc for gas chromatography flame ionization detection (GC-FID) analysis. We analyzed samples on a Shimadzu GC-2010 Plus with 30m InertCap 5MS/NP GC column using a 47 minute run starting at 70 °C and ramping up to 150 °C and then 320 °C running in splitless injection mode with a column flow of 2.07 mL/min. We identified compounds based on retention times relative to a Supelco FAME mix (G4-C24, 18919-1AMP) using the software GC Solution (Shimadzu). We then calculated the content of each compound in µg/g by dividing peak area by the peak area of our C23:0 recovery standard, multiplying this by C23:0 total mass added to the sample, and dividing by mass of dried tissue.

## Statistical Analyses

### How does the fatty acid content of chironomid diets vary across treatments?

We visually compared control versus marine oil-treated chironomids (Figure S1) fed to sticklebacks using non-metric multidimensional scaling using the *metaMDS* (vegan, version 2.6-4) in R. All data was log transformed. We included the following fatty acids: 13:0, 14:0, 15:0, 15:1, 16:0, 16:1, 17:0, 18:0, 18:1n-9t, 18:2n-6c (LIN), 18:2n6t or 18:1n-9c, 18:3n-3 (ALA) or 18:3n-6, 20:0, 20:1, 20:2, 20:4n-6 (ARA), 20:5n-3 (EPA), 21:0, 22:0, 22:2, 22:5n-3 (DPA), 22:5n-6, 22:6n-3 (DHA), 23:3n-3 or 20:3n-6, and 24:0. No marine oil-treated chironomids contained the fatty acid 22:1n-9 so we did not include it in our NMDS visualization even though it was detected in control chironomids.

### How did fads2 copy numbers vary across freshwater populations?

To understand if there was significant variation in *fads2* relative copy number between our four of five freshwater European stickleback populations (i.e., the Reveillon population was excluded due to small sample size), we used a general linear model (GLM, base R) with a Gaussian error distribution and an identity link function to test for differences among populations, between sexes, and their interaction. We performed post-hoc pairwise comparisons between individual populations as well as between sex by population pairs using the R package *emmeans* (Lenth, 2024, version 1.10.5) with Tukey corrections for multiple comparisons.
*Were there initial differences in length, weight, and/or condition between populations?*

We also used GLM with a Gaussian error distribution and an identity link function to examine differences in length, weight, and condition at the start of experiments from the Clittuno, Chessel, Chriesbach, Binnenkanal, and marine populations. Ten fish were randomly measured from each population of mixed clutches.

*How did population and/or treatment influence mortality, size, and body condition?*

We used logistic regression to examine variation in mortality by population, treatment, and their interaction in five of our six experimental populations (i.e., excluding the Reveillon population without replicate tanks). Because some populations had little to no mortality (i.e., Clitunno and Chessel), we used a Firth bias-reduced logistic regression model (*logistf*, version 1.26.1) with a binomial error distribution and logit link function. We modeled mortality as mortality of all fish that originally started in treatments, not including replacement control fish from stock tanks. In cases where population and/or an interaction of population and treatment were significant, we performed post-hoc tests Tukey tests with *emmeans*.

We sought to understand how experimental dietary treatment and population influenced performance in five of our six experimental populations (i.e., excluding the Reveillon population without replicate tanks). We tested for differences in weight and length of sticklebacks using general linear mixed models (GLMM, *lme4* version 1.1-34) with population, treatment, and their interaction as fixed effects and tank as a random effect using models with Gamma error distribution and log link functions. Because body condition (i.e., mean weight of fish in tank / mean length of fish in tank) and changes in mean weight, length, and body condition were measured at the tank-level, we used GLM with a Gamma error distribution and log link function to examine differences in these variables based on population, treatment, and their interaction. Sex was not included in any of these models because fish were only sexed at the end of the experiment. We created separate models of weight and length at each of the three time measurement points (i.e., measurement 1 = approximately 10 weeks in experiments, measurement 2 = approximately 16 weeks in experiments, and measurement 3 = approximately 22 weeks in experiments). In cases where population and/or an interaction of population and treatment were significant, we performed post-hoc tests Tukey tests with *emmeans*.

*How did fatty acid content vary by population, treatment, and/or sex?*

We used GLMMs with a Gaussian family and identity link function to examine variation in the content of palmitic acid (16:0), oleic acid (18:1n-9), linoleic acid (18:2n-6; LIN), ALA, arachidonic acid (20:4n-6; ARA), eicosapentaenoic acid (20:5n-3; EPA), docosapentaenoic acid (22:5n-3; DPA), and DHA in fish from all six experimental populations with sex, population, treatment, and the three-way
interaction of population, treatment, and sex as fixed effects with tank identity as a random effect. In
cases where either population and/or interactions of population and treatment were significant, we
performed post-hoc tests Tukey tests with emmeans. Because sex was unbalanced across populations and
treatments, we subset emmeans models by sex.

All statistical analyses were conducted in R (R Core Team, version 4.5.0) and graphics were
created with the package ggplot2 (Wickham, 2011, version 3.4.4). Model fits were assessed with marginal
R² values and appropriateness of model fits was assessed with residual plots (ggResidpanel version 0.3.0).

**Results**

*fads2 copy numbers vary across European stickleback populations*

As expected due to the location of the *fads2* gene on the X chromosome, XY males had significantly lower *fads2* copy numbers than XX females across all sites (Figure 1a; Table 1). Fish collected from Chriesbach had significantly lower *fads2* copy numbers compared to fish collected from Binnenkanal (Bin - Chr; estimate = 0.798, SE = 0.1459, df = 149, t-ratio = 5.492, p < 0.0001), Chessel (Che - Chr; estimate = 1.103, SE = 0.139, df = 149, t-ratio = 7.955, p < 0.0001), and Clittuno (Chr - Cli; estimate = -0.602, SE = 0.157, df = 149, t-ratio = -3.830, p = 0.001), while fish collected from Chessel had *fads2* copy numbers that were significantly higher than those of fish collected from Chriesbach and Clittuno (Che - Cli; estimate = 0.501, SE = 0.153, df = 149, t-ratio = 3.267, p = 0.007). The four fish collected from the Reveillon River had values that were intermediate between those of Chriesbach and Binnenkanal (Figure 1a).

*Initial variation in condition among populations*

Prior to the onset of the experimental dietary treatment, fish from Grand Canal Chessel were
significantly lighter, shorter, and in lower condition (i.e., ratio of weight to length) compared to those
from other populations (Tables S2-4).

*The significance of diet and population on mortality*

Survival in experiments varied strongly by population. Mortality occurred within two waves: there was an initial period of low mortality from a variety of causes during the first two months of the experiment and a later period of high mortality was due to a widespread *Vibrio* infection across our entire facility. Throughout our experiments, significantly more marine stickleback died compared to other populations (Figure 2; Table 2).
*The significance of diet and population on length, weight, and condition*

Compared to sticklebacks on standard chironomid diets, fish supplemented with marine fats grew significantly heavier and increased their condition significantly more (i.e., heavier per length) throughout the experiment and were longer by the latter part of the experiments (Fig. 3; Tables 3-5). By 10 weeks into experiments, prior to the *Vibrio* outbreak in our facility, fish treated with marine fats were longer, heavier, and in better condition compared to those on control chironomid diets (Figure 3; Tables 3-5). Fish from Binnenkanal and Chessel were also significantly lighter and lower condition than those from Chriesbach or Clitunno. Binnenkanal fish on control diets were also significantly shorter than control fish from Chriesbach while treated Binnenkanal fish were shorter than treated fish from Chriesbach, Clitunno, and marine populations and treated fish from Chessel were shorter than treated fish from Chriesbach and marine populations.

By 16 weeks into experiments, while fish from several populations had become infected with *Vibrio* (i.e., different ages and time in experiment due to differences in hatch dates), overall patterns remained relatively unchanged (Figure 3; Table 3-5). Supplemented fish continued to be heavier, longer, and in better condition compared to those on control diets. Fish from Binnenkanal were still significantly lighter than those from Chriesbach and Clitunno as well as those from our marine population. Clitunno fish were in significantly better condition than those from Binnenkanal, Chessel, and Chriesbach and marginally better condition than marine fish by the second measurement. By this timepoint, marine fish were longer than those from all other populations. Fish from Binnenkanal were also significantly shorter than those from those from Chessel, Chriesbach while fish from Clitunno were significantly shorter than those from Chessel and Chriesbach, and but these effects no longer interacted with treatment. Marine fish were still longer at this point compared to fish from Binnenkanal and Clitunno.

After 22 weeks, after all populations had been exposed to *Vibrio*, fish supplemented with marine oils remained heavier, longer, and in better condition compared to those on control diets (Figure 3; Tables 3-5). Although many marine fish died (see above), those that did survive until the third measurement were significantly longer compared to those from other populations. There were no differences in weight by population by the third measurement. Meanwhile fish from Clitunno remained in significantly better condition compared to fish from Binnenkanal and in marginally better condition compared to fish from Chriesbach and marine populations.

*Fatty acid phenotypes vary with treatment, population, and sex in common garden*

Population, treatment, sex, and their interactions all influenced the content of n-3 and other PUFA (Figure 4) as well as the energy storage fatty acids palmitic and oleic acid. In the case of DHA, marine fish had significantly lower DHA content than other populations, but marine fish supplemented
with marine oils rich in DHA (Figure 1b) were able to achieve higher muscle DHA content than those on control diets (Figure 4c; Table 6; Table S5). Fish those from Clitunno (our oldest estimated population) had significantly higher DHA than other populations regardless of dietary treatment (Figure 4c; Table 6; Table S5).

Although chironomids treated with marine oil contained more of the omega-3 PUFA DHA precursors (Figure 4d), namely ALA, EPA, (Figure 1b) and DPA, fish did not differ in their contents of these fatty acids based on dietary treatment (Figure 4a-b; Tables S6-8). EPA content was significantly lower in Chessel and marine fish compared to those from Binnenkanal, but did not vary based on sex or its interactions (Table S7). ALA and DPA content were also significantly lower in fish from Chessel and marine populations, but these effects varied based on interactions between population, sex, and/or treatment (Tables S6, S8). Specifically, females did not vary in their ALA contents (Table S6), but males from Chessel and marine populations supplemented with DHA had lower ALA contents than those from Binnenkanal (Table S6), while males from Chessel and Chriesbach had higher DPA than males from Binnenkanal (Table S8).

Fish did not differ in their content of the omega-6 PUFA LIN based on diet, sex, population, or their interactions (Table S9-). The n-6 LC-PUFA, ARA was more complex: fish from Chessel had significantly lower ARA content than those from Binnenkanal or marine populations and male marine fish supplemented with DHA had higher ARA than those from Chessel (Table S10).

Contents of energy storage fat palmitic acid were significantly lower in DHA treated marine male fish, but did otherwise not vary based on dietary treatment, population, sex, or their interactions (Table S11). Oleic acid was lower in Chessel fish and well as those from our marine population, but this effect in marine fish varied based on interactions: marine male fish had higher oleic acid contents than females, but treated marine males had lower oleic acid contents than others (Table S12)

## Discussion

We sought to reveal the roles of diet and metabolic capacity in determining nutritional phenotypes and the consequences of this for growth and survival during the freshwater establishment process. To understand capacity for nutrient synthesis and accumulation, we conducted common garden experiments with threespine stickleback populations that varied in their time of freshwater establishment and their *fads2* copy numbers, suggesting among-population differentiation in their putative genetic capacity for n-3 LC-PUFA synthesis. To understand the effects of dietary DHA intake, we also manipulated the DHA content of diets in our experimental treatments. We found that the DHA content of diet had a strong effect on DHA phenotypes in marine fish (i.e., those with the lowest *fads2* copy
numbers). We also observed higher rates of growth and body condition across individuals with diets that were rich in DHA and other LC-PUFA. Both muscle fatty acid contents and performance also varied among populations. Importantly, across dietary treatments, mortality was highest and DHA content was lowest in marine populations. Together, these results suggest that dietary n-3 LC-PUFA availability during an individual's development, together with freshwater establishment history, can strongly influence stickleback fatty-acid phenotypes and survival, with the potential to act as a source of selection as populations establish in freshwater.

*Common garden conditions reveal effects of diet and metabolism on nutritional phenotypes*

Manipulating nutrient availability for stickleback populations with variable *fads2* genotypes in common garden experiments helped elucidate the processes of n-3 LC-PUFA synthesis and n-3 LC-PUFA retention from diet across populations. Supplementation with marine-derived fatty acids, including DHA, resulted in marine fish with higher muscle DHA, demonstrating that marine sticklebacks will preferentially retain dietary DHA in spite of their more limited ability to synthesize it. Interestingly, while the marine oil supplemented diets contained more of the n-6 LC-PUFA, ARA, and the n-3 LC-PUFA, DPA, muscular ARA and DPA contents did not vary with dietary treatment, suggesting that DHA availability was more limiting than ARA or DPA availability. This metabolic plasticity in response to variable nutritional environments might be an adaptive solution in environments with high nutritional heterogeneity (Raubenheimer et al., 2012), such as the streams used as sources for our freshwater populations.

Echoing the findings of previous studies, we found that lineage-based phenotypic differences in
nutritional metabolism became apparent while rearing fish under common garden laboratory conditions.
For instance, the oldest experimental lineage from the Clitunno River in Italy, which had some of the
highest *fads2* copy numbers, had higher DHA content compared to other populations. Previously, Hudson
et al. (2022) found that stickleback from an older Lac Léman population (Grand Canal Chessel)
accumulated more DHA compared to a more recently-established Bodensee population. Here, we found
that fish from Chessel and Chriesbach (i.e., Bodensee) populations, despite differing in growth and
condition (Figure 3), had DHA phenotypes that were similar to each other and intermediate compared to
other populations (Figure 4c). In addition to these differences within freshwater lineages, we found that
marine sticklebacks in experiments had significantly lower muscle DHA content compared to freshwater
lineages, highlighting both their more limited ability to synthesize DHA (control treatments) as well as
their more limited ability to accumulate DHA from diet (marine oil-treated diets). Together, this provides
further evidence that lineages with a longer history of freshwater establishment are able to accumulate
DHA more efficiently than those of marine or more recent freshwater establishment. However, in contrast
to our expectations, freshwater populations with higher *fads2* copy numbers did not consistently have correspondingly higher DHA content, suggesting that other genes (e.g., *elovl*) may underlie the nutritional adaptation process in freshwater.

Intriguingly, sticklebacks from Clitunno and Chessel populations, which had varying degrees of ancestry from pre-glacial Mediterranean lineages, both performed better and, in the case of Clitunno accumulated more DHA, than other freshwater lineages despite differing from each other in *fads2* copy numbers. Chessel fish also experienced the lowest mortality of all populations, echoing findings from previous mesocosm studies (Best et al., 2017; Hudson et al., 2023). Fish from Clitunno, the oldest freshwater population in our experiments, outperformed those from Chessel in terms of DHA accumulation even though fish from Chessel had significantly higher *fads2* copy numbers. Clitunno sticklebacks accumulated more DHA and were also in better condition than fish from other populations regardless of diet. These populations may have additional metabolic adaptations for freshwater nutritional environments beyond increased *fads2* copy numbers. Adaptations beyond increases in *fads2* copy numbers could include changes in *elovl* elongase genes involved with DHA synthesis as seen in flatfish (Matsushita et al., 2020), as well as increased ability to take advantage of DHA when it is available via changes in nutrient retention.

As in previous studies (e.g., Ishikawa et al. 2019, 2021; Hudson et al. 2024), freshwater female sticklebacks in our experimental populations had higher *fads2* copy numbers than their male counterparts (Fig. 1a). As females invest large amounts of lipids, including LC-PUFA into their eggs, it may be especially advantageous for females to have higher *fads2* copy numbers. We found that stickleback muscle tissue varied in ALA and DPA content based on sex and as well as interactions between sex, population, and treatment, such that females had higher ALA and DPA content than males (Fig. 4a). ALA is a precursor to the n-3 LC-PUFA EPA, DPA, and DHA while DPA is an intermediate between EPA and DHA (Fig. 4d). This suggests that within populations, males and females, despite varying in their *fads2* copy numbers, were able to achieve similar EPA and DHA levels in muscle tissue, albeit with potential costs in terms of their reserves of other precursor and/or intermediate fatty acids. Future studies could examine if females preferentially allocate their EPA and DHA to their gonads compared to males and thus have higher LC-PUFA content at the whole-body level.

*Diet determines growth, but population origin determines survival*

Across all populations, sticklebacks supplemented with marine-derived fatty acids, including DHA, grew larger and were in better condition than those on standard DHA-poor chironomid diets (Fig. 3). While diet had strong effects on growth, condition, and fatty acid phenotypes of sticklebacks in our common garden experiments, population origin had the strongest effects on survival. Fish throughout our
aquarium facility were unintentionally exposed to a *Vibrio* infection midway through experiments, allowing us to opportunistically examine effects of diet and population on performance under the challenge of infection. Regardless of dietary treatment, fish from marine populations had significantly higher mortality than those from other populations (Fig. 2, Tables 2). In contrast, populations from lineages that established the earliest in freshwater (e.g. Clitunno and Chessel) had zero mortality on marine supplemented diets and the lowest mortality of all populations on standard treatments (treatment differences not significant). Fish from more recently-established Binnenkanal and Chriesbach populations had intermediate levels of mortality across both dietary treatments. Population-based differences in mortality were not explained by *fads2* copy numbers, suggesting that additional genetic mechanisms, including those unrelated to nutrition, likely influence survival in the face of infection.

## Conclusions

Overall, we found clear evidence of metabolic-based adaptation to freshwater nutritional environments. Multiple European freshwater stickleback lineages accumulated increased genetic capacity for nutrient synthesis (i.e., greater *fads2* copy numbers). These populations, especially those with the longest histories of freshwater establishment, were able to accumulate more DHA from both synthesis and preferential retention compared to marine sticklebacks. In contrast, DHA content in marine sticklebacks varied based on dietary DHA availability, suggesting little use of synthesis. Freshwater populations also suffered lower mortality compared to the marine population. Together, these results suggest that DHA availability in diet may act as a source of selection as fish and other taxa of marine origin establish new populations in freshwater nutritional environments. Threespine stickleback are a classic example of a species whose ancestral form displays developmental plasticity that has ultimately given rise to multiple ecotypes through parallel selection (e.g., Wund et al. 2008). To further evaluate the strength of nutrients on selection, future studies should extend common garden rearing conditions to evaluate the effects of developmental plasticity throughout the stickleback life cycle, including evaluating the effects of diet and *fads2* during early post-larval development (i.e., during artemia feeding, as in Ishikawa et al. 2019), during the pre-reproductive stage (the current study), as well as reproductive investment and success later in life.

In general, our findings on the freshwater nutritional adaptation process echo studies on a diversity of other metabolic adaptations that have allowed species and populations to take advantage of novel nutritional resources. For instance, in humans, the repeated evolution of lactose tolerance likely increased survival on milk products during periods of nutritional stress and disease (Evershed et al., 2022). Likewise, the ability to survive upon freshwater resources and to accumulate DHA via both synthesis and selective retention appears to have repeatedly evolved across threespine sticklebacks,
including across the multiple European lineages included in this study, as well as more broadly (Ishikawa et al., 2019). Future studies should also examine whether *fads2* duplication and phenotypic differences in fatty acid accumulation and performance exist across other closely-related taxa, such as fourspine sticklebacks (*Apeltes quadracus*), brook sticklebacks (*Culaea inconstans*) and ninespine sticklebacks (*Pungitius pungitius*; e.g., Liu et al. 2022), that have encountered similar nutritional challenges while establishing freshwater populations.

## Acknowledgements

We thank Daniel Steiner, Danina Schmidt, Pascal Reichlin, Andreas Taverna, Luka Mrzelj, Ryan Greenway, Leighton King, David Marques, Kay Lucek, and Ole Seehausen for assistance with fish sampling, breeding, and rearing experiments. We thank Salome Mwaiko, Karin Beck, Oliver Hanson, and Nicole Nesvadba for laboratory assistance. We also thank Nadja Pepe and Patricia Achleitner for administrative and logistics assistance. CWT was funded by Marie Skłodowska Curie Action fellowship H2020-MSCA-IF-2020 101025939 from the European Research Council. ZF was supported by the Slovenian Research and Innovation Agency via grant no. P1-0184. Fish collection and breeding in Switzerland was permitted at the Swiss federal level under permits 33583 and LU02/2021. In Italy, sampling was authorized under permit no. 003169_2022 from the Umbria region. French fish were moved across the French-Swiss border under Nagoya protocol agreement TREL2103218S/461.

## Conflict of interest

We have no conflicts of interest to report.

## Author contributions

CWT, CMH, and BM conceived the ideas and CWT, CMH, PGDF, BM, and CLP designed methodology; CWT, CMH, BM, AC, GD, AC, DZ, LZ, ZF, and NB acquired permits and collected samples; CLP and VS provided lab-reared animals; CWT, CMH, and HR collected the data; CWT analysed the data; CWT, CMH, CLP, and BM led the writing of the manuscript. All authors contributed critically to the drafts and gave final approval for publication.

## Statement on inclusion

This study brings together authors from eight countries, including scientists based in the countries where the study was carried out. All authors were engaged early on with the research and study design to ensure that the diverse sets of perspectives they represent was considered from the onset. Whenever relevant, literature published by scientists from the region was cited.
**Data availability statement**

Data is stored on the Eawag institutional repository ERIC and will be made publicly available upon
manuscript acceptance.

