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Partridge, G.J. and Lymbery, A.J. (2009) Effects of manganese on juvenile mulloway (Argyrosomusjaponicus) cultured in water with varying salinity-Implications for inland mariculture. Aquaculture, 290

(3-4). pp. 311-316.

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Effects of manganese on juvenile mulloway (Argyrosomus japonicus) culturedin water with varying salinity – implications for inland mariculture

G.J. Partridge, A.J. Lymbery

PII: S0044-8486(09)00149-5DOI: doi: 10.1016/j.aquaculture.2009.02.020Reference: AQUA 628641

To appear in: Aquaculture

Received date: 27 March 2008Revised date: 9 February 2009Accepted date: 13 February 2009

Please cite this article as: Partridge, G.J., Lymbery, A.J., Effects of manganese on juve-nile mulloway (Argyrosomus japonicus) cultured in water with varying salinity – implica-tions for inland mariculture, Aquaculture (2009), doi: 10.1016/j.aquaculture.2009.02.020

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Effects of manganese on juvenile mulloway (Argyrosomus japonicus) cultured in 1

water with varying salinity – implications for inland mariculture.2

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G. J. Partridge1,2,* and A. J. Lymbery25

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1. Aquaculture Development Unit, Challenger TAFE, Fremantle, Western 7

Australia.8

2. Fish Health Unit, Centre for Fish and Fisheries Research, Murdoch University, 9

Murdoch, Western Australia.10

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* Corresponding author: Telephone: +61 8 9239 803213

Facsimile: +61 8 9239 808114

Email: gavin.partridge@challengertafe.wa.edu.au15

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Abstract22

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The effects of 5 mg/L of dissolved manganese on juvenile mulloway at salinities of 5, 24

15 and 45 ppt were determined by comparing their survival, growth and blood plasma25

and organ chemistry with those of fish grown at the same salinities without 26

manganese addition. Survival of mulloway at 45 ppt in the presence of 5 mg/L of 27

manganese (73 ± 13%) was significantly lower than all other treatments, which 28

achieved 100% survival. Those fish grown in water without manganese exhibited 29

rapid growth, which was not affected by salinity (SGR = 4.05 ± 0.29%/day). Those 30

fish grown at 5 ppt and 45 ppt in the presence of manganese lost weight over the two 31

week trial (SGR -0.17 ± 0.42 and -0.44 ± 0.83%/day, respectively), whilst those at 15 32

ppt gained some weight (SGR 1.70 ± 0.20%/day). Manganese accumulated in the 33

gills, liver and muscle of the fish and significant differences in blood plasma34

chemistry were observed. Blood plasma sodium and chloride of fish exposed to 35

manganese were significantly elevated in hyperosmotic salinity (45ppt) and depressed 36

at hyposmotic salinity (5 ppt) compared with unexposed fish at the same salinity; 37

consistent with manganese causing apoptosis or necrosis to chloride cells. We did not, 38

however, observe any gill epithelial damage under light microscopy. Blood plasma39

potassium was significantly elevated at all salinities in the presence of manganese and 40

liver potassium and glycogen reduced. These findings are consistent with manganese 41

interfering with carbohydrate metabolism. 42

43

Keywords: manganese, inland saline aquaculture, mulloway Argyrosomus japonicus,44

carbohydrate metabolism.45

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1. Introduction47

48

Salt is a natural part of the Australian landscape, having been deposited by rainfall, 49

wind and dust over thousands of years (Zalizniak et al., 2006). In many arid and semi-50

arid areas of southern Australia, human activities including land-clearing and 51

irrigation have led to rising groundwater tables, dissolving stored salts from the soil 52

and bringing them to the surface (Ghassemi et al., 1995; Lambers, 2003). The impacts 53

of such secondary salinisation are many and include the loss of farm production and 54

revenue, the degradation of rural infrastructure and recreational facilities, the 55

salinisation of rivers and associated impacts on irrigation and drinking water56

(Williams, 2001).57

58

As the salt in the Australian landscape is of marine origin, the culture of marine 59

organisms has considerable potential as an adaptive use for saline groundwater 60

(Doupé et al., 2003). Despite its marine origin, however, important deviations in ionic 61

composition occur in saline groundwater compared to seawater (Partridge et al., 62

