marine and coastal fisheries - review pollachius virens

For Peer Review Only

The use of total lipids, fatty acids and trace elements

profiles to assess farming influence on saithe (Pollachius virens L.)

Journal: Marine and Coastal Fisheries

Manuscript ID: UMCF-2014-0013

Manuscript Type: Article

Keywords: Aquafeed, Fish populations, Metabolites, Mariculture, Fisheries < Management, Interactions, Management

URL: http://mc.manuscriptcentral.com/mcf Email: [email protected]

Marine and Coastal Fisheries


1

Abstract 1

2

Saithe (Pollachius virens) aggregates at open cage salmon farms in Norway due to an abundance of 3

uneaten salmon feed underneath the cages. The aggregated saithe have modified their feeding habits as 4

they have switched from wild prey to lost food pellets. This diet switch might lead to physiological 5

changes and biochemical variation. In this study, variations in total lipids, fatty acids and trace 6

elements profiles in saithe liver and muscle were measured to evaluate farming influence on wild 7

saithe populations. Farm-aggregated saithe had higher fat content in liver tissues compared to 8

individuals captured far from farms, but no clear differences were found in muscle tissues. The 9

proportions of fatty acids of terrestrial origin, such as oleic acid, linoleic acid and linolenic acid, were 10

higher in liver and muscle tissues of farm-aggregated saithe. In addition, low proportions of 11

eicosapentaenoic acid and docosahexaenoic acid in farm-aggregated saithe tissues reflected the 12

influence of farms activity. Variations in specific trace elements signatures among fish groups, such as 13

Fe, Mo and Se in liver, and Hg, B and Se in muscle tissues, also revealed farming influence on saithe. 14

However, fatty acids and trace elements profiles varied between fishing methods. Thus, the potential 15

use of specific metabolites as bioindicators to discriminate whether an individual fish has been feeding 16

in the vicinity of the farms is discussed. 17

18

Keywords: Aquafeed; fish populations; metabolites; aquaculture; fisheries; interactions; management. 19

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1. Introduction 20

21

A wide number of fish species are attracted towards natural or artificial floating objects, often referred 22

to as Fish Aggregation Devices (FADs) (Dempster and Taquet 2004). Fish farms may serve as 23

FADs due to an abundance of uneaten fish feed, and possibly also because they provide hiding places 24

and attract smaller prey fish species (Sanchez-Jerez et al. 2011). Since fishing is not allowed in the 25

close vicinity of the farms, these effects of fish farming may create conflicts with local fisheries 26

because wild fish stocks becomes less available for exploitation. Moreover, A diet shift from wild prey 27

to a diet consisting partly of artificial fish feed also may affect fish quality and fish fillet organoleptic 28

characteristics (Carss 1990; Skog et al. 2003; Otter et al. 2009; Dempster et al. 2011). Saithe 29

(Pollachius virens) is an important commercial fish species in Norway, which typically occur in 30

pelagic schools along the coast during part of their migration and range extensively through a wide 31

number of fjords (Bjordal and Skar 1992; Bjordal and Johnstone 1993). In addition, saithe is also the 32

most abundant wild fish around Norwegian salmon farms. 33

34

In presence of fish farms, wild saithe reside in the vicinity of facilities for several months (Bjordal and 35

Skar 1992; Bjordal and Johnstone 1993; Uglem et al. 2009; Dempster et al. 2009, 2010), a sufficient 36

period to cause physiological changes and modification of metabolic profiles due to a diet switch from 37

wild prey to lost feed pellets (Skog et al. 2003; Dempster et al. 2009; Otter et al. 2009; Bustnes et al. 38

2010; Fernandez-Jover et al. 2011a,b). Significant differences in body condition, relative liver size, 39

lipid content and fatty acid composition in both muscle and liver have been reported in previous works 40

for farm-associated and un-associated saithe, but also that saithe fillets from a fjord without salmon 41

farms tasted better than those collected in a fjord with farms (Skog et al. 2003). Besides the nutrient 42

inputs, fish farms may also represent additional sources of a range of trace elements, since fish diets 43

are enriched with various essential elements, including copper (Cu), iron (Fe), zinc (Zn), manganese 44

(Mn), cobalt (Co) and chrome (Cr) among others (CIESM 2007). In addition, Cu is still used as anti-45

fouling treatment (copper-based algaecides) for the net-pens and related equipment (e.g. Solberg et al. 46

2002; Brooks and Manhken 2003; Braithwaite and McEvoy 2005; Braithwaite et al. 2007). Bustnes et 47

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al. (2011) found that Hg concentrations of farm-aggregated saithe were higher, but not at critically 48

elevated levels of public health concern, compared to non-associated fish, suggesting that the 49

distribution of Hg and other elements in saithe and cod in Norwegian coastal waters may be more 50

influenced by habitat use, diet, geochemical conditions and water chemistry than farming activity per 51

se. Nevertheless, the overall knowledge about presence and origin of essential and non-essential trace 52

elements in saithe populations, both influenced or non-influenced by farms, is still sparse. 53

54

To solve conflicts between fish farming and fisheries with respect to aggregation of saithe at fish 55

farms, as well as quality reduction of saithe that feed on waste salmon pellets, a quantitative tool to 56

determine retrospectively if the fish have been eating pellets would be required. In some cases the fish 57

might have been feeding actively on pellets over prolonged periods even though pellets are not found 58

in stomach samples. The evacuation time for gadoids usually varies (Andersen 2001) and stomach 59

analyses would thus only reveal recent feeding on artificial fish feed. Enlarged livers have been used 60

as an indication on active feeding on salmon pellets (Dempster et al. 2009; 2011), but it is also 61

possible that saithe will develop enlarged livers due to feeding on natural prey with high fat content, 62

like herring (Clupea harengus) or capelin (Mallotus villosus). However, as the fatty acid and trace 63

element profiles (here and thereafter: FA and TE respectively) of commercial fish feed differs from 64

natural food it might be possible to use biochemical variation in tissues to decide the dietary pre-65

history of saithe. For instance it has been shown that the FA profiles of liver, muscle and eggs varies 66

between farm associated and un-associated gadoids (Skog et al. 2003; Fernandez Jover et al. 2011b; 67

Uglem et al. 2012). In particular, the FAs from vegetable oils varies between the two groups, since 68

vegetable fats are used to substitute marine fat in artificial fish feed (e.g. Bell et al., 2001, 2003). In the 69

current study we examined if TEs in addition to FAs and lipid content could be used for distinguishing 70

between farm-associated and un-associated saithe. The specific objectives of the study were: i) to 71

compare the total lipids, FAs and TEs composition between saithe captured with bottom net or jigging, 72

both at fish farms and in areas with no farming activity, and ii) to determine the reliability of using 73

these compounds as indicators to detect the farming influence on wild saithe assemblages. 74

75

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76

2. Material and methods 77

78

2.1. Fish sampling and preparation 79

80

A total of 32 saithe were captured between the 19th and 21st of September of 2012 with bottom nets 81

and hooks or jigging (automatic jigging/juksa machines) around salmon farms and control areas away 82

from farms in the vicinity of Hitra Island, Norway (63.603658N / 8.645661E) (Figure 1; Table 1). 83