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Figures and Tables

<figure><img src="image_2.png" /><figcaption>**Fig. 1.** (a) mean, standard deviation, and individual replicates (smaller points) of relative *fads2* copy numbers from field-sampled sticklebacks from populations used in experiments (Chriesbach: n = 24 female, n = 19 male; Réveillon: n = 2 female, n = 2 male; Binnenkanal: n = 21 female, n = 17 male; Clitunno: n = 18 female, n = 12 male; Chessel: n = 21 female, n = 25 male), (b) non-metric multidimensional scaling (NMDS) visualization of chironomid fatty acid content sampled monthly</figcaption></figure>
throughout experiment (n=7 per treatment), (c) stickleback source populations and number of replicated
tanks (n=5 fish per tank) for each of the two chironomid dietary treatments used in common garden
experiments, and (d) locations of sampled European freshwater populations. Populations are ordered
based on mean female parent copy numbers. Chriesbach is estimated to be the youngest populations from
the Baltic Sea lineage, which is estimated to have established freshwater populations within the last 10-5
kya. Binnenkanal, Chessel, and Réveillon are assumed to be of mixed European freshwater lineages
established during the Pleistocene (up to 40 kya) in southern Europe as well as after the Pleistocene (i.e.,
~11.7-5 kya) in Central and Northern Europe. Clitunno is estimated to be from an older (≥ 25 kya) pre-
glacial southern Mediterranean lineage. The *fads2* copy number data for marine sticklebacks from Puget
Sound (USA) is taken from Ishikawa et al., (2019).
<figure><img src="image_3.png" /><figcaption>**Fig. 2.** Mean and standard deviation of mortality (i.e., proportion of fish dying per tank) for each population by dietary treatment combination. Smaller points represent mortality for individual tanks (n = 5 tanks per treatment for Binnenkanal, Chessel, Chriesbach, and marine populations and n = 3 tanks per treatment for Clitunno). The experiment was run a single time in the laboratory. Data were analyzed with bias-reduced logistic regression followed by post-hoc Tukey-adjusted pairwise population comparisons (see methods). Post-hoc comparisons indicated that marine populations had higher mortality at the p = 0.018, 0.018, 0.018, and 0.06 level for Binnenkanal, Chessel, Chriesbach, and Clitunno respectively.</figcaption></figure>
<figure><img src="image_4.png" /><figcaption>(b)</figcaption></figure>

<figure><img src="image_5.png" /><figcaption>(c)</figcaption></figure>

<figure><img src="image_6.png" /></figure>
**Fig. 3.** Mean and standard deviation of stickleback (a) weight, (b) length, and (c) condition (ratio of weight to length) at all three experimental measurement points with onset of *Vibrio*-induced mortality for each population shown (gray shaded areas). Mean weight and length were measured at the individual level and were replicated as follows: Binnenakanal: n = 25/25, 20/24, 19/24; Chriesbach: n = 24/24, 21/21, 16/15; Chessel: n=25/25, 25/25, 25/25; Clitunno: n= 14/15, 14/15, 12/15; marine: n = 15/17, 15/17, 11/12 for control/treated fish at 10 weeks, 16 weeks, and 22 weeks respectively. Condition was calculated at the tank level and was replicated as follows: Binnenkanal, Chriesbach, Chessel: n = 5/5 control/treated tanks throughout the experiment; Clitunno: n = 3/3 control/treated tanks throughout the experiment; marine: n = 5/5 control/treated tanks at 10 weeks, n = 3/4 control/treated tanks at 16 weeks, and n = 2/4 control/treated tanks at 22 weeks. The experiment was run a single time in the laboratory. Data were analyzed with either GLMM (weight and length) or GLM (condition) followed by post-hoc Tukey-adjusted pairwise comparisons (see methods). Throughout the experiment, weight with treatment at the p < 0.001 level and condition at the p < 0.02. Condition and length both varied with population at the p < 0.05 level throughout the experiment while weight only varied with population (p < 0.05) at 10 weeks.
See results and tables for all interactions and pairwise comparisons.
<figure><img src="image_7.png" /><figcaption>**Fig. 4.** Mean and standard deviation of (a) ALA, (b) EPA, and (c) DHA content from fish muscle tissue and (d) the DHA synthesis pathway, with ALA, EPA, and DHA highlighted. Smaller points in (a), (b), and (c) represent individual fish. Sample sizes were as follows: Binnenkanal n=9/11, Chessel n = 10/10, Chriesbach n = 9/9, Clitunno n = 7/7, marine n= 7/5 and Réveillon n = 3/3 control/treated fish. The experiment was run a single time in the laboratory. Data were analyzed with either GLMM (weight and length) followed by post-hoc Tukey-adjusted pairwise comparisons (see methods). ALA varied based on population (p < 0.05), sex (p < 0.05), and their interactions (p = 0.05). EPA varied based on population at the p < 0.05 level. DHA content varied with population (p = 0.001) and interactions of population and treatment (p < 0.05). See results and tables for all interactions and pairwise comparisons.</figcaption></figure>
<table><caption>**Table 1. Fads2 copy numbers by sex and population of parents of experimental fish (Population effects are shown relative to Binnenkanal)**</caption><thead><tr><th>Predictors</th><th>Estimates</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>2.65</td><td>2.38-2.92</td><td>&lt;0.001</td></tr><tr><td>Population [Chessel]</td><td>0.42</td><td>0.03-0.80</td><td>0.034</td></tr><tr><td>Population [Chriesbach]</td><td>-1.11</td><td>-1.49- -0.73</td><td>&lt;0.001</td></tr><tr><td>Population [Clitunno]</td><td>0.09</td><td>-0.32-0.49</td><td>0.673</td></tr><tr><td>Sex [m]</td><td>-1.42</td><td>-1.83- -1.01</td><td>&lt;0.001</td></tr><tr><td>Population [Chessel] x Sex [m]</td><td>-0.23</td><td>-0.78-0.33</td><td>0.432</td></tr><tr><td>Population [Chriesbach] x Sex [m]</td><td>0.63</td><td>0.06-1.20</td><td>0.031</td></tr><tr><td>Population [Clitunno] x Sex [m]</td><td>-0.57</td><td>-1.19-0.06</td><td>0.076</td></tr><tr><td>Observations</td><td colspan="3">157</td></tr><tr><td>R<sup>2</sup></td><td colspan="3">0.640</td></tr></tbody></table>
<table><caption>Table 2. Mortality of fish in experiment (Population effects are shown relative to Binnenkanal)</caption><thead><tr><th>Predictors</th><th>Odds ratio</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>0.02</td><td>0.00 – 0.32</td><td>&lt; 0.001</td></tr><tr><td>Treatment [treated]</td><td>1.00</td><td>0.02 – 52.36</td><td>1.000</td></tr><tr><td>Population [Chessel]</td><td>1.00</td><td>0.02 – 52.36</td><td>1.000</td></tr><tr><td>Population [Chriesbach]</td><td>1.00</td><td>0.02 – 52.36</td><td>1.000</td></tr><tr><td>Population [Clitunno]</td><td>1.65</td><td>0.02 – 87.21</td><td>0.807</td></tr><tr><td>Population [Marine]</td><td>75.29</td><td>4.12 – 1377.06</td><td>&lt; 0.001</td></tr><tr><td>Treatment [treated] x Population [Chessel]</td><td>1.00</td><td>0.00 – 269.80</td><td>1.000</td></tr><tr><td>Treatment [treated] x Population [Chriesbach]</td><td>1.00</td><td>0.00 – 269.80</td><td>1.000</td></tr><tr><td>Treatment [treated] x Population [Clitunno]</td><td>1.00</td><td>0.00 – 274.52</td><td>1.000</td></tr><tr><td>Treatment [treated] x Population [Marine]</td><td>0.18</td><td>0.00 – 11.45</td><td>0.438</td></tr><tr><td>Observations</td><td colspan="3">230</td></tr><tr><td>R<sup>2</sup></td><td colspan="3">0.156</td></tr></tbody></table>

<table><thead><tr><th>Contrast</th><th>Odds Ratio</th><th>SE</th><th>df</th><th>t ratio</th><th>p value</th></tr></thead><tbody><tr><td>Binnenkanal / Chessel</td><td>1.000</td><td>1.428</td><td>36</td><td>0.000</td><td>1.000</td></tr><tr><td>Binnenkanal / Chriesbach</td><td>1.000</td><td>1.428</td><td>36</td><td>0.000</td><td>1.000</td></tr><tr><td>Binnenkanal / Clitunno</td><td>0.608</td><td>0.871</td><td>36</td><td>-0.348</td><td>0.997</td></tr><tr><td>Binnenkanal / Marine</td><td>0.031</td><td>0.033</td><td>36</td><td>-3.282</td><td>0.018</td></tr><tr><td>Chessel / Chriesbach</td><td>1.000</td><td>1.428</td><td>36</td><td>0.000</td><td>1.000</td></tr><tr><td>Chessel / Clitunno</td><td>0.608</td><td>0.871</td><td>36</td><td>-0.348</td><td>0.997</td></tr><tr><td>Chessel / Marine</td><td>0.031</td><td>0.033</td><td>36</td><td>-3.282</td><td>0.018</td></tr><tr><td>Chriesbach / Clitunno</td><td>0.608</td><td>0.871</td><td>36</td><td>-0.348</td><td>0.997</td></tr><tr><td>Chriesbach / Marine</td><td>0.031</td><td>0.033</td><td>36</td><td>-3.282</td><td>0.018</td></tr><tr><td>Clitunno / Marine</td><td>0.051</td><td>0.054</td><td>36</td><td>-2.795</td><td>0.060</td></tr></tbody></table>
<table><caption>Table 3. GLMMs of weights of fish in experiment at time point 1 (10 weeks in experiment), time point 2 (16 weeks in experiment), and time point 3 (22 weeks in experiment) (Population effects are shown relative to Binnenkanal)</caption><thead><tr><th colspan="4">Time Point 1</th></tr><tr><th>Predictors</th><th>Estimates</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>0.37</td><td>0.32 – 0.43</td><td>&lt;0.001</td></tr><tr><td>Treatment [treated]</td><td>0.15</td><td>0.08 – 0.23</td><td>&lt;0.001</td></tr><tr><td>Population [Chessel]</td><td>0.09</td><td>0.01 – 0.16</td><td>0.021</td></tr><tr><td>Population [Chriesbach]</td><td>0.16</td><td>0.09 – 0.24</td><td>&lt;0.001</td></tr><tr><td>Population [Clitunno]</td><td>0.20</td><td>0.12 – 0.29</td><td>&lt;0.001</td></tr><tr><td>Population [Marine]</td><td>0.05</td><td>-0.03 – 0.13</td><td>0.204</td></tr><tr><td>Treatment [treated] x Population [Chessel]</td><td>-0.06</td><td>-0.16 – 0.05</td><td>0.281</td></tr><tr><td>Treatment [treated] x Population [Chriesbach]</td><td>-0.03</td><td>-0.14 – 0.07</td><td>0.536</td></tr><tr><td>Treatment [treated] x Population [Clitunno]</td><td>-0.10</td><td>-0.22 – 0.02</td><td>0.104</td></tr><tr><td>Treatment [treated] x Population [Marine]</td><td>0.05</td><td>-0.06 – 0.16</td><td>0.371</td></tr><tr><th colspan="4">Random Effects</th></tr><tr><td>σ<sup>2</sup> 0.01</td><td>τ<sub>100 Tank</sub> 0.00</td><td>ICC 0.07</td><td>N<sub>Tank</sub> 42</td></tr><tr><td>Observations</td><td>214</td><td></td><td></td></tr><tr><td>Marginal R<sup>2</sup></td><td>0.384</td><td>Conditional R<sup>2</sup></td><td>0.428</td></tr><tr><th colspan="4">Time Point 2</th></tr><tr><th>Predictors</th><th>Estimates</th><th>CI</th><th>p</th></tr><tr><td>(Intercept)</td><td>0.51</td><td>0.43 – 0.58</td><td>&lt;0.001</td></tr><tr><td>Treatment [treated]</td><td>0.16</td><td>0.06 – 0.25</td><td>0.001</td></tr><tr><td>Population [Chessel]</td><td>0.10</td><td>0.01 – 0.20</td><td>0.037</td></tr><tr><td>Population [Chriesbach]</td><td>0.11</td><td>0.01 – 0.21</td><td>0.028</td></tr><tr><td>Population [Clitunno]</td><td>0.20</td><td>0.09 – 0.31</td><td>0.001</td></tr></tbody></table>
<table><tr><td>Population [Marine]</td><td>0.13</td><td>0.00 – 0.25</td><td>0.046</td></tr><tr><td>Treatment [treated] x Population [Chessel]</td><td>-0.06</td><td>-0.19 – 0.07</td><td>0.380</td></tr><tr><td>Treatment [treated] x Population [Chriesbach]</td><td>-0.01</td><td>-0.14 – 0.13</td><td>0.938</td></tr><tr><td>Treatment [treated] x Population [Clitunno]</td><td>-0.13</td><td>-0.29 – 0.02</td><td>0.085</td></tr><tr><td>Treatment [treated] x Population [Marine]</td><td>0.08</td><td>-0.08 – 0.24</td><td>0.326</td></tr></table>

Random Effects

<table><tr><td>σ² 0.02</td><td>τ₁₀₀ Tank 0.00</td><td>ICC 0.03</td><td>N<sub>Tank</sub> 40</td></tr><tr><td>Observations</td><td>192</td><td></td><td></td></tr><tr><td>Marginal R²</td><td>0.273</td><td>Conditional R²</td><td>0.294</td></tr></table>

Time Point 3

<table><thead><tr><th>Predictors</th><th>Estimates</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>0.61</td><td>0.50-0.73</td><td>&lt;0.001</td></tr><tr><td>Treatment [treated]</td><td>0.26</td><td>0.10-0.41</td><td>0.001</td></tr><tr><td>Population [Chessel]</td><td>0.14</td><td>-0.01 – 0.30</td><td>0.070</td></tr><tr><td>Population [Chriesbach]</td><td>0.05</td><td>-0.11 – 0.22</td><td>0.530</td></tr><tr><td>Population [Clitunno]</td><td>0.18</td><td>0.00 – 0.36</td><td>0.051</td></tr><tr><td>Population [Marine]</td><td>0.04</td><td>-0.25 – 0.34</td><td>0.761</td></tr><tr><td>Treatment [treated] x Population [Chessel]</td><td>-0.07</td><td>-0.28 – 0.14</td><td>0.529</td></tr><tr><td>Treatment [treated] x Population [Chriesbach]</td><td>0.10</td><td>-0.13 – 0.33</td><td>0.406</td></tr><tr><td>Treatment [treated] x Population [Clitunno]</td><td>-0.02</td><td>-0.27 – 0.23</td><td>0.875</td></tr><tr><td>Treatment [treated] x Population [Marine]</td><td>0.05</td><td>-0.29 – 0.38</td><td>0.787</td></tr></tbody></table>

Random Effects
<table><tr><td>σ² 0.04</td><td>τ₁₀₀ Tank 0.01</td><td>ICC 0.16</td><td>N<sub>Tank</sub> 41</td></tr><tr><td>Observations</td><td>166</td><td></td><td></td></tr><tr><td>Marginal R²</td><td>0.321</td><td>Conditional R²</td><td>0.428</td></tr></table>

Time Point 1

<table><thead><tr><th>Contrast</th><th>Odds Ratio</th><th>SE</th><th>df</th><th>t ratio</th><th>p value</th></tr></thead><tbody><tr><td>Binnenkanal / Chessel</td><td>-0.059</td><td>0.027</td><td>27.181</td><td>-2.225</td><td>0.201</td></tr><tr><td>Binnenkanal / Chriesbach</td><td>-0.147</td><td>0.026</td><td>30.399</td><td>-5.558</td><td>0.000</td></tr><tr><td>Binnenkanal / Clitunno</td><td>-0.153</td><td>0.030</td><td>31.227</td><td>-5.043</td><td>0.000</td></tr><tr><td>Binnenkanal / Marine</td><td>-0.076</td><td>0.028</td><td>31.297</td><td>-2.676</td><td>0.081</td></tr><tr><td>Chessel / Chriesbach</td><td>-0.088</td><td>0.026</td><td>30.665</td><td>-3.324</td><td>0.018</td></tr><tr><td>Chessel / Clitunno</td><td>-0.094</td><td>0.030</td><td>31.436</td><td>-3.103</td><td>0.031</td></tr><tr><td>Chessel / Marine</td><td>-0.017</td><td>0.028</td><td>31.538</td><td>-0.592</td><td>0.975</td></tr><tr><td>Chriesbach / Clitunno</td><td>-0.007</td><td>0.030</td><td>34.410</td><td>-0.217</td><td>0.999</td></tr><tr><td>Chriesbach / Marine</td><td>0.071</td><td>0.028</td><td>35.025</td><td>2.517</td><td>0.110</td></tr><tr><td>Clitunno / Marine</td><td>0.077s</td><td>0.032</td><td>34.786</td><td>2.425</td><td>0.133</td></tr></tbody></table>

Time Point 2

<table><thead><tr><th>Contrast</th><th>Odds Ratio</th><th>SE</th><th>df</th><th>t ratio</th><th>p value</th></tr></thead><tbody><tr><td>Binnenkanal / Chessel</td><td>-0.072</td><td>0.035</td><td>22.091</td><td>-2.091</td><td>0.259</td></tr><tr><td>Binnenkanal / Chriesbach</td><td>-0.109</td><td>0.035</td><td>34.743</td><td>-3.118</td><td>0.028</td></tr><tr><td>Binnenkanal / Clitunno</td><td>-0.130</td><td>0.039</td><td>31.285</td><td>-3.356</td><td>0.017</td></tr><tr><td>Binnenkanal / Marine</td><td>-0.166</td><td>0.041</td><td>31.155</td><td>-4.087</td><td>0.002</td></tr><tr><td>Chessel / Chriesbach</td><td>-0.037</td><td>0.035</td><td>23.027</td><td>-1.050</td><td>0.830</td></tr><tr><td>Chessel / Clitunno</td><td>-0.058</td><td>0.039</td><td>22.612</td><td>-1.496</td><td>0.575</td></tr><tr><td>Chessel / Marine</td><td>-0.093</td><td>0.040</td><td>23.120</td><td>-2.308</td><td>0.178</td></tr><tr><td>Chriesbach / Clitunno</td><td>-0.021</td><td>0.039</td><td>32.305</td><td>-0.548</td><td>0.981</td></tr><tr><td>Chriesbach / Marine</td><td>-0.057</td><td>0.041</td><td>32.086</td><td>-1.394</td><td>0.636</td></tr><tr><td>Clitunno / Marine</td><td>-0.035</td><td>0.044</td><td>29.991</td><td>-0.803</td><td>0.928</td></tr></tbody></table>
<table><caption>Table 4. GLMs of condition of fish in experiment at time point 1 (10 weeks in experiment), time point 2 (16 weeks in experiment), and time point 3 (22 weeks in experiment) (Population effects are shown relative to Binnenkanal)</caption><thead><tr><th colspan="4">Time Point 1</th></tr><tr><th>Predictors</th><th>Estimates</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>0.12</td><td>0.11 – 0.14</td><td>&lt;0.001</td></tr><tr><td>Treatment [treated]</td><td>0.04</td><td>0.02 – 0.06</td><td>&lt;0.001</td></tr><tr><td>Population [Chessel]</td><td>0.01</td><td>-0.01 – 0.03</td><td>0.234</td></tr><tr><td>Population [Chriesbach]</td><td>0.03</td><td>0.01 – 0.05</td><td>0.005</td></tr><tr><td>Population [Clitunno]</td><td>0.04</td><td>0.02 – 0.06</td><td>0.001</td></tr><tr><td>Population [Marine]</td><td>0.01</td><td>-0.01 – 0.04</td><td>0.221</td></tr><tr><td>Treatment [treated] x Population [Chessel]</td><td>-0.01</td><td>-0.04 – 0.02</td><td>0.568</td></tr><tr><td>Treatment [treated] x Population [Chriesbach]</td><td>-0.02</td><td>-0.05 – 0.01</td><td>0.236</td></tr><tr><td>Treatment [treated] x Population [Clitunno]</td><td>-0.03</td><td>-0.06 – 0.00</td><td>0.078</td></tr><tr><td>Treatment [treated] x Population [Marine]</td><td>0.00</td><td>-0.03 – 0.03</td><td>0.814</td></tr><tr><td>Observations</td><td>43</td><td></td><td></td></tr><tr><td>Marginal R<sup>2</sup></td><td>0.626</td><td></td><td></td></tr><tr><th colspan="4">Time Point 2</th></tr><tr><th>Predictors</th><th>Estimates</th><th>CI</th><th>p</th></tr><tr><td>(Intercept)</td><td>0.15</td><td>0.13 – 0.16</td><td>&lt;0.001</td></tr><tr><td>Treatment [treated]</td><td>0.03</td><td>0.01 – 0.05</td><td>0.002</td></tr><tr><td>Population [Chessel]</td><td>0.02</td><td>0.00 – 0.04</td><td>0.098</td></tr><tr><td>Population [Chriesbach]</td><td>0.02</td><td>0.00 – 0.04</td><td>0.061</td></tr><tr><td>Population [Clitunno]</td><td>0.06</td><td>0.03 – 0.08</td><td>&lt;0.001</td></tr><tr><td>Population [Marine]</td><td>0.02</td><td>0.00 – 0.05</td><td>0.064</td></tr></tbody></table>
<table><tr><td>Treatment [treated] x Population [Chessel]</td><td>-0.01</td><td>-0.04 – 0.02</td><td>0.381</td></tr><tr><td>Treatment [treated] x Population [Chriesbach]</td><td>-0.01</td><td>-0.04 – 0.02</td><td>0.619</td></tr><tr><td>Treatment [treated] x Population [Clitunno]</td><td>-0.04</td><td>-0.07 – 0.00</td><td>0.025</td></tr><tr><td>Treatment [treated] x Population [Marine]</td><td>0.00</td><td>-0.04 – 0.03</td><td>0.821</td></tr><tr><td>Observations</td><td>43</td><td></td><td></td></tr><tr><td>R<sup>2</sup></td><td>0.591</td><td></td><td></td></tr></table>

