Utilization of wastewater-grown zooplankton: Nutritionalquality of zooplankton and performance of silver perchBidyanus bidyanus (Mitchell 1838) (Teraponidae) fedon wastewater-grown zooplankton

G. KIBRIA & D. NUGEGODA Department of Environmental Management, Victoria University of Technology,

MCMC, Melbourne, Australia

R. FAIRCLOUGH Department of Biomedical Sciences, Victoria University of Technology, Melbourne, Australia

P. LAM Chemistry and Biology Department, City University of Hong Kong, Kowloon, Hong Kong

A. BRADLEY Zootech Research Group, Werribee, Victoria, Australia

Abstract

Zooplankton grow in the last stage of water puri®cation at

the Werribee Sewage Treatment Lagoons (WSTL) and the

resource is huge and unutilized. Daphnia carinata and Moina

australiensis are the dominant species at the WSTL. The

protein content of D. carinata and M. australiensis was

54.80% and 64.80%, respectively. Analysis of zooplankton

showed that both essential and nonessential amino acids were

present at a level that was higher than control diet. Silver

perch fed on D. carinata exhibited better growth, better food

conversion ratio (FCR), protein e�ciency ratio (PER) and

apparent net protein utilization (ANPU), which were not

signi®cantly di�erent from the control diet (P > 0.05).

Heavy metals concentrations were at very low levels in ®sh

fed on wastewater-grown zooplankton.

KEYKEY WORDSWORDS: amino acids, heavy metals, performance, prox-

imate composition, silver perch, wastewater reuse, zooplank-

ton

Received 10 February 1997, accepted 14 May 1998

Correspondence: Dr Golam Kibria, 45 Lowson Street, Fawkner, VIC 3060,

Australia. E-mail: humayun@pl.camtech.net.au

Introduction

Zooplankton are the natural food item of many marine and

freshwater ®sh and crustaceans. They have been used

extensively to rear larvae and fry (De Pauw et al. 1981;

Tay et al. 1991). Both live (Dabrowski 1984; Alam et al.

1993) and frozen zooplankton (Brett 1971; Sargent et al.

1979) have been used to rear ®n®sh and crustaceans. Studies

have shown that fry performed better when fed zooplankton

than when fed with arti®cial dry diets (Dabrowski 1984;

Dave 1989). Frozen and freeze-dried zooplankton represent

an excellent food source for hatchlings of white ®sh Coreg-

onus sp., carp Cyprinus carpio, lake char Salvelinus sp. and

species which normally require live plankton for growth

(Bryant & Matty 1980).

Zooplankton are a valuable source of protein, amino

acids, lipid, fatty acids and enzymes (Ogino 1963; Millamena

et al. 1990; Munilla-Moran et al. 1990; Pillay 1990).

Yurkowski & Tabachek (1979) reported that essential amino

acid levels in the cladoceran Daphnia pulex and the copepod

Diaptomus sp. were equal to or greater than ®sh require-

ments. Studies on the nutrient composition of natural

zooplankton are signi®cant since they would aid in deter-

mining the suitability of the organisms as feed ingredients for

aquaculture (Yurkowski & Tabachek 1979). The nutritional

quality of living organisms can be evaluated by determining

their proximate and amino acid composition, and by protein

e�ciency ratio and net protein utilization (Watanabe et al.

1978).

Zooplankton grow abundantly in the nutrient-rich Werri-

bee Sewage Treatment Lagoons (WSTL), Melbourne, Aus-

tralia but the resource is unutilized. To date, there has been

no research on the utilization of this resource for aquaculture

in Australia. Fish meal is the highest quality protein source

for ®sh feeds (Lovell 1989). However, it is becoming

221

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Ó 1999 Blackwell Science Ltd

expensive owing to high demand and reduced supply (Barlow

1989). Therefore, it is essential to evaluate other ingredients

as alternative protein sources for rearing ®sh. Silver perch

Bidyanus bidyanus is an Australian native ®sh of high

aquaculture potential and represents the main freshwater

native aquaculture industry in Australia. This paper exam-

ines the proximate and amino acid composition of wastewa-

ter grown caldocerans (Daphnia carinata and Moina

australiensis) and the preliminary performance of silver perch

fed on wastewater grown zooplankton.

Materials and methods

Zooplankton source and production

Zooplankton grow abundantly in the last stage of the

wastewater treatment process of the WSTL, which are

located 35 km south-west of Melbourne, Australia. They

graze on algae and remove nitrogen and phosphorus (pol-

lutants) from the wastewater. A simpli®ed sequence of

zooplankton production at the WSTL is given in Fig. 1.

