2.5 Food and feeding
Chlorella has probably been the most popularly used algal food for the culture of Brachionus plicatilis (Fig. 2). Hirata and Mori (1967) made use of Chlorella and Dunaliella while Theilacker and McMaster (1971) experimented on Monochrysis lutheri, Nannochloris sp., and Dunaliella. Person-Le Ruyet (1975) noted that Tetraselmis suecica was a more suitable feed than Chlorella based on the high yields of B. plicatilis he obtained. The use of dried and ground blue-green alga Spirulina maxima also proved promising (Preisser, 1979). Hirayama and Nakamura (1976) achieved success using dried Chlorella sp.
Owing, however, to some difficulties encountered by many in mass producing sufficient supply of phytoplankton species for rotifer culture, the possibility of utilizing other types of food was looked into. It was in 1967 when Hirata and Mori introduced the use of baker's yeast for rotifer culture. Further confirmation of the results they obtained led to a number of studies conducted on yeast in Japan (Kawano, 1968; Osawa and Kawano, 1971; Furukawa, 1972; Furukawa and Hidaka, 1973; Hirata, 1974; Ueki, 1974; Fukusho et al., 1976; Hirata, 1977; Endo, 1978; Hirata et al., 1978; and Mochisuki et al., 1968). Other experimental diets were tried like Torula yeast and “food” yeast. In 1973, Hirayama and Watanabe also tested caked yeast, Rhodotorula sp., and a commercialized dried marine yeast (ASY-4011). He concluded that yeast may be used most effectively as a supplemental food when phytoplankton supply is not sufficient. Another marine yeast species, Zygosaccharomyces marinus, which was isolated by Kawano (1968) was also found to be a good food for Brachionus (Furukawa and Hidaka, 1973; Hirata, 1974) and zoeal larvae of P. japonicus (Furukawa, 1972).
2.6 Culture techniques
2.6.1 The “daily-tank-transfer” method
In this method, Brachionus is continuously subcultured using 0.5-m3 tanks. The tanks are initially used for Chlorella cultures. When Chlorella density reaches about 10–20 × 106 cells/ml inoculation of rotifers is done. Once Brachionus yield increases significantly, the rotifers are harvested and transferred to another tank (Fig. 3). The process of transfer or subculturing continues for an indefinite period of time. The main disadvantage of this method is that it is too labour intensive.
2.6.2 The “drain-off” system for large tank outdoor culture
This extensive method of rotifer culture was first developed in Japan using large concrete tanks (Fukusho et al., 1976; Kureha et al., 1977). The procedure involves the following steps:
Culture marine Chlorella using modified Hirata medium (Appendix 1) until density reaches 10–20 × 106 cells/ml.
Fertilize again using double strength of above-mentioned culture medium.
When Chlorella reaches stationary phase of growth, Brachionus is inoculated at an initial density of 10–50 individuals/ml.
Once Chlorella is consumed, introduce additional food or baker's yeast (at 1 gallon/million rotifers/day).
When Brachionus density reaches 120–150 ind/mil., harvest about 20–30 percent of the culture and transfer to another tank.
The procedure is thus repeated until the water becomes polluted with faecal materials. A diagram of the “drain-off” system is shown in Fig. 4.
2.6.3 The “feedback” culture system
Hirata (1979) describes feedback as “feeding resulting in excretion, then excretion converted into feeding again”. In an artificial culture system, “self-purification” which is carried out smoothly in the natural waters, cannot be attained due to the accumulation of biodeposits produced by the cultured organisms. Therefore, a system should be devised to promote energy flow and this is the very principle on which Hirata's feedback system was evolved. A flow chart of the principle of the feedback culture system is presented in Fig. 5, while Fig. 6 shows a conceptual diagram of a feedback system for mass-scale production of rotifers. Fig. 7 presents a schematic diagram of the zigzag stream set-up.
Fig. 2 Population growth of Rotifers fed on marine Chlorella (A); 100% yeast (B); A mixture of Chlorella + yeast (C); and non-fed (D). The experiment were repeated three times in 0.5 m3 transparent polycarbonate tanks under indoor conditions.
Fig. 3 Schematic diagram of the initial culture method for Rotifers, called “daily tank-transfer”. The method was developed in the Yashima station of the Seto inland sea farming fisheries association during 1964 – 1967. Several 0.5 m3 slate tanks were used for producing both marine Chlorella and Rotifers.
