Dietary lysine requirement of juvenile Beluga (Huso huso) M. Mohseni1&2; M. Bahmani2; M. H. Hassani2; H.R. Pourali2; J. H. Lee1 and Sungchul C. Bai1 (1) Department

of Aquaculture/Feeds & Foods Nutrition Research Centre, College of Fisheries Science, Pukyong National University, Nam-gu, Busan 608-737 Rep. of Korea

(2)

International Sturgeon Research Institute, P.O. Box 41635-3464, Rasht, Guilan, Iran

1

 Sturgeon are Chondrosten fishes and belong to :

Acipenseriformes

Chondrosteridae

Acipenseridae

Polydontidae

(Traquair 1877)

(Bonaparte 1831)

(Bonaparte 1838)

Extinct

25 species

2 species

(known as fossils from lower Jurassic)

Caviar production region, Caspian Sea: Russia, Iran, Kazakhstan, Turkmenistan & Azerbaijan are responsible for about 90 percent of the world’s black caviar production

35,000

30,000

25,000

MT

20,000

15,000

10,000

5,000

19 50 19 52 19 54 19 56 19 58 19 60 19 62 19 64 19 66 19 68 19 70 19 72 19 74 19 76 19 78 19 80 19 82 19 84 19 86 19 88 19 90 19 92 19 94 19 96 19 98 20 00 20 02 20 04 20 06

0

Years

Countries: Azerbaijan, Canada, Denmark, France, Germany, Iran, Kazakhstan, Russian Federation, United States of America, etc.

Species: Persian, Russian, Sterlet sturgeons, Beluga, Green sturgeon, paddlefish, White, Lake Sturgeon, etc. FAO statistics

A (Over fishing - Illegal catch & Poaching)

B Construction of dams and bridges across main rivers (Volga River, Kura River, Sefidroud River; Anadromous species, have a problem during migration due to loss of natural spawning grounds)

C Pollution: – Urban & Oil)

(Agricultural - Industrial

Ton

1984

150

1985

150

1986

180

1987

160

1988

1,098

1989

260

1990

328

1991

385

1992

412

1993

495

1994

892

1995

1,135

1996

1,297

1997

2,025

1998

2,034

1999

2,465

2000

3,158

2001

3,091

2002

3,816

2003

14,747

2004

15,551

2005

19,898

2006

21,319

25000

20000

15000

MT

Year

10000

5000

19 84 19 85 19 86 19 87 19 88 19 89 19 90 19 91 19 92 19 93 19 94 19 95 19 96 19 97 19 98 19 99 20 00 20 01 20 02 20 03 20 04 20 05 20 06

0

Years

Countries: Austria, Iran, Azerbaijan, Belgium, Bulgaria, China, Estonia, France, Germany, Hungary, Italy, Latvia, Poland, Romania, Russian Federation, Slovenia, Spain, Ukraine, United States of America, Uruguay, Kazakhstan

Species: White sturgeon, Siberian sturgeon, Sterlet Beluga, Bester, Danube sturgeon (=Osetra), Russian and Ship Sturgeon…….

FAO statistics

Commercial value of Sturgeon Caviar of Sturgeon Most sturgeon species have been highly valued for their eggs, which are processed into caviar * Beluga (Huso huso) : 4000~6000 $/kg * Osetra (Acipenser spp.- gueldenstaedti , persicus) ; 2000~3000$/kg * Sevruga (Acipenser stellatus): 800~1000 $/kg Composition: 40~55% (Water), 25~30% (Protein), 10~14% (Fat), Energy 2,500 kcal/kg

Iran Released 20 to 25 million of 2 to 3g sturgeon fingerlings • About 50 applicants for rearing sturgeon • Five farms working to develop domesticated brood stock • Eight farmers for beluga as food fish

Proteins Fats

Carbohydrates Nutrition

Vitamins

Minerals

Protein is the most expensive component of fish diets, providing essential amino acids, nitrogen to synthesize non-essential amino acids, sulfur & energy

 Amino acids fulfill multiple metabolic

functions in the human body Amino acids act as building blocks for proteins and as precursors of hormones, neurotransmitters, antioxidants, nucleic acids and other complex body constituents  They are also a very important energy source

 Essential amino acids Argenine Leucine Isoleucine

Lysine Methionine Phenylalanine Threonine Tryptophan Valine

Histidine

Non-essential amino acids: Alanine, Asparagine, Aspartate, Cysteine, Glutamate, Glutamine, Glycine, Proline, Serine and Tyrosine (21st and 22nd AAs have been proposed).