Altogether, 4 groups of saithe populations (8 individuals in each group) were thus defined: aggregated 84

fish captured by hooks (U-hooks), un-aggregated fish captured by nets (U-nets), farm-aggregated fish 85

captured by hooks (A-hooks), and farm-aggregated fish captured by nets (A-nets) (Table 1). There 86

were no significant differences in total length and weight among the fish groups (ANOVA, total 87

length: p-value=0.14; weight: p-value=0.74). Muscle and liver tissue samples (around 6 g) were 88

collected from captured fish and stored at -80C for further analyses of total lipids, FAs and TEs. 89

90

2.2. Total lipids and fatty acid analysis 91

92

Extraction and determination of lipids and FA composition of the total lipid fraction on muscle and 93

liver was determined in each sampled individual, after tissue homogenization, by fat extraction 94

following the method of Folch et al. (1957), with a mixture of chloroform and methanol (1:1 95

proportion for the first extraction and 2:1 proportion for the second). FA methyl esters (FAME) 96

samples were analysed according to the method of Stoffel et al. (1959) by high-performance liquid 97

chromatography (HPLC). Individual methyl esters were identified by comparison with known 98

standards. The lipid content was expressed as percentage of ash-free dry matter and individual FA 99

concentrations were expressed as percentages of the total FA composition. 100

101

2.3. Trace elements analysis 102

103

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Approximately 1 g of saithe muscle and liver of each individual was subjected to wet mineralization 104

following homogenization using a mixture of nitric acid and hydrogen peroxide (4:1, w/w) to extract 105

TEs from sample matrix through a vessel microwave digestion system. A total of 26 minority elements 106

were analysed through inductively coupled plasma mass spectrometry (ICP-MS): lithium (Li), 107

beryllium (Be), boron (B), aluminium (Al), vanadium (V), chrome (Cr), manganese (Mn), iron (Fe), 108

cobalt (Co), nickel (Ni), copper (Cu), zinc (Zn), gallium (Ga), arsenic (As), selenium (Se), strontium 109

(Sr), molybdenum (Mo), silver (Ag), cadmium (Cd), indium (In), antimony (Sb), barium (Ba), 110

mercury (Hg), tantalum (Tl), plumb (Pb) and bismuth (Bi). In addition, 4 majority TEs were also 111

assessed: calcium (Ca), phosphate (K), magnesium (Mg) and sodium (Na). Each sample was analysed 112

in triplicate. ICP-MS is the routine method of choice for trace elements determination in 113

environmental studies involving fish farms, which allows simultaneous determination of most 114

elements within the periodic table with limits of detection below one part per billion (p.p.b.) (e.g. 115

Campana et al. 1994; Dean et al. 2007). Nevertheless, minority and majority elements were expressed 116

in parts per billion (p.p.b.) and parts per million (p.p.m.) respectively. TEs were quantified on the basis 117

of peak areas and comparison with a calibrated curve obtained with the corresponding standards. 118

119

2.4. Statistical analysis 120

121

Variation in total lipids proportions, specific FAs and TEs on muscle and liver samples between farm-122

associated (A-saithe) and un-associated saithe (U-saithe) were analysed through analysis of variance 123

(ANOVA, 95%). After performing fourth and square root transformations of FAs and TEs 124

respectively, principal components analyses (PCA) were performed as the ordination method of saithe 125

assemblages with the elements that presented significant differences between groups. Moreover, cross-126

validation discriminant analysis (DA) was applied as a classification method of saithe individuals 127

within groups according to the FAs and TEs profiles. Statistical analyses were performed with IBM-128

SPSS Statistics-20 and PRIMER-6 packages. 129

130

131

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3. Results 132

133

3.1. Total lipids and fatty acid profiles 134

135

Livers from farm-aggregated saithe contained higher proportions of total fats compared to those from 136

non-influenced areas, and saithe captured by nets had a higher lipid proportion compared to fish 137

caught by jigging (Figure 2a). There were no differences in lipid content in muscle between farm 138

associated and un-associated saithe, but saithe captured by jigging had higher lipid proportions than 139

those caught with nets (Figure 2b). 140

141

The FA profiles of liver and muscle differed significantly among fishing methods and origin areas, 142

with palmitic (16:0), oleic (OA, 18:1 n-9), eicosapentaenoic (EPA, 20:5 n-3) and docosahexaenoic 143

(DHA, 22:6 n-3) acids being the most abundant FAs in both tissues (Tables 2 and 3). The percentage 144

of total saturated FAs in liver samples was significantly higher in un-associated fish compared to 145

farm-associated saithe. In contrast, proportions of total unsaturated FAs were significantly higher in 146

livers from farm-associated fish compared to un-associated saithe. (Table 2). Within unsaturated FAs, 147

the proportion of total monounsaturated FAs (MUFAs) in liver of un-aggregated saithe samples from 148

hooks (U-hooks) was significantly lower than the other 3 groups (U-net, A-hook, A-net). Conversely, 149

total long-chain polyunsaturated FAs (PUFAs) proportions in saithe liver were significantly higher in 150

liver samples from un-associated jigged fish compared to the other 3 groups. Accordingly, proportions 151

of pentadecanoic (15:0), vaccenic (18:1 n-7), eicosapentaenoic (EPA, 20:5 n-3), adrenic (DTA, 22:4 n-152

6) and docosahexaenoic (DHA, 22:6 n-3) acids were significantly higher in liver from saithe captured 153

by hooks, farm-aggregated groups presented the lowest proportions of these FAs (Figure 3a). 154

However, the proportions of oleic (OA, 18:1 n-9), linoleic (LA, 18:2 n-6) and linolenic (LNA, 18:3 n-155

3) acids were significantly higher in farm-associated saithe liver compared to un-associated fish 156

(Figure 3a). The OA levels in liver were lower in U-hook samples compared to U-net saithe, while n-157

3/n-6 ratio in liver was higher in U-hook group, and the lowest in both farm-aggregated saithe groups 158

(A-net and A-hook) (Table 2). 159

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160

The percentage of total saturated and unsaturated FAs in saithe muscle were significantly higher and 161

lower respectively in U-net samples compared to the other groups (Table 3). Within unsaturated FAs, 162

there were no significant differences in the proportion of MUFAs in muscle between farm-aggregated 163

and un-aggregated saithe groups, with the highest proportions in A-net samples. Moreover, the 164

percentages of PUFAs were significantly higher in saithe captured by hooks from both areas (U-hook 165

and A-hook) compared to those from nets. However, the highest LC-PUFA proportions in saithe 166

muscle were detected in U-hook, while the lowest was observed in A-net group (Table 3). A high 167

variability was found among different groups within each specific FA proportions. Nevertheless, U-168

hook samples presented the lowest proportions in most of the FAs analysed, while A-net samples 169

showed higher proportions. This is the case of OA, LA, LNA, EPA and clupanodonic (DPA) acids, 170

whose proportions in muscle were also similar between U-net and A-line (Figure 3, Table 3). In 171

addition, palmitoleic acid (PA, 16:1), nervonic acid (NEA, 24:1), eicosatrienoic acid (ETE, 20:3 n-3), 172