During 1995±96 (summer), Zootech (a private company

researching utilization of wastewater zooplankton) used a

proprietary zooplankton harvester called `Baleen' and har-

vested 40±84 kg h)1 of zooplankton from the lagoons.

Zooplankton was harvested every day during the summer

period (January±March). Zootech used carefully chosen

mesh for selective harvesting of D. carinata or M. austral-

iensis from the ponds because there is a distinct size variation

between the two populations (Lai 1994). Harvested zoo-

plankton were kept frozen at )20 C in blocks until used. The

dominant zooplankton at the WSTL were D. carinata King

and M. australiensis Sars (Lai 1994). These natural and

invaluable resources are normally drained into Port Philip

Bay by the Melbourne Water Authority.

Fish husbandry and experimental procedures

Preliminary rearing experiments were conducted at the

Zootech multipurpose hatchery located at the Werribee

Treatment Complex. Silver perch ®ngerlings (612 mg) were

bought from a local native ®sh hatchery. Fish were acclima-

tized in the hatchery environment for 3 weeks and were

subjected to a short salt bath (10 g L)1) for 60 min prior to

the start of feeding trials (Rowland & Ingram 1991).

Rectangular, dark-coloured PVC tanks (41 ´ 84 ´ 27 cm)

were selected randomly for each replicate of the seven feeding

treatments (Table 1). Each tank was stocked with 15 ®sh

with three replicates per treatment. Fish were reared in static

water. Water temperature was kept at 25 °C by a thermo-

statically controlled heater. Water quality was maintained at

pH 7.5±8.0, hardness at 80±100 mg L)1, dissolved oxygen

levels at > 6 mg L)1 and a cycle of 12 h light and 12 h dark

Figure 1 Simpli®ed sequence of zooplankton production at the Werribee Treatment Lagoons. 1. Wastewater (raw sewage) enters the aerobic

ponds. 2. Bacteria digest the organic material. 3. Sludge containing the heavy metals and other chemicals settle out. 4. Wastewater moves into

the aerobic pond. 5. Algae grow in the nutrient-rich water. 6. Bacteria and algae remove nitrogen from the water. 7. Zooplankton grow and feed

on algae and bacteria. 8. Treated wastewater, along with zooplankton, ¯ows into the Port Phillip Bay.

G. Kibria et al.

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Ó 1999 Blackwell Science Ltd Aquaculture Nutrition 5; 221^227

222

was used. Fish were fed frozen zooplankton and dry diet

(control), twice per day at 3% of their body weight, 5 days

per week for 4 weeks (dry matter basis).

Biochemical analysis

Fish and zooplankton samples were stored at )20°C until

analysed. Triplicate samples of zooplankton (D. carinata and

M. australiensis), ®sh and feed were subjected to chemical

analysis. Samples were weighed using an analytical balance

(Mettler AE 200 USA). At the completion of the experiment,

pooled ®sh samples were dried and ground, using mortar and

pestle for crude protein, fat, ash and phosphorus determina-

tion. Dry matter was determined by weight loss after drying

in an oven at 100°C for 24 h (AOAC 1990). Ash was

determined after ignition at 600°C for 12 h (AOAC 1990),

crude protein by digesting samples with Kjeldahl tablets and

5 mL concentrated sulphuric acid (N ´ 6.25) (AOAC 1990),

carbohydrate by the di�erence between the dry matter of the

sample and the sum of the determined crude protein, fat and

ash, lipid by ether extract in a Soxhlet apparatus (New 1987)

and total phosphorus following AOAC (1990). Protein

retention was determined by subtracting the initial protein

content of ®sh from the ®nal protein content. Amino acid

composition of zooplankton and dry feed was analysed in the

State Chemistry Laboratory, Melbourne by high-pressure

liquid chromatography (HPLC).

Heavy metals analysis

Fish carcasses were analysed for heavy metals (zinc,

cadmium and lead). Samples were dried in an oven and

crushed gently in a mortar and pestle. Dried samples were

digested in analytical nitric acid (HNO3) at 100°C in a block

heater for 30 min (Corp & Morgan 1991). Metals were

analysed on a Varian spectra AA-400 Flame Absorption

Spectrophotometer with background correction using pro-

cedural blanks.