Dietary lysine requirements in Fish Species

Source

Lysine (g kg-1 protein)

References

Jian Carp

Coated L-lysine

59

Zhou et al. 2008

Turbot

L-lysine- HCL

56

Peres and Teles 2008

Gilthead sea bream

Crystalline L-lysine- HCL

50.4

Marcouli et al. 2006

Japanese sea bass

Crystalline L-lysine

58 – 60.7

Mai et al. 2006

Beluga

L-lysine- HCL

Sodagar et al. 2006

Grass carp

Crystalline L-lysine

Wang et al. 2005

54.4 – 58.9

Objectives To determine the dietary lysine requirement based on growth, body composition and feed efficiency ratio in sub-year beluga during growout phase

Experimental Design Fish weight: 1.2 ± 0.02 g (mean ± SE) Rearing Density: 270 fish / 60-L tank (triplicates) Feeding Rate: 2~3 %; four times per day (02:00, 08:00, 14:00 and 21:00 h). Temperature: 19 ± 1.5 ℃ Duration: 8 weeks

Weight check: 15-day intervals 6 diets: 1.4, 1.95, 2.51, 3.06, 3.7 and 4.29 g /100 g of dry diet

Formulation of Experimental Diets Ingredients (g kg -1 dry diet)

14

19.5

25.1

30.6

37

42.9

Fish meal Meat meal Wheat gluten

300 90 100

300 90 100

300 90 100

300 90 100

300 90 100

300 90 100

Milk powder

30

30

30

30

30

30

Soybean meal Yeast Wheat meal Molasses

130 50 100 30

130 50 100 30

130 50 100 30

130 50 100 30

130 50 100 30

130 50 100 30

Cellulose

37.5 0 27.5

37.5 5.5 22

37.5 11 16.5

37.5 16.5 11

37.5 22 5.5

37.5 27.5 0

80

80

80

80

80

80

L-lysine – Hcl Glutamic acid Oil (Fish oil/ corn oil 1:1)

Equal amino acid nitrogen was maintained by replacing lysine with glutamic acid Vitamin premixture and mineral premixture mixed 20 & 5 g kg -1 in the diet, respectively (Mohseni et al. 2006)

Diet composition: Dry mater (%)

91.7

92.4

91.9

91.8

91.3

91.5

Crude protein (%)

44.7

45

45.2

45.5

45.2

45.3

Crude lipid (%)

15.3

15.1

15.3

15.3

15.2

15.1

Crude ash (%)

9.5

9.8

9.9

9.7

9.3

9.6

Gross energy (kJ/g diet)

19.9

19.8

19.9

19.8

19.9

19.9

L-crystalline amino acids and salt mixture were thoroughly stirred in hot water (80 ºC) in a steel bowl attached to a mixer for 15 min All ingredients mixed in a mixer and pelleted with a CPM pelletizer

Parameters Growth parameters: WG (%) = 100 × (final weight - initial weight)/initial weight SGR (% BW day-1) = 100 × [(Ln final weight – Ln initial weigh)/day] FE = 100 × weight gain (g) / feed intake (g) PER = wet weight gain (g) / Protein intake (g). LPV = fish lipid gain / lipid intake. PD = 100 × protein gain (g) / protein intake (g)

Whole-body proximate analysis (AOAC method) Amino acid analysis (amino acid analyzer)

Statistical Analysis One-way ANOVA test using SPSS 10.0 software Duncan Multiple range test Treatment effects were considered significant at P < 0.05 Broken line analysis & second order polynomial regression analysis

Results WG, SGR, FE, PER, PPV, PD and LPV of Juvenile beluga fed diets containing six levels of lysine (g lysine kg-1 crude protein)

14

19.5

25.1

30.6

37

42.9

Pooled SEM

777.9c

991.5 b

998.7 b

1235.6 a

1274.4 a

931.6 bc

43.5

SGR (% day-1)