DTA and DHA proportions were similar between un-aggregated and farm-aggregated saithe captured 173

by nets (U-net and A-net), but significant different from those caught by hooks (U-hook and A-hook) 174

(Figure 3, Table 3). 175

176

A combination of two principal components explained 80.8% of the total variation of FA profiles in 177

liver samples (PC1: 62.9%, PC2: 17.9%; Figure 4a). Both PCs comprised mainly the variations in OA, 178

LA, LNA, DHA and EPA. The two latter and both PCs were negatively correlated (Figure 4a). 179

Furthermore, two principal components explained 82.9% of the total variation in muscle samples 180

(PC1: 60.6%, PC2: 21.5%; Figure 4b). PC1 represented the variations in Myristic acid (MA, 14:0), 181

stearic acid (SA, 18:0), OA, LA, LNA, DHA and EPA, all of them with a negative correlation. 182

Moreover, PC2 represented variations in OA, LA, LNA, MA and myristoleic acid (MYA, 14:1). The 183

two latter and PC2 were negatively correlated (Figure 4b). Discriminant analysis with selected FAs in 184

liver and muscle samples (those with significantly different proportions among groups; see Table 2 185

and 3) showed that 62.5% and 76.9% respectively were correctly classified (Table 4). In addition, 186

around the 69% of un-aggregated saithe were correctly classified from selected FAs profile in liver 187

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and muscle, whereas higher percentages were obtained from selected FAs in farm-aggregated saithe 188

liver (93.7%) and muscle (84.6%)(Table 4). 189

190

3.2. Trace elements composition 191

192

From the analysis of minority TEs in liver, the highest concentrations found were Fe, followed by Zn, 193

As and Cu (Table 5). The concentrations of Fe, Co, As, Se and Mo in liver of un-aggregated liver 194

samples were significantly higher than in samples from farm-aggregated saithe (Figure 5a, Table 5). 195

Moreover, concentrations of Mo, Zn, Cd and Sb in U-hook samples were significantly higher than the 196

other 3 groups (Figure 5a, Table 5). Furthermore, Sr was found in higher concentrations in lovers from 197

U-net fish compared to the other 3 groups. Significant differences in K, Mg and Na were also found 198

between saithe assemblages. Concentrations of K and Mg were higher in un-aggregated compared 199

tofarm-aggregated saithe, and the significantly highest value of Na was found in U-net samples 200

compared to the other 3 groups (Table 5). 201

202

The analysis of minority TEs in muscle samples showed that concentrations of As, followed by Fe, Zn 203

and Sr, were the most predominant elements (Table 6). Significantly higher concentrations of Se and 204

Hg in muscle and lower concentrations of Mn were found in un-aggregated samples compared to 205

farm-aggregated saithe (Figure 5b, Table 6). Although, significant differences in B concentrations 206

were also found between un-aggregated and farm-aggregated muscle samples, no differences were 207

detected among U-net, A-hook and A-net samples (Figure 5b, Table 6). Moreover, Li, Fe, Cu, Bi and 208

the majority element K were found in significantly higher concentrations in U-hook samples compared 209

to U-net, A-hook and A-net samples (Figure 5b, Table 6). In fact, no significant differences were 210

found in these concentrations among the latter 3 groups of samples. No differences were detected in 211

CA and Mg concentrations in muscle among groups (Table 6). 212

213

A combination of 2 principal components explained 94.9% of the total variation of TEs profile in liver 214

samples (PC1: 88.9%, PC2: 6%; Figure 6a). PC1 represented the variations in Fe, AS and Zn 215

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concentrations. However, positive correlations between trace elements and PC1 were not found 216

(Figure 6a). Moreover, PC2 represented variations in Fe, As, Zn and Se concentrations. The former 217

and PC2 were positively correlated (Figure 6a). Moreover, 2 principal components explained 74.2% of 218

the total variation in muscle samples (PC1: 58.6%, PC2: 15.6%; Figure 6b). PC1 comprised the 219

variations in Fe, Al and B concentrations, while PC2 comprised mainly the variations in Hg, B, Al, Fe 220

and Mn concentrations in muscle samples (Figure 6b). Discriminant analysis with selected TEs in liver 221

and muscle samples (those with significantly different proportions among groups; see Table 4 and 5) 222

showed that 51.6% and 65.6% respectively were correctly classified (Table 7). In addition, a total of 223

93.3% and 93.7% of un-aggregated saithe were correctly classified from selected TEs profile in liver 224

and muscle respectively, and similar percentages were obtained from selected TEs in farm-aggregated 225

saithe liver (93.7%) and muscle (87.5%)(Table 7). 226

227

228

4. Discussion 229

230

Variations in total lipids, fatty acids and trace elements profiles in saithe liver and muscle were to a 231

large extent associated with presence at fish farms. Saithe can be found in far higher concentrations 232

immediately beside and beneath salmon cages than just 25200 m distance away from the nearest 233

cage, most likely because they feed on waste fish food (Cromey et al. 2002; Tuya et al. 2006; 234

Dempster et al. 2010;). Hence, wild saithe that normally feed on crustaceans and fish situation (e.g. Du 235

Buit 1991; Carruthers et al. 2005), substitute their natural diet by feed pellets when aggregated at 236

farms. This affects the chemical composition of the fish in a similar way as seen in cultured fish 237

species (Skog et al. 2003; Fernandez-Jover et al. 2011a). 238

239

Following a diet switch from natural preys to salmon pellets, it is reasonable to assume that the high 240

lipid content of the salmon feed will result in a higher fat content of the fish (Lopparelli et al. 2004). In 241

this study, higher fat content in liver tissues was detected in farm-aggregated saithe compared to those 242

individuals captured far from farms. Previous studies on saithe, but also on other farm-aggregated 243

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species, such as cod (Gadus morhua), horse mackerel (Trachurus mediterraneus) and bogue (Boops 244

boops), revealed higher condition indices (i.e. Fulton Condition Index, Liverosomatic index) in farm-245

aggregated fish compared to non-influenced individuals (Skog et al. 2003; Fernandez-Jover et al. 246

2007, 2011b; Arechavala-Lopez et al. 2011). The incorporation and storage of FAs in fish tissues 247

strongly depends on the FA profile of the diet (Sargent et al. 2002). The current practice of 248

substituting fish oils with other vegetable lipid sources in farmed marine fish diets leads to notable 249

changes in lipid composition and FA profiles in fish tissues (Fernandez-Jover et al. 2011a). In fact, 250

some studies have demonstrated that wild saithe feeding around Norwegian salmon farms had liver 251

and muscle FA profiles similar to the feed pellets used at the farm (Skog et al. 2003; Fernandez-Jover 252

et al. 2011b). Accordingly, the present study demonstrated that the presence of FAs of terrestrial origin 253

in high proportions in liver and muscle tissues, such as OA, LA and LNA indicate that these fish have 254

been feeding in the vicinity of the farms. In addition, this study also show that low proportions of 255

n3/n6 ratio or EPA and DHA, main responsible of long-chain n-3 PUFAs enrichment in fish lipids 256