Statistical procedures

Mean, standard deviation and standard error of the mean of

speci®c growth rate (SGR), food conversion ratio (FCR),

protein e�ciency ratio (PER) and apparent net protein

utilization (ANPU) were calculated following Zar (1984). All

percentage data were transformed to arcsin values prior to

analysis. One-way analysis of variance (ANOVAANOVA) was used to

compare the means of SGR, FCR, PER and ANPU. All

ANOVAANOVAs were calculated using an IBM compatible Microsoft

EXCELEXCEL programme setting the signi®cance level at 0.05.

Results and discussion

Proximate composition of zooplankton

D. carinata and M. australiensis possessed an attractive

biochemical composition (Table 2). The protein content in

D. carinata and M. australiensis was 54.34% and 64.80%,

respectively, higher than the control commercial silver perch

diet. The protein content of Daphnia sp. has been reported to

be 49.70±70% (Blazka 1966; Yurkowski & Tabachek 1979;

Watanabe et al. 1983) whereas for Moina sp. it varies

between 59.00 and 77.85% (Tay et al. 1991; Creswell 1993).

It appears that D. carinata and M. australiensis are a

potentially valuable protein source.

The lipid content in the two zooplankton studied was in

the range 7.29±7.73% (Table 2). Watanabe et al. (1983)

analysed various zooplankton, with Daphnia containing 13%

and Moina 12±27% lipids. The lipid content in freshwater

zooplankton can be extremely variable (Vijverberg & Frank

1976) and may be in¯uenced by seasonal succession of

Table 1 Di�erent feeding regimes used in the silver perch perfor-

mance study

Feeding regimes Code

1 Control (artificial silver perch diet;100%) C2 Daphnia carinata (100%) D3 Moina australiensis (100%) M4 D. carinata (50%) and M. australiensis (50%) D + M5 D. carinata (33.33%) + M. australiensis (33.33%) +arti¢cial diet (33.33%)

D + M + C

6 D. carinata (50%) + control (50%) D + C7 M. australiensis (50%) + control (50%) M + C

Table 2 Proximate analysis of control feed and zooplankton

(Daphnia carinata and Moina australiensis) harvested from the

Werribee Sewage Treatment Lagoons

Dry diet(control) D. carinata

Moina

australiensis

Carbohydrate (g kg)1) 366.6 þ 10.4 271.0 þ 15.6 206.5 þ 9.7Crude ash (g kg)1) 98.4 þ 0.2 112.6 þ 0.1 68.2 þ 0.7Crude fat (g kg)1) 85.0 þ 4.9 72.9 þ 0.3 77.3 þ 0.6Crude protein (g kg)1) 450.0 þ 15.02 543.4 þ 28.3 648.0 þ 14.8Moisture (g kg)1) 98.8 þ 1.0 928.7 þ 0.9 937.2 þ 1.1Nitrogen (g kg)1) 72.0 þ 5.0 86.9 þ 3.1 103.7 þ 1.7Phosphorus (g kg)1) 11.6 þ 0.8 11.4 þ 0.2 11.1 þ 0.1Digestible energy

(DE, kcal kg)1)15273 5250 5700

DE/P (g kcal)1) 117 97 88

1Gross energy was calculated using kilocaloric values of 5.5 g)1 protein,9.1 g)1 fat and 4.1 g)1 carbohydrate (New 1987).2 Each value represents a mean þ SE; n = 3.

Nutritional quality of wastewater-grown zooplankton and performance of silver perch

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phytoplankton species (Je�ries 1970) or source of food fed to

zooplankton (Proulx & de la Noue 1985). Although the fatty

acid composition of D. carinata and M. australiensis was not

analysed, a study by Tucker (1992) noted that Daphnia sp.

may be de®cient in docosahexaenoic acid (DHA) and Moina

sp. in both DHA and eicosapentaenoic acid (EPA).