Fig. 4 Diagram of mass culture of Rotifers in a “drain-off” system in large outdoor tanks
In a feedback system, marine zooplankters like Brachionus and Trigriopus japonicus (or the harpacticoid copepod, Tisbintra elongata) may be cultured together. These organisms serve as consumers. The microalga, Chlorella and the macroalga Enteromorpha intestinalis are initially cultured as producers. Bacteria which grow in the tank will then serve as decomposers.
There are two tanks in this system - a circular tank which serves as reservoir and a rectangular tank called the zigzag stream unit. The water in the first tank is recirculated to the stream by an airlift pump at the rate of 20 times a day. The macroalga, Enteromorpha intestinalis is grown together with the zooplankton in the zigzag stream. Framed nylon nettings are placed in each of the slots in the stream to serve as attachments for Enteromorpha which is filamentous. Water quality can be maintained for long periods of time in this type of culture system.
In SEAFDEC, both the drain-off and the feedback culture systems are employed for the mass culture of Brachionus plicatilis and the copepod, Tisbintra elongata.
For outdoor cultures of Brachionus, one-ton fiberglass tanks are used and rotifers rely mainly on Chlorella virginica for food. The use of baker's yeast is also employed especially when Chlorella supply is not sufficient. The use of other types of food is presently being experimented on.
Tanks are regularly cleaned and disinfected using dilute (about 10 percent) muriatic acid solution. Thorough rinsing is done prior to use. During rainy months, the tanks are usually provided with framed plastic sheets which serve as covers.
Water utilized for culture purposes undergo filtration provided by improvised sand filters.
Water salinity for rotifer culture is usually within the range of 25–28 ppt.
Water temperature ranges between 27–30°C.
Fig. 5 Flow chart of marine zooplankton culture in a feedback system
Fig. 6 Diagram of mass culture of Rotifers in a “feedback” system. E. Planation in the text
Fig.7 Schematic diagram of main tank and zigzag stream unit for
cultivation of zooplankton (Tigriopus and Brachionus) and
macro alga (Enteromorpha)
A. Side view; B. Top view; a.150 I zigzag stream unit; b.550 I main cultivation tank; c. Recirculation pipe (After Hirata, 1977)
3. CULTURE OF MOINA MACROCOPA STRAUS
3.1 Taxonomic position
Genus Moina Baird
Species Moina macrocopa Straus
3.2 General description
Body wall short and compact, with faint segmentation and often enclosed in a bivalve carapace; with 4–6 pairs of thoracic appendages; first pair of antennae often minute, second pair large, with two branches (exopod and endopod) prominent. These are used for swimming; the abdomen is usually small and bent under the thorax. There is a single median eye. The female individuals have large brood sacs (dorsally situated) in which the eggs develop. Their “winter eggs” are usually enveloped by an additional shell called the ephippium. This shell has two chitinous plates, the edges of which fit together. One, two, or more eggs may be contained in a single ephippium. Most species under this group live in freshwater (Fig. 8).
Carapace enclosing the entire body; the body is contracted; segmentation is very faint; the legs are foliaceaus and respiratory in function; the first two pairs of legs are prehensile, the other are foliaceous; one branch of the second antenna with three segments; the other with four, the first antennae are minute; there is no beak present (only a rudimentary one); the abdomen extends beyond the anus.
Fertilized sexual eggs of M. macrocopa hatches out as a female, which reproduces (through parthenogenesis) young individuals that may either be parthenogenetic females, sexual females, or males; the resulting parthenogenetic females and the succeeding generations may give rise to all three types of individuals (Allen and Banta, 1929). Bellosillo (1937) has given a detailed description of the three individual types in his paper.
Fig. 8 Moina macrocopa
Moina macrocopa is cosmopolitan. Its occurrence has been reported in Europe, Central Asia (Sars, 1903), South Africa (Sars, 1916), and in the United States (Herrick, 1875; Birge, 1918).
Bellosillo (1937) reported Moina macrocopa to be present in practically all temporary standing water areas in Manila, San Pedro (Laguna), Makati, Pasig, Rizal province, and Calamba (Laguna).