(Fernandez-Jover et al. 2011a), also are indicators of residence at fish farms. Therefore, variation in 257

dietary FA profiles or specific FAs can be widely used to detect the influence of farming activity on 258

wild saithe populations. 259

260

Furthermore, and in agreement with previous studies, it was demonstrated that influence of fish farms 261

on wild saithe populations is also reflected by variation in TEs signatures in muscle and liver tissues 262

(Bustnes et al. 2011). Saithe captured far from farms could be seen as representing a higher trophic 263

level as they feed on a wide variability of food-prey items, as compared to farm-aggregated saithe 264

which usually feed on pellets in high abundance (Fernandez-Jover et al. 2011a). Thus, the higher 265

levels of TEs found in their liver and muscle compare to farm-aggregated saithe might be a result of 266

accumulation of TEs from natural prey (e.g. As, Se, Zn, Hg, Fe, etc.). Accordingly, farm-aggregated 267

saithe might have reduced their trophic level, and consequently also their accumulation of TEs, not 268

only due to the direct feeding on pellets, but also due to aquaculture related deposition of elements in 269

the vicinity of the farms (Solberg et al. 2002; Bustnes et al. 2011). However, some discrepancies with 270

previous studies were found for Hg levels in saithe. Bustnes et al. (2011) suggested that salmon feed 271

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was responsible for elevated Hg levels found in farm-aggregated saithe, whereas our results show Hg 272

to be higher in muscle and liver for un-aggregated fish. The lower Fe levels in liver samples from 273

farm-aggregated saithe might also be explained by the current practice of reducing the Fe 274

supplementation of commercial salmon feeds, being sometimes even absent (Lorentzen and Maage 275

1999), since farmed Atlantic salmon have a limited capacity to regulate Fe absorption causing winter 276

ulcers (Salte et al. 1994). Our results show that concentrations of specific TEs (e.g. Fe, Mo and Se in 277

liver, and Hg, B and Se in muscle), might be useful to differentiate between farm-aggregated and un-278

aggregated saithe, but that differences between groups may change over time, perhaps due to changes 279

of TEs in natural prey or in waste pellets. 280

281

In addition to fish origin, FAs and TEs varied between fishing methods. This was a surprising result 282

as both methods were used on the same locations. One possible explanation may be that the different 283

fishing gear select for different sub-populations. The nets were deployed at the bottom on 284

approximately 100-150 meter depth, while jigging took place in the middle of water column. 285

Differences between fishing methods could reflect differences between benthic and pelagic sub-286

populations that may have different diets and perhaps also life histories. Saithe is believed to be a 287

seasonally migratory species (e.g. Bjordal and Skar 1992; Bjordal and Johnstone 1993; Neilson et al. 288

2002, 2003; Armannsson et al. 2007; Homrum et al. 2012, 2013), and movements between the control 289

area and the farms are not unrealistic (Uglem et al. 2009). However, the low number of fish examined 290

in this study could cause some of the variability observed, but also supports the low proportions of fish 291

correctly classified within the 4 studied groups through metabolites profiles. However, a similar 292

previous size study indicated that over 90% of saithe could be classified correctly from fatty acid 293

profile with higher sample, regardless differences in capture methods (Fernandez-Jover et al. 2011a). 294

Thus, further studies taking into account seasonal variability, fishing characteristics and higher sample 295

sizes in different areas are required to increase the accuracy of origin discrimination. 296

297

In sum, the results reported in the present study suggest that variation in specific metabolic elements 298

might have a potential for being used to detect if specific fish have inhabited the surrounding of fish 299

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farms for a certain period of time. However, how long these farm-characteristic parameters persist in 300

the fish body, which will influence their accuracy, remains unknown. Also, the FA and TE content in 301

both natural prey and waste feed may vary in time and space, and it is thus important to correlate 302

variations in FAs and TEs with e.g. potential recent changes fish feed formulation when using such 303

substances for examining farm influence on wild fish. Moreover, little is known about the potential 304

consequences of various FAs and TEs in terms of changed fitness, and food quality of this targeted 305

species. However, the possibility of the existence of several ecologically different saithe assemblages 306

within a meta-population, with different feeding activities or movement patterns, involve that 307

prediction of origin and potential effects due to variations in FAs and TEs might more complex than 308

previously assumed. 309

310

311

5. Acknowledgements 312

313

This study was part of the project Evaluation of actions to promote sustainable coexistence between 314

salmon culture and coastal fisheries ProCoEx funded by The Norwegian Seafood Research Fund 315

FHF. The study was also supported by the Norwegian Research council through the EcoCoast project. 316

317

318

6. References 319

320

Andersen, N.G. 2001. A gastric evacuation model for three predatory gadoids and implications of 321

using pooled field data of stomach contents to estimate food rations. Journal of Fish Biology 59: 322

11981217. 323

324

Arechavala-Lopez, P., Sanchez-Jerez, P., Bayle-Sempere, J., Fernandez-Jover, D., Martinez-Rubio, L., 325

Lopez-Jimenez, J.A. and Martinez-Lopez, F.J. 2011. Direct interaction between wild fish aggregations 326

Page 12 of 32




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at fish farms and fisheries activity at fishing grounds: a case study with Boops boops. Aquaculture 327

Research 42: 115. 328

329

Armannsson, H., Jonsson, S. Th., Marteinsdottir, G. and Neilson, J.D. 2007. Distribution and 330

migration of saithe (Pollachius virens) around Iceland inferred from markrecapture studies. ICES 331

Journal of Marine Science 64: 10061016. 332

333

Bell, J.G., McEvoy, J., Tocher, D.R., McGhee, F., Campbell, P.J. and Sargent, J.R. 2001. Replacement 334

of fish oil with rape seed oil in diets of Atlantic salmon (Salmo salar) affects tissue lipid compositions 335

and hepatocyte fatty acid metabolism. Journal of Nutrition 131: 15351543. 336

337

Bell, J.G., McGhee, F., Campbell, P.J. and Sargent, J.R. 2003. Rapeseed oil as an alternative to marine 338

fish oil in diets of post-smolt Atlantic salmon (Salmo salar): changes in flesh fatty acid composition 339

and effectiveness of subsequent fish oil wash out. Aquaculture 218: 515528. 340

341

Bjordal, . and Johnstone A.D.F. 1993. Local movements of saithe (Pollachius virens L.) in the 342

vicinity of fish cages. ICES Marine Science Symposia 196: 143-146. 343

344

Bjordal, . and Skar, A.B. 1992. Tagging of saithe (Pollachius virens L.) at a Norwegian fish farm: 345

preliminary results on migration. ICES Council Meeting Papers 1992/G:35. 346

347

Braithwaite, R. and McEvoy, L.A. 2005. Marine biofouling on fish farms and its remediation. 348

Advances in Marine Biology 47: 215-252. 349

350

Braithwaite, R., Cadavid Carrascosa, M.C. and McEvoy, L.A. 2007. Biofouling of salmon cage 351

netting and the efficacy of a typical copper-based antifoulant. Aquaculture 262 (2-4): 219-226. 352