Time Point 3

<table><thead><tr><th>Predictors</th><th>Estimates</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>0.16</td><td>0.14 – 0.18</td><td>&lt;0.001</td></tr><tr><td>Treatment [treated]</td><td>0.04</td><td>0.02 – 0.07</td><td>0.002</td></tr><tr><td>Population [Chessel]</td><td>0.02</td><td>-0.01 – 0.05</td><td>0.115</td></tr><tr><td>Population [Chriesbach]</td><td>0.01</td><td>-0.02 – 0.04</td><td>0.552</td></tr><tr><td>Population [Clitunno]</td><td>0.05</td><td>0.02 – 0.08</td><td>0.002</td></tr><tr><td>Population [Marine]</td><td>0.02</td><td>-0.02 – 0.05</td><td>0.350</td></tr><tr><td>Treatment [treated] x Population [Chessel]</td><td>-0.01</td><td>-0.05 – 0.03</td><td>0.513</td></tr><tr><td>Treatment [treated] x Population [Chriesbach]</td><td>0.02</td><td>-0.02 – 0.05</td><td>0.433</td></tr><tr><td>Treatment [treated] x Population [Clitunno]</td><td>0.00</td><td>-0.05 – 0.04</td><td>0.887</td></tr><tr><td>Treatment [treated] x Population [Marine]</td><td>-0.01</td><td>-0.06 – 0.04</td><td>0.662</td></tr><tr><td>Observations</td><td>42</td><td></td><td></td></tr><tr><td>R<sup>2</sup></td><td>0.651</td><td></td><td></td></tr></tbody></table>
Time Point 1

<table><thead><tr><th>Contrast</th><th>Odds Ratio</th><th>SE</th><th>df</th><th>t ratio</th><th>p value</th></tr></thead><tbody><tr><td>Binnenkanal / Chessel</td><td>-0.008</td><td>0.007</td><td>33</td><td>-1.162</td><td>0.772</td></tr><tr><td>Binnenkanal / Chriesbach</td><td>-0.021</td><td>0.007</td><td>33</td><td>-2.928</td><td>0.045</td></tr><tr><td>Binnenkanal / Clitunno</td><td>-0.026</td><td>0.008</td><td>33</td><td>-3.125</td><td>0.028</td></tr><tr><td>Binnenkanal / Marine</td><td>-0.012</td><td>0.008</td><td>33</td><td>-1.539</td><td>0.546</td></tr><tr><td>Chessel / Chriesbach</td><td>-0.013</td><td>0.007</td><td>33</td><td>-1.820</td><td>0.380</td></tr><tr><td>Chessel / Clitunno</td><td>-0.018</td><td>0.008</td><td>33</td><td>-2.160</td><td>0.220</td></tr><tr><td>Chessel / Marine</td><td>-0.003</td><td>0.008</td><td>33</td><td>-0.452</td><td>0.991</td></tr><tr><td>Chriesbach / Clitunno</td><td>-0.005</td><td>0.008</td><td>33</td><td>-0.584</td><td>0.997</td></tr><tr><td>Chriesbach / Marine</td><td>0.010</td><td>0.008</td><td>33</td><td>1.264</td><td>0.714</td></tr><tr><td>Clitunno / Marine</td><td>0.014</td><td>0.009</td><td>33</td><td>1.669</td><td>0.466</td></tr></tbody></table>

Time Point 2

<table><thead><tr><th>Contrast</th><th>Odds Ratio</th><th>SE</th><th>df</th><th>t ratio</th><th>p value</th></tr></thead><tbody><tr><td>Binnenkanal / Chessel (control)</td><td>-0.017</td><td>0.010</td><td>33</td><td>-1.656</td><td>0.474</td></tr><tr><td>Binnenkanal / Chriesbach (control)</td><td>-0.019</td><td>0.010</td><td>33</td><td>-1.871</td><td>0.353</td></tr><tr><td>Binnenkanal / Clitunno (control)</td><td>-0.058</td><td>0.012</td><td>33</td><td>-4.877</td><td>0.000</td></tr><tr><td>Binnenkanal / Marine (control)</td><td>-0.022</td><td>0.012</td><td>33</td><td>-1.850</td><td>0.363</td></tr><tr><td>Chessel / Chriesbach (control)</td><td>-0.002</td><td>0.010</td><td>33</td><td>-0.214</td><td>1.000</td></tr><tr><td>Chessel / Clitunno (control)</td><td>-0.041</td><td>0.012</td><td>33</td><td>-3.442</td><td>0.013</td></tr><tr><td>Chessel / Marine (control)</td><td>-0.005</td><td>0.012</td><td>33</td><td>-0.416</td><td>0.993</td></tr><tr><td>Chriesbach / Clitunno (control)</td><td>-0.039</td><td>0.012</td><td>33</td><td>-3.257</td><td>0.021</td></tr><tr><td>Chriesbach / Marine (control)</td><td>-0.003</td><td>0.012</td><td>33</td><td>-0.230</td><td>0.999</td></tr></tbody></table>
<table><tbody><tr><td>(control)</td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Clitunno / Marine<br>(control)</td><td>0.036</td><td>0.013</td><td>33</td><td>2.707</td><td>0.074</td></tr><tr><td>Binnenkanal / Chessel<br>(treated)</td><td>-0.004</td><td>0.010</td><td>33</td><td>-0.416</td><td>0.993</td></tr><tr><td>Binnenkanal /<br>Chriesbach<br>(treated)</td><td>-0.012</td><td>0.010</td><td>33</td><td>-1.168</td><td>0.769</td></tr><tr><td>Binnenkanal / Clitunno<br>(treated)</td><td>-0.020</td><td>0.012</td><td>33</td><td>-1.707</td><td>0.443</td></tr><tr><td>Binnenkanal / Marine<br>(treated)</td><td>-0.018</td><td>0.011</td><td>33</td><td>-1.681</td><td>0.459</td></tr><tr><td>Chessel / Chriesbach<br>(treated)</td><td>-0.008</td><td>0.010</td><td>33</td><td>-0.752</td><td>0.942</td></tr><tr><td>Chessel / Clitunno<br>(treated)</td><td>-0.016</td><td>0.012</td><td>33</td><td>-1.347</td><td>0.664</td></tr><tr><td>Chessel / Marine<br>(treated)</td><td>-0.014</td><td>0.011</td><td>33</td><td>-1.288</td><td>0.700</td></tr><tr><td>Chriesbach / Clitunno<br>(treated)</td><td>-0.008</td><td>0.012</td><td>33</td><td>-0.696</td><td>0.956</td></tr><tr><td>Chriesbach / Marine<br>(treated)</td><td>-0.006</td><td>0.011</td><td>33</td><td>-0.579</td><td>0.977</td></tr><tr><td>Clitunno / Marine<br>(treated)</td><td>0.002</td><td>0.012</td><td>33</td><td>0.157</td><td>1.000</td></tr></tbody></table>

Time Point 3

<table><thead><tr><th>Contrast</th><th>Odds Ratio</th><th>SE</th><th>df</th><th>t ratio</th><th>p value</th></tr></thead><tbody><tr><td>Binnenkanal / Chessel</td><td>-0.015</td><td>0.010</td><td>32</td><td>-1.574</td><td>0.524</td></tr><tr><td>Binnenkanal /<br>Chriesbach</td><td>-0.016</td><td>0.010</td><td>32</td><td>-1.625</td><td>0.493</td></tr><tr><td>Binnenkanal / Clitunno</td><td>-0.048</td><td>0.011</td><td>32</td><td>-4.252</td><td>0.002</td></tr><tr><td>Binnenkanal / Marine</td><td>-0.012</td><td>0.012</td><td>32</td><td>-1.022</td><td>0.843</td></tr><tr><td>Chessel / Chriesbach</td><td>0.000</td><td>0.010</td><td>32</td><td>-0.051</td><td>1.000</td></tr><tr><td>Chessel / Clitunno</td><td>-0.032</td><td>0.011</td><td>32</td><td>-2.889</td><td>0.050</td></tr></tbody></table>
<table><tr><td>Chessel / Marine</td><td>0.003</td><td>0.012</td><td>32</td><td>0.290</td><td>0.998</td></tr><tr><td>Chriesbach / Clitunno</td><td>-0.032</td><td>0.011</td><td>32</td><td>-2.845</td><td>0.055</td></tr><tr><td>Chriesbach / Marine</td><td>0.004</td><td>0.012</td><td>32</td><td>0.333</td><td>0.997</td></tr><tr><td>Clitunno / Marine</td><td>0.036</td><td>0.013</td><td>32</td><td>2.768</td><td>0.066</td></tr></table>
<table><caption>Table 5. GLMMs of length of fish in experiment at time point 1 (10 weeks in experiment), time point 2 (16 weeks in experiment), and time point 3 (22 weeks in experiment) (Population effects are shown relative to Binnenkanal)</caption><thead><tr><th colspan="4">Time Point 1</th></tr><tr><th>Predictors</th><th>Estimates</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>3.25</td><td>3.11 – 3.39</td><td>&lt;0.001</td></tr><tr><td>Treatment [treated]</td><td>-0.13</td><td>-0.33 – 0.08</td><td>0.230</td></tr><tr><td>Population [Chessel]</td><td>0.17</td><td>-0.03 – 0.36</td><td>0.089</td></tr><tr><td>Population [Chriesbach]</td><td>0.30</td><td>0.10 – 0.49</td><td>0.003</td></tr><tr><td>Population [Clitunno]</td><td>0.28</td><td>0.05 – 0.51</td><td>0.018</td></tr><tr><td>Population [Marine]</td><td>0.21</td><td>0.00 – 0.41</td><td>0.050</td></tr><tr><td>Treatment [treated] x Population [Chessel]</td><td>0.04</td><td>-0.24 – 0.33</td><td>0.763</td></tr><tr><td>Treatment [treated] x Population [Chriesbach]</td><td>0.32</td><td>0.04 – 0.61</td><td>0.026</td></tr><tr><td>Treatment [treated] x Population [Clitunno]</td><td>0.21</td><td>-0.12 – 0.53</td><td>0.218</td></tr><tr><td>Treatment [treated] x Population [Marine]</td><td>0.39</td><td>0.09 – 0.69</td><td>0.012</td></tr><tr><th colspan="4">Random Effects</th></tr><tr><td>σ<sup>2</sup> 0.07</td><td>τ<sub>100 Tank</sub> 0.01</td><td>ICC 0.15</td><td>N<sub>Tank</sub> 43</td></tr><tr><td>Observations</td><td>208</td><td></td><td></td></tr><tr><td>Marginal R<sup>2</sup></td><td>0.322</td><td>Conditional R<sup>2</sup></td><td>0.421</td></tr><tr><th colspan="4">Time Point 2</th></tr><tr><th>Predictors</th><th>Estimates</th><th>CI</th><th>p</th></tr><tr><td>(Intercept)</td><td>3.37</td><td>3.24 – 3.50</td><td>&lt;0.001</td></tr><tr><td>Treatment [treated]</td><td>0.35</td><td>0.18 – 0.52</td><td>&lt;0.001</td></tr><tr><td>Population [Chessel]</td><td>0.32</td><td>0.15 – 0.49</td><td>&lt;0.001</td></tr><tr><td>Population [Chriesbach]</td><td>0.32</td><td>0.14 – 0.49</td><td>0.001</td></tr><tr><td>Population [Clitunno]</td><td>0.10</td><td>-0.10 – 0.30</td><td>0.319</td></tr></tbody></table>
<table><tr><td>Population [Marine]</td><td>0.55</td><td>0.33 – 0.77</td><td>&lt;0.001</td></tr><tr><td>Treatment [treated] x Population [Chessel]</td><td>-0.14</td><td>-0.37 – 0.10</td><td>0.245</td></tr><tr><td>Treatment [treated] x Population [Chriesbach]</td><td>0.00</td><td>-0.24 – 0.25</td><td>0.993</td></tr><tr><td>Treatment [treated] x Population [Clitunno]</td><td>-0.16</td><td>-0.44 – 0.11</td><td>0.248</td></tr><tr><td>Treatment [treated] x Population [Marine]</td><td>0.12</td><td>-0.16 – 0.40</td><td>0.405</td></tr></table>

Random Effects

<table><tr><td>σ² 0.08</td><td>τ₁₀₀ Tank 0.00</td><td>ICC 0.02</td><td>N<sub>Tank</sub> 42</td></tr><tr><td>Observations</td><td>191</td><td></td><td></td></tr><tr><td>Marginal R²</td><td>0.481</td><td>Conditional R²</td><td>0.490</td></tr></table>

Time Point 3

<table><thead><tr><th>Predictors</th><th>Estimates</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>3.70</td><td>3.52 – 3.88</td><td>&lt;0.001</td></tr><tr><td>Treatment [treated]</td><td>0.47</td><td>0.23 – 0.71</td><td>&lt;0.001</td></tr><tr><td>Population [Chessel]</td><td>0.33</td><td>0.09 – 0.57</td><td>0.008</td></tr><tr><td>Population [Chriesbach]</td><td>0.11</td><td>-0.15 – 0.36</td><td>0.414</td></tr><tr><td>Population [Clitunno]</td><td>0.03</td><td>-0.27 – 0.32</td><td>0.867</td></tr><tr><td>Population [Marine]</td><td>0.46</td><td>0.00 – 0.91</td><td>0.049</td></tr><tr><td>Treatment [treated] x Population [Chessel]</td><td>-0.14</td><td>-0.48 – 0.19</td><td>0.398</td></tr><tr><td>Treatment [treated] x Population [Chriesbach]</td><td>0.01</td><td>-0.35 – 0.37</td><td>0.959</td></tr><tr><td>Treatment [treated] x Population [Clitunno]</td><td>-0.14</td><td>-0.53 – 0.26</td><td>0.504</td></tr><tr><td>Treatment [treated] x Population [Marine]</td><td>-0.13</td><td>-0.66 – 0.39</td><td>0.617</td></tr></tbody></table>

Random Effects
<table><tr><td>σ² 0.08</td><td>τ₁₀₀ Tank 0.02</td><td>ICC 0.18</td><td>N<sub>Tank</sub> 41</td></tr><tr><td>Observations</td><td>163</td><td></td><td></td></tr><tr><td>Marginal R²</td><td>0.388</td><td>Conditional R²</td><td>0.501</td></tr></table>

<table><caption>Time Point 1</caption><thead><tr><th>Contrast</th><th>Odds Ratio</th><th>SE</th><th>df</th><th>t ratio</th><th>p value</th></tr></thead><tbody><tr><td>Binnenkanal / Chessel (control)</td><td>-0.168</td><td>0.098</td><td>31.712</td><td>-1.709</td><td>0.443</td></tr><tr><td>Binnenkanal / Chriesbach (control)</td><td>-0.297</td><td>0.099</td><td>32.414</td><td>-3.004</td><td>0.038</td></tr><tr><td>Binnenkanal / Clitunno (control)</td><td>-0.279</td><td>0.117</td><td>34.840</td><td>-2.389</td><td>0.142</td></tr><tr><td>Binnenkanal / Marine (control)</td><td>-0.205</td><td>0.104</td><td>31.712</td><td>-1.969</td><td>0.304</td></tr><tr><td>Chessel / Chriesbach (control)</td><td>-0.129</td><td>0.099</td><td>32.414</td><td>-1.305</td><td>0.690</td></tr><tr><td>Chessel / Clitunno (control)</td><td>-0.111</td><td>0.117</td><td>34.840</td><td>-0.953</td><td>0.874</td></tr><tr><td>Chessel / Marine (control)</td><td>-0.037</td><td>0.104</td><td>31.712</td><td>-0.358</td><td>0.996</td></tr><tr><td>Chriesbach / Clitunno (control)</td><td>0.018</td><td>0.117</td><td>35.376</td><td>0.150</td><td>1.000</td></tr><tr><td>Chriesbach / Marine (control)</td><td>0.092</td><td>0.105</td><td>32.336</td><td>0.875</td><td>0.904</td></tr><tr><td>Clitunno / Marine (control)</td><td>0.074</td><td>0.122</td><td>34.570</td><td>0.608</td><td>0.973</td></tr><tr><td>Binnenkanal / Chessel (treated)</td><td>-0.211</td><td>0.105</td><td>32.336</td><td>-2.016</td><td>0.281</td></tr><tr><td>Binnenkanal / Chriesbach (treated)</td><td>-0.620</td><td>0.105</td><td>32.336</td><td>-5.913</td><td>0.000</td></tr><tr><td>Binnenkanal / Clitunno</td><td>-0.485</td><td>0.119</td><td>31.712</td><td>-4.090</td><td>0.002</td></tr></tbody></table>
<table><tbody><tr><td>(treated)</td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Binnenkanal / Marine (treated)</td><td>-0.594</td><td>0.112</td><td>33.494</td><td>-5.309</td><td>0.000</td></tr><tr><td>Chessel / Chriesbach (treated)</td><td>-0.408</td><td>0.099</td><td>33.121</td><td>-4.105</td><td>0.002</td></tr><tr><td>Chessel / Clitunno (treated)</td><td>-0.274</td><td>0.114</td><td>32.238</td><td>-2.404</td><td>0.140</td></tr><tr><td>Chessel / Marine (treated)</td><td>-0.382</td><td>0.107</td><td>34.324</td><td>-3.587</td><td>0.009</td></tr><tr><td>Chriesbach / Clitunno (treated)</td><td>0.134</td><td>0.114</td><td>32.238</td><td>1.178</td><td>0.764</td></tr><tr><td>Chriesbach / Marine (treated)</td><td>0.026</td><td>0.107</td><td>34.324</td><td>0.243</td><td>0.999</td></tr><tr><td>Clitunno / Marine (treated)</td><td>-0.108</td><td>0.121</td><td>33.239</td><td>-0.899</td><td>0.895</td></tr></tbody></table>

## Time Point 2

<table><thead><tr><th>Contrast</th><th>Odds Ratio</th><th>SE</th><th>df</th><th>t ratio</th><th>p value</th></tr></thead><tbody><tr><td>Binnenkanal / Chessel</td><td>-0.250</td><td>0.060</td><td>29.417</td><td>-4.200</td><td>0.002</td></tr><tr><td>Binnenkanal / Chriesbach</td><td>-0.317</td><td>0.062</td><td>33.250</td><td>-5.105</td><td>0.000</td></tr><tr><td>Binnenkanal / Clitunno</td><td>-0.019</td><td>0.070</td><td>30.807</td><td>-0.275</td><td>0.999</td></tr><tr><td>Binnenkanal / Marine</td><td>-0.612</td><td>0.072</td><td>29.632</td><td>-8.506</td><td>0.000</td></tr><tr><td>Chessel / Chriesbach</td><td>-0.067</td><td>0.060</td><td>30.820</td><td>-1.108</td><td>0.801</td></tr><tr><td>Chessel / Clitunno</td><td>0.231</td><td>0.068</td><td>28.965</td><td>3.405</td><td>0.015</td></tr><tr><td>Chessel / Marine</td><td>-0.362</td><td>0.070</td><td>27.953</td><td>-5.150</td><td>0.000</td></tr><tr><td>Chriesbach / Clitunno</td><td>0.298</td><td>0.070</td><td>31.880</td><td>4.250</td><td>0.002</td></tr><tr><td>Chriesbach / Marine</td><td>-0.296</td><td>0.072</td><td>30.597</td><td>-4.079</td><td>0.003</td></tr><tr><td>Clitunno / Marine</td><td>0.593</td><td>0.079</td><td>29.258</td><td>-7.511</td><td>0.000</td></tr></tbody></table>

