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Animal-protein-free feeds for hybrid tilapia (Oreochromis niloticus × O. aureus) in intensive culture
Aquaculture, 75 (1988) 115-125 Elsevier Science Publishers B.V., Amsterdam -
115 Printed in The Netherlands
Animal-Protein-Free Feeds for Hybrid Tilapia (Oreochromis niloticusx 0. aureus) in Intensive Culture S. VIOLAl, Y. ARIELI’ and G. ZOHAR’ ‘Israeli Feedmills Association, Miloubar Central Feedmill, D.N. Ashrat 25201 (Israel) ‘Intensive Fishculture Station, Department of Fisheries, P.O.B. 33, Tiberias (Israel) (Accepted 15 February 1988)
ABSTRACT Viola, S., Arieli, Y. and Zohar, G., 1988. Animal-protein-free feeds for hybrid tilapia (Oreochromis niloticus x 0. aureus) in intensive culture. Aquaculture, 75: 115-125. Growth trials were carried out with tilapia in experimental ponds to develop an animal-proteinfree diet of nutritive value equal to a standard commercial fish-meal feed. A diet based on soybean meal and supplemented with amino acids, oil and di-calcium phosphate resulted in growth performances and body compositions equal to those on a fish-meal diet. Successive withdrawal of the supplements resulted in a final fish-meal-free diet, with 3% di-calcium phosphate and 2% oil as the only supplements necessary to achieve growth performances equal to those obtained with a fully supplemented diet.
INTRODUCTION
Many studies have been made to determine optimal protein levels for tilapia. Jauncey (1982) found 40% to be optimal for fish of 2 g in aquaria. Winfree and Stickney (1981) recommended 34% protein for fish of 7.5 g, and Jackson et al. (1982) 30% for fish of 30 g. In our studies with market-size fish in ponds we found 30% optimal (Viola and Zohar, 1984a). Tilapia are known to be much more herbivorous than catfish or carp (Jauncey and Ross, 1982). However, in many countries feeds for tilapia include a substantial proportion of animal proteins such as fish meal, poultry meal, meat meal or similar offal meals. These animal proteins usually increase the cost of the feed, yet low production costs are essential, especially for industrial processing. Jackson et al. (1982) tried to substitute different plant proteins for fish meal. At low levels of replacement (25%) growth rates were similar, but at higher inclusions the performance was considerably lower. They did not try to compensate for any imbalances of amino acids or minerals. Appler and
0044-8486/88/$03.50
0 1988 Elsevier Science Publishers B.V.
116
Jauncey (1983) replaced fish meal with increasing proportions of algal meal. Growth rates decreased proportionally when more than 5% algae were included. Under practical culture conditions in Israel, replacement of half the fish meal by soybean meal has been found nutritionally equivalent, without any supplementation (Viola and Arieli, 1983a). Attempts to determine essential amino acid requirements of tilapia fry have been reported (Jackson and Capper, 1982; Jauncey et al., 1984). However, the requirements of market-size tilapia for amino acids in relation to energy or the bioavailabilities of nutrients in diverse feedstuffs have not been clarified sufficiently. Thus the application of linear programming of feeds is not yet possible for tilapia, as it is for catfish (Robinette, 1984). Therefore, substitution and supplementation studies are the next best method to pinpoint critical nutrient levels under practical intensive culture conditions. Our preliminary trials of complete replacement of fish meal by soybean meal in tilapia feeds, on the basis of protein alone, were performed in cages of 1 m3, suspended in ponds, 100 fishes per cage, receiving daily rations of equal weight. These trials yielded inferior growth rates and feed conversions - in inverse relation to the degree of replacement - indicating nutrient imbalances (Table 1). Previous work with carp (Viola et al., 1982) had shown that replacement of fish meal by soybean meal created imbalances of amino acids and energy. When TABLE 1 Preliminary trials in cages: replacement of fish meal by soybean meal on the basis of protein alone (25% protein, means of 4 replicates, 100 fish/cage, 30-31 days) First trial Diet
25% Fish meal
40% Soybean meal Alone
+ Oil + Lysine 165 208 43b 1.4 3.2
Initial weight of fish (g) Final weight of fish (g) Gain in weight of fish (g) Daily gain (g ) Feed conversion
165 218 53” 1.7 2.6
165 205 4oh 1.3 3.4
Second trial Diet
Fish meal
Soybean meal
Initial weight of fish (g) Final weight of fish (g ) Gain in weight of fish (g) Daily gain (g)
240 307 67” 2.2
250 290 5ob 1.7
Values followed by the same superscript are not significantly different (Newman-Keul’s P
test,
117
these deficiencies were balanced by supplements of lysine, methionine and oil, isonutritious carp feeds without animal protein could be compounded. On the other hand, studies with tilapia had shown that they responded to phosphorus supplementation to feeds that contained plant proteins (Viola et al., 1986a). From this work the conclusion was drawn that plant phosphorus (phytin) has very low availability for tilapia and that supplementation of available phosphorus was essential in animal-protein-free diets. In order to assess the essentiality of further supplements, e.g. lysine, methionine, oil or others, a program was designed for the progressive omission of one supplement after another from a fully supplemented positive control diet. The final diet was to contain as few supplements as necessary to achieve equally good growth, feed conversion and retentions of protein and energy. METHODS
AND MATERIALS
Hybrids of Oreochromis niloticus and 0. aureus, 95% males, were raised in 12 experimental ponds of 200 m2 and depth of 1 m, at a density of 400 fish per pond. All the fish were counted and weighed at the start and end of each trial. TABLE 2 Composition of feeds for the first series of trials: complete balance of nutrients in low-fish-meal diets (% air-dry matter) Diet
Fish protein 50%
Ingredients Fish meal (72% protein) Soybean meal (44% protein ) Wheat Milocorn Oil (poultry) Di-calcium phosphate Lysine-HCl Methionine Nutrients Protein Fat Ash Phosphorus Methionine’ Lysinel Gross energy (kcal/kg)’ 1 Estimated according to NRC-NAS
20 24 10 46 _ _
29.5 4.0 5.2 0.75 0.7 1.8 4150 (1977,1983).