353

Page 13 of 32




14

Brooks, K.M. and Mahnken, C.V.W. 2003. Interactions of Atlantic salmon in the Pacific Northwest 354

environment: III. Accumulation of zinc and copper. Fisheries Research 62: 295305. 355

356

Bustnes, J.O., Herske, D., Dempster, T., Bjrn, P.A., Nygrd, T., Lie, E. and Uglem, I. 2010. Salmon 357

farms as a source of organohalogenated contaminants in wild fish. Environmental Science and 358

Technology 44 (22): 8736-8743. 359

360

Bustnes, J.O., Nygrd, T., Dempster, T., Ciesielski, T., Jensen, B.M., Bjrn, P.A. and Uglem, I., 2011. 361

Do salmon farms increase the concentrations of mercury and other elements in wild fish? Journal of 362

Environmental Monitoring 13: 1687-1694. 363

364

Campana, S.E., Fowler, A.J. and Jones, C.M. 1994. Otolith elemental fingerprinting for stock 365

identification of Atlantic cod (Gadus morhua) using laser ablation ICPMS. Canadian Journal of 366

Fisheries and Aquatic Sciences 51: 1942-1950. 367

368

Carruthers, E.H., Neilson, J.D., Waters, C. and Perley, P. 2005. Long-term changes in the feeding of 369

Pollachius virens on the Scotian Shelf: responses to a dynamic ecosystem. Journal of Fish Biology 66: 370

327347. 371

372

Carss D.N. 1990. Concentrations of wild and escaped fishes immediately adjacent to fish farms. 373

Aquaculture 90: 29-40. 374

375

CIESM 2007. Impact of mariculture on coastal ecosystems. CIESM Whorkshop Monographs, 32. 376

Mnaco. 120 pp. 377

378

Cromey, C.J., Nickell, T.D. and Black, K.D. 2002. DEPOMOD - modelling the deposition and 379

biological effects of waste solids from marine cage farms. Aquaculture 214 (14): 211239. 380

381

Page 14 of 32




15

Dean, R.J., Shimmield, T.M. and Black, K.D. 2007. Copper, zinc and cadmium in marine cage fish 382

farm sediments: An extensive survey. Environmental Pollution 145: 84-95. 383

384

Dempster, T., Sanchez-Jerez, P., Uglem, I. And Bjrn, P.-A. 2010. Species-specific patterns of 385

aggregation of wild fish around fish farms. Estuarine Coastal and Shelf Science 86: 271-275. 386

387

Dempster, T., Sanchez-Jerez, P., Fernandez-Jover, D., Bayle-Sempere, J.T., Uglem, I., Nilsen, R. and 388

Bjrn, P.-A. 2011. Proxy measures of fitness suggest coastal fish farms can act as population sources 389

and not ecological traps for wild gadoid fish. PLoS ONE 6(1): e15646. doi:10.1371/ 390

journal.pone.0015646. 391

392

Dempster T. and Taquet M. 2004. Fish aggregation device (FAD) research: gaps in current knowledge 393

and future directions for ecological studies. Reviews in Fish Biology and Fisheries 14(1): 21 42. 394

395

Dempster, T., Uglem, I., Sanchez-Jerez, P., Fernandez-Jover, D., Bayle-Sempere, J., Nilsen, R. and 396

Bjrn, P.A. 2009. Coastal salmon farms attract large and persistent aggregations of wild fish: an 397

ecosystem effect. Marine Ecology Progress Series 385: 1-14. 398

399

Du Buit, M.H. 1991. Food and feeding of saithe (Pollachius virens L) of Scotland. Fisheries Research 400

12: 307323. 401

402

Fernandez-Jover, D., Lopez-Jimenez, J.A., Sanchez-Jerez, P., Bayle-Sempere, J., Gimenez-403

Casalduero, F., Martinez-Lopez, F.J. and Dempster, T. 2007. Changes in body condition and fatty acid 404

composition of wild Mediterranean horse mackerel (Trachurus mediterraneus, Steindachner, 1868) 405

associated with sea cage fish farms. Marine Environmental Research 63: 1-18. 406

407

Fernandez-Jover, D., Arechavala-Lopez, P., Martinez-Rubio, L., Tocher, D.R., Bayle-Sempere, J.T., 408

Lopez-Jimenez, J.A., Martinez-Lopez, F.J. and Sanchez-Jerez, P. 2011a. Monitoring the influence of 409

Page 15 of 32




16

marine aquaculture on wild fish communities: benefits and limitations of fatty acid profiles. 410

Aquaculture Environment Interactions 2: 39-47. 411

412

Fernandez-Jover, D., Martinez-Rubio, L., Sanchez-Jerez, P., Bayle-Sempere, J.T., Lopez Jimenez, 413

J.A., Martnez Lopez, F.J., Bjrn, P.-A., Uglem, I. and Dempster, T. 2011b. Waste feed from coastal 414

fish farms: a trophic subsidy with compositional side-effects for wild gadoids. Estuarine Coastal and 415

Shelf Science 91: 559568. 416

417

Folch, J., Lees, M. and Stanley, G.A. 1957. A simple method for the isolation and purification of total 418

lipids from animal tissues. Journal of Biological Chemistry 226: 497-509. 419

420

Homrum, E. , Hansen, B., Steingrund, P., and Htn, H. 2012. Growth, maturation, diet and 421

distribution of saithe (Pollachius virens) in Faroese waters (NE Atlantic). Marine Biology Research 8: 422

246254. 423

424

Homrum, E. , Hansen, B., Jnsson, S., Michalsen, K., Burgos, J., Righton, D., Steingrund, P., 425

Jakobsen, T., Mouritsen, R., Htn, H., Armannsson, H. and Joensen, J.S. 2013. Migration of saithe 426

(Pollachius virens) in the Northeast Atlantic. ICES Journal of Marine Science 427

doi:10.1093/icesjms/fst048. 428

429

Lopparelli, R.M., Segato, S., Corato, A., Fasolato, L. and Andrighetto, I. 2004. Sensory evaluation of 430

sea bass (Dicentrarchus labrax L.) fed two diets differing in fat content. Veterinary Research 431

Communications 28: 225227. 432

433

Lorentzen, M. and Maage, A. 1999. Trace element status of juvenile Atlantic salmon Salmo salar L. 434

fed a fish-meal based diet with or without supplementation of zinc, iron, manganese and copper from 435

first feeding. Aquaculture Nutrition 5: 163-171. 436

437

Page 16 of 32




17

Neilson, J. D., Annis, L., Perley, P., Clay, A. and OConnor, M. 2002. Seasonal aggregations of 438

Canadian east coast pollock as inferred from the commercial fishery and hydroacoustic observations. 439

Journal of Fish Biology 61: 10671084. 440

441

Neilson, J. D., Clark, D., Melvin, G. D., Perley, P. and Stevens, S. 2003. The diel vertical distribution 442

and characteristics of pre-spawning aggregations of pollock (Pollachius virens) as inferred from 443

hydroacoustic observations: the implications for survey design. ICES Journal of Marine Science 60: 444

