The work reported incorporates group training of counterparts and the latters' participation in all aspects of the programme. Training included: use and care of equipment; analytical methodology and techniques; fish nutrition basics; diet formulation, preparation and testing; design of experiments in fish nutrition.
Equipment received under Projects IND/75/031 and RAS/76/031 included testing and measuring instruments employed in fish nutrition research. These were checked, assembled and tested before final installation. A significant portion of this newly received equipment had arrived with damaged or missing components. Broken or missing parts were identified and replacement orders sent out to suppliers. Equipment with minor damage was repaired by the author.
Fish nutrition research may be divided into four general areas of activity. These are: feed analysis; diet formulation and preparation; diet testing; biochemical studies. The new equipment was installed in four separate rooms according to work to be carried out. Some existing equipment was also recommissioned after minor repairs, where necessary, to complement the new equipment. Major items of the Nutrition Laboratory are listed in Table 1.
Feed analysis involves the determination of the chemical composition of feedstuffs. Usually, this is confined to analysis of gross nutrient content such as: crude protein, crude fat, crude fibre, ash and moisture. However, when further quality specifications of the materials are required, it includes assays on amino acids, vitamins, minerals, fatty acids and caloric value. When an ingredient is suspected of containing harmful levels of adventitious toxins such as urease, thioglucosides, or aflatoxins, and adulterants such as urea, assays on these are also carried out.
Equipment for proximate analysis of feeds was installed in the feed analysis laboratory. This included: a Kjeldahl digestion unit designed for high sample throughput; an automatic Soxhlet extraction unit for crude fat analysis; a semi-automatic crude fibre analysis unit; a muffler furnace for ash determination; and a drying oven for moisture determination. The spectrophotometer used for measurement of ammonia in the analysis of crude protein was installed in a separate room. This equipment is now fully operational and analysis on a variety of local feedstuffs received by the Centre has been carried out.
Other new feed analysis equipment that were tested and installed included four bomb calorimeters and an atomic absorption spectrophotometer. An amino acid analyzer and a gas chromatograph unit arrived during the latter part of the author's visit. Installation of these two pieces of equipment was to be carried out by the suppliers' agents. These and all other optical instruments were to be installed in a properly air-conditioned instruments room.
Diet formulation uses information obtained from chemical analysis of individual ingredients to determine the proportion in which such ingredients should be constituted in a diet in order to meet specified nutrient levels for that diet. The diet is then prepared according to the manner in which it is to be fed; for example, a diet formulated for production of moist feed should not be processed into dry feed, and vice versa.
At the time of the visit, equipment for feed preparation purchased for the project had not yet been received. In preparation for this, however, a diet processing room was specially installed with high-current electrical outlets necessary for operation of the machines, which were to include: a laboratory model pelleting machine, and mixing and grinding equipment. In the meantime, experimental diets were prepared using a manually operated meat grinder fitted with a 3 mm die, and ordinary kitchen utensils.
It is anticipated that a mini-computer system purchased for processing aquaculture information will be made available, as will also be the necessary software, for least-cost formulation of diets. This computer system will be installed in a separate air-conditioned room.
Diet testing embodies the experimental phase of nutrition research. Experiments are designed to answer specific questions relating to nutrient requirements and the suitability of selected feed ingredients as dietary components. Such experiments are conducted under specified conditions over a fixed period during which observations are made and weight data recorded.
Pending completion of proper indoor diet testing facilities at the Centre, one of the rooms adjacent to the feed analysis laboratory was converted for temporary use in diet testing work. Equipment installed included plastic-lined circular pools of 1 m diameter and 70 cm height. An aeration system consisting of an air pump, that had been recommissioned after minor repairs, and plastic tubings, was set up. This system was demonstrated to provide adequate aeration for 10 pools filled to a water depth of 40 cm and stocked with up to 50 rohu fingerlings of 7 cm length, maintaining near-saturation levels of dissolved oxygen (7.5 ppm) at 24°C. Two units of dissolved oxygen and water temperature monitoring instruments were installed and used in this verification. The pools were filled by means of a hose connected to one of the water faucets, and emptying was by siphoning.
Biochemical studies in nutrition research aim at more complete understanding of the physiological relationships between nutrition and growth. Examples are studies on the effects of feeding on digestive enzyme secretion and nutrient absorption, and the influence of temperature and oxygen supply. Results from such studies not only contribute to knowledge of nutrient requirements of fish but also provide important information concerning biological as well as physical constraints affecting food intake and growth.
The basic equipment tested and installed in the biochemistry laboratory was an automatic sample analyzer system. This system consisted of: a full-range spectrophotometer; sample-changer immersed in a thermostatically controlled water bath; and, a computer that controls the entire analysis operation and automatically computes raw numerical data before printing out the results of the analysis. During its installation, the system was found to be lacking in the electronic circuit board controlling sample advance in the automatic sample-changer. The equipment supplier was duly notified and a request was made for the missing component. Other parts of the system were tested out and found to function normally. The instrument was then used for ammonia analysis in the determination of crude protein in feed samples described in the next section of this report. A spectrofluorometer, used mainly for vitamin assay work, was badly damaged during transit. However, after the necessary repair, part of it to its internal wiring, the instrument also functioned normally. These delicate optical instruments were installed in the air-conditioned Instruments Room that will also accommodate the atomic absorption spectrophotometer, amino acid analyzer and gas chromatograph.