25% 10 42 10 33.5 3 1 0.25 0.15 29.7 6.0 5.1 0.75 0.7 l-8 4150
0%
60 10 21 6 2 0.5 0.3 29.2 7.5 5 0.75 0.7 1.8 4150
118 TABLE 3 Composition of feeds for the final trial (No. 6) in the second series: progressive omission of supplements from fish-meal-free diets (0 = oil, M = methionine, L = Lysine, P = DCP) (% air-dry matter) Diet
Supplements: Omission:
Ingredients Soybean meal (44% protein) Wheat Milocorn Di-calcium phosphate Oil Methionine Lysine-HCl Nutrients Protein Fat Ash Calcium Phosphorus Methioninel Lysinel Gross energy (kcal/kg)’
O+M+L+P (Balanced) 60 10 21.25 3 5 0.25 0.5 30.3 6.3 6.4 1.0 1.0 0.7 2.3 4100
M+L+P 0 60 10 24.25 3 2 0.25 0.5 29.5 3.4 6.2 N.D. N.D. 0.7 2.3
L+P
P
GM
O,M,L
60 10 24.25 3 2
60 10 25.0 3 2
0.5 29.5 3.4 6.0 N.D. N.D. 0.45 2.3 3950
29.2 3.5 6.2 N.D. N.D. 0.45 1.8
‘See Table 2. N.D. - not determined.
Duration of a trial was 55-65 days. Feeds were dispensed in equal amounts to each pond by automatic feeders. Samples of 150-200 fishes were weighed biweekly for adjustment of the ration. At the start of a trial, the average weight was equal in all ponds, between 100 and 300 g. Each dietary treatment was replicated in three ponds. Fresh water was added only to replace losses by evaporation and seepage. Water temperatures at 6 a.m. were between 21 and 23”C, rising to 24-26” C at noon. The ponds were aerated by paddle wheels. Oxygen levels were monitored and never below 4 ppm; the pH of the water was around 7.5. At the start and at the end of each trial, five fishes from each treatment were sampled for laboratory analysis. Moisture was determined at 105°C for 24 h. protein by Kjeldahl’s method (Kjeltec ) , fat by extraction with perchloro-ethylene (Fosslet), ash by incineration at 550” C, phosphorus by molybdovanadate spectrophotometry, and calcium by EDTA titration (AOAC, 1984). The feeds were pelleted in a Lister laboratory pellet mill (4 mm pellets) and analysed by the same methods. The contents of amino acids and gross energy were estimated from tables (NRC-NAS, 1977,1983).
119 TABLE 4 Composition of feeds for third series of trials Diet Ingredients ( % ) Fish meal (65% protein) Soybean meal (44% protein) Wheat Milocorn Wheat bran Di-calcium phosphate Poultry oil Binder Nutrients ( % ) Crude protein Ether extract Crude fiber Ash Calcium Phosphorus Lysinel Methionine and cysteinel Gross energy (kcal/kg)’
Fish meal
Mixed
Soybean meal
Soybean meal +DCP
_
_
20 32 10
23 20 20 24 10
55 20 12 10
1 2
1 2
1 2
55 20 8 10 4 1 2
30.2 5.0 2.7 7.6 1.5 1.3 2.0 1.15 4000
31.1 4.6 3.5 7.1 1.35 1.0 1.9 1.1 4050
28.8 2.8 5.7 4.6 0.45 0.6 i.75 0.9 4050
30.0 2.9 5.5 9.3 1.65 1.6 1.75 0.9 3850
35
‘See Table 2.
The research plan comprised three series of trials. In the first series, soybean meal was increased progressively to replace fish meal. The latter provided 50%) 25 % and 0% of the total protein. Deficiencies of amino acids, energy and phosphorus were compensated by synthetic lysine, methionine, oil and di-calcium phosphate (DCP). The composition of the diets is shown in Table 2. The trial was repeated three times (trials 1,2,3). The second series was to show which of the four supplements were really essential for equal performances. This series also contained three trials: no. 4, omission of one supplement (for oil reduction from 5% to 2% ); no. 5, omission of two supplements; no. 6, omission of all supplements except DCP, since phosphorus supplementation had been found essential in previous studies, as mentioned before. The reduction of oil to 2% minimum was intended to safeguard essential fatty acids. The composition of the diets for the last trial (no, 6) is shown in Table 3. The diets for the intermediate trials no. 4 and 5 were composed correspondingly. The findings of the second series were put to a final test in the third series. This contained two trials of 54 days: animal-protein-free diets with and without di-calcium phosphate were compared with mixed (standard) diets and with
120
full fish meal diet without soybean meal. Table 4 shows the composition of the diets. RESULTS
The performance of the fishes in trial no. 1 is shown in Table 5. All three diets yielded almost equal growth rates, feed conversions, and retentions of protein and energy. The other two trials (nos. 2 and 3)) under slightly different conditions of initial weights and temperature, gave the same general result: equal performances with all three diets. This series proved that no imbalances other than lysine, methionine, oil and phosphorus were incurred under the conditions of the trial. The results of the second series are shown in Tables 6 and 7. Again, no differences were apparent between the diet groups in any trial of this series. Body analyses were carried out in the last trial only. The conclusion from the second series, that phosphorus supplementation alone was critical, was finally tested in the third series. Both trials of this series yielded similar results and only one of them is reported here in Table 8. The soybean meal diet without DCP caused severe retardation of growth and bone mineralization, accompanied by a slight accumulation of fat. All these symptoms were prevented by the addition of 4% DCP. The full fish meal diet did not excel the other diets in any performance parameter. TABLE 5 Performance of tilapia in the first trial (no. 1) of the first series (means and S.D. of 3 ponds X 400 fish, 60 days, 2.5% feeding rate) Diet
Initial weight (g ) Final weight (g) Gain (g) S.G.R (%) Daily gain (g ) Feed conversion Protein in body (% ) Fat in body (% ) Energy retention ( % )2 Protein retention (% )”
Fish protein 50%
25%
0%
77 188 111+2 1.47 1.85 1.65 17.6 8.5 42.0 42.0
77 190 113-t3 1.48 1.88 1.65 17.3 8.0 42.0 42.0
77 188 111*4 1.47 1.85 1.70 17.5 8.5 41.0 41.0
‘S.G.R. ( =specific growth rate) = (In final weight-ln initial weight)/days 2Retention = body gain/intake ( % ). Factors for calculation: protein = 5.7 kcal/g; fat = 9.5 kcal/g (NRC-NAS,
(%). 1977).