## Time Point 3

<table><thead><tr><th>Contrast</th><th>Odds Ratio</th><th>SE</th><th>df</th><th>t ratio</th><th>p value</th></tr></thead><tbody><tr><td>Binnenkanal / Chessel</td><td>-0.256</td><td>0.085</td><td>25.644</td><td>-3.014</td><td>0.042</td></tr><tr><td>Binnenkanal / Chriesbach</td><td>-0.111</td><td>0.091</td><td>32.472</td><td>-1.217</td><td>0.742</td></tr></tbody></table>
<table><tr><td>Binnenkanal / Clitunno</td><td>0.043</td><td>0.101</td><td>29.458</td><td>0.412</td><td>0.993</td></tr><tr><td>Binnenkanal / Marine</td><td>-0.389</td><td>0.134</td><td>37.286</td><td>-2.912</td><td>0.045</td></tr><tr><td>Chessel / Chriesbach</td><td>0.144</td><td>0.089</td><td>30.821</td><td>1.620</td><td>0.497</td></tr><tr><td>Chessel / Clitunno</td><td>0.298</td><td>0.099</td><td>28.175</td><td>3.010</td><td>0.040</td></tr><tr><td>Chessel / Marine</td><td>-0.133</td><td>0.132</td><td>36.481</td><td>-1.009</td><td>0.850</td></tr><tr><td>Chriesbach / Clitunno</td><td>0.154</td><td>0.105</td><td>33.507</td><td>1.468</td><td>0.590</td></tr><tr><td>Chriesbach / Marine</td><td>-0.277</td><td>0.136</td><td>40.069</td><td>-2.036</td><td>0.268</td></tr><tr><td>Clitunno / Marine</td><td>-0.431</td><td>0.143</td><td>37.259</td><td>-3.016</td><td>0.035</td></tr></table>
<table><caption>**Table 6.** GLMM of DHA content of fish in experiment at end of experiment (Population effects are shown relative to Binnenkanal)</caption><thead><tr><th colspan="4">Time Point 1</th></tr><tr><th>Predictors</th><th>Estimates</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>3.201</td><td>2.87 – 3.18</td><td>&lt;0.001</td></tr><tr><td>Treatment [treated]</td><td>0.05</td><td>-0.18 – 0.28</td><td>0.648</td></tr><tr><td>Population [Chessel]</td><td>-0.16</td><td>-0.37 – 0.04</td><td>0.122</td></tr><tr><td>Population [Chriesbach]</td><td>0.06</td><td>-0.16 – 0.27</td><td>0.599</td></tr><tr><td>Population [Clitunno]</td><td>0.34</td><td>0.14 – 0.54</td><td>0.001</td></tr><tr><td>Population [Marine]</td><td>-0.66</td><td>-0.91 – -0.42</td><td>&lt;0.001</td></tr><tr><td>Population [Reveillon]</td><td>0.10</td><td>-0.15 – 0.36</td><td>0.415</td></tr><tr><td>Sex [m]</td><td>0.09</td><td>-0.13 – 0.32</td><td>0.411</td></tr><tr><td>Treatment [treated] x Population [Chessel]</td><td>0.13</td><td>-0.18 – 0.44</td><td>0.415</td></tr><tr><td>Treatment [treated] x Population [Chriesbach]</td><td>-0.13</td><td>-0.50 – 0.23</td><td>0.466</td></tr><tr><td>Treatment [treated] x Population [Clitunno]</td><td>0.13</td><td>-0.18 – 0.44</td><td>0.407</td></tr><tr><td>Treatment [treated] x Population [Marine]</td><td>0.46</td><td>0.07 – 0.85</td><td>0.020</td></tr><tr><td>Treatment [treated] x Population [Reveillon]</td><td>-0.01</td><td>-0.40 – 0.38</td><td>0.953</td></tr><tr><td>Treatment [treated] x Sex [m]</td><td>-0.05</td><td>-0.36 – 0.27</td><td>0.756</td></tr><tr><td>Population [Chessel] x Sex [m]</td><td>-0.03</td><td>-0.35 – 0.29</td><td>0.848</td></tr><tr><td>Population [Chriesbach] x Sex [m]</td><td>-0.26</td><td>-0.58 – 0.06</td><td>0.108</td></tr><tr><td>Population [Clitunno] x Sex [m]</td><td>-0.07</td><td>-0.43 – 0.29</td><td>0.700</td></tr></tbody></table>
<table><tbody><tr><td>Population [Marine] x<br>Sex [m]</td><td>0.32</td><td>-0.02 – 0.67</td><td>0.063</td></tr><tr><td>Population [Reveillon]<br>x Sex [m]</td><td>0.24</td><td>-0.23 – 0.71</td><td>0.310</td></tr><tr><td>Treatment [treated] x<br>Population [Chessel] x<br>Sex [m]</td><td>0.00</td><td>-0.43 – 0.44</td><td>0.983</td></tr><tr><td>Treatment [treated] x<br>Population<br>[Chriesbach] x Sex [m]</td><td>0.10</td><td>-0.37 – 0.58</td><td>0.674</td></tr><tr><td>Treatment [treated] x<br>Population [Marine] x<br>Sex [m]</td><td>-0.29</td><td>-0.80 – 0.22</td><td>0.254</td></tr><tr><td colspan="4"><strong>Random Effects</strong></td></tr><tr><td>σ<sup>2</sup> 0.03</td><td>τ<sub>100 Tank</sub> 0.00</td><td>ICC 0.03</td><td>N<sub>Tank</sub> 44</td></tr><tr><td>Observations</td><td>89</td><td></td><td></td></tr><tr><td>Marginal R<sup>2</sup></td><td>0.662</td><td>Conditional R<sup>2</sup></td><td>0.672</td></tr></tbody></table>
<figure><img src="image_8.png" /><figcaption>**Fig. S1.** Mean and standard deviation of chironomid fatty acid content. Smaller points represent individual chironomid feed replicates at the monthly level (n=7 per treatment). Control chironomids were the standard husbandry fed in our facility and treated chironomids were first dipped in a marine oil treatment.</figcaption></figure>
<table><caption>Table S1. Study site information</caption><thead><tr><td>Stream Name:</td><td>Latitude, Longitude</td><td>Town, Country</td><td>Dominant Lineage(s)</td><td>Data from Site (number of individuals per dataset)</td></tr></thead><tbody><tr><td>Chriesbach, Switzerland</td><td>47.404491, 8.609296</td><td>Dübendorf, Switzerland</td><td>Baltic</td><td>fads2 (41), fish in experiment (5 crosses, 50 fish total, 18 for fatty acid analysis [FA])</td></tr><tr><td>Réveillon, France</td><td>48.720650, 2.585725</td><td>Villecresnes, France</td><td>NW Europe</td><td>fads2 (4), fish in experiment (1.5 crosses, 10 fish total, 6 for FA)</td></tr><tr><td>Binnenkanal, Switzerland</td><td>47.590724, 8.223962</td><td>Leuggern, Switzerland</td><td>Rhein</td><td>fads2 (40), fish in experiment (5 crosses, 50 fish total, 20 for FA)</td></tr><tr><td>Grand Canal Chessel, Switzerland</td><td>46.347484, 6.910664</td><td>Chessel, Switzerland</td><td>Rhône, Mediterranean</td><td>fads2 (47), fish in experiment (8 crosses, 50 fish total, 20 for FA)</td></tr><tr><td>Clitunno, Italy</td><td>42.833630, 12.764389</td><td>Campello sul Clitunno, Italy</td><td>Tyrrhenian Basin, Mediterranean</td><td>Fads2 (30), fish in experiment (2 crosses, 30 fish total, 14 for FA)</td></tr><tr><td>Puget Sound Marine, USA</td><td>47.571300, - 122.546800</td><td>Clam Bay, Puget Sound, Washington, USA</td><td>Pacific Northwest Marine</td><td>fish in experiment (9 crosses, 50 fish total, 12 for FA)</td></tr></tbody></table>
<table><caption>**Table S2. GLM of initial length of fish before start of experiment (Population effects are shown relative to Chriesbach)**</caption><thead><tr><th rowspan="2">Predictors</th><th colspan="3">Standard_Length Initial</th></tr><tr><th>Estimates</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>2.64</td><td>2.49 - 2.79</td><td><strong>&lt;0.001</strong></td></tr><tr><td>Population [Chessel]</td><td>-0.27</td><td>-0.48 - -0.06</td><td><strong>0.013</strong></td></tr><tr><td>Population [Marine]</td><td>0.20</td><td>-0.01 - 0.41</td><td>0.066</td></tr><tr><td>Population [Binnenkanal]</td><td>-0.17</td><td>-0.38 - -0.04</td><td>0.118</td></tr><tr><td>Observations</td><td colspan="3">40</td></tr><tr><td>R<sup>2</sup></td><td colspan="3">0.374</td></tr></tbody></table>

<table><caption>**Table S3. GLM of initial weight of fish before start of experiment (Population effects are shown relative to Chriesbach)**</caption><thead><tr><th rowspan="2">Predictors</th><th colspan="3">Wet_Weight Initial</th></tr><tr><th>Estimates</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>0.23</td><td>0.19 - 0.27</td><td><strong>&lt;0.001</strong></td></tr><tr><td>Population [Chessel]</td><td>-0.06</td><td>-0.12 - -0.01</td><td><strong>0.028</strong></td></tr><tr><td>Population [Marine]</td><td>0.02</td><td>-0.04 - 0.07</td><td>0.480</td></tr><tr><td>Population [Binnenkanal]</td><td>-0.02</td><td>-0.08 - -0.03</td><td>0.414</td></tr><tr><td>Observations</td><td colspan="3">40</td></tr><tr><td>R<sup>2</sup></td><td colspan="3">0.205</td></tr></tbody></table>
<table><caption>**Table S4. GLM of initial condition of fish before start of experiment (Population effects are shown relative to Chriesbach)**</caption><thead><tr><th rowspan="2">Predictors</th><th rowspan="2">Estimates</th><th colspan="2">Condition Initial</th></tr><tr><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>0.09</td><td>0.08 - 0.10</td><td><strong>&lt;0.001</strong></td></tr><tr><td>Population [Chessel]</td><td>-0.02</td><td>-0.03 - -0.00</td><td><strong>0.039</strong></td></tr><tr><td>Population [Marine]</td><td>0.00</td><td>-0.01 - 0.02</td><td>0.898</td></tr><tr><td>Population [Binnenkanal]</td><td>-0.00</td><td>-0.02 - 0.01</td><td>0.649</td></tr><tr><td>Observations</td><td>40</td><td></td><td></td></tr><tr><td>R<sup>2</sup></td><td>0.145</td><td></td><td></td></tr></tbody></table>
<table><caption>Table S5. Post-hoc tests of GLMM of DHA content of fish in experiment at end of experiment (NA represent population and/or treatment combinations where only one sex was present)</caption><caption>Pairwise Population Contrasts Within Treatments - Female DHA</caption><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal - Chessel</td><td>0.099</td><td>0.079</td><td>55.438</td><td>1.266</td><td>0.802</td></tr><tr><td>Binnenkanal - Chriesbach</td><td>0.010</td><td>0.094</td><td>60.444</td><td>0.109</td><td>1.000</td></tr><tr><td>Binnenkanal - Clitunno</td><td>-0.408</td><td>0.078</td><td>37.676</td><td>-5.206</td><td>0.000</td></tr><tr><td>Binnenkanal - Marine</td><td>0.434</td><td>0.098</td><td>62.830</td><td>4.419</td><td>0.001</td></tr><tr><td>Binnenkanal - Reveillon</td><td>-0.098</td><td>0.099</td><td>27.609</td><td>-0.996</td><td>0.915</td></tr><tr><td>Chessel - Chriesbach</td><td>-0.089</td><td>0.090</td><td>60.112</td><td>-0.993</td><td>0.918</td></tr><tr><td>Chessel - Clitunno</td><td>-0.507</td><td>0.074</td><td>34.885</td><td>-6.881</td><td>0.000</td></tr><tr><td>Chessel - Marine</td><td>0.334</td><td>0.095</td><td>62.733</td><td>3.537</td><td>0.010</td></tr><tr><td>Chessel - Reveillon</td><td>-0.198</td><td>0.095</td><td>25.871</td><td>-2.078</td><td>0.329</td></tr><tr><td>Chriesbach - Clitunno</td><td>-0.418</td><td>0.090</td><td>46.568</td><td>-4.662</td><td>0.000</td></tr><tr><td>Chriesbach - Marine</td><td>0.424</td><td>0.107</td><td>64.239</td><td>3.944</td><td>0.003</td></tr><tr><td>Chriesbach - Reveillon</td><td>-0.109</td><td>0.108</td><td>33.607</td><td>-1.006</td><td>0.913</td></tr><tr><td>Clitunno - Marine</td><td>0.841</td><td>0.094</td><td>51.526</td><td>8.918</td><td>0.000</td></tr><tr><td>Clitunno - Reveillon</td><td>0.309</td><td>0.095</td><td>19.658</td><td>3.254</td><td>0.040</td></tr><tr><td>Marine - Reveillon</td><td>-0.532</td><td>0.112</td><td>37.348</td><td>-4.754</td><td>0.000</td></tr></tbody></table>
Table S5 continued

<table><caption>Pairwise Population Contrasts Within Treatments - Male DHA</caption><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal - Chessel</td><td>0.127</td><td>0.080</td><td>58.509</td><td>1.584</td><td>0.396</td></tr><tr><td>Binnenkanal - Chriesbach</td><td>0.223</td><td>0.078</td><td>49.412</td><td>2.842</td><td>0.032</td></tr><tr><td>Binnenkanal - Clitunno</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Binnenkanal - Marine</td><td>0.256</td><td>0.086</td><td>63.558</td><td>2.990</td><td>0.020</td></tr><tr><td>Binnenkanal - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel - Chriesbach</td><td>0.095</td><td>0.080</td><td>46.654</td><td>1.187</td><td>0.638</td></tr><tr><td>Chessel - Clitunno</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel - Marine</td><td>0.129</td><td>0.087</td><td>61.547</td><td>1.475</td><td>0.458</td></tr><tr><td>Chessel - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach - Clitunno</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach - Marine</td><td>0.034</td><td>0.085</td><td>54.763</td><td>0.396</td><td>0.979</td></tr><tr><td>Chriesbach - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno - Marine</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Marine - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr></tbody></table>
Table S5 continued

Pairwise Population Contrasts Within Treatments - Female DHA

<table><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal control - Chessel control</td><td>0.163</td><td>0.105</td><td>54.268</td><td>1.559</td><td>0.916</td></tr><tr><td>Binnenkanal control - Chriesbach control</td><td>-0.057</td><td>0.109</td><td>58.727</td><td>-0.524</td><td>1.000</td></tr><tr><td>Binnenkanal control - Clitunno control</td><td>-0.343</td><td>0.101</td><td>41.324</td><td>-3.392</td><td>0.059</td></tr><tr><td>Binnenkanal control - Marine control</td><td>0.665</td><td>0.125</td><td>65.113</td><td>5.298</td><td>0.000</td></tr><tr><td>Binnenkanal control - Reveillon control</td><td>-0.104</td><td>0.127</td><td>21.676</td><td>-0.821</td><td>0.999</td></tr><tr><td>Binnenkanal control - Binnenkanal treated</td><td>0.053</td><td>0.117</td><td>56.375</td><td>-0.451</td><td>1.000</td></tr><tr><td>Binnenkanal control - Chessel treated</td><td>-0.017</td><td>0.108</td><td>64.904</td><td>-0.157</td><td>1.000</td></tr><tr><td>Binnenkanal control - Chriesbach treated</td><td>0.025</td><td>0.145</td><td>65.536</td><td>0.171</td><td>1.000</td></tr><tr><td>Binnenkanal control - Clitunno treated</td><td>-0.525</td><td>0.111</td><td>42.006</td><td>-4.745</td><td>0.001</td></tr><tr><td>Binnenkanal control - Marine treated</td><td>0.150</td><td>0.144</td><td>65.218</td><td>1.041</td><td>0.996</td></tr><tr><td>Binnenkanal control - Reveillon treated</td><td>-0.145</td><td>0.144</td><td>36.496</td><td>-1.007</td><td>0.997</td></tr><tr><td>Chessel control - Chriesbach control</td><td>-0.220</td><td>0.106</td><td>45.219</td><td>-2.084</td><td>0.636</td></tr><tr><td>Chessel control - Clitunno control</td><td>-0.506</td><td>0.097</td><td>27.468</td><td>-5.198</td><td>0.001</td></tr><tr><td>Chessel control - Marine control</td><td>0.502</td><td>0.122</td><td>58.011</td><td>4.099</td><td>0.007</td></tr><tr><td>Chessel control - Reveillon control</td><td>-0.267</td><td>0.124</td><td>16.709</td><td>-2.155</td><td>0.597</td></tr><tr><td>Chessel control - Binnenkanal treated</td><td>-0.216</td><td>0.114</td><td>44.188</td><td>-1.895</td><td>0.756</td></tr><tr><td>Chessel control - Chessel treated</td><td>-0.180</td><td>0.105</td><td>54.268</td><td>-1.721</td><td>0.850</td></tr><tr><td>Chessel control - Chriesbach treated</td><td>-0.138</td><td>0.143</td><td>60.904</td><td>-0.969</td><td>0.998</td></tr><tr><td>Chessel control - Clitunno treated</td><td>-0.687</td><td>0.107</td><td>29.778</td><td>-6.420</td><td>0.000</td></tr><tr><td>Chessel control - Marine treated</td><td>-0.013</td><td>0.141</td><td>60.316</td><td>-0.091</td><td>1.000</td></tr><tr><td>Chessel control - Reveillon treated</td><td>-0.308</td><td>0.142</td><td>29.977</td><td>-2.175</td><td>0.578</td></tr><tr><td>Chriesbach control - Clitunno control</td><td>-0.286</td><td>0.102</td><td>33.213</td><td>-2.796</td><td>0.227</td></tr><tr><td>Chriesbach control - Marine control</td><td>0.722</td><td>0.126</td><td>60.874</td><td>5.712</td><td>0.000</td></tr><tr><td>Chriesbach control - Reveillon control</td><td>-0.047</td><td>0.128</td><td>19.115</td><td>-0.368</td><td>1.000</td></tr><tr><td>Chriesbach control - Binnenkanal treated</td><td>0.004</td><td>0.118</td><td>49.022</td><td>0.037</td><td>1.000</td></tr><tr><td>Chriesbach control - Chessel treated</td><td>0.040</td><td>0.109</td><td>58.727</td><td>0.369</td><td>1.000</td></tr><tr><td>Chriesbach control - Chriesbach treated</td><td>0.082</td><td>0.146</td><td>62.707</td><td>0.561</td><td>1.000</td></tr></tbody></table>
<table>
  <tr>
    <td>Chriesbach control - Clitunno treated</td>
    <td>-0.467</td>
    <td>0.112</td>
    <td>34.848</td>
    <td>-4.188</td>
    <td>0.009</td>
  </tr>
  <tr>
    <td>Chriesbach control - Marine treated</td>
    <td>0.207</td>
    <td>0.145</td>
    <td>62.212</td>
    <td>1.431</td>
    <td>0.953</td>
  </tr>
  <tr>
    <td>Chriesbach control - Reveillon treated</td>
    <td>-0.088</td>
    <td>0.145</td>
    <td>32.779</td>
    <td>-0.607</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Clitunno control - Marine control</td>
    <td>1.008</td>
    <td>0.120</td>
    <td>48.850</td>
    <td>8.430</td>
    <td>0.000</td>
  </tr>
  <tr>
    <td>Clitunno control - Reveillon control</td>
    <td>0.239</td>
    <td>0.121</td>
    <td>13.411</td>
    <td>1.972</td>
    <td>0.703</td>
  </tr>
  <tr>
    <td>Clitunno control - Binnenkanal treated</td>
    <td>0.291</td>
    <td>0.111</td>
    <td>33.690</td>
    <td>2.621</td>
    <td>0.308</td>
  </tr>
  <tr>
    <td>Clitunno control - Chessel treated</td>
    <td>0.326</td>
    <td>0.101</td>
    <td>41.324</td>
    <td>3.225</td>
    <td>0.088</td>
  </tr>
  <tr>
    <td>Clitunno control - Chriesbach treated</td>
    <td>0.368</td>
    <td>0.140</td>
    <td>54.694</td>
    <td>2.626</td>
    <td>0.291</td>
  </tr>
  <tr>
    <td>Clitunno control - Clitunno treated</td>
    <td>-0.181</td>
    <td>0.104</td>
    <td>21.520</td>
    <td>-1.745</td>
    <td>0.829</td>
  </tr>
  <tr>
    <td>Clitunno control - Marine treated</td>
    <td>0.493</td>
    <td>0.139</td>
    <td>53.970</td>
    <td>3.550</td>
    <td>0.035</td>
  </tr>
  <tr>
    <td>Clitunno control - Reveillon treated</td>
    <td>0.198</td>
    <td>0.139</td>
    <td>25.123</td>
    <td>1.420</td>
    <td>0.948</td>
  </tr>
  <tr>
    <td>Marine control - Reveillon control</td>
    <td>-0.769</td>
    <td>0.142</td>
    <td>27.660</td>
    <td>-5.411</td>
    <td>0.000</td>
  </tr>
  <tr>
    <td>Marine control - Binnenkanal treated</td>
    <td>-0.717</td>
    <td>0.133</td>
    <td>59.019</td>
    <td>-5.380</td>
    <td>0.000</td>
  </tr>
  <tr>
    <td>Marine control - Chessel treated</td>
    <td>-0.682</td>
    <td>0.125</td>
    <td>65.113</td>
    <td>-5.434</td>
    <td>0.000</td>
  </tr>
  <tr>
    <td>Marine control - Chriesbach treated</td>
    <td>-0.640</td>
    <td>0.159</td>
    <td>65.559</td>
    <td>-4.035</td>
    <td>0.007</td>
  </tr>
  <tr>
    <td>Marine control - Clitunno treated</td>
    <td>-1.189</td>
    <td>0.128</td>
    <td>48.568</td>
    <td>-9.321</td>
    <td>0.000</td>
  </tr>
  <tr>
    <td>Marine control - Marine treated</td>
    <td>-0.514</td>
    <td>0.158</td>
    <td>65.296</td>
    <td>-3.267</td>
    <td>0.069</td>
  </tr>
  <tr>
    <td>Marine control - Reveillon treated</td>
    <td>-0.810</td>
    <td>0.158</td>
    <td>41.141</td>
    <td>-5.132</td>
    <td>0.000</td>
  </tr>
  <tr>
    <td>Reveillon control - Binnenkanal treated</td>
    <td>0.051</td>
    <td>0.135</td>
    <td>20.150</td>
    <td>0.381</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Reveillon control - Chessel treated</td>
    <td>0.087</td>
    <td>0.127</td>
    <td>21.676</td>
    <td>0.687</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Reveillon control - Chriesbach treated</td>
    <td>0.129</td>
    <td>0.160</td>
    <td>34.319</td>
    <td>0.807</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Reveillon control - Clitunno treated</td>
    <td>-0.420</td>
    <td>0.129</td>
    <td>15.011</td>
    <td>-3.254</td>
    <td>0.133</td>
  </tr>
  <tr>
    <td>Reveillon control - Marine treated</td>
    <td>0.254</td>
    <td>0.159</td>
    <td>33.880</td>
    <td>1.602</td>
    <td>0.897</td>
  </tr>
  <tr>
    <td>Reveillon control - Reveillon treated</td>
    <td>-0.041</td>
    <td>0.159</td>
    <td>18.955</td>
    <td>-0.259</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Binnenkanal treated - Chessel treated</td>
    <td>0.036</td>
    <td>0.117</td>
    <td>56.375</td>
    <td>0.306</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Binnenkanal treated - Chriesbach treated</td>
    <td>0.078</td>
    <td>0.152</td>
    <td>61.306</td>
    <td>0.510</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Binnenkanal treated - Clitunno treated</td>
    <td>-0.472</td>
    <td>0.119</td>
    <td>35.096</td>
    <td>-3.949</td>
    <td>0.016</td>
  </tr>
  <tr>
    <td>Binnenkanal treated - Marine treated</td>
    <td>0.203</td>
    <td>0.151</td>
    <td>60.795</td>
    <td>1.343</td>
    <td>0.970</td>
  </tr>
  <tr>
    <td>Binnenkanal treated - Reveillon treated</td>
    <td>-0.093</td>
    <td>0.151</td>
    <td>33.054</td>
    <td>-0.612</td>
    <td>1.000</td>
  </tr>
</table>
<table><tr><td>Chessel treated - Chriesbach treated</td><td>0.042</td><td>0.145</td><td>65.536</td><td>0.287</td><td>1.000</td></tr><tr><td>Chessel treated - Clitunno treated</td><td>-0.508</td><td>0.111</td><td>42.006</td><td>-4.591</td><td>0.002</td></tr><tr><td>Chessel treated - Marine treated</td><td>0.167</td><td>0.144</td><td>65.218</td><td>1.159</td><td>0.990</td></tr><tr><td>Chessel treated - Reveillon treated</td><td>-0.128</td><td>0.144</td><td>36.496</td><td>-0.889</td><td>0.999</td></tr><tr><td>Chriesbach treated - Clitunno treated</td><td>-0.549</td><td>0.147</td><td>54.010</td><td>-3.735</td><td>0.021</td></tr><tr><td>Chriesbach treated - Marine treated</td><td>0.125</td><td>0.174</td><td>65.555</td><td>0.721</td><td>1.000</td></tr><tr><td>Chriesbach treated - Reveillon treated</td><td>-0.170</td><td>0.174</td><td>45.974</td><td>-0.978</td><td>0.997</td></tr><tr><td>Clitunno treated - Marine treated</td><td>0.675</td><td>0.146</td><td>53.341</td><td>4.622</td><td>0.001</td></tr><tr><td>Clitunno treated - Reveillon treated</td><td>0.379</td><td>0.146</td><td>26.375</td><td>2.591</td><td>0.332</td></tr><tr><td>Marine treated - Reveillon treated</td><td>-0.295</td><td>0.173</td><td>45.511</td><td>-1.708</td><td>0.855</td></tr></table>
Table S5 continued