It is usually spotted in filthy, dirty environment, particularly in freshwater pools polluted with decaying materials. It may also be found in stagnant pools and ditches near or under houses, where food particles, dead grass, decaying wood, manures, and other refuse are disposed (Bellosillo, 1937).
Many aquaculturists have emphasized the importance of cladocerans as food for the young and adult stages of various fishes. Day (1958) stated that cladocerans can serve as food for the various freshwater fish in India, particularly those of the Cyprinidae and Siluridae. Moina has also been extensively utilized as live food in many hatcheries and in the maintenance and culture of aquarium fishes of commercial importance (Bellosillo, 1937; Alikunhi, 1952; Alikunhi et al., 1955; Cooper and Grasslight, 1963; Free, 1966).
It has also been noted that cladocerans play an important role in the stabilization of sewage as shown by Moina dubia (Loedolff, 1964). The biological changes are chiefly due to the mutually beneficial interaction between bacteria, algae, protozoa, and other planktonic and benthic organisms.
Grazing of phytoplankton by Moina and other crustaceans have been reported by various workers (Lakshminarayana, 1964; Loedolff, 1964; Elaster, 1964; Hall, 1964; Zhukova, 1963; and Macan, 1961). Parabrahman et al. (1967) stated that there were clear indications showing the grazing of Moina on Chlorella. Ventura and Enderez (1980) also noted the direct relationship between population increase in Moina and the abundance of several algae like Anabaena, Scenedesmus, Chlorella, Schroederia, and Staurastrum. Nandy et al. (1977) screened several types of feed for Daphnia lumholtzi (also a cladoceran) and used dried brewer's yeast, Chlorella vulgaris, and poultry excreta. Samingin (unpublished) did a similar work on Moina macrocopa using baker's yeast, Scenedesmus, and chicken manure extract. Bellosillo (1937) also tested ricebran and soybean for Moina macrocopa.
3.6 Culture methods
So much has been written on the physiology of Moina and its vital role in the food chain, but little is known on its culture aspects.
A few culture methods will be described with special reference to the various culture media employed by several workers.
1. Yeast-Chlorella - poultry excreta (Nandy et al., 1977)
0.1 percent suspension of yeast granules were fed every other day
Chlorella vulgaris was also given as food (no feeding level reported)
Poultry excreta is also used as nutrient source. A polyethylene tube for regulating the flow of poultry manure extract was used; nutrients were replaced every 7–9 days
culture tanks were covered with nets to prevent breeding of insects
peak population was attained in 7 days
2. Soil-manure culture medium (Bellosillo, 1937)
100 g garden soil and 10 g of dried horse manure are placed in glass jars
Place 4–5 liters of water into the jars; stir vigorously
Allow the solution to stand for a day or two
Remove floating debris on water surface
Utilize solution leaving bottom sediments behind
3. Sack-method of fertilization (Ventura and Enderez, 1980)
Place previously dried chicken manure in a sack (no quantity of manure reported)
Submerge sack in water for 1–2 hours
Transfer sack to the productivity tank
Dipping of sack is carried out for 2–4 days
NOTE: No inoculation of Moina starter is done
4. Yeast-Scenedesmus-chicken manure media (Villegas and Tech, unpublished)
(a) The following components are combined:
2 000 ppm chicken manure extract
500 ppm baker's yeast suspension
5 × 104 cells/ml Scenedesmus
(b) Inoculate with Moina individuals at 5 ind/ml
5. Ito's method
Cattledung and hen's droppings are placed in a cotton fabric bag
Dip bag into a tank filled with water until water turns brown
Expose tank to sunlight till water turns green (algal propagation has set in)
Inoculate Moina or Daphnia
6. Leaf juice method (Matsudaira, 1943)
A handful of leaves (clover, lettuce, radish, or cabbage) is ground into small pieces
Place in 10 liters of water; allow to stand for a few days until propagation of algae starts
Inoculate with cladocerans
3.7 Factors affecting growth
Water - rainwater is excellent for Moina culture; in the absence of rainwater, filtered tap water (not newly chlorinated) will suffice.
Oxygen - Moina can normally live on little oxygen supply although dense populations should be given sufficient supply since oxygen requirement becomes greater.
Temperature - Hall (1964) states that temperature tolerance can be utilized to predict the frequency of molting, reproduction, and duration of egg development. Bellosillo (1937) reported a temperature range of 26–31°C to be optimum for laboratory and outdoor cultures of Moina macrocopa.