Other major pieces of equipment tested out and installed were: a high capacity glass water distilling plant; a tray-type freeze dryer; and, a plankton centrifuge separator. The water still was put to use at half capacity pending installation of high current electrical outlets in the room where it is to be permanently installed. The freeze dryer lost its digital display and will have to be sent for repairs. The plankton centrifuge was used in preliminary studies on the natural productivity of one of the Centre's holding reservoirs.
Fish diet development involves not only the application of knowledge on nutritional requirements for growth of the specified species but also identification and development of ready sources of feed ingredients that ultimately make up the diet. These ingredients have first to be evaluated to determine their suitability and safety for use in compounded feed. Precise information on the chemical composition of individual feed ingredients is indispensable, not only for formulation of efficient diets that meet desired nutrient specifications but also for setting restrictions for their safe usage when anti-nutritional factors are known to be present.
The programme for diet development, therefore, began with a survey of available feed ingredients considered suitable for feeding to fish. Samples of these were then evaluated on the basis of results obtained through chemical analysis in the newly established feed analysis laboratory and on available information in the literature concerning possible presence of anti-nutritional factors. Experimental diets were then formulated, prepared and tested to observe growth response of the fish.
A survey conducted among a few feed stores in Bhubaneswar showed a rather limited variety of feedstuffs available in the area. One large store offered the following: rice bran, wheat bran, maize meal, groundnut oil cakes (two types), gingelli cake, and an assortment of grams and gram brans. Other smaller stores were each selling a few of the items mentioned. The feed stores appeared to cater primarily to homestead dairies. However, judging from the ready availability of poultry eggs in market stalls, there appeared to be a faily large poultry farming industry in the area. This would mean that compounded poultry feed was also available even though the author did not see these products sold in the stores visited.
No estimate was available on the annual production and consumption of individual feed materials. Considering the importance of dairy products in the local diet, the quantities must be fairly large.
Prices generally reflected the relative availability of individual ingredients. For example, maize meal cost I. Rs. 2.10/kg compared to I. Rs. 1.80/kg for groundnut oil cake1. This was due more to maize being a scarce item than to low economic value of groundnut oil cake. Cost comparison among other ingredients listed in Table 2 is more normal and reflects the direct relationship between nutritive value, particularly protein content, of a feed and its cost.
Product quality appeared to vary considerably among certain types of ingredients, especially rice bran and oilseed cakes. This quality inconsistency is due to processing methods rather than to deliberate adulteration by unscrupulous entrepreneurs. For example, the efficiency by which oil could be extracted using the expeller process determines the percentage of protein in the oil cake, since less efficient extraction would result in more residual oil and, therefore, lower overall protein content (see Table 2 for such a comparison). Furthermore, some mills remove the shell of nuts before oil extraction, especially when chemical solvents are used in the extraction process instead of the screw expeller. This invariably results in lower fibre and higher protein content in the final product (compare composition of groundnut oil cakes in Tables 2 and 3). Quality variation in rice bran can be attributed to similar reasons. The lack of standardization of milling processes is inevitable since a significant quantity of grains and seeds are processed by small country mills.
Finally, a search of the literature suggests a far more extensive list of feedstuffs traditionally available to the livestock industry in the country as a whole; those considered most suitable for compound fish diet manufacture are listed in Table 3.
Concurrent with the feed survey, a work programme to analyse all feed samples received by the Centre was instituted. Proximate analysis was conducted on a few selected feed ingredients for immediate formulation of test diets for comparative feeding trials. The Weende method for feed analysis was adopted, which provided information on levels of: crude protein, crude fat, crude fibre, ash and moisture. Nitrogen-free extract, the non-fibre carbohydrate fraction, was obtained by difference. All analytical work was carried out by counterpart staff with the assistance of the author. The results of this work appear in Table 2.
During a meeting with heads of regional CIFRI sub-stations who attended the 5th Workshop of the All-India Coordinated Project on Composite Fish Culture and Training at FARTC on 19–20 January 1981, the author proposed that each station submitted samples, together with pertinent commercial information, of feed ingredients available in their respective geographic regions for analysis and for least-cost test diet formulation at FARTC. Response was good, and samples from a few stations were received during the latter half of the author's stay.