121 TABLE 6 Performance of tilapia in trials with progressive omission of supplements (second series) Trial no. 4 - omission of one supplement (means and S.D. of 3 ponds X 400 fish, 62 days) Diet designation: Omitted supplement: Initial weight (g ) Final weight (g ) Total gain (g)
M+L+O+P -
M+L+P 0
M+O+P L
84 187 10326
84 183 99t-12
84 182 9821
o+L+P M 84 183 99 + 8 N.S.
Trial No. 5 - Omission of two supplements (means and SD. of 3 ponds x 400 fish, 54 days) Diet designation: Omitted supplement: Initial weight (g ) Final weight (g ) Total gain (g )
M-tL+O+P -
M+P
175 295 120+4
175 289 114+5
0,L
o+p M,L 175 287 112+5
L+P M,O 175 300 125 k 3 N.S.
N.S. - differences not significant. TABLE 7 Performance of tilapia in the last trial (no. 6) of the second series: progressive omission of supplements (means and S.D. of 3 ponds X 400 fish, 56 days, 2% feeding rate) Dietdesignation: O+M+L+P Omitted supplement: Initial weight (g) Final weight (g ) Gain (g) S.G.R. (%)l Daily gain (g ) Feed conversion Protein in body ( % ) Fat in body (% ) Energy retention (% )’ Protein retention (g ) 1
275 469 194k 11 0.95 3.5 2.35 17.3 12.0 26.7 27.0
Mi-L+P 0
L+P
P
0,M
O,M,L
275 459 184-tlO 0.9 3.3 2.45 17.8 10.5 23.0 25.9
275 464 189+8 0.92 3.4 2.4 17.8 10.5 24.2 27.3
275 467 192 k 7 N.S. 0.95 3.45 2.35 18.2 12.0 25.4 27.6
‘See Table 5. DISCUSSION
The final conclusion arising from this study is that the only essential supplement was phosphorus. This result is strikingly different from our previous study with carp and raises several points for discussion.
122 TABLE 8
Growth performance of tilapia in the third series of trials (means and standard deviations of 3 ponds x 400 fish, 196 g at start, 54 days, 2.5% feeding rate) Diet
Fish meal
Mixed
Soybean meal
Soybean meal +DCP
Total weight gain (g ) Standard deviation (g ) Average daily gain (g ) Specific growth ( % ) ’ Feed conversion
175.2” 15.5 3.2 1.18 1.9
175.2” 9.4 3.25 1.2 1.85
137.3b 10.0 2.5 1.0 2.35
167.4” 3.8 3.1 1.15 2.0
Body composition: Protein (% ) FaL (W) Ash (%) Calcium (%) Phosphorus ( % ) Energy retention (% )” Protein retention (% )”
17.2 9.7 3.6 2.1 0.55 29.3 31.4
17.3 9.7 3.2 2.0 0.52 29.3 31.1
17.8 10.5 2.8 1.6 0.45 26.8 28.4
18.8 9.2 3.5 2.0 0.55 29.7 33.3
‘,‘See Table 5. Values followed by the same superscript are not significantly different.
Oil as source
of energy
Carp had been found very responsive to oil supplementation (Viola et al., 1981), yet the reduction of oil in the tilapia diets had no effect at all on the performance of the fishes. This confirms previous findings on the poor utilization of oil by tilapia (Viola and Arieli, 198313). Consequently, the oil content of fish meal and other meat meals, which are valued highly for poultry, carp or catfish, has very little worth for tilapia. Hemicelluloses and oligosaccharides as sources
of energy
On the other hand, 60% soybean meal in the diets contributed a great amount of cellulose, hemicellulose and oligosaccharides. Crude fiber in the soybean meal amounted to 6.5% and nitrogen-free extract to 30%. According to Smith and Circle (1978), sucrose may reach 6% and starch is negligible, leaving 24% oligosaccharides and hemicelluloses. These are considered almost indigestible for monogastric animals. Lodhi et al. (1969) found a digestibility of 14% for the nitrogen-free extract for poultry - which again leads to an estimate of 26% indigestible carbohydrates. Together with the crude fiber, 60% soybean meal contained, therefore, about 18% complex carbohydrates. In the nutrition of carp, these polysaccharides are almost indigestible and created large deficits
123
of energy that could be balanced by oil (Viola et al., 1982). However, in the first series of our trials with tilapia, feeding rates were the same and therefore any deficit of energy would have asserted itself in substantially decreased growth performance. Thus it may be concluded that these polysaccharides were at least partly utilized for energy. Similar conclusions were found in previous experiments with high-fiber feeds: 60% wheatbran caused only slight impairment of growth (Viola and Zohar, 198413). Popma (1982)) too, found increased carbohydrate digestibility of fibrous feeds for tilapia, as compared to carp. Lysine In our diets without supplemental lysine, the total lysine was higher than the requirements established by Jackson and Capper (1982) - 1.62% - or by Jauncey et al. (1984) - 1.51% - with synthetic or semisynthetic diets. In carp feeds, when fish meal was replaced by soybean meal, lysine was found to be deficient, although the total lysine was higher than the requirements (Viola et al., 1982). Probably the stomachless carp is at a disadvantage in utilizing lysine complexes created by Maillard condensations during the thermal treatment in soybean meal production. In contrast, we were not able to prove that lysine was critical for tilapia in any practical combination of feedstuffs and protein levels. Be it due to the lower specific growth rate or to their stronger gastric digestion, tilapia appear to derive all their necessary lysine from soybean meal or even from low-gossypol cottonseed meal (Viola and Zohar, 1984a). Phosphorus - the neglected nutrient Ogino et al. (1979) found that both bone phosphorus and plant phosphorus were of very low availability to stomachless carp. Therefore, replacement of one by the other does not change the supply. Actually, neglect of supplementation of soluble mineral phosphorus to both kinds of diet for carp has serious consequences of growth depression (Hepher and Sandbank, 1984; Viola et al., 1986b). Tilapia do produce acid gastric juice and therefore are able to utilize bone phosphorus to a much higher extent (Viola et al., 1986a), but not phytin. Thus, substitution of animal meals, which contain bones, by seed proteins creates a phosphorus deficiency, which must be balanced by suitable mineral supplements. This turned out to be the only really critical nutrient when fish meal was replaced by soybean meal. This feature has escaped the attention of many fish nutritionists and might explain unsatisfactory results, such as those of Jackson et al. (1982), in which fish meal was replaced completely or almost completely by plant proteins.
