Dietary Vitamin B 6 and Vitamin C
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Dietary Vitamin B, and Vitamin C
Influence on Immune Response and Disease
Resistance in Atlantic Salmon (Salmo salar)"
R. WAAGBP), K. SANDNES,b J. GLETTE,' E . R. N I L S E N ,
AND S. ALBREKTSEN
Institute of Nutrition
Directorate of Fisheries
P . O . Box 1900-Nordnes
N-5024 Bergen, Norway
blnstitute of Marine Research
Division of Aquaculture
P . O . Box 1870
N-5024 Bergen, Norway
University of Bergen
Center for Biotechnology
N-5020 Bergen, Norway
Vitamins C and B, are important for normal growth and development in Atlantic
salmon, and the minimum dietary requirement for young fish has recently been
established to be in the order of 10-20 mg/kg and 6-8 mglkg, respectively.'.?
The aim of the present study was to evaluate the effect of increasing dietary
vitamin supplementation on immunology and disease resistance in Atlantic salmon.
Growth, mortality, hematology (red blood cell count, hemoglobin, hematocrit),
serum-specific antibody level, serum hemolytic complement activity, and pronephros lysozyme activity were analyzed in both experiments, as well as challenge with
the pathogen bacteria Aeromonas salmonicida causing furunculosis in salm~nids.~.~
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STUDY I
Presmolt Atlantic salmon were fed a diet containing 70% fish meal supplemented with 0, 10, 20, 40, 80, and 160 mg/kg pyridoxine.HC1 (PN.HCI) for 5
months. The basal vitamin B, level was 2.3 mg/kg (PN). Vitamin B, in selected
organs determined after 17 weeks difFered significantly between the 0 and 10 mgl
kg groups only. Growth, mortality, and hematology were not affected by dietary
vitamin B,. Serum-specific antibody levels may indicate an optimum P N . H C L
level of 20-40 mglkg in Atlantic salmon feed, whereas serum hemolytic complement activity and head kidney lysozyme activity showed no differences according
to dietary treatment. Bacterial challenge showed n o clear effects of feeding different levels of vitamin B,. The results are summarized in TABLEI .
This project was kindly supported by F. Hoffmann-La Roche Ltd, Switzerland.
379
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TABLE 1.
Immunological Analyses of Atlantic Salmon Fed Different Levels of Vitamin Bh for Five Months
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Feeding Group (mg/kg)
Analysis
Hemoglobin (g/ 100 mL)
Hematocrit (percent)
Red blood cell count (*lo’’
cells/L)
Serum antibody level ( I :500 dilution; OD,)
Serum protein (g/L)
Serum complement activity (1 : 20
dilution; percent hemolysis)
Pronephros lysozyme activity (U/g
tissue)
Muscle aspartate amino transferase (U/g)
Percent cumulative mortality after
bacterial challenge
0
20
10
40
9.6 (0.2)a.‘
51 (2)”
I .35 (0.02)”
9.6 (0.2)a
49 (3)”
1.39 (0.04)”
9.6 (0.2)”
45 (I)”
1.31 (0.02)”
44
0.464 (0.048)”
0.334 (0.052)b
53.6 (2.7)”
54.3 (0.9)”
54.8 (I.8)”b
55.8 (1.1)”
8 856 (576)”
9 464 ( I 708)”
17
39
43
55
80
160
1.30 (0.03)”
9.7 (0.3)a
49 (2)a
1.30 (0.03)”
9.6 (0.3)a
49 (1)”
1.34 (0.02)”
0.387 (0.045)c
0.370 (0.038)‘
0.319 (0.048)b
0.270 (0.045)d
58.1 (1.4)b
56.7 (1.7)”
53.3 (1.8)”
56.0 (1 .OF
54.6 (4.6)ab
58.4 ( I .6)a
57.9 (4.6)h
57.4 (4.4)a
9 166 (758)a
10 386 (227)”
8 710 (271)”
40
39
38
44
51
45
43
47
8 612 (230)a
9.4 (0.2)a
‘ Mean values (SEM) with different superscript letters indicate significant differences (p < 0.05).
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TABLE 2. Immunological Analyses of Atlantic Salmon F e d Different Levels of Vitamin
C in t h e Form of
Ascorbate-2-Monophosphate (AP) f o r Six Months
Feedine Grouo (me / k d
Analysis
40
400
2000
4000
Hemoglobin (g/lOO mL)
Hematocrit (percent)
Red blood cell count (*lo” cellsiL)
Serum antibody level ( I : 500 dilution; OD,92)
Serum complement activity ( I : 20 dilution, percent hemolysis)
Pronephros lysozyme activity (Uig tissue)
Serum iron (mg/L)
Percent cumulative mortality after bacterial challenge
10.2 (0 .2)&d
53 (l)d
1.35 (0.03)a
0 630 (0.070)a
43.7 (4.2)a
10.0 (0.4)a
47 (2)b
1.28 (0.04)ab
0.654 (0.057)a
38.7 (3.8)d
10.5 (0.5)a
46 (2)b
1.22 (0.02)b
0.638 (0.061)a
41.8 (5.O)a
10.5 (0.6)”
48 (2)b
1.29 (O.OS)ab
0.778 (0.063)b
51.0 (2.2)b
6 734 (313)a
0.93 (0.11)”
20
9 518 (647)h
9 500 (697)b
I .29 (0. 18Yb
22
14 375 ( 1 462)’
1.75 (0.20)b
12
1.25 (0.