3.6 c

4b

4b

4.3 a

4.4 a

3.9 b

0.063

FE (%)

69.1 b

69.6 ab

73.1 ab

74.2 a

73.4 ab

69.8 ab

0.678

PER

1.9 c

2.5 b

2.5 b

3.1 a

3.2 a

2.3 bc

0.109

131.3 b

148.3ab

153.2 ab

173.2 a

169.9 a

146.9 ab

4.52

9.61 a

8.26 ab

6.71 bc

5.06 c

5.96 c

8.69 a

0.44

WG (%)

PD LPV

Figure 1. Optimum dietary Lysine requirement of juvenile beluga, based on the broken-line regression model of percent weight gain 1600

y = 14.715x + 749.07

R2 = 0.7746

WG (%)

1200

800

400

0 0

10

20

30

Dietary Lysine (g kg -1)

40

50

1,400

y = -146.2x2 + 921.6x - 258.4 R² = 0.763

1,200

Weight gain (%)

1,000 800 600 400

3.1 200 0

0

0.5

1

1.5

2

2.5

3

3.5

4

4.5

5

Lysine in diet (%)

Figure 2. The second order polynomial regression (solid curved line) of mean weight gain and dietary lysine levels for juvenile beluga fed different levels of dietary lysine for 8 weeks. Values are mean±SEM, n=3

Whole Body Proximate Composition Unit : % (g lysine kg-1 crude protein)

14

19.5

25.1

30.6

37

42.9

Pooled SEM

Moisture

75.14ab

72.65b

73.51b

76.21a

72.61 b

73.15b

0.446

Crude protein

14.22b

14.91ab 15.10ab 15.91a

15.77 a

14.85ab

0.182

Crude lipid

5.6 a

5.61a

5.2 ab

4.59 b

5.1ab

5.63 a

0.118

Ash

2.91

2.97

3.31

3.74

3.82

3.45

0.141

HSI %2

2.36

2.41

2.41

2.39

2.38

2.37

0.01

Essential amino acid profiles in the muscle of Beluga fed the experimental diets for 8 weeks (g per 16 g N) 14

19.5

25.1

30.6

37

42.9

Pooled SEM

Arginine

4.98

4.96

4.95

4.99

5.04

5.01

0.015

Histidine

3.30

3.28

3.31

3.34

3.31

3.29

0.008

Isoleucine

3.24

3.25

3.33

3.27

3.32

3.31

0.015

Leucine

6.62

6.63

6.70

6.67

6.67

6.71

0.011

(g lysine kg-1 crude protein)

Lysine Methionine

7.22d 7.63 cd 7.85 bc 8.37 a 8.41 a 8.20 ab 0.115 0.011

2.54

2.56

2.55

2.61

2.54

2.60

2.96

2.98

2.99

3.09

2.98

2.98

Threonine

3.57

3.60

3.62

3.63

3.59

3.62

0.011

Valine

3.72

3.73

3.75

3.74

3.73

3.73

0.006

Phenylalaine

0.008

Discussion & Conclusions  WG is the most frequently chosen response criterion for evaluating lysine requirements in fish (Hauler and Carter, 2001; Peres and Teles 2008) and the same criterion was used in the present study

 Growth of beluga fingerling responded positively to dietary lysine levels

 Fingerling Beluga fed 30.6 and 37 g lysine kg-1 diet for 8 wks showed significantly higher WG, SGR & PER than other groups

FE was unaffected by dietary treatment, except in fish fed 14 g lysine kg-1 of diet that was lower than in the other experimental groups

According to these results, the optimum lysine requirement in beluga was estimated to be 6.42

g/100 g of dietary protein

 kaushik et al. (1991) used the daily whole body increment to estimate

the

lysine

requirement

of

22

g

Siberian

sturgeon to be 5.4 g/100g dietary protein.

 Ng and Hung (1995) using a similar method, found the lysine requirement of 67 g white sturgeon to be 5.4 g/100g dietary protein

Whole body proximate analysis Whole body protein content was positively correlated with dietary lysine levels, while lipid content was negatively correlated with it.

Lysine content of fish muscle was significantly affected by dietary lysine levels (P