2008). These deviations depend primarily on the nature of the groundwater systems 63

and the regolith with which they are associated (Mazor and George, 1992). One 64

common deviation in Australian saline groundwater is an elevated concentration of 65

manganese relative to seawater. A recent survey of 315 saline water sources in the 66

wheatbelt of Western Australia, for example, found dissolved manganese 67

concentrations as high as 83 mg/L, with an average value of 3.1 mg/L (George, 68

unpublished data). Sources of dissolved manganese include anaerobic reduction of 69

manganese oxides or the pedogenic weathering of manganese-bearing rock (Howe et 70

al., 2004; Hardie et al., 2007).71

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72

Manganese is an essential nutrient for all organisms, including fishes (Underwood, 73

1977; Knox et al., 1981; Gatlin and Wilson, 1984; Maage et al., 2000) and plays 74

critical physiological roles as a constituent of several metaloenzymes and as a75

coactivator of many other enzymes (Underwood, 1977; Cossarini-Dunier et al., 1988). 76

Excess concentrations of manganese, however, can have negative effects including77

the disruption of sodium balance (Gonzalez et al., 1990), the impairment of calcium 78

uptake and mineralisation (Reader et al., 1988), impacts on carbohydrate metabolism 79

(Nath and Kumar, 1987; Barnhoorn et al., 1999), effects on immune response 80

(Cossarini-Dunier et al., 1988; Hernroth et al., 2004) and neurotoxicity (Newland, 81

1999; Gunter et al., 2006). Of those studies on fishes, only freshwater fishes have 82

been investigated, presumably due to the susceptibility of freshwater sources to 83

anthropogenic sources of manganese pollution. Seawater contains a very low 84

concentration of manganese (0.002 mg/L) (Spotte, 1992) and there appears to be no 85

documented studies on the effect of elevated manganese on marine or estuarine fishes.86

Unpublished data from our laboratory collected during bioassays investigating the 87

aquaculture potential of saline groundwater sources suggested that an elevated 88

concentration of manganese may have a chronic effect on the growth of mulloway 89

(Argyrosomus japonicus).90

91

The aim of this study was therefore to investigate the effects of an elevated dissolved 92

manganese (II) concentration (5 mg/L) on the survival, growth, pathology, and blood 93

plasma and organ chemistry of mulloway. Mulloway is a temperate species 94

considered a suitable candidate for inland saline aquaculture based on its euryhalinity,95

well-documented hatchery production techniques, reasonably rapid growth rates and a 96

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temperate habitat compatible with the climatic zones where most secondary salinity in 97

Australia occurs (Partridge et al., 2008). Because of the wide range of groundwater 98

salinities found throughout Australia, the effect of elevated manganese in this trial 99

was investigated across the range of salinities that mulloway are known to tolerate, 100

namely 45, 15 and 5 ppt. 101

102

2. Materials and Methods103

104

At each of the three salinities investigated (45, 15 and 5 ppt), the effects of an 105

elevated manganese concentration (5 mg/L) were compared against a control (0 mg/L) 106

in triplicate, flow-through 180 L tanks at 23°C over a period of two weeks. Five 107

juvenile mulloway (9.4 ± 0.2 g) were stocked into each tank with flowing seawater 108

(33 ppt), and then acclimated to their respective salinity over a period of three days. 109

Once the target salinities had been reached, fish were acclimated to the experimental 110

conditions for a further five days, then anaesthetised and reweighed. Prior to 111

transferring fish back to their respective tank, those tanks randomly assigned to a 112

treatment of 5 mg Mn/L were supplemented with laboratory grade MnCl2.6H2O by 113

dissolving the required quantity of salt into 1 litre of tank water. Water flowed 114

through each tank at 8 L/hour from header tanks containing the treatment water.115

Header tanks were emptied and cleaned daily and new treatment water prepared by 116

dissolving the required quantity of MnCl2.6H2O into 1 L of water of the appropriate 117

salinity before mixing this into the header tank. Water of 45 ppt was prepared by the 118

addition of artificial seasalt (Ocean Nature®; Aquasonic Pty Ltd, Wauchope, 119

Australia) to seawater, whilst 5 and 15 ppt water were prepared by diluting seawater 120

with dechlorinated tap water. Seawater (33 ppt) was sourced from a marine bore at 18 121