24
Mean values (SEM) with different superscript letters indicate significant differences (p < 0.05).
382
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ANNALS NEW YORK ACADEMY OF SCIENCES
STUDY I1
Presmolt Atlantic salmon were fed the same diet supplemented with graded
levels of ascorbate-2-monophosphate (AP), equivalent to 40,400,2000, and 4000
mg ascorbic acid/kg for 6 months. Growth, mortality, and hematology were not
affected by dietary vitamin C. Production of specific antibodies, serum hemolytic
complement activity, and lysozyme activity in pronephros tissue were higher in fish
fed the highest vitamin C level. Challenge with Aeromonas salmonicida showed
increased survival in the 4000 AP group. The results show that high dietary levels
of AP can favorably affect health in Atlantic salmon. The disease resistance was
not correlated with the ascorbic acid status in the liver. The results are summarized
in TABLE2 .
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REFERENCES
1. S A N t m E s , K., 0. J. TORRISSEN
& R. WAAGB@.
1992. The minimum dietary requirement
of vitamin C in Atlantic salmon (Salmo salar) fry using Ca ascorbate-2-monophosphate
as dietary source. Fish Physiol. Biochem. In press.
2. ALBREKTSEN,
S., R. WAAGB@
& K. SANDNES.1992. Vitamin B6 requirement of Atlantic
salmon (Salmo salar). Submitted.
3. ALBREKTSEN,
S., K. SANDNES,
J. GLETTE& R. W A A G B ~1992.
.
Influence of dietary
vitamin B, on immune response and disease resistance in Atlantic salmon (Salrno
salar). Submitted.
4. WAAGBB,R., J. GLETTE,E. R. NILSEN& K. SANDNES.1992. Dietary vitamin C,
immunity and disease resistance in Atlantic salmon (Salrno salar). Submitted.
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Dietary Vitamin B, and Vitamin C
Influence on Immune Response and Disease
Resistance in Atlantic Salmon (Salmo salar)"
R. WAAGBP), K. SANDNES,b J. GLETTE,' E . R. N I L S E N ,
AND S. ALBREKTSEN
Institute of Nutrition
Directorate of Fisheries
P . O . Box 1900-Nordnes
N-5024 Bergen, Norway
blnstitute of Marine Research
Division of Aquaculture
P . O . Box 1870
N-5024 Bergen, Norway
University of Bergen
Center for Biotechnology
N-5020 Bergen, Norway
Vitamins C and B, are important for normal growth and development in Atlantic
salmon, and the minimum dietary requirement for young fish has recently been
established to be in the order of 10-20 mg/kg and 6-8 mglkg, respectively.'.?
The aim of the present study was to evaluate the effect of increasing dietary
vitamin supplementation on immunology and disease resistance in Atlantic salmon.
Growth, mortality, hematology (red blood cell count, hemoglobin, hematocrit),
serum-specific antibody level, serum hemolytic complement activity, and pronephros lysozyme activity were analyzed in both experiments, as well as challenge with
the pathogen bacteria Aeromonas salmonicida causing furunculosis in salm~nids.~.~
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STUDY I
Presmolt Atlantic salmon were fed a diet containing 70% fish meal supplemented with 0, 10, 20, 40, 80, and 160 mg/kg pyridoxine.HC1 (PN.HCI) for 5
months. The basal vitamin B, level was 2.3 mg/kg (PN). Vitamin B, in selected
organs determined after 17 weeks difFered significantly between the 0 and 10 mgl
kg groups only. Growth, mortality, and hematology were not affected by dietary
vitamin B,. Serum-specific antibody levels may indicate an optimum P N . H C L
level of 20-40 mglkg in Atlantic salmon feed, whereas serum hemolytic complement activity and head kidney lysozyme activity showed no differences according
to dietary treatment. Bacterial challenge showed n o clear effects of feeding different levels of vitamin B,. The results are summarized in TABLEI .
This project was kindly supported by F. Hoffmann-La Roche Ltd, Switzerland.
379
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TABLE 1.