Pairwise Population Contrasts Within Treatments - Male DHA

<table><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal control - Chessel control</td><td>0.193</td><td>0.123</td><td>57.089</td><td>1.574</td><td>0.855</td></tr><tr><td>Binnenkanal control - Chriesbach control</td><td>0.205</td><td>0.123</td><td>57.049</td><td>1.673</td><td>0.805</td></tr><tr><td>Binnenkanal control - Clitunno control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Binnenkanal control - Marine control</td><td>0.340</td><td>0.121</td><td>65.039</td><td>2.807</td><td>0.156</td></tr><tr><td>Binnenkanal control - Reveillon control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Binnenkanal control - Binnenkanal treated</td><td>0.004</td><td>0.111</td><td>61.240</td><td>-0.034</td><td>1.000</td></tr><tr><td>Binnenkanal control - Chessel treated</td><td>0.058</td><td>0.115</td><td>65.053</td><td>0.503</td><td>1.000</td></tr><tr><td>Binnenkanal control - Chriesbach treated</td><td>0.236</td><td>0.109</td><td>49.755</td><td>2.166</td><td>0.493</td></tr><tr><td>Binnenkanal control - Clitunno treated</td><td>-0.406</td><td>0.150</td><td>65.481</td><td>-2.705</td><td>0.194</td></tr><tr><td>Binnenkanal control - Marine treated</td><td>0.169</td><td>0.131</td><td>65.020</td><td>1.290</td><td>0.953</td></tr><tr><td>Binnenkanal control - Reveillon treated</td><td>-0.335</td><td>0.191</td><td>64.958</td><td>-1.753</td><td>0.761</td></tr><tr><td>Chessel control - Chriesbach control</td><td>0.012</td><td>0.124</td><td>46.532</td><td>0.098</td><td>1.000</td></tr><tr><td>Chessel control - Clitunno control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel control - Marine control</td><td>0.147</td><td>0.122</td><td>56.846</td><td>1.200</td><td>0.969</td></tr><tr><td>Chessel control - Reveillon control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel control - Binnenkanal treated</td><td>-0.197</td><td>0.113</td><td>49.141</td><td>-1.751</td><td>0.761</td></tr><tr><td>Chessel control - Chessel treated</td><td>-0.135</td><td>0.116</td><td>55.741</td><td>-1.165</td><td>0.975</td></tr><tr><td>Chessel control - Chriesbach treated</td><td>0.043</td><td>0.110</td><td>37.100</td><td>0.387</td><td>1.000</td></tr><tr><td>Chessel control - Clitunno treated</td><td>-0.599</td><td>0.151</td><td>60.818</td><td>-3.967</td><td>0.007</td></tr><tr><td>Chessel control - Marine treated</td><td>-0.025</td><td>0.132</td><td>58.228</td><td>-0.186</td><td>1.000</td></tr><tr><td>Chessel control - Reveillon treated</td><td>-0.529</td><td>0.192</td><td>62.217</td><td>-2.753</td><td>0.176</td></tr><tr><td>Chriesbach control - Clitunno control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach control - Marine control</td><td>0.135</td><td>0.122</td><td>56.805</td><td>1.102</td><td>0.983</td></tr><tr><td>Chriesbach control - Reveillon control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach control - Binnenkanal treated</td><td>-0.209</td><td>0.112</td><td>49.090</td><td>-1.859</td><td>0.695</td></tr><tr><td>Chriesbach control - Chessel treated</td><td>-0.148</td><td>0.116</td><td>55.695</td><td>-1.269</td><td>0.956</td></tr><tr><td>Chriesbach control - Chriesbach treated</td><td>0.031</td><td>0.110</td><td>37.054</td><td>0.278</td><td>1.000</td></tr></tbody></table>
<table><tbody><tr><td>Chriesbach control - Clitunno treated</td><td>-0.611</td><td>0.151</td><td>60.794</td><td>-4.049</td><td>0.005</td></tr><tr><td>Chriesbach control - Marine treated</td><td>-0.037</td><td>0.132</td><td>58.194</td><td>-0.278</td><td>1.000</td></tr><tr><td>Chriesbach control - Reveillon treated</td><td>-0.541</td><td>0.192</td><td>62.204</td><td>-2.816</td><td>0.154</td></tr><tr><td>Clitunno control - Marine control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Reveillon control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Binnenkanal treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Chessel treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Chriesbach treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Clitunno treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Marine treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Reveillon treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Marine control - Reveillon control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Marine control - Binnenkanal treated</td><td>-0.344</td><td>0.111</td><td>60.990</td><td>-3.104</td><td>0.079</td></tr><tr><td>Marine control - Chessel treated</td><td>-0.282</td><td>0.115</td><td>64.904</td><td>-2.464</td><td>0.307</td></tr><tr><td>Marine control - Chriesbach treated</td><td>-0.104</td><td>0.109</td><td>49.426</td><td>-0.960</td><td>0.993</td></tr><tr><td>Marine control - Clitunno treated</td><td>-0.746</td><td>0.150</td><td>65.402</td><td>-4.982</td><td>0.000</td></tr><tr><td>Marine control - Marine treated</td><td>-0.171</td><td>0.131</td><td>64.904</td><td>-1.314</td><td>0.947</td></tr><tr><td>Marine control - Reveillon treated</td><td>-0.676</td><td>0.191</td><td>64.904</td><td>-3.536</td><td>0.024</td></tr><tr><td>Reveillon control - Binnenkanal treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Reveillon control - Chessel treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Reveillon control - Chriesbach treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Reveillon control - Clitunno treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Reveillon control - Marine treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Reveillon control - Reveillon treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Binnenkanal treated - Chessel treated</td><td>0.062</td><td>0.104</td><td>60.344</td><td>0.592</td><td>1.000</td></tr><tr><td>Binnenkanal treated - Chriesbach treated</td><td>0.240</td><td>0.097</td><td>37.686</td><td>2.465</td><td>0.319</td></tr><tr><td>Binnenkanal treated - Clitunno treated</td><td>-0.402</td><td>0.142</td><td>63.423</td><td>-2.836</td><td>0.147</td></tr><tr><td>Binnenkanal treated - Marine treated</td><td>0.173</td><td>0.121</td><td>61.749</td><td>1.422</td><td>0.916</td></tr><tr><td>Binnenkanal treated - Reveillon treated</td><td>-0.332</td><td>0.185</td><td>63.696</td><td>-1.794</td><td>0.736</td></tr></tbody></table>
<table><tr><td>Chessel treated - Chriesbach treated</td><td>0.178</td><td>0.102</td><td>46.832</td><td>1.754</td><td>0.759</td></tr><tr><td>Chessel treated - Clitunno treated</td><td>-0.464</td><td>0.145</td><td>65.435</td><td>-3.203</td><td>0.061</td></tr><tr><td>Chessel treated - Marine treated</td><td>0.111</td><td>0.125</td><td>64.904</td><td>0.889</td><td>0.996</td></tr><tr><td>Chessel treated - Reveillon treated</td><td>-0.393</td><td>0.187</td><td>64.904</td><td>-2.101</td><td>0.534</td></tr><tr><td>Chriesbach treated - Clitunno treated</td><td>-0.642</td><td>0.140</td><td>57.420</td><td>-4.584</td><td>0.001</td></tr><tr><td>Chriesbach treated - Marine treated</td><td>-0.067</td><td>0.119</td><td>52.563</td><td>-0.564</td><td>1.000</td></tr><tr><td>Chriesbach treated - Reveillon treated</td><td>-0.571</td><td>0.184</td><td>60.635</td><td>-3.113</td><td>0.077</td></tr><tr><td>Clitunno treated - Marine treated</td><td>0.575</td><td>0.158</td><td>65.355</td><td>3.645</td><td>0.018</td></tr><tr><td>Clitunno treated - Reveillon treated</td><td>0.071</td><td>0.211</td><td>65.161</td><td>0.335</td><td>1.000</td></tr><tr><td>Marine treated - Reveillon treated</td><td>-0.504</td><td>0.197</td><td>64.904</td><td>-2.555</td><td>0.260</td></tr></table>
<caption>Table S6. GLM of ALA content of fish in experiment at end of experiment and post-hoc tests</caption>

(Population effects are shown relative to Binnenkanal; NA represent population and/or treatment combinations where only one sex was present)

<table><thead><tr><th rowspan="2">Predictors</th><th colspan="3">Log10_FA ALA</th></tr><tr><th>Estimates</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>3.25</td><td>3.00 - 3.51</td><td><strong>&lt;0.001</strong></td></tr><tr><td>Treatment [treated]</td><td>-0.03</td><td>-0.39 - 0.33</td><td>0.867</td></tr><tr><td>Pop [Chessel]</td><td>-0.36</td><td>-0.71 - -0.01</td><td><strong>0.046</strong></td></tr><tr><td>Pop [Chriesbach]</td><td>-0.20</td><td>-0.57 - -0.16</td><td>0.269</td></tr><tr><td>Pop [Clitunno]</td><td>0.07</td><td>-0.28 - 0.41</td><td>0.704</td></tr><tr><td>Pop [Marine]</td><td>-0.42</td><td>-0.83 - -0.01</td><td><strong>0.047</strong></td></tr><tr><td>Pop [Reveillon]</td><td>-0.04</td><td>-0.50 - 0.42</td><td>0.860</td></tr><tr><td>Sex [m]</td><td>-0.45</td><td>-0.80 - -0.10</td><td><strong>0.013</strong></td></tr><tr><td>Treatment [treated] × Pop [Chessel]</td><td>0.15</td><td>-0.36 - 0.66</td><td>0.558</td></tr><tr><td>Treatment [treated] × Pop [Chriesbach]</td><td>-0.00</td><td>-0.60 - 0.60</td><td>0.995</td></tr><tr><td>Treatment [treated] × Pop [Clitunno]</td><td>-0.10</td><td>-0.61 - 0.41</td><td>0.699</td></tr><tr><td>Treatment [treated] × Pop [Marine]</td><td>0.20</td><td>-0.44 - 0.83</td><td>0.538</td></tr><tr><td>Treatment [treated] × Pop [Reveillon]</td><td>-0.01</td><td>-0.69 - 0.67</td><td>0.978</td></tr><tr><td>Treatment [treated] × Sex [m]</td><td>0.43</td><td>-0.05 - 0.91</td><td>0.081</td></tr><tr><td>Pop [Chessel] × Sex [m]</td><td>0.47</td><td>-0.04 - 0.98</td><td>0.069</td></tr><tr><td>Pop [Chriesbach] × Sex [m]</td><td>0.25</td><td>-0.27 - 0.77</td><td>0.336</td></tr><tr><td>Pop [Clitunno] × Sex [m]</td><td>-0.53</td><td>-1.10 - 0.03</td><td>0.063</td></tr><tr><td>Pop [Marine] × Sex [m]</td><td>0.53</td><td>0.00 - 1.07</td><td><strong>0.050</strong></td></tr><tr><td>Pop [Reveillon] × Sex [m]</td><td>-0.49</td><td>-1.21 - 0.22</td><td>0.173</td></tr><tr><td>(Treatment [treated] × Pop [Chessel]) × Sex [m]</td><td>-0.81</td><td>-1.50 - -0.12</td><td><strong>0.022</strong></td></tr><tr><td>(Treatment [treated] × Pop [Chriesbach]) × Sex [m]</td><td>-0.30</td><td>-1.05 - 0.45</td><td>0.428</td></tr></tbody></table>
(Treatment [treated] × -1.09 -1.88 -0.30 **0.008**
Pop [Marine]) × Sex [m]

**Random Effects**

σ² 0.07
τ₀₀ Tank 0.01
ICC 0.16
N Tank 44

Observations 90
Marginal R² / Conditional R² 0.378 / 0.479
Table S6 continued

<table><caption>Pairwise Population Contrasts Within Treatments - Female ALA</caption><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal - Chessel</td><td>0.284</td><td>0.128</td><td>55.672</td><td>2.227</td><td>0.242</td></tr><tr><td>Binnenkanal - Chriesbach</td><td>0.204</td><td>0.152</td><td>62.576</td><td>1.345</td><td>0.759</td></tr><tr><td>Binnenkanal - Clitunno</td><td>-0.017</td><td>0.130</td><td>40.212</td><td>-0.127</td><td>1.000</td></tr><tr><td>Binnenkanal - Marine</td><td>0.322</td><td>0.160</td><td>63.332</td><td>2.018</td><td>0.344</td></tr><tr><td>Binnenkanal - Reveillon</td><td>0.045</td><td>0.171</td><td>28.919</td><td>0.264</td><td>1.000</td></tr><tr><td>Chessel - Chriesbach</td><td>-0.080</td><td>0.150</td><td>60.902</td><td>-0.530</td><td>0.995</td></tr><tr><td>Chessel - Clitunno</td><td>-0.301</td><td>0.128</td><td>37.068</td><td>-2.352</td><td>0.200</td></tr><tr><td>Chessel - Marine</td><td>0.038</td><td>0.158</td><td>61.928</td><td>0.241</td><td>1.000</td></tr><tr><td>Chessel - Reveillon</td><td>-0.239</td><td>0.169</td><td>27.459</td><td>-1.414</td><td>0.719</td></tr><tr><td>Chriesbach - Clitunno</td><td>-0.221</td><td>0.152</td><td>49.548</td><td>-1.452</td><td>0.696</td></tr><tr><td>Chriesbach - Marine</td><td>0.118</td><td>0.178</td><td>64.918</td><td>0.661</td><td>0.986</td></tr><tr><td>Chriesbach - Reveillon</td><td>-0.159</td><td>0.188</td><td>35.253</td><td>-0.847</td><td>0.956</td></tr><tr><td>Clitunno - Marine</td><td>0.339</td><td>0.160</td><td>51.966</td><td>2.119</td><td>0.294</td></tr><tr><td>Clitunno - Reveillon</td><td>0.062</td><td>0.171</td><td>22.812</td><td>0.361</td><td>0.999</td></tr><tr><td>Marine - Reveillon</td><td>-0.277</td><td>0.194</td><td>37.445</td><td>-1.425</td><td>0.712</td></tr></tbody></table>
Table S6 continued