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m depth. This water is filtered to 1 µm and is free of particulate and dissolved organic 122

carbon. Fish were fed daily to satiety and the amount of food consumed was recorded. 123

Water quality parameters including temperature, dissolved oxygen and pH were 124

measured daily. Dissolved manganese was measured four times throughout the trial in 125

all tanks using inductively coupled plasma atomic emission spectroscopy (Vista AX 126

CCD Simultaneous ICP-AES; Varian, Mulgrave, Australia) after filtering water 127

samples to 0.45 µm (Whatman GFC).128

129

At the completion of the two week trial, all fish were anaesthetised and weighed. 130

Blood was immediately taken from the caudal vessels of three fish per replicate and 131

pooled for the determination of plasma sodium, chloride and potassium using an ion-132

specific electrode analyser (Vetlyte, IDEXX, USA). Samples of dorsal muscle, liver 133

and gills were taken for analysis of their ionic composition. These samples were134

freeze-dried, weighed to determine water content, then ground and digested in a 135

combination of hydrogen peroxide, concentrated nitric acid and hydrochloric acid at 136

120°C. Digests were subsequently analysed for manganese, sodium, potassium, 137

calcium, magnesium and iron via ICP-AES. 138

139

Samples of gill and liver were preserved in 10% formalin prepared in seawater, before 140

embedding in paraffin then cutting 5 µm sections. Gill and liver sections were stained 141

with haematoxylin and eosin (H&E) and liver sections with the periodic acid-Schiff142

(PAS) method for glycogen.143

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2.1. Statistical Analysis 145

146

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Growth was expressed as specific growth rate (SGR) using the following equation:147

148

100(days)Time

)Ln(-)(Ln (%/day) SGR x

WiWf

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150

Where Wf and Wi were the final and initial wet weights of the fish, averaged over all 151

fish in a replicate. Feed intake was calculated as a percentage of the fish’s initial body 152

weight. 153

154

The effects of dissolved manganese concentration and salinity on survival, specific 155

growth rate, feed intake, and blood plasma and organ chemistry were compared using 156

two way ANOVA followed by a post hoc comparison of group LSD (Least Square 157

Difference) means using Tukey’s HSD (Honestly Significant Differences) test. 158

Statements of statistical significance refer to the 0.05 level. Arcsine transformations 159

were performed where necessary to normalise data prior to ANOVA160

161

3. Results162

3.1. Water Quality163

164

Actual dissolved manganese concentrations were slightly less than the target value of 165

5 mg/L (Table 1). Despite concentrations between treatments differing by no more 166

than 0.15 mg/L, the differences were found to be significant (P < 0.01), with the 167

concentration at 15 ppt (4.13 ± 0.01 mg/L) being significantly lower than at 5 ppt168

(4.28 ± 0.01 mg/L) and 45 ppt (4.24 ± 0.01 mg/L). Manganese concentration in the 169

control treatments were below the detectable limit of 0.01 mg/L. 170

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171

Water pH (Table 1) was significantly affected by salinity (P < 0.001), but not 172

manganese concentration (P = 0.24), or the interaction of manganese and salinity (P = 173

0.23). Water pH was significantly higher at 45 ppt (8.13 ± 0.01) than at 15 ppt (8.01 ± 174

0.01) and 5 ppt (8.03 ± 0.01). 175

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3.2. Survival, Growth, Food Intake and Conversion Ratio177

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Survival of mulloway over the two week experimental period was 100% in all 179

treatments except 45ppt-5mg Mn/L, with a survival of 73 ± 13%; significantly lower 180

than all other treatments (Table 1).181

182

At all salinities, those fish exposed to manganese grew at a significantly slower rate 183

than those without manganese (Table 1). Two way analysis of variance revealed that 184

salinity did not affect SGR (P = 0.09) but both manganese concentration (P<0.0001) 185

and the interaction of manganese concentration and salinity did (P = 0.03). Those 186

mulloway grown in the presence of manganese at salinities of 5 and 45 ppt lost weight 187

over the 2 week period (SGR -0.17 ± 0.42 and -0.44 ± 0.83%/day, respectively), 188

whereas those at 15 ppt gained some weight (1.70 ± 0.20%/day). Those mulloway 189

grown in the control treatments grew rapidly with an average SGR of 4.05 ± 190

0.29%/day (pooled across three salinities). 191

192

Food intake was affected by both salinity (P = 0.02) and manganese concentration (P193