Immunological Analyses of Atlantic Salmon Fed Different Levels of Vitamin Bh for Five Months
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Feeding Group (mg/kg)
Analysis
Hemoglobin (g/ 100 mL)
Hematocrit (percent)
Red blood cell count (*lo’’
cells/L)
Serum antibody level ( I :500 dilution; OD,)
Serum protein (g/L)
Serum complement activity (1 : 20
dilution; percent hemolysis)
Pronephros lysozyme activity (U/g
tissue)
Muscle aspartate amino transferase (U/g)
Percent cumulative mortality after
bacterial challenge
0
20
10
40
9.6 (0.2)a.‘
51 (2)”
I .35 (0.02)”
9.6 (0.2)a
49 (3)”
1.39 (0.04)”
9.6 (0.2)”
45 (I)”
1.31 (0.02)”
44
0.464 (0.048)”
0.334 (0.052)b
53.6 (2.7)”
54.3 (0.9)”
54.8 (I.8)”b
55.8 (1.1)”
8 856 (576)”
9 464 ( I 708)”
17
39
43
55
80
160
1.30 (0.03)”
9.7 (0.3)a
49 (2)a
1.30 (0.03)”
9.6 (0.3)a
49 (1)”
1.34 (0.02)”
0.387 (0.045)c
0.370 (0.038)‘
0.319 (0.048)b
0.270 (0.045)d
58.1 (1.4)b
56.7 (1.7)”
53.3 (1.8)”
56.0 (1 .OF
54.6 (4.6)ab
58.4 ( I .6)a
57.9 (4.6)h
57.4 (4.4)a
9 166 (758)a
10 386 (227)”
8 710 (271)”
40
39
38
44
51
45
43
47
8 612 (230)a
9.4 (0.2)a
‘ Mean values (SEM) with different superscript letters indicate significant differences (p < 0.05).
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TABLE 2. Immunological Analyses of Atlantic Salmon F e d Different Levels of Vitamin
C in t h e Form of
Ascorbate-2-Monophosphate (AP) f o r Six Months
Feedine Grouo (me / k d
Analysis
40
400
2000
4000
Hemoglobin (g/lOO mL)
Hematocrit (percent)
Red blood cell count (*lo” cellsiL)
Serum antibody level ( I : 500 dilution; OD,92)
Serum complement activity ( I : 20 dilution, percent hemolysis)
Pronephros lysozyme activity (Uig tissue)
Serum iron (mg/L)
Percent cumulative mortality after bacterial challenge
10.2 (0 .2)&d
53 (l)d
1.35 (0.03)a
0 630 (0.070)a
43.7 (4.2)a
10.0 (0.4)a
47 (2)b
1.28 (0.04)ab
0.654 (0.057)a
38.7 (3.8)d
10.5 (0.5)a
46 (2)b
1.22 (0.02)b
0.638 (0.061)a
41.8 (5.O)a
10.5 (0.6)”
48 (2)b
1.29 (O.OS)ab
0.778 (0.063)b
51.0 (2.2)b
6 734 (313)a
0.93 (0.11)”
20
9 518 (647)h
9 500 (697)b
I .29 (0. 18Yb
22
14 375 ( 1 462)’
1.75 (0.20)b
12
1.25 (0.
24
Mean values (SEM) with different superscript letters indicate significant differences (p < 0.05).
382
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ANNALS NEW YORK ACADEMY OF SCIENCES
STUDY I1
Presmolt Atlantic salmon were fed the same diet supplemented with graded
levels of ascorbate-2-monophosphate (AP), equivalent to 40,400,2000, and 4000
mg ascorbic acid/kg for 6 months. Growth, mortality, and hematology were not
affected by dietary vitamin C. Production of specific antibodies, serum hemolytic
complement activity, and lysozyme activity in pronephros tissue were higher in fish
fed the highest vitamin C level. Challenge with Aeromonas salmonicida showed
increased survival in the 4000 AP group. The results show that high dietary levels
of AP can favorably affect health in Atlantic salmon. The disease resistance was
not correlated with the ascorbic acid status in the liver. The results are summarized
in TABLE2 .
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REFERENCES
1. S A N t m E s , K., 0. J. TORRISSEN
& R. WAAGB@.
1992. The minimum dietary requirement
of vitamin C in Atlantic salmon (Salmo salar) fry using Ca ascorbate-2-monophosphate
as dietary source. Fish Physiol. Biochem. In press.
2. ALBREKTSEN,
S., R. WAAGB@
& K. SANDNES.1992. Vitamin B6 requirement of Atlantic
salmon (Salmo salar). Submitted.
3. ALBREKTSEN,
S., K. SANDNES,
J. GLETTE& R. W A A G B ~1992.
.
Influence of dietary
vitamin B, on immune response and disease resistance in Atlantic salmon (Salrno
salar). Submitted.
4. WAAGBB,R., J. GLETTE,E. R. NILSEN& K. SANDNES.1992. Dietary vitamin C,
immunity and disease resistance in Atlantic salmon (Salrno salar). Submitted.