Light - diffused light was observed to be best
Food - the far most important growth factor
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|Ammonium sulfate||100–200 g/ton|
|Superphosphate of lime||10–20|
* Clewat 32 is a commercial name given to aformulation consisting of Fe, Zn, Mn, Co, Cuand Bo with EDTA
Nutrient medium for Chlorella virginica (SEAFDEC)
|Ammonium sulfate||100 g/ton|
BIOLOGY, USE AND CULTURE OF ARTEMIA
N.A. Jumalon, R.F. Figueroa, A.G. Mabaylan
and D.G. Estenor1
1. THE USE AND IMPORTANCE OF ARTEMIA IN AQUACULTURE
Artemia is widely-used in aquaculture as an excellent food source for fish and crustaceans. In many instances, it is the only food available for the larval stages of cultured species. Feeding tests with milkfish (Chanos chanos) and shrimp (Penaeus monodon) larvae showed that significantly higher growth and survival is obtained with Artemia as food than with artificial diets or other live food.
Artemia has several characteristics which make it ideal for aquaculture use. It is easy to handle, adaptable to a wide-range of environmental conditions, non-selective as a filter-feeder and is capable of growing at very high densities. Artemia also has a high nutritive value (40–60 percent protein, rich amino acid composition), an unchanging food requirement, high conversion efficiency, short generation time, high fecundity rate and long lifespan. The whole animal (even adult stage) may be consumed without previous processing by many aquaculture organisms.
With the expansion of aquaculture, the yearly demand of Artemia far exceeds its harvest. More attention should be given, therefore, to increase production of Artemia and maximize its use.
1 Tigbauan Research Station, SEAFDEC Aquaculture Department, Iloilo, Philippines
2. BIOLOGY OF ARTEMIA
2.1 Systematic classification
2.2 Life cycle
Artemia undergoes various stages in its life cycle. The discussion of each stage that follows is based on the works of Sorgeloos (1977) and Vos (1979).
The dehydrated dormant eggs of Artemia are called cysts. These minute brown particles (200–300 um in diameter) are found floating in many salt lakes and brine ponds or carried to the shore by wind and wave action. When kept dry or under oxygen-free conditions, they remain inactive, without a decrease in viability and hatching efficiency even after several years.
Upon immersion in seawater (5–70 ppt), the bean-shaped cyst becomes spherical and within the shell the metabolism of the embryo is activated. Hours later, the cyst's outer membrane bursts (breaking stage) and the embryo emerges still enclosed by the hatching membrane.
A short period after emergence of the embryo, the hatching membrane ruptures and the free-swimming nauplius (0.4 mm, 0.002 mg) is released. The first instar larva is brownish orange due to the presence of yolk. It has three pairs of appendages and an unpaired red ocellus (eye) in the head region.
The larva grows and differentiates through about 15 molts during which the trunk and abdomen elongate and the digestive tract becomes functional. Food particles are collected from the setae of the antennae. Lateral complex eyes develop at both sides of the ocellus.
Important changes in the morphology of Artemia take place from the 10th instar on. Antennae lose their primitive function and undergo sexual differentiation. Thoracopods also differentiate into three functional parts: telopodites as filter, oar-like endopodites for locomotion and membranous expodites as gills.
Adult Artemia (usually 8–10 mm long) are characterized by their stalked lateral eyes, sensorial antenullae, linear digestive tract and 11 pairs of thoracopods. In male Artemia, antennae are transformed into muscular graspers. A paired penis is found in the posterior part of the trunk region. In female Artemia, antennae have degenerated into sensorial appendages. Paired ovaries are found on both sides of the digestive tract, behind the thoracopods. Ripe eggs are transported from the ovaries into an unpaired brood or uterus.
A sketch of a male and female Artemia is shown in Fig. 1.
In some Artemia strains, parthenogensis occurs, i.e., no males are present and embryos develop from unfertilized eggs in females.
Fig. 1 Male and Female artemia
(Based from Schlosser's drawing in Sorgeloos, 1977)
2.1.5 Riding pairs
Precopulation in Artemia starts with the male grasping the female with its antennae between the uterus and the last pair of thoracopods. The couples can swim around in this position for several days. Copulation is very fast, with the male abdomen bent forward and one penis introduced into the uterus opening.