< 0.0001), but not their interaction (P = 0.63) (Table 1). The post hoc comparison of 194

group LSD means showed that those fish at 45 ppt ate significantly less food than 195

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those at 15 ppt and those fish exposed to 5 mg/L of manganese at all salinities ate196

significantly less food than those in the control treatments. 197

198

Due to mulloway’s loss of weight at 5 and 45 ppt in the presence of manganese, we 199

were unable to calculate food conversion ratios for these fish. Fish grown in the 200

presence of manganese at 15 ppt had a significantly higher (poorer) FCR (1.55 ± 0.22) 201

than those cultured at the same salinity without manganese addition (0.81 ± 0.06). 202

203

3.3. Histology204

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No epithelial damage to the gills was observed in histological sections. PAS-positive 206

cells in the liver were clearly more abundant in manganese exposed fish at all 207

salinities, however, quantification could not be conducted due to a lack of uniform 208

distribution within sections. 209

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3.4. Blood plasma ionic composition211

212

Concentrations of the electrolytes in the blood plasma of mulloway are shown in 213

Figure 1. Blood plasma potassium and chloride were significantly affected by 214

manganese concentration (P<0.0001 and 0.0073 for K and Cl, respectively), salinity 215

(P = 0.04 and <0.0001, respectively) and the interaction of these terms (P = 0.006 and 216

<0.0001, respectively). Blood plasma sodium concentration was not affected by 217

manganese concentration (P = 0.27) but was affected by salinity (P = 0.0005) and the 218

interaction of manganese concentration and salinity (P = 0.002). 219

220

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Because of the significant interaction terms, we conducted one-way analysis of 221

variance, comparing blood plasma ionic concentrations between fish exposed and not 222

exposed to manganese at each salinity. Blood plasma sodium was significantly greater 223

in fish exposed to manganese at 45 ppt (P = 0.002) but significantly lower in those224

exposed at 5 ppt (P = 0.04). There was no difference in blood plasma sodium 225

concentrations between exposed and unexposed fish at 15 ppt (P = 0.11). The same 226

pattern can be seen with blood plasma chloride at the various salinities and manganese 227

concentrations, i.e. a significantly greater chloride concentration in manganese 228

exposed fish at 45 ppt compared with unexposed fish (P = 0.006), with the opposite 229

effect at 5 ppt (P = 0.016) and no significant difference at 15 ppt (P = 0.51). Blood 230

plasma potassium was significantly higher in exposed fish compared with unexposed 231

fish at all salinities (P<0.02).232

233

In order to test the hypothesis that mulloway were osmoregulating well in the absence 234

of manganese across the range of salinities tested, one-way analyses of variance were235

also conducted on the electrolyte data from unexposed fish. These ANOVAs revealed 236

that salinity had no significant effect on blood plasma sodium (P = 0.97), chloride (P 237

= 0.13) or potassium (P = 0.69). 238

239

3.5. Organ Chemistry240

241

Exposure to manganese resulted in significantly higher concentrations of this metal in 242

the gills, muscle and liver of the fish (Table 2). The manganese content of the gills 243

and muscle were both affected by water manganese concentration (P<0.0001 for both 244

organs), salinity (P = 0.001 and 0.01, respectively) and the interaction (P = 0.0008 and 245

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0.015, respectively). Despite fish exposed to manganese at 45 ppt showing the poorest 246

performance in terms of survival and growth, gill and muscle manganese 247

concentrations at this salinity were lower than at the other salinities; significantly 248

more so than at 5 ppt. Liver manganese concentration was not significantly affected 249

by salinity (P = 0.65) or the interaction of salinity and manganese concentration (P = 250

0.70), but was significantly affected by water manganese concentration (P < 0.0001). 251

252

There were no clear trends in other organ chemistry parameters with salinity or 253

manganese concentration, with the exception of the potassium content of liver, which 254

was significantly lower in those fish reared in the presence of manganese (P < 0.0001)255

(Table 2). Salinity (P = 0.30) or the interaction of manganese concentration and 256

salinity (P = 0.41) did not affect liver potassium.257

258

4. Discussion259

260

Dissolved manganese had a significant, detrimental effect on the growth of juvenile 261

mulloway at all salinities tested, with this effect being greater at salinities of 5 and 262