860871. 445

446

Otter, H., Karlsen, , Slinde, E. and Olsen, R.E. 2009. Quality of wild-captured saithe (Pollachius 447

virens L.) fed formulated diets for 8 months. Aquaculture Research 40: 1310-1319. 448

449

Sanchez-Jerez, P., Dempster, T., Fernandez-Jover, D., Uglem, I., Bayle-Sempere, J., Bjrn, P.A., 450

Arechavala-Lopez, P., Valle, C. and Nilsen, R. 2011. Coastal fish farms as Fish Aggregation Devices 451

(FADs): potential effects on fisheries. In: Bortone, S. (Ed.), Artificial Reefs in Fisheries Management. 452

Taylor and Francis. 453

454

Salte, R., Rrvik, K.A., Reed, E. and Nordberg, K. 1994. Winter ulcers of the skin in Atlantic salmon, 455

Salmo salar L. Pathogenesis and possible aetiology. Journal of Fish Diseases 17: 661-665. 456

457

Sargent, J., Tocher, D. and Bell, G. 2002. The lipids. In: Halver, J.E., Hardy, R.W. (Eds.), Fish 458

Nutrition. Elsevier Academic Press, U.S.A., pp. 181-257. 459

460

Skog, T.E., Hylland, K., Torstensen, B.E. and Berntssen, M.H.G. 2003. Salmon farming affects the 461

fatty acid composition and taste of wild saithe Pollachius virens L. Aquaculture Research 34: 999-462

1007. 463

464

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Solberg, C.B., Sthre, L. and Julshamn, K. 2002. The effect of copper-treated net pens on farmed 465

salmon (Salmo salar) and other marine organisms and sediments. Marine Pollution Bulletin 45: 126466

132. 467

468

Stoffel, W., Chu, F. and Edward, H. 1959. Analysis of long-chain fatty acids by gaseliquid 469

chromatography. Micromethod for preparation of methyl esters. Analytical Chemistry 31: 307-308. 470

471

Tuya, F., Sanchez-Jerez, P., Dempster, T., Boyra, A. and Haroun, R. 2006. Changes in demersal wild 472

fish aggregations beneath a sea-cage fish farm after the cessation of farming. Journal of Fish Biology 473

69: 682-697. 474

475

Uglem, I., Dempster, T., Bjorn, P.A. and SanchezJerez, P. 2009. High connectivity of salmon farms 476

revealed by aggregation, residence and repeated migrations of wild saithe (Pollachius virens) among 477

farms. Marine Ecology Progress Series 384: 251260. 478

479

Uglem, I., Knutsen, ., Kjesbu, O.S., Hansen, .J., Mork, J., Bjrn, P.A., Varne, R., Nilsen, R., 480

Ellingsen, I. and Dempster, T. 2012. Extent and ecological importance of escape through spawning in 481

sea-cages for Atlantic cod. Aquaculture Environment Interactions 3: 3349. 482

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19

FIGURE CAPTIONS

Figure 1. Map of the study area around Hitra Island, Norway. Salmon farming and control sampling

areas are shown with a black spot.

Figure 2. Total fats (percentage) in the liver and muscle of un-aggregated (U) and farm-aggregated (A)

saithe captured by hooks and nets. Whiskers show standard error. Different lowercase letters means

significant differences (p0.2 are plotted.

Figure 5. Proportion of selected trace elements in liver (A) and muscle (B) of saithe among the four

studied groups. Whiskers show standard error.

Figure 6. Principal components analysis of those minority elements on saithe liver and muscle samples

(square roots transformed) that showed significant differences between fish groups. Saithe from

control areas in grey, farm-aggregated saithe in black; circles mean long-line captures and triangles

mean trammel-net captures. Only vectors with a correlation >0.1 are plotted.

Page 19 of 32




20

TABLES

Table 1. Characteristics (number, length and weight) of sampled saithe for each treatment, location

and fishing gear used in this study. SE: standard error.

Locality Treatment Gear Code N Length SE (mm) Weight SE (g)

Control Un-aggregated Hooks U-hook 8 693.7 34.3 2466.2 271.9

Nets U-net 8 635.0 37.5 2485.0 468.9

664.4 25.7 2475.6 261.8

Farm Aggregated Hooks A-hook 8 698.7 17.5 3569.1 286.3

Nets A-net 8 620.0 20.7 2662.5 224.3

659.4 16.6 3115.8 211.1

Page 20 of 32




21

Table 2. Proportions of total fatty acids in liver of un-aggregated and farm-aggregated saithe

assemblages. Data are expressed as mean SE (standard error); nd: non detected. Significance level:

*0.05, **0.01. Means in the same row with different superscript letters are significantly different.