Amino acid content in zooplankton

Amino acid analysis revealed that both D. carinata and

M. australiensis contained appreciable levels of both essen-

tial and nonessential amino acids (Table 3). Compared

with the commercial silver perch diet (control), both

zooplankton appeared slightly superior. However, values

of some of the essential amino acids in D. carinata and M.

australiensis were lower than those previously cited for

Daphnia sp. (Yurkowski & Tabachek 1979; Hepher 1988)

and Moina sp. (Hepher 1988; Creswell 1993). The lower

values could have been due to seasonal variation in amino

acid content (Holm & Walther 1988), foods consumed by

zooplankton (Watanabe et al. 1983) or culture medium

(Allen & Allen 1981). The essential amino acid content of

zooplankton can meet the requirements of ®sh, for example

the essential amino acid content of Daphnia pulex was

found to be equal to or greater than the requirement of

Chinook salmon (Yurkowski & Tabachek 1979) (see also

Table 3). Both D. carinata and M. australiensis had a

relatively similar essential and nonessential amino acids

composition (average 51.5% essential amino acids). Holm

& Walther (1988) reported that 50.6% of amino acids in

daphnids were essential. The cladocerans, D. carinata and

M. australiensis, contained relatively similar proportions of

polar to nonpolar amino acids, (0.89 for D. carinata and

0.80 for M. australiensis). A higher proportion (1.57) was

calculated for daphnids by Holm & Walther (1988), which

may indicate a better nutritional status of those species.

Performance of silver perch fed on zooplankton

The control silver perch diet (treatment 1) resulted the best

growth among the seven feeding regimes, followed by ®sh fed

D. carinata (treatment 2) (Fig. 2). Gain in weight was not

signi®cantly di�erent between treatments 1 and 2 (P > 0.05).

Among the zooplankton-based treatments, ®sh fed

D. carinata (treatment 2) and a combination treatment of

D. carinata plus control diet (treatment 6) showed better

growth (P < 0.05). Silver perch fed M. australiensis showed

comparatively less growth. The results may indicate that

silver perch have a preference for D. carinata over

M. australiensis. Confer & Lake (1987) established that

yellow perch Perca ¯avescens has a greater a�nity for

Diaptomus sicilis than for other species of zooplankton.

Minimum essentialamino acid requirementsfor finfish1

Dry diet(control) D. carinata M. australiensis Omnivorous Carnivorous

Essential amino acidsArginine (Arg) 2.42 3.37 3.01 1.81 2.24Histidine (His) 1.29 1.65 1.37 0.76 0.95Isoleucine (Iso) 1.66 2.60 2.67 1.18 1.46Leucine (Leu) 3.25 5.22 4.94 2.15 2.66Lysine (Lys) 2.65 3.35 3.34 2.48 3.08Methionine (Met) 0.80 1.46 1.13 0.81 1.00Phenylalanine (Phe) 1.86 2.52 2.67 1.22 1.51Threonine (Thr) 1.68 2.86 2.79 1.35 1.67Tryptophan (Try) 0.43 0.71 0.76 0.25 0.31Valine (Val) 2.36 3.27 3.56 1.40 1.73

Nonessential amino acidsAlanine (Ala) 2.27 3.23 3.77Aspartic acid (Asp) 2.42 4.70 5.12Cystine 0.44 0.71 0.60Glutamic acid (Glu) 6.36 6.39 6.57Glycine (Gly) 2.03 2.48 2.59Proline (Pro) 2.03 2.20 2.36Serine (Ser) 1.88 2.82 2.62Tyrosine (Tyr) 1.17 2.14 2.38

1Tacon (1990).

Table 3 Essential and nonessential

amino acid composition of dry diet

(control) and zooplankton (Daphnia

carinata and Moina australiensis)

harvested from the Sewage Treatment

Lagoons and used in feeding trials

(g 100 g)1). Essential amino acid

requirements of ®sh (omnivorous and

carnivorous fry, 0.05 g) are also given

for comparison

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Similarly, Atlantic salmon Salmo salar showed a preference

for daphnids (Holm & Walther 1988). Furthermore, the

species and their origin (Watanabe et al. 1978), and seasonal

changes in the biochemical composition in zooplankton can

in¯uence their nutritional value to ®sh (Sadykhov et al. 1975;

Je�ries 1979). SGR, FCR, PER and ANPU were signi®-

cantly better with treatments 1 and 2 (Table 4). D. carinata

again gave the best FCR, PER and ANPU values compared

with M. australiensis. The maximum PER values obtained

with D. carinata were 1.57±1.64 (treatments 6 and 2). Trout

fed on zooplankton-krill (Euphsusia sp.) had a PER of 1.92

(Koops et al. 1979), whereas channel cat®sh had a PER of

1.03 (Hilge 1979). However, Watanabe et al. (1983) reported

a PER of 3.9 with Daphnia sp. and 2.6 with Moina sp. fed to

rainbow trout. Those zooplankton were raised indoors and

supplied with nutritionally enriched foods which might have

led to the better PER in ®sh. Nevertheless, D. carinata did

show comparable results for SGR, FCR, PER, ANPU in

comparison with the control commercial silver perch diet.