1 US$ 1.00 = I.Rs. 8.00 (February 1981)
As had been stated earlier, artificial feeding of Indian major carps in polyculture at present involves a mixture of groundnut oil cake and rice bran in roughly equal proportions. Intended as a supplemental diet, its effectiveness depends mainly on the natural productivity of the pond, especially with regard to the supply of animal protein in the form of zooplankton. The inadequcy of such a feed, unbalanced as it was with regard to nutrients, will become most apparent when the supply of zooplankton is low. An enhancement in feeding value, through improvement in the balance of nutrients without greatly increasing the cost of the feed, is possible. An initial effort in diet development at the FARTC, therefore, was to seek such an improvement without radically changing the nature and increasing the cost of the diet mixture now popular among farmers. To determine if an improvement could be obtained by supplementing the mixture with minerals and vitamins, two diets were formulated, based on groundnut oil cake and rice bran, to contain 28 percent crude protein. One of the diets also contained the vitamin and mineral supplements1.
Other test diets were also formulated to determine if improvement in feed efficiency could be obtained by substituting groundnut oil cake with sesame oil cake, the latter being richer in methionine, an essential amino acid generally lacking in vegetable proteins but present at high levels in fish meal. The cost effectiveness of substituting wheat bran for rice bran was also to be studied. These diets, formulated for testing on fingerlings in aquaria, may also be tested on juvenile fish in pond polyculture. The formulations appear in Table 4.
Test diets for fry and brood fish were also formulated (see Tables 5 and 6). Test diets for fry compare two levels of fish meal and the use of synthetic lysine as a substitute for animal protein in the diet. Test diets for brood stock are intended to evaluate prawn head meal as a dietary protein source.
Artificial feed for fish should be in a form that can be consumed with minimum wastage. Compounded feed especially should be processed so as to ensure that ingredient components do not separate in water during feeding. Such processing usually involves some form of heat treatment which brings out the natural adhesiveness of starchy ingredients almost always included in compounded feed. Sometimes the addition of chemical or extraneous natural binders is necessary. In commercial production of compound feed for fish, pelleting with the aid of steam is most common. Depending on the equipment used, such processing methods result in either sinking or floating-type dry pellets. Diets for fish may, however, be prepared in moist or semi-moist form if the feed is fed fresh. To prepare such types of feed, heat processing is usually not required if fresh fish or slaughterhouse waste, which both contain animal glues, constitute a significant portion of the diet mixture. Where such materials are absent, heat processing becomes necessary.
Pending arrival and installation of the Centre's new pellet mill, wet processing methods for the preparation of experimental diets were employed. Each of the two diets mentioned in the previous section were prepared as follows: the rice bran was cooked in water constituting 120%2 of total weight of the diet mixture. The remaining component (in the case of the fortified diet, plus the vitamins and minerals) was then blended into the hot pasty slurry to form a dough. This was then extruded into pellets by means of a meat mincer fitted with a 3 mm die. The extruded pellets contained about 55 percent moisture and, if fed without drying, represented a moist type of feed. In order to impart a longer shelf life to the product the pellets were spread out over a fine screen and dried in the sun. The dried pellets, which then contained about 11 percent moisture, were found to have a good water stability and keeping qualities.
To determine the value of vitamin and mineral fortification of the traditional supplementary diet used in carp polyculture, a comparative feeding trial was conducted on rohu fingerlings using the two test diets mentioned in the previous section.
Fingerlings weighing an average of 3 g were kept outdoors in plastic-lined pools of 1 m diameter. Water was to a depth of 30 cm. The test was carried out in three replicate pools containing 10 fish each for each dietary treatment. Fish were fed only once daily at mid-day due to rather cool temperatures during that time of the year (water temperatures ranged between 16°C in the early hours and 25°C in the afternoon). Fifty percent of the water in the pool was changed every morning. During this time, unconsumed feed and faecal matter were removed by siphoning1.
Fish were weighed once fortnightly. Observations on growth during the first month indicated significantly better growth in fish fed the fortified diet, with specific weight gain of about 1 percent/day as against no gain by fish receiving the unfortified diet. Unfortunately, the experiment had to be stopped after a month due to unexpected high mortality among fish fed the unfortified diet following treatment (with 50 ppm potassium dichromate) of all fish for a mild fungal infection affecting populations in both groups. Weight and length measurements were taken of surviving fish. The fish were then oven-dried and preserved for later chemical analysis.
Comparative feeding trials are now underway using rohu fingerlings to evaluate two vegetable protein sources available in relative abundance in most parts of the country. These are groundnut oil cake and gingelli (sesame) oil cake. The formulations for these diets appear in Table 4. Isonitrogenous diets containing either protein source in combination with rice bran or wheat bran, together with vitamin and mineral supplements, are being fed to the fish in plastic-lined pools kept indoors. The pools are continuously aerated by means of a common air line, and dissolved oxygen monitored periodically. With 10 fish per pool, dissolved oxygen level could be maintained at 8 ppm all day. Indoor temperatures ranged between 22° to 26°C during the month of February. Water ammonia level will be monitored as a criterion for water change. Effects on growth and feed conversion will be measured over a 3-month period. Proximate analysis will be performed on carcasses of experimental fish at the close of the experiment.
Tests have also been planned to evaluate diets specially designed to accelerate sexual maturity in juvenile fish to determine the feasibility of off-season spawning. Diets for these tests are shown in Table 6.