Fig. 2 illustrates the life cycle of Artemia.
2.3 Modes of reproduction
Two main modes of reproduction are found in Artemia. In ovoviviparous reproduction, the fertilized eggs develop to free-swimming nauplii which are released by the mother while in oviparous reproduction, eggs develop to the gastrula stage, become surrounded by a thick shell and are deposited as cysts. The latter type of reproduction is predominant at high salinities (150–200 ppt) and lower dissolved oxygen concentration.
One female produces about 50–200 cysts/nauplii per reproductive cycle. The shortest period observed between the consecutive cycles is four days.
3. QUALITY EVALUATION, DECAPSULATION AND HATCHING OF ARTEMIA CYSTS
3.1 Quality evaluation of Artemia cysts
The quality of Artemia cysts often varies according to strain or commercial brand. Lack of information, especially in the hatching performance of cysts could lead to wasteful hatching. Determination of the cyst quality is, therefore, necessary to maximize the use of Artemia.
Cyst quality may be expressed using the following indices: hatching percentage, hatching efficiency and hatching output.
|Hatching percentage||-||ratio of hatched cysts over the unhatched full cysts.|
|Hatching efficiency||-||quantity of cysts that should be incubated to produce a million nauplii.|
|Hatching output||-||product of naupliar biomass and hatching efficiency.|
3.2 Decapsulation and hatching of Artemia cysts
To produce nauplii, Artemia cysts are either hatched naturally by incubation in seawater for 24–48 hours or hatched after decapsulation.
Figure 2. Life cycle of Artemia
Decapsulation is the removal of the outer membrane of a cyst called the chorion by dissolution in hypochlorite, without affecting the viability of the embryo. The outer shell often causes problems when not removed since it can harbour bacteria and other organisms which may be harmful to the species feeding on Artemia. Also, non-hatched cysts and their shells cannot be digested and may cause blockage of the gut in fish and crustaceans.
3.2.1 The decapsulation/hatching process
The following procedure on cyst decapsulation and hatching is presently used at the SEAFDEC Aquaculture Department and is based on the work of Brugeman et al. (1979) and Sorgeloos et al. (1977).
Hydrate the dry cysts in natural seawater. Use a transparent conical tank or funnel-shaped container (e.g., glass or plastic cylinder, thick plastic bags formed into the desired shape) and keep the cysts in continuous suspension by aerating from the bottom of the apparatus for one hour.
Upon hydration, the dry cysts which are deflated like bean seeds become round-shaped. Full hydration is necessary to insure that the inner part of the indented dry cyst shell will be completely exposed when the decapsulation solution is added.
B. Reaction with decapsulation solution (hypochlorite)
Prepare the decapsulation solution using 1N NaOH, sodium hypochlorite (NaOCI) and seawater (see Laboratory Exercise No. 9 for details).
Allow the hydrated cysts to react with the decapsulation solution (hypochlorite) for 7–15 minutes. To prevent damage of embryo, keep the temperature below 40°C by adding ice cubes to the suspension or by using a water bath. A change in colour of the cysts from brown to white to orange usually indicates that the reaction is complete. Check under the microscope if possible.
C. Sieving and washing
Drain the suspension of decapsulated cysts into a fine-mesh sieve and rinse immediately with seawater 6–10 times or until the smell of the hypochlorite is removed.
Decapsulated cysts may be fed directly to the cultured fish/ crustacean or stored in saturated brine solution at low temperature for future use.
Incubate the decapsulated cysts for 24–48 hours in natural seawater at a density not greater than 5 grams cysts per liter of incubation medium. For optimum hatching, keep the temperature at 30°C and the pH at 8–9 (Sorgeloos, 1980). Provide sufficient light at least during the first two hours or preferably continuous illumination of about 1 000 lux (attained with 40-watt flourescent light tube, 20 cm away from the hatching container). Maintain the dissolved oxygen at levels close to saturation, with cysts kept in suspension throughout the incubation period.
If culture to the adult stage is not intended, it is preferable to feed the Artemia to the fish or shrimp larvae immediately upon hatching to take advantage of the yolk in the nauplii.
Fig. 3 illustrates the decapsulation and hatching process.