Effect of different feeding regimes on the chemicalcomposition of fish

The carcass composition of ®sh is shown in Table 5. The

moisture level decreased whereas protein, lipid and ash

content increased from initial values. Fish fed the control diet

had more lipids whereas ®sh fed on zooplankton had higher

ash and phosphorus contents. Rainbow trout fed on the

zooplankton Euphsusia paci®ca showed higher ash and lower

fat content (Spenelli 1979). Koops et al. (1979) also found a

higher phosphorus content in ®sh fed on krillmeal than on

®shmeal.

Heavy metals in fish carcass

Zinc, cadmium and lead were present at very low levels in ®sh

carcasses (Table 6). Fish fed mixed zooplankton (treatment

Figure 2 Gain in weight (%) of silver perch fed di�erent combina-

tions of zooplankton-based diets. Fish were fed at 3% body weight,

twice per day, 5 days per week for 4 weeks. The control diet (1) was a

commercial silver perch feed (mean � SE; n � 3); C � control,

D � Daphnia; M � Moina.

Table 4 Mean performance of silver perch juveniles fed either single zooplankton or a mixed zooplankton, or a combination of zooplankton

plus arti®cial diet. The ®sh were reared at 25°C for 4 weeks and fed at 3% body weight, twice per day, 5 days per week

C1 D M D + M D + M + C D + C M + C1 2 3 4 5 6 7

IAW2 610 609 611 613 608 617 619FAW3 1105 1104 901 863 904 1012 892SGR4,5 2.99 þ 0.04a 2.97 þ 0.16a 1.94 þ 0.09b 1.70 þ 0.05bc 1.99þ 0.11b 2.50 þ 0.04bdef 1.85 þ 0.02bcg

SGR as % of C 100.0 99.30 64.90 56.86 66.55 83.61 61.87FCR6 1.02 þ 0.07a 1.18 þ 0.12a 1.38 þ 0.04b 1.94 þ 0.47bcd 1.68þ 0.09b 1.23 þ 0.12ae 1.71 þ 0.13b

PER7 2.17 þ 0.14a 1.57 þ 0.11a 1.12 þ 0.09b 0.86 þ 0.11bc 1.10 þ 0.06b 1.64 þ 0.02adef 1.06 þ 0.05 bg

ANPU8 58.07 þ 2.1a 47.48 þ 4.0a 37.68 þ 3.10b 23.38 þ 2.13bcd 38.16 þ 0.48b 33.86 þ 1.77bd 23.37 þ 0.91bcdef

1SeeTable1for codes.2 IAW = Initial average weight (mg).3 FAW = Final average weight (mg).4 Each value represents a mean � SE; n = 3; numbers within a row not sharing the same superscript (a^g) are signi¢cantly di¡erent (P < 0.05).5 Speci¢c growth rate (SGR) = ln ¢nal body weight ) ln initial body weight/time (days) ´ 100 (Haiqing & Xiqin1994).6 Feed conversion ratio (FCR) = dry weight fed/wet weight gain (Haiqing & Xiqin1994).7 Protein e¤ciency ratio (PER) = wet weight gain/protein consumed (Haiqing & Xiqin1994).8 Apparent net protein utilization (ANPU) = body nitrogen at the end of the test ) body nitrogen at the beginning of the test/amount of nitrogenconsumed.(Jauncey1982).

Nutritional quality of wastewater-grown zooplankton and performance of silver perch

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5) had signi®cantly higher levels of zinc than those of other

treatments (P < 0.05). Cadmium levels were negligible, with

no signi®cant di�erences among the treatments (P > 0.05).

Heavy metals were within the safe levels as set out by the

National Health and Medical Research Council (1987) for

human consumption. These preliminary data indicate that

zooplankton from the wastewater treatment lagoons were

not contaminated with the metals tested.

Conclusions

It is concluded that zooplankton from the Werribee waste-

water treatment facilities are rich in protein, essential amino

acids, lipid and phosphorus. Moreover, silver perch per-

formed signi®cantly better with D. carinata than with

M. australiensis (P < 0.05). SGR, FCR and PER were not

signi®cantly di�erent for ®sh fed a control commercial silver

perch diet or D. carinata. Further studies are needed on the

proximate, amino acid and fatty acid composition to

establish any seasonal variations in nutrient composition in

wastewater zooplankton. Research is also needed on the

possible loss of nutrients from frozen and freeze-dried

zooplankton, and on appropriate ways to reduce such losses.