45 ppt. Dissolved manganese only affected survival at the highest salinity.263

264

Our finding of equivalent survival, growth and blood plasma electrolyte 265

concentrations across the control salinity treatments demonstrates that mulloway are 266

capable of efficient osmoregulation across this salinity range. Mulloway’s ability to 267

tolerate 5 ppt has been described previously (Fielder and Bardsley, 1999), however 268

this appears to be the first published data on their performance at a salinity greater 269

than seawater. 270

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271

The poor growth performance of fish exposed to manganese was concomitant with a 272

significantly lower food intake. Intakes in the control treatments were similar to that 273

recommended for mulloway of similar size by Collett et al. (2008). Food intake of 274

manganese-exposed fish was approximately half this rate. Similar inhibitory effects of 275

excess manganese on appetite have been described for pigs, cows and sheep 276

(Cunningham et al., 1966; Underwood, 1977). The significantly higher (poorer) FCR 277

in fish exposed to manganese at 15 ppt, compared with unexposed fish at this same 278

salinity, indicates that the reduced growth of manganese exposed fish was contributed 279

to by poor feed utilisation efficiency as well as reduced food intake.280

281

The mode of action of manganese toxicity to fishes has not been well defined. Nyberg 282

et al. (1995) suggested that its toxic action is probably similar to other metals whereby 283

it is deposited onto the gills, causing physiological disruption to iono- and or osmo-284

regulatory functions. Manganese has been shown to deposit on the gills of lobsters 285

(Baden et al., 1990), however, we found no evidence of precipitation of manganese on 286

the gill surface or damage to the gill epithelium. Although chemical analysis of gills 287

in the current study revealed a high concentration of manganese, we are unable to 288

determine whether this was precipitated on the gill surface, within the gills filaments 289

or associated with the cartilaginous gill arches, which have been proposed as a long 290

term fixation site for manganese (Adam et al., 1997).291

292

Rouleau et al. (1995) suggested that uptake of manganese probably occurs mainly via 293

the gills and is then transported by the blood to various organs and tissues, where it 294

accumulates most in those tissues rich in mitochondria such as liver, pancreas and 295

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kidney. In their study, Rouleau et al. (1995) found that manganese accumulation in296

freshwater brown trout was greatest in the liver, then gills, then muscle. In our study, 297

manganese concentration was greatest in the gills, followed by the liver then muscle. 298

Our findings are consistent with Capelli et al. (1987) who found higher concentrations 299

of (naturally occurring) manganese in the gills of marine Atlantic bonito, Sarda sarda,300

compared with muscle and liver. These differences in the major target organs for 301

manganese accumulation suggest that different uptake mechanisms occur between 302

freshwater and marine species. These differences may be caused by differences in 303

metal speciation between freshwater and seawater as described in further detail below.304

305

It is well documented that manganese uptake and toxicity in freshwater fishes306

increases with decreasing pH (Bendell-Young and Harvey, 1986; Rouleau et al., 307

1996) and with decreasing freshwater hardness (Reader et al., 1988; Gonzalez et al., 308

1990; Adam et al., 1997; Stubblefield et al., 1997; Reimer, 1999). At low pH, 309

particularly at pH values below 7, a greater proportion of manganese in freshwater310

exists as aqueous, toxic Mn(II), whereas high pH values favour the oxidation of 311

insoluble and inert colloidal oxides (Florence et al., 1992; Rouleau et al., 1996; Zaw 312

and Chiswell, 1999). Although there is a lack of data on the effects of pH on 313

manganese speciation in seawater, our findings are consistent with this freshwater 314

model, given that the pH of 45 ppt water in the current study was significantly higher 315

than at 15 and 5 ppt, and that uptake of manganese at this salinity was significantly 316

lower than at the lower salinities. However, although the difference in pH was 317

significant, it amounted to less than 0.12 pH units and the pH at all salinities was well 318

above the ‘critical’ range of 6 - 7 described by Rouleau et al. (1996). It seems 319

unlikely, therefore, that differences in pH among salinity treatments had a major 320