124 ACKNOWLEDGMENT
Our thanks to Mrs. D. Yishar and the staff of the “Milouda” their technical assistance.
laboratory for
REFERENCES AOAC, 1984. Official Methods of Analysis, 14th edn. S. Williams (Editor). Association of Official Analytical Chemists, Arlington, VA 1102 pp. Appler, H.N. and Jauncey, K., 1983. The utilization of a filamentous green alga (Cladophora glomerata (L. ) Kutzin) as a protein source in pelleted feeds for Surotherodon (Tilapia) niloticus fingerlings. Aquaculture, 30: 21-30. Hepher, B. and Sandbank, S., 1984. The effects of phosphorus supplementation to common carp diets on fish growth. Aquaculture, 36: 323-332. Jackson, A.J. and Capper, B.S., 1982. Investigations into the requirements of the tilapia Sarotherodon mossambicus for dietary methionine, lysine and arginine in semi-synthetic diets. Aquaculture, 29: 289-297. Jackson, A.J., Capper, B.S. and Matty, A.J., 1982. Evaluation of some plant proteins in complete diets for the tilapia Surotherodon mossumbicus. Aquaculture, 27: 97-109. Jauncey, K., 1982. The effects of varying dietary protein level on the growth, food conversion, protein utilization and body composition of juvenile tilapia (Surotherodon mossumbicus). Aquaculture, 27: 43-54. Jauncey, K. and Ross, B., 1982. A Guide to Tilapia Feeds and Feeding. Institute of Aquaculture, Univ. Stirling, Scotland, 111 pp. Jauncey, K., Tacon, A.C.J. andJackson, A.J., i984. The quantitative essential amino acid requirements of Oreochromis (Sarotherodon) mossambicus. Int. Symp. Tilapia in Aquaculture, TelAviv Univ., pp. 328-337. Lodhi, G.N., Renner, R. and Clandinine, D.R., 1969. Available carbohydrate in rapeseed meal and soybean meal as determined by a chemical method and a chick bio-assay. J. Nutr., 99: 413418. NRC-NAS (National Research Council-National Academy of Sciences), 1977. Nutrient Requirements of Warmwater Fishes. National Academy Press, Washington, DC, 77 pp. NRC-NAS (National Research Council-National Academy of Sciences), 1983. Nutrient Requirements of Warmwater Fishes and Shellfishes. National Academy Press, Washington, DC, 94 PP. Ogino, C., Takeuchi L., Takeda, H. and Watanabe, T., 1979. Availability of dietary phosphorus in carp and rainbow trout. Bull. Jpn. Sot. Sci. Fish., 45 (12): 1527-1532. Popma, T.J., 1982. Digestibility of selected feedstuffs and naturally occurring algae by tilapia. D.Sc. Thesis, Auburn Univ., Auburn, AL, 78 pp. Robinette, H.R., 1984. Feed formulation and processing. In: E.H. Robinson and R.T. Love11 (Editors), Nutrition and Feeding of Channel Catfish. South. Coop. Ser. Bull., No. 296, Auburn Univ., Auburn. AL, pp. 29-39. Smith, A.K. and Circle, S.J., 1978. Soybeans: Chemistry and Technology, Vol. 1. Avi Publ., Westport, CT, p. 81. Viola, S. and Arieli, Y., 1983A. Nutrition studies with tilapia (Sarotherodon). 1. Replacement of fish meal by soybean meal in feeds for intensive tilapia culture. Bamidgeh, 35 (1): 9-17. Viola, S. and Arieli, Y., 1983B. Nutrition studies with tilapia hybrids. 2. The effects of oil supplements to practical diets for intensive aquaculture. Bamidgeh, 35 (2): 44-52.
125 Viola, S. and Zohar, G., 1984a. Nutntion studies with market size hybrids of tilapia (Oreochromis) in intensive culture. 3. Protein levels and sources. Bamidgeh, 36 (1): 3-15. Viola, S. and Zohar, G., 198413.Report to the Ministry of Science in Israel (in Hebrew, unpublished). Viola, S., Rappaport, U., Arieli, Y., Amidan, G. and Mokady, S., 1981. The effects of oil-coated pellets on carp (Cyprinus carpio ) in intensive culture, Aquaculture, 26: 49-65. Viola, S., Mokady, S., Rappaport, U. and Arieli, Y., 1982. Partial and complete replacement of fish meal by soybean meal in feeds for intensive culture of carp. Aquaculture, 26: 223-236. Viola, S., Zohar, G. and Arieli, Y., 1986a. Phosphorus requirements and its availability from different sources for intensive pond culture species in Israel. Part I. Tilapia. Bamidgeh, 38 (1): 3-12. Viola, S., Zohar, G. and Arieli, Y., 1986b. Requirements of phosphorus and ita availability from different sources for intensive pond culture species in Israel. Part II. Carp culture. Bamidgeh, 38 (2): 44-54. Winfree, R.A. and Stickney, R.R., 1981. Effects of dietary protein and energy on growth, feed conversion efficiency and body composition of Tilapia aurea J. Nutr., 111: 1001-1012.