LIVER Un-Aggregated Farm-Aggregated

Fatty Acids U-hook U-net A-hook A-net P-value

14:0 3.07 0.17 2.98 0.55 2.46 0.22 2.42 0.18 0.352

15:0 0.41 0.02a 0.23 0.03

b 0.23 0.01

b 0.14 0.04

b 0.001**

16:0 11.91 0.81ab

12.75 0.91a 10.64 0.33

ab 10.14 0.47

b 0.042*

17:0 0.22 0.03 0.14 0.02 0.18 0.02 0.17 0.01 0.12

18:0 3.88 0.32 3.87 0.312 3.91 0.18 3.91 0.23 0.999

20:0 0.13 0.01a 0.20 0.02

a 0.17 0.01

a 0.28 0.03

b 0.001**

22:0 0.12 0.01 0.27 0.22 0.06 0.01 0.02 0.01 0.411

24:0 0.06 0.01 0.03 0.02 0.03 0.01 0.01 0.0 0.126

Total Saturated 28.13 0.82a 25.99 1.28

a 21.09 1.02

b 21.86 1.11

b 0.001**

14:1 0.01 0.01 0.01 0.01 0.01 0.01 0.01 0.01 0.606

15:1 nd 0.01 0.01 Nd nd 0.037*

16:1 5.60 0.31a 4.81 0.28

a 3.79 0.27

b 3.81 0.27

b 0.001**

17:1 0.34 0.02ab

0.12 0.05c 0.41 0.02

b 0.23 0.06

ac 0.001**

18:1 n-7 5.66 0.36a 4.47 0.41

b 4.21 0.17

b 4.39 0.23

b 0.010**

18:1 n-9 11.29 0.58a 18.92 2.66

b 27.68 2.32

c 23.36 2.01

bc 0.001**

20:1 6.22 0.27 6.92 1.19 5.08 0.70 5.06 0.50 0.243

22:1 n-9 0.55 0.09a 0.15 0.04

b 0.53 0.06

a 0.64 0.11

a 0.001**

24:1 0.61 0.04a 0.64 0.08

a 0.41 0.04

b 0.39 0.05

b 0.005**

Total MUFA 41.23 0.88a 44.53 1.83

b 48.38 0.84

b 46.69 0.59

b 0.001**

18:2 n-6 1.03 0.16a 2.50 1.22

a 8.86 1.03

b 7.78 0.96

b 0.001**

18:3 n-3 0.76 0.10a 1.13 0.33

a 3.36 0.43

b 3.01 0.39

b 0.001**

20:2 1.47 0.11 1.04 0.18 1.42 0.09 1.47 0.08 0.065

20:3 n-3 0.05 0.02 0.08 0.02 0.02 0.01 0.05 0.02 0.162

20:4 n-6 0.63 0.03 0.77 0.27 0.37 0.02 0.36 0.02 0.122

20:5 n-3 8.34 0.52a 5.89 0.44

b 5.13 0.36

b 5.23 0.27

b 0.001**

22:2 0.42 0.01 0.35 0.02 0.43 0.03 0.45 0.03 0.076

22:4 n-6 0.37 0.01a 0.27 0.03

b 0.26 0.02

b 0.26 0.022

b 0.004**

22:5 n-3 0.77 0.03 0.79 0.06 0.87 0.06 0.82 0.08 0.65

22:6 n-3 9.45 0.44a 7.87 0.71

b 6.28 0.43

b 6.50 0.45

b 0.001**

Total LC-PUFA 29.27 1.07a 21.21 1.39

b 17.12 1.31

b 18.71 1.01

b 0.001**

Total PUFA 31.80 1.09a 25.64 1.19

b 31.05 0.51

a 31.79 0.92

a 0.001**

Total Unsaturated 73.03 0.82a 70.17 1.8

a 79.43 0.98

b 78.48 1.09

b 0.001**

n-3/n-6 1.68 0.03a 1.41 0.09

b 1.01 0.05

c 1.06 0.05

c 0.001**

Page 21 of 32




22

Table 3. Proportions of total fatty acids in muscle of un-aggregated and farm-aggregated saithe

assemblages. Data are expressed as mean SE; nd: non detected. Significance un-aggregated and

farm-aggregated level: *0.05, **0.01. Means in the same row with different superscript letters are

significantly different.

MUSCLE Un-Aggregated Farm-Aggregated Fatty Acids U-hook U-net A-hook A-net P-value

14:0 0.12 0.02a 1.52 0.15

b 0.17 0.05

b 1.37 0.43

a 0.003**

15:0 nd 0.01 0.01 nd nd 0.225

16:0 2.92 0.41a 6.18 0.21

b 5.48 0.79

b 7.23 0.85

b 0.001**

17:0 0.01 0.01 0.05 0.03 0.01 0.01 0.03 0.01 0.335

18:0 0.93 0.13a 3.13 0.13

bc 1.91 0.21

ab 3.93 0.64

c 0.001**

20:0 0.01 0.01a 0.01 0.01

ab 0.01 0.01

ab 0.03 0.01

b 0.025*

22:0 nd 0.01 0.01 nd nd 0.548

24:0 0.01 0.01 nd 0.01 0.01 0 0 0.308

Total Saturated 37.55 0.99a 41.79 1.31

b 34.94 0.31

a 36.93 1.17

a 0.003**

14:1 nd 0.03 0.01 nd 0.01 0.01 0.047*

15:1 nd 0.01 0.01 nd 0.01 0.01 0.451

16:1 0.15 0.03a 0.56 0.04

b 0.18 0.05

a 0.67 0.09

b 0.001**

17:1 0.03 0.01 0.27 0.05 0.02 0.01 0.46 0.19 0.070

18:1 n-7 0.99 0.21a 1.49 0.06

ab 1.24 0.19

ab 1.68 0.20

b 0.052

18:1 n-9 0.87 0.17a 2.39 0.24

b 3.18 0.35

b 5.21 0.67

c 0.001**

20:1 0.24 0.05 0.55 0.08 0.31 0.07 0.57 0.10 0.036

22:1 n-9 0.01 0.01 0.04 0.01 0.01 0.01 0.03 0.01 0.075

24:1 0.07 0.01a 0.22 0.03

b 0.12 0.03

ab 0.20 0.02

b 0.007**

Total MUFA 15.78 0.85a 16.21 0.43

a 18.06 0.63

a 20.24 0.70

b 0.001**

18:2 n-6 0.11 0.02

a 0.51 0.22

ab 1.21 0.16

bc 1.63 0.29

c 0.001**

18:3 n-3 0.04 0.01a 0.11 0.02

a 0.19 0.02

a 0.39 0.07

b 0.001**

20:2 0.04 0.01a 0.09 0.01

ab 0.09 0.03

ab 0.18 0.05

b 0.042*

20:3 n-3 0.01 0.01a 0.13 0.02

b 0.01 0.01

a 0.15 0.03

b 0.001**

20:4 n-6 0.36 0.05 0.79 0.13 0.60 0.09 0.79 0.10 0.052

20:5 n-3 1.91 0.32a 3.61 0.13

b 3.25 0.53

b 4.68 0.58

b 0.003**

22:2 0.05 0.01a 0.11 0.01

a 0.09 0.02

a 0.17 0.02

b 0.002**

22:4 n-6 0.03 0.01a 0.06 0.01

ab 0.04 0.01

a 0.08 0.02

b 0.020**

22:5 n-3 0.15 0.05a 0.43 0.03

b 0.45 0.04

b 0.71 0.10

c 0.001**

22:6 n-3 5.76 0.91a 12.04 0.60

b 9.72 1.23

b 12.91 1.50

b 0.002**

Total LC-PUFA 55.92 0.47a 49.89 0.75

b 49.91 0.70

b 45.20 1.15

c 0.001**

Total PUFA 56.94 0.43a 51.63 0.91

b 55.11 0.32

a 50.96 0.84

b 0.001**

Total Unsaturated 72.72 0.68

a 67.84 0.73

b 73.17 0.55

a 71.20 0.55

a 0.001**

n-3/n-6 1.25 0.01a 1.21 0.01

ab 1.16 0.03

b 1.18 0.01

b 0.024*

Page 22 of 32




23

Table 4. Classification through cross-validation (DA) of saithe groups according to selected liver and

muscle FAs (p


24

Table 5. Minority and majority trace elements in liver of un-aggregated and farm-aggregated saithe

assemblages. Data are expressed as mean SE. Significance level: *0.05, **0.01. nd: non detected.

Means in the same row with different superscript letters are significantly different.