<table><caption>Pairwise Population Contrasts Within Treatments - Male ALA</caption><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal - Chessel</td><td>0.219</td><td>0.135</td><td>57.555</td><td>1.614</td><td>0.379</td></tr><tr><td>Binnenkanal - Chriesbach</td><td>0.104</td><td>0.133</td><td>50.176</td><td>0.784</td><td>0.861</td></tr><tr><td>Binnenkanal - Clitunno</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Binnenkanal - Marine</td><td>0.331</td><td>0.143</td><td>61.116</td><td>2.314</td><td>0.106</td></tr><tr><td>Binnenkanal - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel - Chriesbach</td><td>-0.114</td><td>0.137</td><td>48.461</td><td>-0.834</td><td>0.838</td></tr><tr><td>Chessel - Clitunno</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel - Marine</td><td>0.113</td><td>0.147</td><td>59.433</td><td>0.770</td><td>0.868</td></tr><tr><td>Chessel - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach - Clitunno</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach - Marine</td><td>0.227</td><td>0.145</td><td>53.463</td><td>1.570</td><td>0.404</td></tr><tr><td>Chriesbach - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno - Marine</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Marine - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr></tbody></table>
<table><caption>Table S6 continued</caption><caption>Pairwise Population Contrasts Within Treatments - Female ALA</caption><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal control - Chessel control</td><td>0.359</td><td>0.177</td><td>52.506</td><td>2.024</td><td>0.675</td></tr><tr><td>Binnenkanal control - Chriesbach control</td><td>0.204</td><td>0.184</td><td>57.114</td><td>1.109</td><td>0.993</td></tr><tr><td>Binnenkanal control - Clitunno control</td><td>-0.067</td><td>0.175</td><td>39.969</td><td>-0.381</td><td>1.000</td></tr><tr><td>Binnenkanal control - Marine control</td><td>0.420</td><td>0.209</td><td>63.862</td><td>2.015</td><td>0.682</td></tr><tr><td>Binnenkanal control - Reveillon control</td><td>0.040</td><td>0.228</td><td>23.788</td><td>0.177</td><td>1.000</td></tr><tr><td>Binnenkanal control - Binnenkanal treated</td><td>0.031</td><td>0.184</td><td>58.514</td><td>0.167</td><td>1.000</td></tr><tr><td>Binnenkanal control - Chessel treated</td><td>0.240</td><td>0.180</td><td>62.168</td><td>1.333</td><td>0.971</td></tr><tr><td>Binnenkanal control - Chriesbach treated</td><td>0.236</td><td>0.240</td><td>66.450</td><td>0.984</td><td>0.998</td></tr><tr><td>Binnenkanal control - Clitunno treated</td><td>0.064</td><td>0.189</td><td>43.666</td><td>0.338</td><td>1.000</td></tr><tr><td>Binnenkanal control - Marine treated</td><td>0.255</td><td>0.239</td><td>64.660</td><td>1.065</td><td>0.995</td></tr><tr><td>Binnenkanal control - Reveillon treated</td><td>0.080</td><td>0.252</td><td>35.482</td><td>0.320</td><td>1.000</td></tr><tr><td>Chessel control - Chriesbach control</td><td>-0.155</td><td>0.181</td><td>46.815</td><td>-0.858</td><td>0.999</td></tr><tr><td>Chessel control - Clitunno control</td><td>-0.425</td><td>0.172</td><td>30.282</td><td>-2.477</td><td>0.390</td></tr><tr><td>Chessel control - Marine control</td><td>0.062</td><td>0.206</td><td>57.418</td><td>0.299</td><td>1.000</td></tr><tr><td>Chessel control - Reveillon control</td><td>-0.318</td><td>0.226</td><td>20.214</td><td>-1.410</td><td>0.948</td></tr><tr><td>Chessel control - Binnenkanal treated</td><td>-0.328</td><td>0.181</td><td>48.324</td><td>-1.816</td><td>0.802</td></tr><tr><td>Chessel control - Chessel treated</td><td>-0.119</td><td>0.177</td><td>52.506</td><td>-0.669</td><td>1.000</td></tr><tr><td>Chessel control - Chriesbach treated</td><td>-0.123</td><td>0.238</td><td>62.945</td><td>-0.517</td><td>1.000</td></tr><tr><td>Chessel control - Clitunno treated</td><td>-0.295</td><td>0.187</td><td>34.622</td><td>-1.578</td><td>0.905</td></tr><tr><td>Chessel control - Marine treated</td><td>-0.104</td><td>0.237</td><td>60.238</td><td>-0.438</td><td>1.000</td></tr><tr><td>Chessel control - Reveillon treated</td><td>-0.278</td><td>0.250</td><td>31.058</td><td>-1.115</td><td>0.992</td></tr><tr><td>Chriesbach control - Clitunno control</td><td>-0.270</td><td>0.178</td><td>35.340</td><td>-1.515</td><td>0.926</td></tr><tr><td>Chriesbach control - Marine control</td><td>0.217</td><td>0.212</td><td>60.415</td><td>1.024</td><td>0.996</td></tr><tr><td>Chriesbach control - Reveillon control</td><td>-0.163</td><td>0.231</td><td>22.375</td><td>-0.707</td><td>1.000</td></tr><tr><td>Chriesbach control - Binnenkanal treated</td><td>-0.173</td><td>0.187</td><td>53.195</td><td>-0.925</td><td>0.999</td></tr><tr><td>Chriesbach control - Chessel treated</td><td>0.037</td><td>0.184</td><td>57.114</td><td>0.199</td><td>1.000</td></tr><tr><td>Chriesbach control - Chriesbach treated</td><td>0.032</td><td>0.242</td><td>64.547</td><td>0.134</td><td>1.000</td></tr></tbody></table>
<table>
  <tr>
    <td>Chriesbach control - Clitunno treated</td>
    <td>-0.140</td>
    <td>0.193</td>
    <td>39.241</td>
    <td>-0.724</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Chriesbach control - Marine treated</td>
    <td>0.051</td>
    <td>0.242</td>
    <td>62.240</td>
    <td>0.212</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Chriesbach control - Reveillon treated</td>
    <td>-0.123</td>
    <td>0.254</td>
    <td>33.457</td>
    <td>-0.484</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Clitunno control - Marine control</td>
    <td>0.487</td>
    <td>0.204</td>
    <td>47.996</td>
    <td>2.388</td>
    <td>0.433</td>
  </tr>
  <tr>
    <td>Clitunno control - Reveillon control</td>
    <td>0.107</td>
    <td>0.224</td>
    <td>17.045</td>
    <td>0.478</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Clitunno control - Binnenkanal treated</td>
    <td>0.097</td>
    <td>0.178</td>
    <td>36.570</td>
    <td>0.545</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Clitunno control - Chessel treated</td>
    <td>0.307</td>
    <td>0.175</td>
    <td>39.969</td>
    <td>1.755</td>
    <td>0.831</td>
  </tr>
  <tr>
    <td>Clitunno control - Chriesbach treated</td>
    <td>0.303</td>
    <td>0.236</td>
    <td>56.717</td>
    <td>1.283</td>
    <td>0.978</td>
  </tr>
  <tr>
    <td>Clitunno control - Clitunno treated</td>
    <td>0.131</td>
    <td>0.184</td>
    <td>26.289</td>
    <td>0.709</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Clitunno control - Marine treated</td>
    <td>0.321</td>
    <td>0.235</td>
    <td>53.464</td>
    <td>1.366</td>
    <td>0.965</td>
  </tr>
  <tr>
    <td>Clitunno control - Reveillon treated</td>
    <td>0.147</td>
    <td>0.248</td>
    <td>26.740</td>
    <td>0.593</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Marine control - Reveillon control</td>
    <td>-0.380</td>
    <td>0.251</td>
    <td>29.328</td>
    <td>-1.513</td>
    <td>0.925</td>
  </tr>
  <tr>
    <td>Marine control - Binnenkanal treated</td>
    <td>-0.390</td>
    <td>0.212</td>
    <td>61.380</td>
    <td>-1.842</td>
    <td>0.788</td>
  </tr>
  <tr>
    <td>Marine control - Chessel treated</td>
    <td>-0.180</td>
    <td>0.209</td>
    <td>63.862</td>
    <td>-0.864</td>
    <td>0.999</td>
  </tr>
  <tr>
    <td>Marine control - Chriesbach treated</td>
    <td>-0.184</td>
    <td>0.262</td>
    <td>66.623</td>
    <td>-0.704</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Marine control - Clitunno treated</td>
    <td>-0.356</td>
    <td>0.217</td>
    <td>50.184</td>
    <td>-1.644</td>
    <td>0.884</td>
  </tr>
  <tr>
    <td>Marine control - Marine treated</td>
    <td>-0.166</td>
    <td>0.261</td>
    <td>65.218</td>
    <td>-0.633</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Marine control - Reveillon treated</td>
    <td>-0.340</td>
    <td>0.273</td>
    <td>40.120</td>
    <td>-1.246</td>
    <td>0.981</td>
  </tr>
  <tr>
    <td>Reveillon control - Binnenkanal treated</td>
    <td>-0.010</td>
    <td>0.231</td>
    <td>22.785</td>
    <td>-0.042</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Reveillon control - Chessel treated</td>
    <td>0.200</td>
    <td>0.228</td>
    <td>23.788</td>
    <td>0.876</td>
    <td>0.999</td>
  </tr>
  <tr>
    <td>Reveillon control - Chriesbach treated</td>
    <td>0.196</td>
    <td>0.278</td>
    <td>36.850</td>
    <td>0.704</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Reveillon control - Clitunno treated</td>
    <td>0.024</td>
    <td>0.236</td>
    <td>19.232</td>
    <td>0.100</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Reveillon control - Marine treated</td>
    <td>0.214</td>
    <td>0.277</td>
    <td>34.994</td>
    <td>0.774</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Reveillon control - Reveillon treated</td>
    <td>0.040</td>
    <td>0.288</td>
    <td>21.579</td>
    <td>0.139</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Binnenkanal treated - Chessel treated</td>
    <td>0.209</td>
    <td>0.184</td>
    <td>58.514</td>
    <td>1.141</td>
    <td>0.991</td>
  </tr>
  <tr>
    <td>Binnenkanal treated - Chriesbach treated</td>
    <td>0.205</td>
    <td>0.242</td>
    <td>65.100</td>
    <td>0.847</td>
    <td>0.999</td>
  </tr>
  <tr>
    <td>Binnenkanal treated - Clitunno treated</td>
    <td>0.033</td>
    <td>0.193</td>
    <td>40.418</td>
    <td>0.173</td>
    <td>1.000</td>
  </tr>
  <tr>
    <td>Binnenkanal treated - Marine treated</td>
    <td>0.224</td>
    <td>0.242</td>
    <td>62.916</td>
    <td>0.927</td>
    <td>0.999</td>
  </tr>
  <tr>
    <td>Binnenkanal treated - Reveillon treated</td>
    <td>0.050</td>
    <td>0.254</td>
    <td>34.016</td>
    <td>0.196</td>
    <td>1.000</td>
  </tr>
</table>
<table><tr><td>Chessel treated - Chriesbach treated</td><td>-0.004</td><td>0.240</td><td>66.450</td><td>-0.017</td><td>1.000</td></tr><tr><td>Chessel treated - Clitunno treated</td><td>-0.176</td><td>0.189</td><td>43.666</td><td>-0.929</td><td>0.998</td></tr><tr><td>Chessel treated - Marine treated</td><td>0.015</td><td>0.239</td><td>64.660</td><td>0.061</td><td>1.000</td></tr><tr><td>Chessel treated - Reveillon treated</td><td>-0.160</td><td>0.252</td><td>35.482</td><td>-0.634</td><td>1.000</td></tr><tr><td>Chriesbach treated - Clitunno treated</td><td>-0.172</td><td>0.247</td><td>57.657</td><td>-0.696</td><td>1.000</td></tr><tr><td>Chriesbach treated - Marine treated</td><td>0.019</td><td>0.287</td><td>66.637</td><td>0.066</td><td>1.000</td></tr><tr><td>Chriesbach treated - Reveillon treated</td><td>-0.156</td><td>0.297</td><td>46.434</td><td>-0.523</td><td>1.000</td></tr><tr><td>Clitunno treated - Marine treated</td><td>0.191</td><td>0.246</td><td>54.706</td><td>0.774</td><td>1.000</td></tr><tr><td>Clitunno treated - Reveillon treated</td><td>0.016</td><td>0.258</td><td>28.866</td><td>0.063</td><td>1.000</td></tr><tr><td>Marine treated - Reveillon treated</td><td>-0.174</td><td>0.297</td><td>44.465</td><td>-0.587</td><td>1.000</td></tr></table>
Table S6 continued

<table><caption>Pairwise Population Contrasts Within Treatments - Male ALA</caption><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal control - Chessel control</td><td>-0.112</td><td>0.207</td><td>57.647</td><td>-0.541</td><td>1.000</td></tr><tr><td>Binnenkanal control - Chriesbach control</td><td>-0.047</td><td>0.207</td><td>57.434</td><td>-0.227</td><td>1.000</td></tr><tr><td>Binnenkanal control - Clitunno control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Binnenkanal control - Marine control</td><td>-0.115</td><td>0.202</td><td>63.276</td><td>-0.568</td><td>1.000</td></tr><tr><td>Binnenkanal control - Reveillon control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Binnenkanal control - Binnenkanal treated</td><td>0.395</td><td>0.186</td><td>59.577</td><td>-2.119</td><td>0.522</td></tr><tr><td>Binnenkanal control - Chessel treated</td><td>0.154</td><td>0.191</td><td>63.394</td><td>0.805</td><td>0.998</td></tr><tr><td>Binnenkanal control - Chriesbach treated</td><td>-0.139</td><td>0.185</td><td>49.945</td><td>-0.752</td><td>0.999</td></tr><tr><td>Binnenkanal control - Clitunno treated</td><td>0.171</td><td>0.248</td><td>66.030</td><td>0.690</td><td>0.999</td></tr><tr><td>Binnenkanal control - Marine treated</td><td>0.383</td><td>0.218</td><td>63.123</td><td>1.756</td><td>0.759</td></tr><tr><td>Binnenkanal control - Reveillon treated</td><td>0.149</td><td>0.319</td><td>62.621</td><td>0.467</td><td>1.000</td></tr><tr><td>Chessel control - Chriesbach control</td><td>0.065</td><td>0.212</td><td>49.987</td><td>0.307</td><td>1.000</td></tr><tr><td>Chessel control - Clitunno control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel control - Marine control</td><td>-0.003</td><td>0.207</td><td>56.319</td><td>-0.014</td><td>1.000</td></tr><tr><td>Chessel control - Reveillon control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel control - Binnenkanal treated</td><td>-0.283</td><td>0.192</td><td>50.655</td><td>-1.478</td><td>0.895</td></tr><tr><td>Chessel control - Chessel treated</td><td>0.266</td><td>0.196</td><td>55.636</td><td>1.353</td><td>0.936</td></tr><tr><td>Chessel control - Chriesbach treated</td><td>-0.027</td><td>0.190</td><td>41.262</td><td>-0.144</td><td>1.000</td></tr><tr><td>Chessel control - Clitunno treated</td><td>0.283</td><td>0.252</td><td>62.421</td><td>1.123</td><td>0.980</td></tr><tr><td>Chessel control - Marine treated</td><td>0.494</td><td>0.222</td><td>57.193</td><td>2.224</td><td>0.453</td></tr><tr><td>Chessel control - Reveillon treated</td><td>0.261</td><td>0.322</td><td>59.924</td><td>0.810</td><td>0.998</td></tr><tr><td>Chriesbach control - Clitunno control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach control - Marine control</td><td>-0.068</td><td>0.207</td><td>56.102</td><td>-0.328</td><td>1.000</td></tr><tr><td>Chriesbach control - Reveillon control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach control - Binnenkanal treated</td><td>-0.348</td><td>0.192</td><td>50.394</td><td>-1.817</td><td>0.721</td></tr><tr><td>Chriesbach control - Chessel treated</td><td>0.201</td><td>0.196</td><td>55.394</td><td>1.023</td><td>0.990</td></tr><tr><td>Chriesbach control - Chriesbach treated</td><td>-0.092</td><td>0.190</td><td>41.025</td><td>-0.485</td><td>1.000</td></tr></tbody></table>
<table><tr><td>Chriesbach control - Clitunno treated</td><td>0.218</td><td>0.252</td><td>62.300</td><td>0.865</td><td>0.997</td></tr><tr><td>Chriesbach control - Marine treated</td><td>0.429</td><td>0.222</td><td>57.008</td><td>1.932</td><td>0.648</td></tr><tr><td>Chriesbach control - Reveillon treated</td><td>0.196</td><td>0.322</td><td>59.843</td><td>0.608</td><td>1.000</td></tr><tr><td>Clitunno control - Marine control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Reveillon control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Binnenkanal treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Chessel treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Chriesbach treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Clitunno treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Marine treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Reveillon treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Marine control - Reveillon control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Marine control - Binnenkanal treated</td><td>-0.280</td><td>0.186</td><td>58.025</td><td>-1.507</td><td>0.884</td></tr><tr><td>Marine control - Chessel treated</td><td>0.269</td><td>0.191</td><td>62.168</td><td>1.407</td><td>0.921</td></tr><tr><td>Marine control - Chriesbach treated</td><td>-0.025</td><td>0.185</td><td>48.246</td><td>-0.133</td><td>1.000</td></tr><tr><td>Marine control - Clitunno treated</td><td>0.286</td><td>0.248</td><td>65.505</td><td>1.154</td><td>0.977</td></tr><tr><td>Marine control - Marine treated</td><td>0.497</td><td>0.218</td><td>62.168</td><td>2.286</td><td>0.413</td></tr><tr><td>Marine control - Reveillon treated</td><td>0.263</td><td>0.318</td><td>62.168</td><td>0.827</td><td>0.998</td></tr><tr><td>Reveillon control - Binnenkanal treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Reveillon control - Chessel treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Reveillon control - Chriesbach treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Reveillon control - Clitunno treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Reveillon control - Marine treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Reveillon control - Reveillon treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Binnenkanal treated - Chessel treated</td><td>0.549</td><td>0.175</td><td>57.428</td><td>3.141</td><td>0.073</td></tr><tr><td>Binnenkanal treated - Chriesbach treated</td><td>0.256</td><td>0.168</td><td>39.561</td><td>1.523</td><td>0.875</td></tr><tr><td>Binnenkanal treated - Clitunno treated</td><td>0.566</td><td>0.235</td><td>63.953</td><td>2.406</td><td>0.340</td></tr><tr><td>Binnenkanal treated - Marine treated</td><td>0.777</td><td>0.203</td><td>58.748</td><td>3.823</td><td>0.011</td></tr><tr><td>Binnenkanal treated - Reveillon treated</td><td>0.544</td><td>0.309</td><td>60.750</td><td>1.760</td><td>0.757</td></tr></table>
<table><tr><td>Chessel treated - Chriesbach treated</td><td>-0.293</td><td>0.174</td><td>46.269</td><td>-1.690</td><td>0.795</td></tr><tr><td>Chessel treated - Clitunno treated</td><td>0.017</td><td>0.239</td><td>65.699</td><td>0.071</td><td>1.000</td></tr><tr><td>Chessel treated - Marine treated</td><td>0.228</td><td>0.208</td><td>62.168</td><td>1.098</td><td>0.983</td></tr><tr><td>Chessel treated - Reveillon treated</td><td>-0.005</td><td>0.312</td><td>62.168</td><td>-0.017</td><td>1.000</td></tr><tr><td>Chriesbach treated - Clitunno treated</td><td>0.310</td><td>0.234</td><td>58.944</td><td>1.323</td><td>0.944</td></tr><tr><td>Chriesbach treated - Marine treated</td><td>0.522</td><td>0.202</td><td>50.692</td><td>2.579</td><td>0.254</td></tr><tr><td>Chriesbach treated - Reveillon treated</td><td>0.288</td><td>0.308</td><td>57.576</td><td>0.934</td><td>0.995</td></tr><tr><td>Clitunno treated - Marine treated</td><td>0.212</td><td>0.261</td><td>65.222</td><td>0.811</td><td>0.998</td></tr><tr><td>Clitunno treated - Reveillon treated</td><td>-0.022</td><td>0.349</td><td>63.970</td><td>-0.064</td><td>1.000</td></tr><tr><td>Marine treated - Reveillon treated</td><td>-0.234</td><td>0.329</td><td>62.168</td><td>-0.711</td><td>0.999</td></tr></table>
Table S7. GLM of EPA content of fish in experiment at end of experiment and post-hoc tests

(Population effects are shown relative to Binnenkanal; NA represent population and/or treatment combinations where only one sex was present)

<table><thead><tr><th rowspan="2">Predictors</th><th colspan="3">Log10_FA EPA</th></tr><tr><th>Estimates</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>3.58</td><td>3.46 - 3.70</td><td><strong>&lt;0.001</strong></td></tr><tr><td>Treatment [treated]</td><td>-0.02</td><td>-0.19 - -0.15</td><td>0.799</td></tr><tr><td>Pop [Chessel]</td><td>-0.26</td><td>-0.43 - -0.10</td><td><strong>0.002</strong></td></tr><tr><td>Pop [Chriesbach]</td><td>-0.14</td><td>-0.31 - -0.03</td><td>0.113</td></tr><tr><td>Pop [Clitunno]</td><td>-0.07</td><td>-0.23 - -0.10</td><td>0.430</td></tr><tr><td>Pop [Marine]</td><td>-0.20</td><td>-0.39 - -0.01</td><td><strong>0.039</strong></td></tr><tr><td>Pop [Reveillon]</td><td>-0.14</td><td>-0.36 - -0.08</td><td>0.207</td></tr><tr><td>Sex [m]</td><td>0.07</td><td>-0.09 - -0.23</td><td>0.361</td></tr><tr><td>Treatment [treated] × Pop [Chessel]</td><td>-0.03</td><td>-0.26 - -0.21</td><td>0.827</td></tr><tr><td>Treatment [treated] × Pop [Chriesbach]</td><td>-0.03</td><td>-0.31 - -0.25</td><td>0.817</td></tr><tr><td>Treatment [treated] × Pop [Clitunno]</td><td>0.00</td><td>-0.24 - -0.25</td><td>0.983</td></tr><tr><td>Treatment [treated] × Pop [Marine]</td><td>0.20</td><td>-0.09 - -0.50</td><td>0.168</td></tr><tr><td>Treatment [treated] × Pop [Reveillon]</td><td>-0.03</td><td>-0.35 - -0.30</td><td>0.873</td></tr><tr><td>Treatment [treated] × Sex [m]</td><td>-0.13</td><td>-0.34 - -0.09</td><td>0.248</td></tr><tr><td>Pop [Chessel] × Sex [m]</td><td>0.08</td><td>-0.15 - -0.31</td><td>0.501</td></tr><tr><td>Pop [Chriesbach] × Sex [m]</td><td>0.02</td><td>-0.21 - -0.26</td><td>0.857</td></tr><tr><td>Pop [Clitunno] × Sex [m]</td><td>-0.06</td><td>-0.31 - -0.20</td><td>0.656</td></tr><tr><td>Pop [Marine] × Sex [m]</td><td>0.16</td><td>-0.08 - -0.40</td><td>0.198</td></tr><tr><td>Pop [Reveillon] × Sex [m]</td><td>0.23</td><td>-0.09 - -0.55</td><td>0.149</td></tr><tr><td>(Treatment [treated] × Pop [Chessel]) × Sex [m]</td><td>0.05</td><td>-0.26 - -0.36</td><td>0.740</td></tr><tr><td>(Treatment [treated] × Pop [Chriesbach]) × Sex [m]</td><td>0.03</td><td>-0.31 - -0.38</td><td>0.840</td></tr></tbody></table>
(Treatment [treated] × -0.17 -0.53 -0.18 0.330
Pop [Marine]) × Sex [m]

Random Effects

σ² 0.01
τ₀₀ Tank 0.00
ICC 0.23
N Tank 44

Observations 89
Marginal R² / Conditional R² 0.373 / 0.515
Table S7 continued

<table><caption>Pairwise Population Contrasts Within Treatments - Female EPA</caption><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal - Chessel</td><td>0.278</td><td>0.059</td><td>53.917</td><td>4.670</td><td>0.000</td></tr><tr><td>Binnenkanal - Chriesbach</td><td>0.153</td><td>0.070</td><td>61.609</td><td>2.172</td><td>0.266</td></tr><tr><td>Binnenkanal - Clitunno</td><td>0.064</td><td>0.062</td><td>43.910</td><td>1.032</td><td>0.904</td></tr><tr><td>Binnenkanal - Marine</td><td>0.100</td><td>0.074</td><td>61.798</td><td>1.350</td><td>0.756</td></tr><tr><td>Binnenkanal - Reveillon</td><td>0.152</td><td>0.081</td><td>28.981</td><td>1.873</td><td>0.438</td></tr><tr><td>Chessel - Chriesbach</td><td>-0.125</td><td>0.070</td><td>59.944</td><td>-1.792</td><td>0.479</td></tr><tr><td>Chessel - Clitunno</td><td>-0.214</td><td>0.061</td><td>40.973</td><td>-3.485</td><td>0.014</td></tr><tr><td>Chessel - Marine</td><td>-0.178</td><td>0.073</td><td>60.331</td><td>-2.428</td><td>0.163</td></tr><tr><td>Chessel - Reveillon</td><td>-0.125</td><td>0.081</td><td>27.648</td><td>-1.553</td><td>0.635</td></tr><tr><td>Chriesbach - Clitunno</td><td>-0.089</td><td>0.072</td><td>52.391</td><td>-1.234</td><td>0.818</td></tr><tr><td>Chriesbach - Marine</td><td>-0.053</td><td>0.082</td><td>63.873</td><td>-0.644</td><td>0.987</td></tr><tr><td>Chriesbach - Reveillon</td><td>-0.001</td><td>0.089</td><td>35.177</td><td>-0.006</td><td>1.000</td></tr><tr><td>Clitunno - Marine</td><td>0.036</td><td>0.075</td><td>53.587</td><td>0.473</td><td>0.997</td></tr><tr><td>Clitunno - Reveillon</td><td>0.088</td><td>0.083</td><td>25.156</td><td>1.067</td><td>0.890</td></tr><tr><td>Marine - Reveillon</td><td>0.053</td><td>0.092</td><td>36.820</td><td>0.572</td><td>0.992</td></tr></tbody></table>
Table S7 continued