115 Printed in The Netherlands
Animal-Protein-Free Feeds for Hybrid Tilapia (Oreochromis niloticusx 0. aureus) in Intensive Culture S. VIOLAl, Y. ARIELI’ and G. ZOHAR’ ‘Israeli Feedmills Association, Miloubar Central Feedmill, D.N. Ashrat 25201 (Israel) ‘Intensive Fishculture Station, Department of Fisheries, P.O.B. 33, Tiberias (Israel) (Accepted 15 February 1988)
ABSTRACT Viola, S., Arieli, Y. and Zohar, G., 1988. Animal-protein-free feeds for hybrid tilapia (Oreochromis niloticus x 0. aureus) in intensive culture. Aquaculture, 75: 115-125. Growth trials were carried out with tilapia in experimental ponds to develop an animal-proteinfree diet of nutritive value equal to a standard commercial fish-meal feed. A diet based on soybean meal and supplemented with amino acids, oil and di-calcium phosphate resulted in growth performances and body compositions equal to those on a fish-meal diet. Successive withdrawal of the supplements resulted in a final fish-meal-free diet, with 3% di-calcium phosphate and 2% oil as the only supplements necessary to achieve growth performances equal to those obtained with a fully supplemented diet.
INTRODUCTION
Many studies have been made to determine optimal protein levels for tilapia. Jauncey (1982) found 40% to be optimal for fish of 2 g in aquaria. Winfree and Stickney (1981) recommended 34% protein for fish of 7.5 g, and Jackson et al. (1982) 30% for fish of 30 g. In our studies with market-size fish in ponds we found 30% optimal (Viola and Zohar, 1984a). Tilapia are known to be much more herbivorous than catfish or carp (Jauncey and Ross, 1982). However, in many countries feeds for tilapia include a substantial proportion of animal proteins such as fish meal, poultry meal, meat meal or similar offal meals. These animal proteins usually increase the cost of the feed, yet low production costs are essential, especially for industrial processing. Jackson et al. (1982) tried to substitute different plant proteins for fish meal. At low levels of replacement (25%) growth rates were similar, but at higher inclusions the performance was considerably lower. They did not try to compensate for any imbalances of amino acids or minerals. Appler and
0044-8486/88/$03.50
0 1988 Elsevier Science Publishers B.V.
116
Jauncey (1983) replaced fish meal with increasing proportions of algal meal. Growth rates decreased proportionally when more than 5% algae were included. Under practical culture conditions in Israel, replacement of half the fish meal by soybean meal has been found nutritionally equivalent, without any supplementation (Viola and Arieli, 1983a). Attempts to determine essential amino acid requirements of tilapia fry have been reported (Jackson and Capper, 1982; Jauncey et al., 1984). However, the requirements of market-size tilapia for amino acids in relation to energy or the bioavailabilities of nutrients in diverse feedstuffs have not been clarified sufficiently. Thus the application of linear programming of feeds is not yet possible for tilapia, as it is for catfish (Robinette, 1984). Therefore, substitution and supplementation studies are the next best method to pinpoint critical nutrient levels under practical intensive culture conditions. Our preliminary trials of complete replacement of fish meal by soybean meal in tilapia feeds, on the basis of protein alone, were performed in cages of 1 m3, suspended in ponds, 100 fishes per cage, receiving daily rations of equal weight. These trials yielded inferior growth rates and feed conversions - in inverse relation to the degree of replacement - indicating nutrient imbalances (Table 1). Previous work with carp (Viola et al., 1982) had shown that replacement of fish meal by soybean meal created imbalances of amino acids and energy. When TABLE 1 Preliminary trials in cages: replacement of fish meal by soybean meal on the basis of protein alone (25% protein, means of 4 replicates, 100 fish/cage, 30-31 days) First trial Diet
25% Fish meal
40% Soybean meal Alone
+ Oil + Lysine 165 208 43b 1.4 3.2
Initial weight of fish (g) Final weight of fish (g) Gain in weight of fish (g) Daily gain (g ) Feed conversion
165 218 53” 1.7 2.6
165 205 4oh 1.3 3.4
Second trial Diet
Fish meal
Soybean meal
Initial weight of fish (g) Final weight of fish (g ) Gain in weight of fish (g) Daily gain (g)
240 307 67” 2.2
250 290 5ob 1.7
Values followed by the same superscript are not significantly different (Newman-Keul’s P
test,
117
these deficiencies were balanced by supplements of lysine, methionine and oil, isonutritious carp feeds without animal protein could be compounded. On the other hand, studies with tilapia had shown that they responded to phosphorus supplementation to feeds that contained plant proteins (Viola et al., 1986a). From this work the conclusion was drawn that plant phosphorus (phytin) has very low availability for tilapia and that supplementation of available phosphorus was essential in animal-protein-free diets. In order to assess the essentiality of further supplements, e.g. lysine, methionine, oil or others, a program was designed for the progressive omission of one supplement after another from a fully supplemented positive control diet. The final diet was to contain as few supplements as necessary to achieve equally good growth, feed conversion and retentions of protein and energy. METHODS
AND MATERIALS
Hybrids of Oreochromis niloticus and 0. aureus, 95% males, were raised in 12 experimental ponds of 200 m2 and depth of 1 m, at a density of 400 fish per pond. All the fish were counted and weighed at the start and end of each trial. TABLE 2 Composition of feeds for the first series of trials: complete balance of nutrients in low-fish-meal diets (% air-dry matter) Diet
Fish protein 50%
Ingredients Fish meal (72% protein) Soybean meal (44% protein ) Wheat Milocorn Oil (poultry) Di-calcium phosphate Lysine-HCl Methionine Nutrients Protein Fat Ash Phosphorus Methionine’ Lysinel Gross energy (kcal/kg)’ 1 Estimated according to NRC-NAS
20 24 10 46 _ _
29.5 4.0 5.2 0.75 0.7 1.8 4150 (1977,1983).