Un-Aggregated Farm-Aggregated

TEs U-hook U-net A-hook A-net p-value

Li 0.028 0.012 0.021 0.018 nd nd 0.183

Be 0.001 0.001 0.002 0.001 nd 0.001 0.001 0.423

B 0.215 0.124 0.112 0.087 nd nd 0.159

Al 2.371 0.376 2.345 0.288 1.880 0.181 2.754 0.412 0.328

V 0.243 0.0753 0.142 0.089 0.072 0.024 0.053 0.018 0.133

Cr 0.128 0.0213 0.134 0.022 0.111 0.005 0.142 0.020 0.676

Mn 1.615 0.124 1.562 0.402 1.184 0.164 1.649 0.387 0.669

Fe 147.737 31.871a 142.118 46.619a 43.481 11.170b 35.080 5.335b 0.011**

Co 0.125 0.0169a 0.084 0.0234a 0.026 0.005b 0.023 0.004b 0.001**

Ni 0.145 0.022 0.108 0.0225 0.074 0.0173 0.075 0.014 0.043*

Cu 17.630 2.156 10.203 1.670 12.106 1.784 11.627 3.293 0.140

Zn 46.705 3.561a 35.286 4.955ab 27.159 2.912b 26.434 5.071b 0.007**

Ga 0.065 0.022 0.066 0.021 0.101 0.015 0.139 0.027 0.080

As 20.850 3.331a 18.110 4.615a 4.988 0.772b 4.851 0.596b 0.001**

Se 2.625 0.239a 2.243 0.384a 0.555 0.089b 0.486 0.055b 0.001**

Sr 0.718 0.0867a 1.119 0.217b 0.342 0.056a 0.492 0.134a 0.003**

Mo 0.469 0.054a 0.298 0.079b 0.116 0.021c 0.102 0.011c 0.001**

Ag 0.125 0.021 0.122 0.015 0.135 0.018 0.105 0.011 0.664

Cd 1.017 0.186a 0.284 0.109b 0.148 0.043b 0.073 0.012b 0.001**

In 0.001 0.001 0.001 0.001 0.012 0.006 0.005 0.001 0.057

Sb 0.005 0.002a ndb 0.001 0.001b 0.001 0.001b 0.001**

Ba 0.202 0.069 0.198 0.0619 0.306 0.042 0.425 0.085 0.073

Hg 0.012 0.011 0.070 0.052 nd nd 0.214

Tl 0.003 0.001 0.002 0.001 0.007 0.003 0.003 0.001 0.127

Pb 0.251 0.129 0.317 0.131 0.104 0.014 0.314 0.116 0.488

Bi 0.004 0.001 0.003 0.001 0.057 0.041 0.007 0.001 0.222

Ca 40.33 5.70 31.09 8.26 21.24 7.51 32.75 13.6 0.549

K 1459.03 152.76a 1150.25 156.93ab 835.08 106.05b 841.49 91.52b 0.005**

Mg 111.37 14.89a 115.20 23.07a 43.13 6.19b 44.67 4.86b 0.001**

Na 850.97 94.82a 1248.89 215.52b 337.41 55.94c 516.87 59.64ac 0.001**

Page 24 of 32




25

Table 6. Minority and majority trace elements in muscle of un-aggregated and farm-aggregated saithe

assemblages. Data are expressed as mean SE. Significance level: *0.05, **0.01. nd: non detected.

Means in the same row with different superscript letters are significantly different.

Un-Aggregated Farm-Aggregated

TEs U-hook U-net A-hook A-net p-value

Li 0.148 0.035a 0.025 0.009b 0.009 0.009b 0.014 0.006b 0.001**

Be nd nd nd 0.001 0.001 0.132

B 1.267 0.296a 0.776 0.199ab 0.241 0.139b 0.382 0.1256b 0.006**

Al 1.810 0.285a 0.695 0.270ab 0.786 0.282ab 0.591 0.120b 0.045*

V 0.022 0.013 0.003 0.001 0.008 0.002 0.009 0.004 0.275

Cr 0.227 0.122 0.007 0.004 0.015 0.006 0.308 0.235 0.290

Mn 0.458 0.0560a 0.455 0.064a 0.722 0.091ab 0.856 0.115b 0.004**

Fe 16.046 3.934a 7.441 1.026b 5.935 0.601b 9.516 1.782b 0.018*

Co 0.006 0.001 0.002 0.001 0.002 0.001 0.005 0.001 0.120

Ni 0.026 0.012 0.055 0.0321 0.002 0.001 0.119 0.105 0.491

Cu 1.457 0.114a 0.820 0.079b 0.885 0.081b 0.804 0.032b 0.001**

Zn 15.812 0.698 14.181 0.567 14.961 0.538 17.163 1.567 0.164

Ga 0.058 0.015 0.020 0.0165 0.036 0.016 0.021 0.005 0.225

As 18.746 2.002 26.708 11.903 8.137 1.436 6.222 0.645 0.082

Se 1.467 0.053a 1.527 0.082a 0.975 0.052b 0.977 0.032b 0.001**

Sr 2.927 0.460 2.123 0.462 1.731 0.430 1.762 0.171 0.147

Mo 0.016 0.002 0.006 0.001 0.006 0.001 0.013 0.007 0.216

Ag 0.073 0.009 0.065 0.012 0.080 0.016 0.071 0.019 0.936

Cd 0.007 0.001 0.005 0.001 0.003 0.001 0.019 0.010 0.138

In 0.006 0.001 0.007 0.001 0.002 0.001 0.006 0.004 0.266

Sb 0.021 0.002 0.005 0.001 0.004 0.001 0.009 0.004 0.001**

Ba 0.384 0.052 0.202 0.065 0.322 0.065 0.223 0.024 0.087

Hg 0.664 0.123a 0.761 0.201a 0.217 0.052b 0.111 0.014b 0.001**

Tl 0.010 0.007 0.003 0.001 0.002 0.001 0.004 0.002 0.395

Pb 0.194 0.056 0.121 0.035 0.185 0.033 0.062 0.005 0.089

Bi 0.107 0.039a 0.012 0.001b 0.011 0.002b 0.039 0.028ab 0.030*

Ca 329.87 43.52 245.56 20.20 282.63 38.90 284.08 22.47 0.364

K 11461.63 316.18a 10308.01 445.01b 10625.93 88.31b 9791.27 100.83b 0.002**

Mg 955.38 41.32 907.93 29.71 846.18 24.49 846.55 15.63 0.036*

Na 2335.16 412.16ab 2573.87 472.34a 1184.21 148.57b 2032.29 226.18ab 0.040*

Page 25 of 32




26

Table 7. Classification through cross-validation (DA) of saithe groups according to selected liver and

muscle TEs (p


Figure 1. Map of the study area around Hitra Island, Norway. Salmon farming and control sampling areas are shown with a black spot. 240x129mm (300 x 300 DPI)

Page 27 of 32




Figure 2. Total fats (percentage) in the liver and muscle of un-aggregated (U) and farm-aggregated (A) saithe captured by hooks and nets. Whiskers show standard error. Different lowercase letters means

significant differences (p


Figure 3. Proportion of selected fatty acids in liver (A) and muscle (B) of saithe among the four studied groups. Whiskers show standard error.

149x185mm (300 x 300 DPI)

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Figure 4. Principal components analysis of those fatty acids on saithe liver and muscle samples (fourth roots transformed) that showed significant differences between fish groups. Saithe from control areas in grey,

farm-aggregated saithe in black; circles mean long-line captures and triangles mean trammel-net captures.

Only vectors with a correlation >0.2 are plotted. 254x321mm (96 x 96 DPI)

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Figure 5. Proportion of selected trace elements in liver (A) and muscle (B) of saithe among the four studied groups. Whiskers show standard error.

166x190mm (300 x 300 DPI)

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Figure 6. Principal components analysis of those minority elements on saithe liver and muscle samples (square roots transformed) that showed significant differences between fish groups. Saithe from control

areas in grey, farm-aggregated saithe in black; circles mean long-line captures and triangles mean trammel-

net captures. Only vectors with a correlation >0.1 are plotted. 254x366mm (96 x 96 DPI)

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