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effect on manganese uptake. As we discuss in further detail below, we believe the far 321

greater differences in salinity between treatments is responsible for the observed 322

differences in toxicity. 323

324

In freshwater, decreases in metal toxicity with increasing hardness have been 325

attributed to the ability of calcium (and to a lesser extent magnesium) to compete with 326

toxic metals for ligand sites on the gill surface, which reduces the permeability of the 327

paracellular pathways to these metals (McDonald et al., 1989). In seawater, the effects 328

of increasing salinity on metal ion toxicity appear to be more complicated and 329

dependent on how the salinity affects metal ion speciation. The freshwater model 330

implies that the higher calcium and magnesium concentrations in seawater should 331

provide even greater protection than hard freshwater and that this protection should 332

increase with increasing salinity. Although supporting data are lacking for manganese, 333

the uptake by fishes of certain other metals, including copper and lead, is inversely 334

proportional to salinity (Somero et al., 1977; Jezierska and Witeska, 2006) and the 335

uptake of manganese by salt water mussels (Mytilus edulis) has been shown to follow 336

this trend (Struck et al., 1997). There are examples, however, where metal 337

bioavailability can increase with salinity. For example, when freshwater containing 338

relatively non-toxic aluminium colloids mixes with seawater, the calcium and 339

magnesium in seawater can displace the aluminium and thereby increase the 340

concentration of toxic, low molecular mass cationic aluminium species (Bjerknes et 341

al., 2003; Teien et al., 2006). Our data showing decreasing manganese concentration342

in gills and muscle with increasing salinity are consistent with the former model, 343

however more research is required to determine the effect of salinity (and pH) on 344

manganese speciation.345

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346

Our data also suggest, however, that despite reduced uptake at 45 ppt, toxicity at this 347

salinity was greater compared with lower salinities. We suggest that the greater 348

toxicity of manganese at higher salinity, despite its reduced uptake, is due to the 349

effects of manganese on osmoregulation.350

351

Gonzalez et al. (1990) found that freshwater brook char, Salvelinus fontinalis,352

exposed to 600 mg/L of dissolved manganese experienced a 40% reduction in plasma 353

sodium concentration. Studies have also shown that copper exposure induces 354

hyponatremia in freshwater fishes caused by apoptosis and necrosis of chloride cells 355

(Dang et al., 2000). Given that mulloway in hyposmotic salinities face the same 356

osomotic challenges as freshwater fishes; our findings of hyponatremia in mulloway 357

at 5 ppt are consistent with these studies and suggest that manganese may also induce 358

apoptosis and necrosis of chloride cells. Mulloway held at hyperosmotic salinity 359

(45 ppt) face the opposite osmotic challenges to those in hyposmotic salinity and their 360

chloride cells are responsible for the elimination (rather than uptake) of monovalent 361

ions (Evans et al., 2005). Our data showing elevated blood plasma sodium and 362

chloride in manganese-exposed fish at 45 ppt is therefore consistent with manganese 363

causing apoptosis and necrosis of chloride cells, as the loss of such cells would result 364

in reduced elimination of these ions. Seemingly inconsistent with this theory of 365

manganese causing increased apoptosis and necrosis of chloride cells is the fact that 366

we did not observe any histological changes in H&E sections. Cells dying from 367

apoptosis, however, remain intact, and Dang et al (2000) only observed these changes 368

using immunocytochemistry and electron microscopy.369

370

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Given that 15 ppt is closer to iso-osmotic than the other two salinities, uptake and 371

elimination of monovalent ions will be less at this salinity and the effects of chloride 372

cell apoptosis not as detrimental as at the more extreme salinities tested. Indeed the 373

differences in plasma sodium and chloride concentrations between exposed and 374

unexposed fish were smaller and statistically non-significant at this salinity, compared 375

with 5 and 45 ppt. We therefore suggest that the reduced osmotic demands at 15 ppt 376

are the likely reason these fish tolerated manganese exposure better than at 5 and 377

45 ppt. We cannot rule out the possibility that the greater tolerance for manganese 378

exposure at a salinity of 15 ppt was due to the small, but significantly lower 379

manganese concentration at this experimental treatment, compared to the 5 ppt and 45 380

ppt treatments. There are, however, two reasons why we believe that this lower 381

manganese concentration in the 15 ppt treatment was not biologically significant. 382