3.2.2 Advantages of cyst decapsulation
(Based from Bruggeman et al., 1979)
Disinfection of cysts through the hypochlorite treatment,
Increased energetic content,
Separation of the nauplii from the hatching debris not necessary since only thin, transparent membranes are left,
Decapsulated eggs may be fed directly to some fish and crustacean larvae.
Lower threshold for light triggering of the cysts' metabolism
4. INDOOR CULTURE OF ARTEMIA
In the absence of natural brine shrimp populations in the locality, various stages of Artemia may be obtained by culture in indoor systems. The batch culturing method recommended by Bossuyt and Sorgeloos (1980) and presently used at the SEAFDEC Aquaculture Department, Tigbauan, Iloilo, is described.
4.1 Culture system
The air-water-lift operated raceway (AWL-raceway) is found to be an effective culture system for growing Artemia larvae in batch cultures from nauplii to adults. This system consists of a rectangular tank with a central partitioning where air-water-lifts are attached (see Fig. 4).
Fig. 3 Steps in decapsulation and hatching of Artemia cysts
Advantages in using the AWL-raceway include the following:
Continuous aeration of the medium.
Nearly homogeneous circulation of the culturing medium.
Nearly all particulate matter is kept in suspension.
Feed added at one place is distributed all over the tank within a few minutes.
The system can be extended as long as necessary; only the height/width ratio of the culturing tank is critical.
At high densities, accumulation of faeces in the culture tank should be minimized. This is done by using a plate separator as shown in Fig. 5.
4.2 Food preparation and distribution
The important factors to be considered in choosing a diet for Artemia are particle size (should be less than 60 μm), food digestibility, nutritional value and solubility in the culturing medium (should be minimal).
Ricebran has been found to be an excellent food for Artemia. It is cheap and widely available and its particle size can be adjusted manually or by industrial grinding techniques. The manual method of preparing ricebran feed consists of wet homogenization in a kitchen blender at a ratio of 1 kg RB:3L seawater, followed by screening of the mixture through a 60-micron filter bag. Concentration of ricebran in the sieved mixture is approximately 100 g/L. The ricebran extract may be stored for 2–3 weeks in brine or at low temperature.
Food should be distributed frequently (at least every 2 hours) to attain the fastest growth and most efficient food conversion for the continuously filter-feeding Artemia. The amount of food to be distributed can be determined by the use of a turbidistick (Fig. 6): 15–20 cm water turbidity appears to be the optimal range for growth and survival of Artemia.
4.3 Culture procedure
Fill the AWL-raceway with natural seawater. Add 50 g CaO per ton of seawater.
Stock naturally-hatched Artemia nauplii (Instar I) in the late afternoon at a density of 15 per ml. Wash newly-hatched nauplii gently with seawater prior to stocking. For indoor culture, Great Salt Lake Artemia is found to be the most suitable strain.
No feeding is necessary overnight while yolk reserves of the Artemia are consumed.
Fig. 4 An air-water-lift operated raceway for indoor culture of Artemia
(Based from Sorgeloos, 1978)
Fig. 5 Plate separator for faeces removal in A-W-L operated raceways
(Base from Bossuyt & Sorgeloos, 1980)
Fig. 6 A Turbidistick
(Based from Bossuyt & Sorgeloos, 1980)
Start food distribution the following morning (after stocking), either manually or automatically. The daily food quantity has to be increased as the larvae grow.
On the third day, attach a plate separator into the AWL-raceway. The mesh-size of the filter screen should be varied according to the size of the Artemia.
After three days, reduce the stocking density of Artemia to 10 per ml.
To prevent accumulation of the ricebran waste, install a flocule trap whenever necessary.
Maintain the following conditions throughout the culture period:
|Dissolved oxygen||:||Higher than 4–5 ppm|
|Ammonia concentration||:||Below 80–90 ppm|
To check the growth and survival of Artemia, population sampling may be done daily using 40-ml bottles while length measurements can be taken every three days. Biomass determination may be made every five days using a 20-liter bucket.
A production of at least 5 kg wet weight Artemia per cu. m. is obtained after a two-week culture in AWL-raceway at optimal condition.
5. CULTURE OF ARTEMIA IN SALTPONDS
Although natural brine shrimp populations are not locally found, production of Artemia in existing saltponds is a distinct possibility. Periodic (dry season) culture of Artemia in the evaporation areas of these ponds will not only maximize utilization of space but will also be an added source of income for the operator. Furthermore, it minimizes dependence on costly imported cysts.