<table><caption>Pairwise Population Contrasts Within Treatments - Male EPA</caption><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal - Chessel</td><td>0.173</td><td>0.063</td><td>56.042</td><td>2.750</td><td>0.039</td></tr><tr><td>Binnenkanal - Chriesbach</td><td>0.114</td><td>0.062</td><td>49.486</td><td>1.834</td><td>0.270</td></tr><tr><td>Binnenkanal - Clitunno</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Binnenkanal - Marine</td><td>0.031</td><td>0.067</td><td>58.584</td><td>0.469</td><td>0.966</td></tr><tr><td>Binnenkanal - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel - Chriesbach</td><td>-0.059</td><td>0.064</td><td>48.317</td><td>-0.922</td><td>0.793</td></tr><tr><td>Chessel - Clitunno</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel - Marine</td><td>-0.142</td><td>0.068</td><td>57.287</td><td>-2.086</td><td>0.170</td></tr><tr><td>Chessel - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach - Clitunno</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach - Marine</td><td>-0.083</td><td>0.068</td><td>51.860</td><td>-1.232</td><td>0.610</td></tr><tr><td>Chriesbach - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno - Marine</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Marine - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr></tbody></table>
<caption>Table S8. GLM of DPA Content of fish in experiment at end of experiment and post-hoc tests</caption>

(Population effects are shown relative to Binnenkanal; NA represent population and/or treatment combinations where only one sex was present)

<table><thead><tr><th rowspan="2">Predictors</th><th colspan="3">Log10_FA DPA</th></tr><tr><th>Estimates</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>2.74</td><td>2.10 - 3.39</td><td><strong>&lt;0.001</strong></td></tr><tr><td>Treatment [treated]</td><td>-0.42</td><td>-1.34 - 0.50</td><td>0.365</td></tr><tr><td>Pop [Chessel]</td><td>-0.97</td><td>-1.86 - -0.08</td><td><strong>0.033</strong></td></tr><tr><td>Pop [Chriesbach]</td><td>-0.80</td><td>-1.72 - -0.13</td><td>0.090</td></tr><tr><td>Pop [Clitunno]</td><td>-0.21</td><td>-1.09 - -0.67</td><td>0.636</td></tr><tr><td>Pop [Marine]</td><td>-0.32</td><td>-1.37 - -0.73</td><td>0.543</td></tr><tr><td>Pop [Reveillon]</td><td>-0.10</td><td>-1.26 - -1.06</td><td>0.865</td></tr><tr><td>Sex [m]</td><td>-0.94</td><td>-1.83 - -0.05</td><td><strong>0.038</strong></td></tr><tr><td>Treatment [treated] × Pop [Chessel]</td><td>1.04</td><td>-0.25 - 2.32</td><td>0.112</td></tr><tr><td>Treatment [treated] × Pop [Chriesbach]</td><td>1.06</td><td>-0.46 - 2.57</td><td>0.169</td></tr><tr><td>Treatment [treated] × Pop [Clitunno]</td><td>0.30</td><td>-1.00 - -1.61</td><td>0.644</td></tr><tr><td>Treatment [treated] × Pop [Marine]</td><td>0.65</td><td>-0.95 - -2.25</td><td>0.418</td></tr><tr><td>Treatment [treated] × Pop [Reveillon]</td><td>0.31</td><td>-1.42 - -2.04</td><td>0.719</td></tr><tr><td>Treatment [treated] × Sex [m]</td><td>0.55</td><td>-0.66 - -1.76</td><td>0.370</td></tr><tr><td>Pop [Chessel] × Sex [m]</td><td>1.36</td><td>0.08 - 2.65</td><td><strong>0.038</strong></td></tr><tr><td>Pop [Chriesbach] × Sex [m]</td><td>1.69</td><td>0.39 - 2.99</td><td><strong>0.012</strong></td></tr><tr><td>Pop [Clitunno] × Sex [m]</td><td>-1.04</td><td>-2.46 - -0.38</td><td>0.148</td></tr><tr><td>Pop [Marine] × Sex [m]</td><td>1.34</td><td>-0.01 - 2.69</td><td>0.051</td></tr><tr><td>Pop [Reveillon] × Sex [m]</td><td>0.50</td><td>-1.31 - -2.30</td><td>0.584</td></tr><tr><td>(Treatment [treated] × Pop [Chessel]) × Sex [m]</td><td>-1.21</td><td>-2.95 - -0.53</td><td>0.169</td></tr><tr><td>(Treatment [treated] × Pop [Chriesbach]) × Sex [m]</td><td>-1.76</td><td>-3.66 - -0.14</td><td>0.069</td></tr></tbody></table>
(Treatment [treated] x -1.14 -3.14 -0.85 0.257
Pop [Marine]) x Sex [m]

**Random Effects**

02 0.43
T₀₀ Tank 0.09
ICC 0.17
N Tank 44

Observations 90

Marginal R² /Conditional R² 0.209 /0.343
Table S8 continued

<table><caption>Pairwise Population Contrasts Within Treatments - Female DPA</caption><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal - Chessel</td><td>0.452</td><td>0.323</td><td>55.531</td><td>1.399</td><td>0.727</td></tr><tr><td>Binnenkanal - Chriesbach</td><td>0.268</td><td>0.385</td><td>62.546</td><td>0.695</td><td>0.982</td></tr><tr><td>Binnenkanal - Clitunno</td><td>0.059</td><td>0.330</td><td>40.232</td><td>0.177</td><td>1.000</td></tr><tr><td>Binnenkanal - Marine</td><td>-0.005</td><td>0.404</td><td>63.238</td><td>-0.013</td><td>1.000</td></tr><tr><td>Binnenkanal - Reveillon</td><td>-0.057</td><td>0.434</td><td>28.949</td><td>-0.132</td><td>1.000</td></tr><tr><td>Chessel - Chriesbach</td><td>-0.185</td><td>0.380</td><td>60.865</td><td>-0.486</td><td>0.996</td></tr><tr><td>Chessel - Clitunno</td><td>-0.394</td><td>0.324</td><td>37.103</td><td>-1.214</td><td>0.827</td></tr><tr><td>Chessel - Marine</td><td>-0.457</td><td>0.400</td><td>61.818</td><td>-1.144</td><td>0.861</td></tr><tr><td>Chessel - Reveillon</td><td>-0.509</td><td>0.429</td><td>27.500</td><td>-1.186</td><td>0.840</td></tr><tr><td>Chriesbach - Clitunno</td><td>-0.209</td><td>0.386</td><td>49.596</td><td>-0.542</td><td>0.994</td></tr><tr><td>Chriesbach - Marine</td><td>-0.273</td><td>0.451</td><td>64.884</td><td>-0.604</td><td>0.990</td></tr><tr><td>Chriesbach - Reveillon</td><td>-0.325</td><td>0.478</td><td>35.274</td><td>-0.680</td><td>0.983</td></tr><tr><td>Clitunno - Marine</td><td>-0.064</td><td>0.405</td><td>51.909</td><td>-0.157</td><td>1.000</td></tr><tr><td>Clitunno - Reveillon</td><td>-0.116</td><td>0.434</td><td>22.928</td><td>-0.266</td><td>1.000</td></tr><tr><td>Marine - Reveillon</td><td>-0.052</td><td>0.493</td><td>37.401</td><td>-0.106</td><td>1.000</td></tr></tbody></table>
Table S8 continued

<table><caption>Pairwise Population Contrasts Within Treatments - Male DPA</caption><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal - Chessel</td><td>-0.305</td><td>0.343</td><td>57.440</td><td>-0.890</td><td>0.810</td></tr><tr><td>Binnenkanal - Chriesbach</td><td>-0.541</td><td>0.338</td><td>50.143</td><td>-1.602</td><td>0.387</td></tr><tr><td>Binnenkanal - Clitunno</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Binnenkanal - Marine</td><td>-0.775</td><td>0.363</td><td>60.903</td><td>-2.137</td><td>0.153</td></tr><tr><td>Binnenkanal - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel - Chriesbach</td><td>-0.236</td><td>0.347</td><td>48.485</td><td>-0.679</td><td>0.904</td></tr><tr><td>Chessel - Clitunno</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel - Marine</td><td>-0.469</td><td>0.371</td><td>59.254</td><td>-1.265</td><td>0.589</td></tr><tr><td>Chessel - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach - Clitunno</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach - Marine</td><td>-0.234</td><td>0.366</td><td>53.331</td><td>-0.639</td><td>0.919</td></tr><tr><td>Chriesbach - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno - Marine</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Marine - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr></tbody></table>
Table S8 continued

<table><caption>Pairwise Population Contrasts Within Treatments - Female DPA</caption><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal control - Chessel control</td><td>0.970</td><td>0.449</td><td>52.359</td><td>2.160</td><td>0.584</td></tr><tr><td>Binnenkanal control - Chriesbach control</td><td>0.795</td><td>0.465</td><td>56.960</td><td>1.711</td><td>0.856</td></tr><tr><td>Binnenkanal control - Clitunno control</td><td>0.210</td><td>0.443</td><td>39.848</td><td>0.475</td><td>1.000</td></tr><tr><td>Binnenkanal control - Marine control</td><td>0.321</td><td>0.528</td><td>63.716</td><td>0.609</td><td>1.000</td></tr><tr><td>Binnenkanal control - Reveillon control</td><td>0.099</td><td>0.580</td><td>23.859</td><td>0.171</td><td>1.000</td></tr><tr><td>Binnenkanal control - Binnenkanal treated</td><td>0.421</td><td>0.465</td><td>58.386</td><td>0.905</td><td>0.999</td></tr><tr><td>Binnenkanal control - Chessel treated</td><td>0.355</td><td>0.456</td><td>61.926</td><td>0.779</td><td>1.000</td></tr><tr><td>Binnenkanal control - Chriesbach treated</td><td>0.160</td><td>0.607</td><td>66.429</td><td>0.264</td><td>1.000</td></tr><tr><td>Binnenkanal control - Clitunno treated</td><td>0.328</td><td>0.480</td><td>43.656</td><td>0.682</td><td>1.000</td></tr><tr><td>Binnenkanal control - Marine treated</td><td>0.089</td><td>0.605</td><td>64.559</td><td>0.147</td><td>1.000</td></tr><tr><td>Binnenkanal control - Reveillon treated</td><td>0.208</td><td>0.639</td><td>35.392</td><td>0.325</td><td>1.000</td></tr><tr><td>Chessel control - Chriesbach control</td><td>-0.174</td><td>0.458</td><td>46.830</td><td>-0.381</td><td>1.000</td></tr><tr><td>Chessel control - Clitunno control</td><td>-0.760</td><td>0.436</td><td>30.369</td><td>-1.743</td><td>0.835</td></tr><tr><td>Chessel control - Marine control</td><td>-0.648</td><td>0.522</td><td>57.324</td><td>-1.243</td><td>0.983</td></tr><tr><td>Chessel control - Reveillon control</td><td>-0.871</td><td>0.574</td><td>20.360</td><td>-1.517</td><td>0.919</td></tr><tr><td>Chessel control - Binnenkanal treated</td><td>-0.549</td><td>0.458</td><td>48.358</td><td>-1.199</td><td>0.987</td></tr><tr><td>Chessel control - Chessel treated</td><td>-0.615</td><td>0.449</td><td>52.359</td><td>-1.369</td><td>0.964</td></tr><tr><td>Chessel control - Chriesbach treated</td><td>-0.810</td><td>0.601</td><td>62.967</td><td>-1.346</td><td>0.969</td></tr><tr><td>Chessel control - Clitunno treated</td><td>-0.642</td><td>0.474</td><td>34.784</td><td>-1.356</td><td>0.964</td></tr><tr><td>Chessel control - Marine treated</td><td>-0.881</td><td>0.600</td><td>60.170</td><td>-1.467</td><td>0.944</td></tr><tr><td>Chessel control - Reveillon treated</td><td>-0.762</td><td>0.634</td><td>31.070</td><td>-1.203</td><td>0.985</td></tr><tr><td>Chriesbach control - Clitunno control</td><td>-0.585</td><td>0.452</td><td>35.381</td><td>-1.294</td><td>0.975</td></tr><tr><td>Chriesbach control - Marine control</td><td>-0.474</td><td>0.536</td><td>60.323</td><td>-0.885</td><td>0.999</td></tr><tr><td>Chriesbach control - Reveillon control</td><td>-0.696</td><td>0.587</td><td>22.502</td><td>-1.187</td><td>0.985</td></tr><tr><td>Chriesbach control - Binnenkanal treated</td><td>-0.375</td><td>0.474</td><td>53.198</td><td>-0.791</td><td>1.000</td></tr><tr><td>Chriesbach control - Chessel treated</td><td>-0.440</td><td>0.465</td><td>56.960</td><td>-0.946</td><td>0.998</td></tr><tr><td>Chriesbach control - Chriesbach treated</td><td>-0.635</td><td>0.613</td><td>64.562</td><td>-1.035</td><td>0.996</td></tr></tbody></table>
<table><tr><td>Chriesbach control - Clitunno treated</td><td>-0.468</td><td>0.489</td><td>39.366</td><td>-0.957</td><td>0.998</td></tr><tr><td>Chriesbach control - Marine treated</td><td>-0.706</td><td>0.612</td><td>62.174</td><td>-1.153</td><td>0.991</td></tr><tr><td>Chriesbach control - Reveillon treated</td><td>-0.588</td><td>0.645</td><td>33.444</td><td>-0.911</td><td>0.999</td></tr><tr><td>Clitunno control - Marine control</td><td>0.111</td><td>0.517</td><td>47.886</td><td>0.215</td><td>1.000</td></tr><tr><td>Clitunno control - Reveillon control</td><td>-0.111</td><td>0.569</td><td>17.202</td><td>-0.195</td><td>1.000</td></tr><tr><td>Clitunno control - Binnenkanal treated</td><td>0.210</td><td>0.452</td><td>36.624</td><td>0.466</td><td>1.000</td></tr><tr><td>Clitunno control - Chessel treated</td><td>0.145</td><td>0.443</td><td>39.848</td><td>0.327</td><td>1.000</td></tr><tr><td>Clitunno control - Chriesbach treated</td><td>-0.050</td><td>0.597</td><td>56.722</td><td>-0.084</td><td>1.000</td></tr><tr><td>Clitunno control - Clitunno treated</td><td>0.117</td><td>0.468</td><td>26.461</td><td>0.251</td><td>1.000</td></tr><tr><td>Clitunno control - Marine treated</td><td>-0.121</td><td>0.596</td><td>53.368</td><td>-0.203</td><td>1.000</td></tr><tr><td>Clitunno control - Reveillon treated</td><td>-0.003</td><td>0.629</td><td>26.781</td><td>-0.004</td><td>1.000</td></tr><tr><td>Marine control - Reveillon control</td><td>-0.222</td><td>0.638</td><td>29.368</td><td>-0.349</td><td>1.000</td></tr><tr><td>Marine control - Binnenkanal treated</td><td>0.099</td><td>0.535</td><td>61.306</td><td>0.185</td><td>1.000</td></tr><tr><td>Marine control - Chessel treated</td><td>0.034</td><td>0.528</td><td>63.716</td><td>0.064</td><td>1.000</td></tr><tr><td>Marine control - Chriesbach treated</td><td>-0.161</td><td>0.662</td><td>66.612</td><td>-0.243</td><td>1.000</td></tr><tr><td>Marine control - Clitunno treated</td><td>0.006</td><td>0.549</td><td>50.174</td><td>0.011</td><td>1.000</td></tr><tr><td>Marine control - Marine treated</td><td>-0.232</td><td>0.661</td><td>65.147</td><td>-0.351</td><td>1.000</td></tr><tr><td>Marine control - Reveillon treated</td><td>-0.114</td><td>0.692</td><td>40.022</td><td>-0.165</td><td>1.000</td></tr><tr><td>Reveillon control - Binnenkanal treated</td><td>0.322</td><td>0.586</td><td>22.916</td><td>0.549</td><td>1.000</td></tr><tr><td>Reveillon control - Chessel treated</td><td>0.256</td><td>0.580</td><td>23.859</td><td>0.442</td><td>1.000</td></tr><tr><td>Reveillon control - Chriesbach treated</td><td>0.061</td><td>0.704</td><td>36.904</td><td>0.087</td><td>1.000</td></tr><tr><td>Reveillon control - Clitunno treated</td><td>0.229</td><td>0.599</td><td>19.408</td><td>0.382</td><td>1.000</td></tr><tr><td>Reveillon control - Marine treated</td><td>-0.010</td><td>0.703</td><td>35.000</td><td>-0.014</td><td>1.000</td></tr><tr><td>Reveillon control - Reveillon treated</td><td>0.109</td><td>0.732</td><td>21.679</td><td>0.148</td><td>1.000</td></tr><tr><td>Binnenkanal treated - Chessel treated</td><td>-0.065</td><td>0.465</td><td>58.386</td><td>-0.141</td><td>1.000</td></tr><tr><td>Binnenkanal treated - Chriesbach treated</td><td>-0.260</td><td>0.613</td><td>65.123</td><td>-0.425</td><td>1.000</td></tr><tr><td>Binnenkanal treated - Clitunno treated</td><td>-0.093</td><td>0.489</td><td>40.560</td><td>-0.191</td><td>1.000</td></tr><tr><td>Binnenkanal treated - Marine treated</td><td>-0.332</td><td>0.612</td><td>62.865</td><td>-0.542</td><td>1.000</td></tr><tr><td>Binnenkanal treated - Reveillon treated</td><td>-0.213</td><td>0.645</td><td>34.008</td><td>-0.331</td><td>1.000</td></tr></table>
<table><tr><td>Chessel treated - Chriesbach treated</td><td>-0.195</td><td>0.607</td><td>66.429</td><td>-0.321</td><td>1.000</td></tr><tr><td>Chessel treated - Clitunno treated</td><td>-0.028</td><td>0.480</td><td>43.656</td><td>-0.058</td><td>1.000</td></tr><tr><td>Chessel treated - Marine treated</td><td>-0.266</td><td>0.605</td><td>64.559</td><td>-0.440</td><td>1.000</td></tr><tr><td>Chessel treated - Reveillon treated</td><td>-0.148</td><td>0.639</td><td>35.392</td><td>-0.231</td><td>1.000</td></tr><tr><td>Chriesbach treated - Clitunno treated</td><td>0.167</td><td>0.625</td><td>57.730</td><td>0.268</td><td>1.000</td></tr><tr><td>Chriesbach treated - Marine treated</td><td>-0.071</td><td>0.726</td><td>66.628</td><td>-0.098</td><td>1.000</td></tr><tr><td>Chriesbach treated - Reveillon treated</td><td>0.047</td><td>0.754</td><td>46.380</td><td>0.063</td><td>1.000</td></tr><tr><td>Clitunno treated - Marine treated</td><td>-0.238</td><td>0.624</td><td>54.689</td><td>-0.382</td><td>1.000</td></tr><tr><td>Clitunno treated - Reveillon treated</td><td>-0.120</td><td>0.656</td><td>28.935</td><td>-0.183</td><td>1.000</td></tr><tr><td>Marine treated - Reveillon treated</td><td>0.118</td><td>0.753</td><td>44.358</td><td>0.157</td><td>1.000</td></tr></table>
Table S8 continued