Analysis of a wide range of contaminants, such as other

heavy metals and pesticides, would provide pertinent infor-

mation on the suitability of wastewater-grown zooplankton

for use in aquaculture.

Acknowledgements

The authors are indebted to the Department of Environ-

mental Management, Victoria University of Technology for

allowing the use of the equipment and chemicals required to

carry out the analysis. The Zootech Research Group

provided zooplankton and hatchery space to Golam Kibria.

We are grateful to two anonymous referees whose sugges-

tions helped to improve the manuscript.

References

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(de Man) as an Artemia substitute in the production of Macro-

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Allen, W.A. & Allen, G.H. (1981) Amino acids in the food web of a

wastewater aquaculture system for rearing juvenile salmon. Prog.

Fish. Cult., 43 (4), 178±182.

AOAC (1990) O�cial methods of analysis of the Association of

O�cial Analytical Chemists (AOAC) (Helrich, K. ed.), 15th edn,

Vol. 1. AOAC, Arlington, VA.

Barlow, S. (1989) Fish meal Ð world outlook to the year 2000. Fish

Farmer, Sept./Oct., 40±43.

Blazka, P. (1966) The ratio of crude protein, glycogen and fat in the

individual steps of the production chain. In: Hydrobiological

Studies (Hrbacck, J. ed.), pp. 395±409. Academia Publishing

House of the Czechoslovak Academy of Sciences, Prague,

Czechoslovakia.

Table 5 E�ect of zooplankton-based feeding regimes on the chemical composition1 of silver perch juveniles. The ®sh were reared at 25°C for

4 weeks and fed at 3% body weight, twice per day, 5 days per week

Feeding regime2 Moisture (g kg)1) Crude protein (g kg)1) Crude lipid (g kg)1) Ash (g kg)1) Phosphorus (g kg)1)

1. C 759.7 þ 6.13 648.1 þ 1.8 116.2 þ 0.9 185.0 þ 0.9 25.6 þ 1.62. D 763.3 þ 1.9 647.5 þ 1.9 99.2 þ 0.3 200.5 þ 0.3 30.4 þ 1.23. M 781.2 þ 2.4 650.3 þ 2.8 93.6 þ 1.2 214.0 þ 0.6 29.5 þ 0.54. D + M 790.2 þ 3.4 634.3 þ 0.3 90.6 þ 1.4 243.4 þ 0.3 34.5 þ 2.75. D + M+ C 780.2 þ 0.9 652.2 þ 0.3 98.3 þ 0.8 225.6 þ 0.5 30.5 þ 0.16. D + C 775.4 þ 2.5 626.2 þ 0.4 106.3 þ 1.1 176.2 þ 0.4 24.9 þ 0.47. M + C 785.0 þ 2.9 624.0 þ 0.2 95.7 þ 1.1 188.6 þ 0.1 28.0 þ 1.1

1Body composition at the beginning of the experiment was: moisture 79.52%, protein 61.95%, lipid 9.07%, ash17.63%, phosphorus 2.51%.2 SeeTable1for codes.3 Each value represents a mean þ SE; n = 3.

Table 6 Metal concentrations (lg g)1, dry weight basis) in the

carcass of silver perch juveniles reared for 4 weeks and fed on

zooplankton and zooplankton plus arti®cial diet. The zooplankton

was harvested from the Werribee Sewage Treatment Lagoons

Feeding regimes1 Zinc2 Cadmium Lead

1C 207.6 þ 15.43a 3.77 þ 0.06a ND3

2 D 226.37 þ 8.05a 3.91 þ 0.03a ND3 M 214.84 þ 15.96a 3.91 þ 0.02a ND4 D + M 287.27 þ 8.10bc 3.96 þ 0.06a ND5 D + M+ C 278.54 þ 8.09a 3.84 þ 0.79a ND6 D + C 233.69 þ 0.00ad 3.85 þ 0.02a ND7 M + C 238.69 þ 7.64ae 3.82 þ 0.76a NDMaximum permitted conc. 75000.00 1000.00 1200.5

in fish as food (lg g)1)4

1SeeTable1for codes.2 Each value represents a mean þ SE; n = 3; numbers within the samecolumn not sharing the same superscript (a^e) are signi¢cantly di¡erent(P < 0.05).3 ND = not detected.4 National Health & Medical Research Council (1987).

G. Kibria et al.

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