First, there was a reduction of only 0.15 mg/L at this treatment, 3% of the target value 383

of 5 mg/L. Second, there was no relationship between manganese concentration and 384

fish survival and growth rate in the three replicates within treatments.385

386

Even though the fish exposed to manganese at 15 ppt performed better than those at 5 387

and 45 ppt, they still exhibited significantly slower growth compared with control fish 388

at the same salinity. Blood plasma potassium was significantly elevated in these fish,389

suggesting that there was some disruption to their osmoregulatory capacity. The 390

reduced feed conversion efficiency seen in these fish, compared with unexposed fish 391

at the same salinity, may therefore be the result of the repartitioning of a greater 392

proportion of dietary energy into osmoregulation or, alternatively, the result of 393

impaired carbohydrate metabolism described below. 394

395

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We found significant reduction in liver potassium and reduced liver glycogen in fish 396

exposed to manganese. Manganese has been implicated in impaired carbohydrate 397

metabolism in both fishes (Nath and Kumar, 1987; Nath and Kumar, 1988; Barnhoorn 398

et al., 1999) and mammals (Underwood, 1977) and observations of decreased liver 399

glycogen and hyperglycemia in manganese exposed fishes have led to the hypothesis 400

that manganese exposure may inhibit insulin release (Nath and Kumar, 1987) in a 401

similar fashion to that seen in cadmium exposed rats (Ghafghazi and Mennear, 1973). 402

Although the reduced liver glycogen that we observed in manganese exposed fish is 403

consistent with these studies, it would also be the expected response of fish that are 404

losing weight or growing at a sub-optimum rate. The significant reduction in liver 405

potassium of manganese exposed fish, however, is consistent with a lack of insulin, as 406

such a deficiency inhibits potassium absorption by liver cells in rats (Fehlmann and 407

Freychet, 1981).408

409

Cardeilhac et al. (1979) and Cardeilhac and Hall (1977) attributed the death of the 410

marine fishes sheepshead (Archosargus probatocephalus) and pinfish (Lagodon 411

rhomboides) exposed to excess copper to osmoregulatory failure and specifically to an 412

elevated concentration of blood plasma potassium. We found significantly elevated 413

blood plasma potassium in fish exposed to manganese at all salinities. The blood 414

plasma potassium of manganese exposed mulloway at 45 ppt was within the toxic 415

range reported for these two species and is therefore a possible cause of death of 416

mulloway in this treatment.417

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418

5. Conclusion419

420

Our results demonstrate that 5 mg/L of dissolved manganese, a concentration typical 421

of many saline groundwater sources in Western Australia, has a significant, 422

detrimental effect on the growth, health and survival of mulloway. It appears that 423

selecting a water source with salinity close to iso-osmotic will minimise the effects of 424

dissolved manganese, however, further studies are required to determine the minimum 425

concentrations at which growth is not affected. Given that this study was conducted at 426

relatively high pH values, the negative impacts of a similar dissolved manganese 427

concentration can be expected to be far greater in aquaculture systems or for 428

groundwater sources which have lower values of pH.429

430

Given that both salinity and the pH values typical of groundwater result in very slow 431

oxidation rates of manganese, its removal from saline groundwater is more 432

complicated than iron; another metal often found in high concentrations in saline 433

groundwater. Significant chemical pre-treatment with strong oxidising agents, or the 434

temporary elevation of pH to above 9,, would be required to oxidise manganese prior 435

to its physical removal; a process that will add significant cost and complexity to the 436

delivery of water, particularly for those production systems using large volumes. 437

Although oxidation is slow, once it does occur, precipitation of manganese oxides 438

onto tanks, pipes and pumps is likely to be problematic and impose a significant 439

management issue if dissolved manganese is not oxidised and removed prior to use. 440

441

442

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6. Acknowledgements443

444

This study was funded by Challenger TAFE, the Australian Department of Fisheries, 445

Forestry and Agriculture and the Department of Conservation and Land Management. 446

447

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592

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Table 1: Water quality and performance parameters for juvenile mulloway 595

grown in the presence or absence or manganese at various 596

salinities for two weeks.597

598

Table 2: Organ chemistry data for juvenile mulloway grown in the presence 599

or absence or manganese at various salinities for two weeks.600

601

Figure 1: Concentrations of (a) sodium, (b) chloride and (c) potassium in the 602

blood plasma of juvenile mulloway grown in the presence or absence603

or manganese at various salinities for two weeks. Columns within 604

salinity sharing the same letter are not significantly different 605

(P>0.05).606

607

608

609

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