25% 10 42 10 33.5 3 1 0.25 0.15 29.7 6.0 5.1 0.75 0.7 l-8 4150
0%
60 10 21 6 2 0.5 0.3 29.2 7.5 5 0.75 0.7 1.8 4150
118 TABLE 3 Composition of feeds for the final trial (No. 6) in the second series: progressive omission of supplements from fish-meal-free diets (0 = oil, M = methionine, L = Lysine, P = DCP) (% air-dry matter) Diet
Supplements: Omission:
Ingredients Soybean meal (44% protein) Wheat Milocorn Di-calcium phosphate Oil Methionine Lysine-HCl Nutrients Protein Fat Ash Calcium Phosphorus Methioninel Lysinel Gross energy (kcal/kg)’
O+M+L+P (Balanced) 60 10 21.25 3 5 0.25 0.5 30.3 6.3 6.4 1.0 1.0 0.7 2.3 4100
M+L+P 0 60 10 24.25 3 2 0.25 0.5 29.5 3.4 6.2 N.D. N.D. 0.7 2.3
L+P
P
GM
O,M,L
60 10 24.25 3 2
60 10 25.0 3 2
0.5 29.5 3.4 6.0 N.D. N.D. 0.45 2.3 3950
29.2 3.5 6.2 N.D. N.D. 0.45 1.8
‘See Table 2. N.D. - not determined.
Duration of a trial was 55-65 days. Feeds were dispensed in equal amounts to each pond by automatic feeders. Samples of 150-200 fishes were weighed biweekly for adjustment of the ration. At the start of a trial, the average weight was equal in all ponds, between 100 and 300 g. Each dietary treatment was replicated in three ponds. Fresh water was added only to replace losses by evaporation and seepage. Water temperatures at 6 a.m. were between 21 and 23”C, rising to 24-26” C at noon. The ponds were aerated by paddle wheels. Oxygen levels were monitored and never below 4 ppm; the pH of the water was around 7.5. At the start and at the end of each trial, five fishes from each treatment were sampled for laboratory analysis. Moisture was determined at 105°C for 24 h. protein by Kjeldahl’s method (Kjeltec ) , fat by extraction with perchloro-ethylene (Fosslet), ash by incineration at 550” C, phosphorus by molybdovanadate spectrophotometry, and calcium by EDTA titration (AOAC, 1984). The feeds were pelleted in a Lister laboratory pellet mill (4 mm pellets) and analysed by the same methods. The contents of amino acids and gross energy were estimated from tables (NRC-NAS, 1977,1983).
119 TABLE 4 Composition of feeds for third series of trials Diet Ingredients ( % ) Fish meal (65% protein) Soybean meal (44% protein) Wheat Milocorn Wheat bran Di-calcium phosphate Poultry oil Binder Nutrients ( % ) Crude protein Ether extract Crude fiber Ash Calcium Phosphorus Lysinel Methionine and cysteinel Gross energy (kcal/kg)’
Fish meal
Mixed
Soybean meal
Soybean meal +DCP
_
_
20 32 10
23 20 20 24 10
55 20 12 10
1 2
1 2
1 2
55 20 8 10 4 1 2
30.2 5.0 2.7 7.6 1.5 1.3 2.0 1.15 4000
31.1 4.6 3.5 7.1 1.35 1.0 1.9 1.1 4050
28.8 2.8 5.7 4.6 0.45 0.6 i.75 0.9 4050
30.0 2.9 5.5 9.3 1.65 1.6 1.75 0.9 3850
35
‘See Table 2.
The research plan comprised three series of trials. In the first series, soybean meal was increased progressively to replace fish meal. The latter provided 50%) 25 % and 0% of the total protein. Deficiencies of amino acids, energy and phosphorus were compensated by synthetic lysine, methionine, oil and di-calcium phosphate (DCP). The composition of the diets is shown in Table 2. The trial was repeated three times (trials 1,2,3). The second series was to show which of the four supplements were really essential for equal performances. This series also contained three trials: no. 4, omission of one supplement (for oil reduction from 5% to 2% ); no. 5, omission of two supplements; no. 6, omission of all supplements except DCP, since phosphorus supplementation had been found essential in previous studies, as mentioned before. The reduction of oil to 2% minimum was intended to safeguard essential fatty acids. The composition of the diets for the last trial (no, 6) is shown in Table 3. The diets for the intermediate trials no. 4 and 5 were composed correspondingly. The findings of the second series were put to a final test in the third series. This contained two trials of 54 days: animal-protein-free diets with and without di-calcium phosphate were compared with mixed (standard) diets and with
120
full fish meal diet without soybean meal. Table 4 shows the composition of the diets. RESULTS
The performance of the fishes in trial no. 1 is shown in Table 5. All three diets yielded almost equal growth rates, feed conversions, and retentions of protein and energy. The other two trials (nos. 2 and 3)) under slightly different conditions of initial weights and temperature, gave the same general result: equal performances with all three diets. This series proved that no imbalances other than lysine, methionine, oil and phosphorus were incurred under the conditions of the trial. The results of the second series are shown in Tables 6 and 7. Again, no differences were apparent between the diet groups in any trial of this series. Body analyses were carried out in the last trial only. The conclusion from the second series, that phosphorus supplementation alone was critical, was finally tested in the third series. Both trials of this series yielded similar results and only one of them is reported here in Table 8. The soybean meal diet without DCP caused severe retardation of growth and bone mineralization, accompanied by a slight accumulation of fat. All these symptoms were prevented by the addition of 4% DCP. The full fish meal diet did not excel the other diets in any performance parameter. TABLE 5 Performance of tilapia in the first trial (no. 1) of the first series (means and S.D. of 3 ponds X 400 fish, 60 days, 2.5% feeding rate) Diet
Initial weight (g ) Final weight (g) Gain (g) S.G.R (%) Daily gain (g ) Feed conversion Protein in body (% ) Fat in body (% ) Energy retention ( % )2 Protein retention (% )”
Fish protein 50%
25%
0%
77 188 111+2 1.47 1.85 1.65 17.6 8.5 42.0 42.0
77 190 113-t3 1.48 1.88 1.65 17.3 8.0 42.0 42.0
77 188 111*4 1.47 1.85 1.70 17.5 8.5 41.0 41.0
‘S.G.R. ( =specific growth rate) = (In final weight-ln initial weight)/days 2Retention = body gain/intake ( % ). Factors for calculation: protein = 5.7 kcal/g; fat = 9.5 kcal/g (NRC-NAS,
(%). 1977).