Pairwise Population Contrasts Within Treatments - Male DPA

<table><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal control - Chessel control</td><td>-0.392</td><td>0.524</td><td>57.616</td><td>-0.749</td><td>0.999</td></tr><tr><td>Binnenkanal control - Chriesbach control</td><td>-0.893</td><td>0.524</td><td>57.395</td><td>-1.705</td><td>0.788</td></tr><tr><td>Binnenkanal control - Clitunno control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Binnenkanal control - Marine control</td><td>-1.019</td><td>0.511</td><td>63.097</td><td>-1.996</td><td>0.605</td></tr><tr><td>Binnenkanal control - Reveillon control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Binnenkanal control - Binnenkanal treated</td><td>0.127</td><td>0.472</td><td>59.415</td><td>-0.268</td><td>1.000</td></tr><tr><td>Binnenkanal control - Chessel treated</td><td>-0.345</td><td>0.484</td><td>63.222</td><td>-0.712</td><td>0.999</td></tr><tr><td>Binnenkanal control - Chriesbach treated</td><td>-0.315</td><td>0.469</td><td>49.873</td><td>-0.671</td><td>1.000</td></tr><tr><td>Binnenkanal control - Clitunno treated</td><td>0.822</td><td>0.627</td><td>65.982</td><td>1.311</td><td>0.948</td></tr><tr><td>Binnenkanal control - Marine treated</td><td>-0.657</td><td>0.551</td><td>62.935</td><td>-1.191</td><td>0.971</td></tr><tr><td>Binnenkanal control - Reveillon treated</td><td>-0.837</td><td>0.807</td><td>62.404</td><td>-1.038</td><td>0.989</td></tr><tr><td>Chessel control - Chriesbach control</td><td>-0.501</td><td>0.536</td><td>50.104</td><td>-0.935</td><td>0.994</td></tr><tr><td>Chessel control - Clitunno control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel control - Marine control</td><td>-0.627</td><td>0.523</td><td>56.233</td><td>-1.198</td><td>0.970</td></tr><tr><td>Chessel control - Reveillon control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel control - Binnenkanal treated</td><td>0.266</td><td>0.485</td><td>50.667</td><td>0.548</td><td>1.000</td></tr><tr><td>Chessel control - Chessel treated</td><td>0.047</td><td>0.498</td><td>55.572</td><td>0.095</td><td>1.000</td></tr><tr><td>Chessel control - Chriesbach treated</td><td>0.077</td><td>0.483</td><td>41.397</td><td>0.160</td><td>1.000</td></tr><tr><td>Chessel control - Clitunno treated</td><td>1.214</td><td>0.637</td><td>62.419</td><td>1.906</td><td>0.665</td></tr><tr><td>Chessel control - Marine treated</td><td>-0.264</td><td>0.563</td><td>57.079</td><td>-0.470</td><td>1.000</td></tr><tr><td>Chessel control - Reveillon treated</td><td>-0.445</td><td>0.815</td><td>59.732</td><td>-0.546</td><td>1.000</td></tr><tr><td>Chriesbach control - Clitunno control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach control - Marine control</td><td>-0.126</td><td>0.523</td><td>56.008</td><td>-0.241</td><td>1.000</td></tr><tr><td>Chriesbach control - Reveillon control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach control - Binnenkanal treated</td><td>0.767</td><td>0.485</td><td>50.397</td><td>1.580</td><td>0.851</td></tr><tr><td>Chriesbach control - Chessel treated</td><td>0.548</td><td>0.498</td><td>55.321</td><td>1.102</td><td>0.982</td></tr><tr><td>Chriesbach control - Chriesbach treated</td><td>0.578</td><td>0.483</td><td>41.152</td><td>1.198</td><td>0.969</td></tr></tbody></table>
<table><tr><td>Chriesbach control - Clitunno treated</td><td>1.715</td><td>0.637</td><td>62.294</td><td>2.692</td><td>0.200</td></tr><tr><td>Chriesbach control - Marine treated</td><td>0.237</td><td>0.563</td><td>56.887</td><td>0.420</td><td>1.000</td></tr><tr><td>Chriesbach control - Reveillon treated</td><td>0.056</td><td>0.814</td><td>59.647</td><td>0.069</td><td>1.000</td></tr><tr><td>Clitunno control - Marine control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Reveillon control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Binnenkanal treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Chessel treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Chriesbach treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Clitunno treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Marine treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno control - Reveillon treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Marine control - Reveillon control</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Marine control - Binnenkanal treated</td><td>0.893</td><td>0.471</td><td>57.792</td><td>1.895</td><td>0.672</td></tr><tr><td>Marine control - Chessel treated</td><td>0.674</td><td>0.484</td><td>61.926</td><td>1.394</td><td>0.924</td></tr><tr><td>Marine control - Chriesbach treated</td><td>0.704</td><td>0.468</td><td>48.111</td><td>1.504</td><td>0.884</td></tr><tr><td>Marine control - Clitunno treated</td><td>1.841</td><td>0.626</td><td>65.428</td><td>2.939</td><td>0.116</td></tr><tr><td>Marine control - Marine treated</td><td>0.363</td><td>0.551</td><td>61.926</td><td>0.658</td><td>1.000</td></tr><tr><td>Marine control - Reveillon treated</td><td>0.182</td><td>0.806</td><td>61.926</td><td>0.226</td><td>1.000</td></tr><tr><td>Reveillon control - Binnenkanal treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Reveillon control - Chessel treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Reveillon control - Chriesbach treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Reveillon control - Clitunno treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Reveillon control - Marine treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Reveillon control - Reveillon treated</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Binnenkanal treated - Chessel treated</td><td>-0.218</td><td>0.443</td><td>57.199</td><td>-0.493</td><td>1.000</td></tr><tr><td>Binnenkanal treated - Chriesbach treated</td><td>-0.188</td><td>0.426</td><td>39.571</td><td>-0.442</td><td>1.000</td></tr><tr><td>Binnenkanal treated - Clitunno treated</td><td>0.949</td><td>0.595</td><td>63.892</td><td>1.594</td><td>0.846</td></tr><tr><td>Binnenkanal treated - Marine treated</td><td>-0.530</td><td>0.515</td><td>58.510</td><td>-1.030</td><td>0.989</td></tr><tr><td>Binnenkanal treated - Reveillon treated</td><td>-0.710</td><td>0.782</td><td>60.506</td><td>-0.908</td><td>0.996</td></tr></table>
<table><tr><td>Chessel treated - Chriesbach treated</td><td>0.030</td><td>0.440</td><td>46.164</td><td>0.068</td><td>1.000</td></tr><tr><td>Chessel treated - Clitunno treated</td><td>1.167</td><td>0.605</td><td>65.632</td><td>1.928</td><td>0.651</td></tr><tr><td>Chessel treated - Marine treated</td><td>-0.312</td><td>0.527</td><td>61.926</td><td>-0.592</td><td>1.000</td></tr><tr><td>Chessel treated - Reveillon treated</td><td>-0.492</td><td>0.790</td><td>61.926</td><td>-0.623</td><td>1.000</td></tr><tr><td>Chriesbach treated - Clitunno treated</td><td>1.137</td><td>0.593</td><td>58.914</td><td>1.917</td><td>0.658</td></tr><tr><td>Chriesbach treated - Marine treated</td><td>-0.342</td><td>0.513</td><td>50.523</td><td>-0.667</td><td>1.000</td></tr><tr><td>Chriesbach treated - Reveillon treated</td><td>-0.522</td><td>0.781</td><td>57.338</td><td>-0.669</td><td>1.000</td></tr><tr><td>Clitunno treated - Marine treated</td><td>-1.479</td><td>0.660</td><td>65.132</td><td>-2.240</td><td>0.441</td></tr><tr><td>Clitunno treated - Reveillon treated</td><td>-1.659</td><td>0.884</td><td>63.817</td><td>-1.876</td><td>0.685</td></tr><tr><td>Marine treated - Reveillon treated</td><td>-0.180</td><td>0.833</td><td>61.926</td><td>-0.216</td><td>1.000</td></tr></table>
<table><caption>**Table S9. GLM of LIN Content of fish in experiment at end of experiment (Population effects are shown relative to Binnenkanal; NA represent population and/or treatment combinations where only one sex was present)**</caption><thead><tr><th rowspan="2">Predictors</th><th colspan="3">Log10_FA LIN</th></tr><tr><th>Estimates</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>3.91</td><td>3.64 - 4.18</td><td><strong>&lt;0.001</strong></td></tr><tr><td>Treatment [treated]</td><td>0.08</td><td>-0.30 - 0.47</td><td>0.662</td></tr><tr><td>Pop [Chessel]</td><td>-0.22</td><td>-0.59 - 0.14</td><td>0.227</td></tr><tr><td>Pop [Chriesbach]</td><td>-0.14</td><td>-0.52 - 0.24</td><td>0.469</td></tr><tr><td>Pop [Clitunno]</td><td>0.15</td><td>-0.21 - 0.50</td><td>0.408</td></tr><tr><td>Pop [Marine]</td><td>-0.27</td><td>-0.72 - 0.17</td><td>0.220</td></tr><tr><td>Pop [Reveillon]</td><td>-0.12</td><td>-0.56 - 0.33</td><td>0.602</td></tr><tr><td>Sex [m]</td><td>-0.08</td><td>-0.48 - 0.33</td><td>0.707</td></tr><tr><td>Treatment [treated] × Pop [Chessel]</td><td>0.01</td><td>-0.52 - 0.54</td><td>0.965</td></tr><tr><td>Treatment [treated] × Pop [Chriesbach]</td><td>-0.13</td><td>-0.76 - 0.51</td><td>0.694</td></tr><tr><td>Treatment [treated] × Pop [Clitunno]</td><td>-0.36</td><td>-0.88 - 0.16</td><td>0.174</td></tr><tr><td>Treatment [treated] × Pop [Marine]</td><td>0.30</td><td>-0.37 - 0.98</td><td>0.370</td></tr><tr><td>Treatment [treated] × Pop [Reveillon]</td><td>-0.04</td><td>-0.71 - 0.64</td><td>0.909</td></tr><tr><td>Treatment [treated] × Sex [m]</td><td>-0.11</td><td>-0.65 - 0.44</td><td>0.695</td></tr><tr><td>Pop [Chessel] × Sex [m]</td><td>0.29</td><td>-0.27 - 0.85</td><td>0.311</td></tr><tr><td>Pop [Chriesbach] × Sex [m]</td><td>0.19</td><td>-0.38 - 0.76</td><td>0.508</td></tr><tr><td>Pop [Clitunno] × Sex [m]</td><td>-0.05</td><td>-0.67 - 0.57</td><td>0.873</td></tr><tr><td>Pop [Marine] × Sex [m]</td><td>0.28</td><td>-0.33 - 0.90</td><td>0.358</td></tr><tr><td>Pop [Reveillon] × Sex [m]</td><td>-0.01</td><td>-0.84 - 0.81</td><td>0.971</td></tr><tr><td>(Treatment [treated] × Pop [Chessel]) × Sex [m]</td><td>-0.18</td><td>-0.95 - 0.59</td><td>0.647</td></tr><tr><td>(Treatment [treated] × Pop [Chriesbach]) × Sex [m]</td><td>-0.02</td><td>-0.86 - 0.82</td><td>0.961</td></tr></tbody></table>
(Treatment [treated] × -0.57 -1.47 -0.33 0.213
Pop [Marine]) × Sex [m]

Random Effects

σ² 0.09
τ₀₀ Tank 0.00
ICC 0.01
N Tank 44

Observations 90
Marginal R² / Conditional R² 0.154 / 0.163
Table S10. ARA Content of fish in experiment at end of experiment and post-hoc tests

(Population effects are shown relative to Binnenkanal; NA represent population and/or treatment combinations where only one sex was present)

<table><thead><tr><th rowspan="2">Predictors</th><th colspan="3">Log10_FA ARA</th></tr><tr><th>Estimates</th><th>CI</th><th>p</th></tr></thead><tbody><tr><td>(Intercept)</td><td>3.64</td><td>3.53 - 3.75</td><td><strong>&lt;0.001</strong></td></tr><tr><td>Treatment [treated]</td><td>0.04</td><td>-0.11 - 0.20</td><td>0.576</td></tr><tr><td>Pop [Chessel]</td><td>-0.18</td><td>-0.33 - -0.02</td><td><strong>0.025</strong></td></tr><tr><td>Pop [Chriesbach]</td><td>-0.07</td><td>-0.23 - 0.09</td><td>0.401</td></tr><tr><td>Pop [Clitunno]</td><td>-0.00</td><td>-0.15 - 0.15</td><td>0.996</td></tr><tr><td>Pop [Marine]</td><td>-0.08</td><td>-0.27 - 0.10</td><td>0.353</td></tr><tr><td>Pop [Reveillon]</td><td>-0.11</td><td>-0.32 - 0.09</td><td>0.273</td></tr><tr><td>Sex [m]</td><td>-0.02</td><td>-0.17 - 0.13</td><td>0.754</td></tr><tr><td>Treatment [treated] × Pop [Chessel]</td><td>0.01</td><td>-0.22 - 0.23</td><td>0.956</td></tr><tr><td>Treatment [treated] × Pop [Chriesbach]</td><td>-0.02</td><td>-0.28 - 0.24</td><td>0.861</td></tr><tr><td>Treatment [treated] × Pop [Clitunno]</td><td>-0.04</td><td>-0.28 - 0.19</td><td>0.704</td></tr><tr><td>Treatment [treated] × Pop [Marine]</td><td>0.24</td><td>-0.03 - 0.52</td><td>0.084</td></tr><tr><td>Treatment [treated] × Pop [Reveillon]</td><td>-0.08</td><td>-0.39 - 0.22</td><td>0.587</td></tr><tr><td>Treatment [treated] × Sex [m]</td><td>0.01</td><td>-0.19 - 0.21</td><td>0.921</td></tr><tr><td>Pop [Chessel] × Sex [m]</td><td>0.08</td><td>-0.13 - 0.30</td><td>0.446</td></tr><tr><td>Pop [Chriesbach] × Sex [m]</td><td>0.06</td><td>-0.17 - 0.28</td><td>0.617</td></tr><tr><td>Pop [Clitunno] × Sex [m]</td><td>-0.18</td><td>-0.43 - 0.06</td><td>0.136</td></tr><tr><td>Pop [Marine] × Sex [m]</td><td>0.17</td><td>-0.06 - 0.40</td><td>0.145</td></tr><tr><td>Pop [Reveillon] × Sex [m]</td><td>0.02</td><td>-0.29 - 0.32</td><td>0.919</td></tr><tr><td>(Treatment [treated] × Pop [Chessel]) × Sex [m]</td><td>-0.05</td><td>-0.34 - 0.25</td><td>0.738</td></tr><tr><td>(Treatment [treated] × Pop [Chriesbach]) × Sex [m]</td><td>-0.02</td><td>-0.35 - 0.30</td><td>0.879</td></tr></tbody></table>
(Treatment [treated]) × -0.40 -0.73 -0.06 **0.022**
Pop [Marine]) × Sex [m]

**Random Effects**

<table><tr><td>σ<sup>2</sup></td><td>0.01</td></tr><tr><td>τ<sub>00</sub> Tank</td><td>0.00</td></tr><tr><td>ICC</td><td>0.21</td></tr><tr><td>N<sub>Tank</sub></td><td>44</td></tr></table>

Observations 89
Marginal R<sup>2</sup> / Conditional R<sup>2</sup> 0.290 / 0.438
Table S10 continued

<table><caption>Pairwise Population Contrasts Within Treatments - Female ARA</caption><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal - Chessel</td><td>0.174</td><td>0.056</td><td>54.306</td><td>3.120</td><td>0.033</td></tr><tr><td>Binnenkanal - Chriesbach</td><td>0.079</td><td>0.066</td><td>61.734</td><td>1.192</td><td>0.839</td></tr><tr><td>Binnenkanal - Clitunno</td><td>0.023</td><td>0.058</td><td>44.163</td><td>0.386</td><td>0.999</td></tr><tr><td>Binnenkanal - Marine</td><td>-0.037</td><td>0.070</td><td>62.071</td><td>-0.525</td><td>0.995</td></tr><tr><td>Binnenkanal - Reveillon</td><td>0.154</td><td>0.076</td><td>28.904</td><td>2.028</td><td>0.352</td></tr><tr><td>Chessel - Chriesbach</td><td>-0.095</td><td>0.066</td><td>60.076</td><td>-1.454</td><td>0.694</td></tr><tr><td>Chessel - Clitunno</td><td>-0.152</td><td>0.057</td><td>41.165</td><td>-2.644</td><td>0.109</td></tr><tr><td>Chessel - Marine</td><td>-0.211</td><td>0.069</td><td>60.630</td><td>-3.060</td><td>0.037</td></tr><tr><td>Chessel - Reveillon</td><td>-0.020</td><td>0.075</td><td>27.532</td><td>-0.269</td><td>1.000</td></tr><tr><td>Chriesbach - Clitunno</td><td>-0.056</td><td>0.068</td><td>52.510</td><td>-0.836</td><td>0.959</td></tr><tr><td>Chriesbach - Marine</td><td>-0.116</td><td>0.078</td><td>63.982</td><td>-1.489</td><td>0.672</td></tr><tr><td>Chriesbach - Reveillon</td><td>0.075</td><td>0.083</td><td>35.138</td><td>0.901</td><td>0.944</td></tr><tr><td>Clitunno - Marine</td><td>-0.059</td><td>0.071</td><td>53.905</td><td>-0.833</td><td>0.960</td></tr><tr><td>Clitunno - Reveillon</td><td>0.132</td><td>0.077</td><td>24.962</td><td>1.706</td><td>0.541</td></tr><tr><td>Marine - Reveillon</td><td>0.191</td><td>0.086</td><td>36.923</td><td>2.216</td><td>0.255</td></tr></tbody></table>
Table S10 continued

<table><caption>Pairwise Population Contrasts Within Treatments - Male ARA</caption><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal - Chessel</td><td>0.115</td><td>0.059</td><td>56.344</td><td>1.941</td><td>0.223</td></tr><tr><td>Binnenkanal - Chriesbach</td><td>0.036</td><td>0.059</td><td>49.590</td><td>0.609</td><td>0.929</td></tr><tr><td>Binnenkanal - Clitunno</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Binnenkanal - Marine</td><td>-0.006</td><td>0.063</td><td>59.166</td><td>-0.103</td><td>1.000</td></tr><tr><td>Binnenkanal - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel - Chriesbach</td><td>-0.079</td><td>0.060</td><td>48.270</td><td>-1.321</td><td>0.555</td></tr><tr><td>Chessel - Clitunno</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chessel - Marine</td><td>-0.122</td><td>0.064</td><td>57.760</td><td>-1.896</td><td>0.241</td></tr><tr><td>Chessel - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach - Clitunno</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Chriesbach - Marine</td><td>-0.042</td><td>0.063</td><td>52.206</td><td>-0.664</td><td>0.910</td></tr><tr><td>Chriesbach - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno - Marine</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Clitunno - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr><tr><td>Marine - Reveillon</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td><td>NA</td></tr></tbody></table>
Table S10 continued

Pairwise Population Contrasts Within Treatments - Female ARA

<table><thead><tr><th>contrast</th><th>estimate</th><th>SE</th><th>df</th><th>t.ratio</th><th>p.value</th></tr></thead><tbody><tr><td>Binnenkanal control - Chessel control</td><td>0.177</td><td>0.078</td><td>51.181</td><td>2.281</td><td>0.502</td></tr><tr><td>Binnenkanal control - Chriesbach control</td><td>0.068</td><td>0.080</td><td>55.652</td><td>0.840</td><td>0.999</td></tr><tr><td>Binnenkanal control - Clitunno control</td><td>0.000</td><td>0.077</td><td>38.935</td><td>0.005</td><td>1.000</td></tr><tr><td>Binnenkanal control - Marine control</td><td>0.085</td><td>0.091</td><td>62.247</td><td>0.930</td><td>0.999</td></tr><tr><td>Binnenkanal control - Reveillon control</td><td>0.113</td><td>0.102</td><td>24.070</td><td>1.105</td><td>0.991</td></tr><tr><td>Binnenkanal control - Binnenkanal treated</td><td>0.045</td><td>0.080</td><td>57.147</td><td>-0.558</td><td>1.000</td></tr><tr><td>Binnenkanal control - Chessel treated</td><td>0.126</td><td>0.079</td><td>59.983</td><td>1.608</td><td>0.900</td></tr><tr><td>Binnenkanal control - Chriesbach treated</td><td>0.046</td><td>0.104</td><td>65.499</td><td>0.439</td><td>1.000</td></tr><tr><td>Binnenkanal control - Clitunno treated</td><td>-0.000</td><td>0.086</td><td>51.053</td><td>-0.002</td><td>1.000</td></tr><tr><td>Binnenkanal control - Marine treated</td><td>-0.203</td><td>0.104</td><td>63.306</td><td>-1.942</td><td>0.729</td></tr><tr><td>Binnenkanal control - Reveillon treated</td><td>0.151</td><td>0.112</td><td>34.696</td><td>1.351</td><td>0.965</td></tr><tr><td>Chessel control - Chriesbach control</td><td>-0.110</td><td>0.080</td><td>46.579</td><td>-1.382</td><td>0.961</td></tr><tr><td>Chessel control - Clitunno control</td><td>-0.177</td><td>0.076</td><td>30.623</td><td>-2.321</td><td>0.484</td></tr><tr><td>Chessel control - Marine control</td><td>-0.093</td><td>0.090</td><td>56.344</td><td>-1.029</td><td>0.996</td></tr><tr><td>Chessel control - Reveillon control</td><td>-0.065</td><td>0.101</td><td>20.985</td><td>-0.639</td><td>1.000</td></tr><tr><td>Chessel control - Binnenkanal treated</td><td>-0.222</td><td>0.079</td><td>48.150</td><td>-2.799</td><td>0.213</td></tr><tr><td>Chessel control - Chessel treated</td><td>-0.051</td><td>0.078</td><td>51.181</td><td>-0.655</td><td>1.000</td></tr><tr><td>Chessel control - Chriesbach treated</td><td>-0.132</td><td>0.104</td><td>62.426</td><td>-1.272</td><td>0.980</td></tr><tr><td>Chessel control - Clitunno treated</td><td>-0.178</td><td>0.085</td><td>43.094</td><td>-2.088</td><td>0.633</td></tr><tr><td>Chessel control - Marine treated</td><td>-0.380</td><td>0.104</td><td>59.260</td><td>-3.667</td><td>0.024</td></tr><tr><td>Chessel control - Reveillon treated</td><td>-0.027</td><td>0.111</td><td>30.932</td><td>-0.241</td><td>1.000</td></tr><tr><td>Chriesbach control - Clitunno control</td><td>-0.067</td><td>0.079</td><td>35.347</td><td>-0.850</td><td>0.999</td></tr><tr><td>Chriesbach control - Marine control</td><td>0.017</td><td>0.092</td><td>59.265</td><td>0.185</td><td>1.000</td></tr><tr><td>Chriesbach control - Reveillon control</td><td>0.045</td><td>0.103</td><td>23.010</td><td>0.438</td><td>1.000</td></tr><tr><td>Chriesbach control - Binnenkanal treated</td><td>-0.112</td><td>0.082</td><td>52.753</td><td>-1.370</td><td>0.964</td></tr><tr><td>Chriesbach control - Chessel treated</td><td>0.059</td><td>0.080</td><td>55.652</td><td>0.733</td><td>1.000</td></tr><tr><td>Chriesbach control - Chriesbach treated</td><td>-0.022</td><td>0.106</td><td>63.917</td><td>-0.207</td><td>1.000</td></tr></tbody></table>