121 TABLE 6 Performance of tilapia in trials with progressive omission of supplements (second series) Trial no. 4 - omission of one supplement (means and S.D. of 3 ponds X 400 fish, 62 days) Diet designation: Omitted supplement: Initial weight (g ) Final weight (g ) Total gain (g)
M+L+O+P -
M+L+P 0
M+O+P L
84 187 10326
84 183 99t-12
84 182 9821
o+L+P M 84 183 99 + 8 N.S.
Trial No. 5 - Omission of two supplements (means and SD. of 3 ponds x 400 fish, 54 days) Diet designation: Omitted supplement: Initial weight (g ) Final weight (g ) Total gain (g )
M-tL+O+P -
M+P
175 295 120+4
175 289 114+5
0,L
o+p M,L 175 287 112+5
L+P M,O 175 300 125 k 3 N.S.
N.S. - differences not significant. TABLE 7 Performance of tilapia in the last trial (no. 6) of the second series: progressive omission of supplements (means and S.D. of 3 ponds X 400 fish, 56 days, 2% feeding rate) Dietdesignation: O+M+L+P Omitted supplement: Initial weight (g) Final weight (g ) Gain (g) S.G.R. (%)l Daily gain (g ) Feed conversion Protein in body ( % ) Fat in body (% ) Energy retention (% )’ Protein retention (g ) 1
275 469 194k 11 0.95 3.5 2.35 17.3 12.0 26.7 27.0
Mi-L+P 0
L+P
P
0,M
O,M,L
275 459 184-tlO 0.9 3.3 2.45 17.8 10.5 23.0 25.9
275 464 189+8 0.92 3.4 2.4 17.8 10.5 24.2 27.3
275 467 192 k 7 N.S. 0.95 3.45 2.35 18.2 12.0 25.4 27.6
‘See Table 5. DISCUSSION
The final conclusion arising from this study is that the only essential supplement was phosphorus. This result is strikingly different from our previous study with carp and raises several points for discussion.
122 TABLE 8
Growth performance of tilapia in the third series of trials (means and standard deviations of 3 ponds x 400 fish, 196 g at start, 54 days, 2.5% feeding rate) Diet
Fish meal
Mixed
Soybean meal
Soybean meal +DCP
Total weight gain (g ) Standard deviation (g ) Average daily gain (g ) Specific growth ( % ) ’ Feed conversion
175.2” 15.5 3.2 1.18 1.9
175.2” 9.4 3.25 1.2 1.85
137.3b 10.0 2.5 1.0 2.35
167.4” 3.8 3.1 1.15 2.0
Body composition: Protein (% ) FaL (W) Ash (%) Calcium (%) Phosphorus ( % ) Energy retention (% )” Protein retention (% )”
17.2 9.7 3.6 2.1 0.55 29.3 31.4
17.3 9.7 3.2 2.0 0.52 29.3 31.1
17.8 10.5 2.8 1.6 0.45 26.8 28.4
18.8 9.2 3.5 2.0 0.55 29.7 33.3
‘,‘See Table 5. Values followed by the same superscript are not significantly different.
Oil as source
of energy
Carp had been found very responsive to oil supplementation (Viola et al., 1981), yet the reduction of oil in the tilapia diets had no effect at all on the performance of the fishes. This confirms previous findings on the poor utilization of oil by tilapia (Viola and Arieli, 198313). Consequently, the oil content of fish meal and other meat meals, which are valued highly for poultry, carp or catfish, has very little worth for tilapia. Hemicelluloses and oligosaccharides as sources
of energy
On the other hand, 60% soybean meal in the diets contributed a great amount of cellulose, hemicellulose and oligosaccharides. Crude fiber in the soybean meal amounted to 6.5% and nitrogen-free extract to 30%. According to Smith and Circle (1978), sucrose may reach 6% and starch is negligible, leaving 24% oligosaccharides and hemicelluloses. These are considered almost indigestible for monogastric animals. Lodhi et al. (1969) found a digestibility of 14% for the nitrogen-free extract for poultry - which again leads to an estimate of 26% indigestible carbohydrates. Together with the crude fiber, 60% soybean meal contained, therefore, about 18% complex carbohydrates. In the nutrition of carp, these polysaccharides are almost indigestible and created large deficits
123
of energy that could be balanced by oil (Viola et al., 1982). However, in the first series of our trials with tilapia, feeding rates were the same and therefore any deficit of energy would have asserted itself in substantially decreased growth performance. Thus it may be concluded that these polysaccharides were at least partly utilized for energy. Similar conclusions were found in previous experiments with high-fiber feeds: 60% wheatbran caused only slight impairment of growth (Viola and Zohar, 198413). Popma (1982)) too, found increased carbohydrate digestibility of fibrous feeds for tilapia, as compared to carp. Lysine In our diets without supplemental lysine, the total lysine was higher than the requirements established by Jackson and Capper (1982) - 1.62% - or by Jauncey et al. (1984) - 1.51% - with synthetic or semisynthetic diets. In carp feeds, when fish meal was replaced by soybean meal, lysine was found to be deficient, although the total lysine was higher than the requirements (Viola et al., 1982). Probably the stomachless carp is at a disadvantage in utilizing lysine complexes created by Maillard condensations during the thermal treatment in soybean meal production. In contrast, we were not able to prove that lysine was critical for tilapia in any practical combination of feedstuffs and protein levels. Be it due to the lower specific growth rate or to their stronger gastric digestion, tilapia appear to derive all their necessary lysine from soybean meal or even from low-gossypol cottonseed meal (Viola and Zohar, 1984a). Phosphorus - the neglected nutrient Ogino et al. (1979) found that both bone phosphorus and plant phosphorus were of very low availability to stomachless carp. Therefore, replacement of one by the other does not change the supply. Actually, neglect of supplementation of soluble mineral phosphorus to both kinds of diet for carp has serious consequences of growth depression (Hepher and Sandbank, 1984; Viola et al., 1986b). Tilapia do produce acid gastric juice and therefore are able to utilize bone phosphorus to a much higher extent (Viola et al., 1986a), but not phytin. Thus, substitution of animal meals, which contain bones, by seed proteins creates a phosphorus deficiency, which must be balanced by suitable mineral supplements. This turned out to be the only really critical nutrient when fish meal was replaced by soybean meal. This feature has escaped the attention of many fish nutritionists and might explain unsatisfactory results, such as those of Jackson et al. (1982), in which fish meal was replaced completely or almost completely by plant proteins.
124 ACKNOWLEDGMENT
Our thanks to Mrs. D. Yishar and the staff of the “Milouda” their technical assistance.
laboratory for
REFERENCES AOAC, 1984. Official Methods of Analysis, 14th edn. S. Williams (Editor). Association of Official Analytical Chemists, Arlington, VA 1102 pp. Appler, H.N. and Jauncey, K., 1983. The utilization of a filamentous green alga (Cladophora glomerata (L. ) Kutzin) as a protein source in pelleted feeds for Surotherodon (Tilapia) niloticus fingerlings. Aquaculture, 30: 21-30. Hepher, B. and Sandbank, S., 1984. The effects of phosphorus supplementation to common carp diets on fish growth. Aquaculture, 36: 323-332. Jackson, A.J. and Capper, B.S., 1982. Investigations into the requirements of the tilapia Sarotherodon mossambicus for dietary methionine, lysine and arginine in semi-synthetic diets. Aquaculture, 29: 289-297. Jackson, A.J., Capper, B.S. and Matty, A.J., 1982. Evaluation of some plant proteins in complete diets for the tilapia Surotherodon mossumbicus. Aquaculture, 27: 97-109. Jauncey, K., 1982. The effects of varying dietary protein level on the growth, food conversion, protein utilization and body composition of juvenile tilapia (Surotherodon mossumbicus). Aquaculture, 27: 43-54. Jauncey, K. and Ross, B., 1982. A Guide to Tilapia Feeds and Feeding. Institute of Aquaculture, Univ. Stirling, Scotland, 111 pp. Jauncey, K., Tacon, A.C.J. andJackson, A.J., i984. The quantitative essential amino acid requirements of Oreochromis (Sarotherodon) mossambicus. Int. Symp. Tilapia in Aquaculture, TelAviv Univ., pp. 328-337. Lodhi, G.N., Renner, R. and Clandinine, D.R., 1969. Available carbohydrate in rapeseed meal and soybean meal as determined by a chemical method and a chick bio-assay. J. Nutr., 99: 413418. NRC-NAS (National Research Council-National Academy of Sciences), 1977. Nutrient Requirements of Warmwater Fishes. National Academy Press, Washington, DC, 77 pp. NRC-NAS (National Research Council-National Academy of Sciences), 1983. Nutrient Requirements of Warmwater Fishes and Shellfishes. National Academy Press, Washington, DC, 94 PP. Ogino, C., Takeuchi L., Takeda, H. and Watanabe, T., 1979. Availability of dietary phosphorus in carp and rainbow trout. Bull. Jpn. Sot. Sci. Fish., 45 (12): 1527-1532. Popma, T.J., 1982. Digestibility of selected feedstuffs and naturally occurring algae by tilapia. D.Sc. Thesis, Auburn Univ., Auburn, AL, 78 pp. Robinette, H.R., 1984. Feed formulation and processing. In: E.H. Robinson and R.T. Love11 (Editors), Nutrition and Feeding of Channel Catfish. South. Coop. Ser. Bull., No. 296, Auburn Univ., Auburn. AL, pp. 29-39. Smith, A.K. and Circle, S.J., 1978. Soybeans: Chemistry and Technology, Vol. 1. Avi Publ., Westport, CT, p. 81. Viola, S. and Arieli, Y., 1983A. Nutrition studies with tilapia (Sarotherodon). 1. Replacement of fish meal by soybean meal in feeds for intensive tilapia culture. Bamidgeh, 35 (1): 9-17. Viola, S. and Arieli, Y., 1983B. Nutrition studies with tilapia hybrids. 2. The effects of oil supplements to practical diets for intensive aquaculture. Bamidgeh, 35 (2): 44-52.
125 Viola, S. and Zohar, G., 1984a. Nutntion studies with market size hybrids of tilapia (Oreochromis) in intensive culture. 3. Protein levels and sources. Bamidgeh, 36 (1): 3-15. Viola, S. and Zohar, G., 198413.Report to the Ministry of Science in Israel (in Hebrew, unpublished). Viola, S., Rappaport, U., Arieli, Y., Amidan, G. and Mokady, S., 1981. The effects of oil-coated pellets on carp (Cyprinus carpio ) in intensive culture, Aquaculture, 26: 49-65. Viola, S., Mokady, S., Rappaport, U. and Arieli, Y., 1982. Partial and complete replacement of fish meal by soybean meal in feeds for intensive culture of carp. Aquaculture, 26: 223-236. Viola, S., Zohar, G. and Arieli, Y., 1986a. Phosphorus requirements and its availability from different sources for intensive pond culture species in Israel. Part I. Tilapia. Bamidgeh, 38 (1): 3-12. Viola, S., Zohar, G. and Arieli, Y., 1986b. Requirements of phosphorus and ita availability from different sources for intensive pond culture species in Israel. Part II. Carp culture. Bamidgeh, 38 (2): 44-54. Winfree, R.A. and Stickney, R.R., 1981. Effects of dietary protein and energy on growth, feed conversion efficiency and body composition of Tilapia aurea J. Nutr., 111: 1001-1012.