Successful inoculations have been reported in Brazil, India, Philippines, and Thailand. Since the climate of the Philippines is characterized to have a generally distinct rainy and dry seasons, the feasibility of integrating Artemia with salt production is also high. The discussion that follows is based on the studies conducted at the Leganes Station of the SEAFDEC Aquaculture Department and the works of Vos (1979), Vos and dela Rosa (1980) and Vos and Tansutapanit (1979).
5.1 Site selection
Artemia production may be undertaken in regions with distinct rainy and dry seasons or in places with low rainfall and high evaporation rate. These areas usually have existing salt production ponds.
Water supply in the area must be free of pollutants like industrial wastes and pesticides from agricultural run-offs. Water from mangrove areas is preferred since it is normally rich in nutrients and suspended food particles.
5.2 Pond design
Ponds must be so constructed that water depth of at least 30 cm can be maintained. Facilities for draining and filling the pond with either high salinity water or fresh seawater anytime the need arises should be provided. This is important for effectively managing the pond to bring in/replenish natural food and to manipulate the salinity to induce cyst or nauplii production.
Orientation of the Artemia ponds should take into consideration the predominant wind direction during the dry season so that cyst collection will be facilitated.
5.3 Pond preparation
All predators in the pond such as gobies, copepods and mosquito wrigglers must be eliminated prior to Artemia inoculation. This can be done by installing nylon screens (0.5–1.0 mm mesh) at the water intake gates or more effectively by raising the salinity to 90–100 ppt. Under optimal weather conditions, it takes 2–3 weeks to obtain the desired salinity at a water depth of about 30 cm.
Leakages and seepages should be checked.
Coconut fronds may be installed in the pond to serve as sun shades for Artemia during very hot days and to help prevent sudden temperature increases.
Application of organic (chicken manure) and inorganic (urea and 16-20-0) fertilizers a week or two before inoculation will help stimulate production of natural food for Artemia. Growth of phytoplankton is preferred over lab-lab since the latter may complicate the cyst collection process and seriously affect the Artemia when not broken down into very fine particles.
5.4 Artemia inoculation
Artemia may be stocked in the ponds early in the morning, late in the afternoon or preferably at night when temperature is relatively low.
Artemia nauplii are preferred over the adults as inoculum since they are easy to transport and less subject to thermal and salinity shock when directly transferred to the pond water.
Although the best stocking density is not yet established, inoculations at 20–40 000 nauplii per cubic meter have had good growth and survival.
Selection of a suitable strain for a particular area has to be determined by inoculation trials with various strains.
5.5 Pond management
If natural food is initially abundant in the pond, water may be allowed to evaporate further for a week after inoculation. Periodically thereafter, water is partially drained and replenished or replenished only with fresh or high salinity water. If the incoming water is not rich, natural food production may be enhanced by follow-up application of organic/inorganic fertilizer. The use of chicken manure is considered advantageous for it can also serve as direct food source for Artemia. Fertilizers must be applied immediately before or after the ponds are replenished with new/fresh seawater.
5.6 Cyst collection/harvest of Artemia
Regular collection of cysts floating at the surface of the pond should be made to prevent loss into the dikes. Installation of barriers or cyst collectors along the side or corner where the wind blows is also recommended. Barriers/collectors may be made of cloth, fine-mesh nylon screen or plastic sheet attached to bamboo stakes and pegged to the pond bottom. These collectors are partially submerged, with at least 15 cm allowance above the water surface. Accumulated cysts are periodically scooped with a two-layered dip net that concentrates the cysts in the inner layer. Harvested cysts are processed and stored in brine or in vacuum-packed/ nitrogen-flushed cans.
Occasional harvest of adult and pre-adult Artemia may be done using scoop nets or buckets. Total harvest of the cultured Artemia at the end of the dry season is recommended to avoid loss by predation when the rains come and salinity decreases. The harvested Artemia can be fed directly to fish/crustaceans or processed (dried or frozen) and stored for future use.
Bossuyt, E. and P. Sorgeloos. 1980 Technological aspects of the batch culturing of Artemia in high densities. Pages 133–152 in G. Persoone et al., eds. The brine shrimp Artemia. Vol. 3. Ecology Culturing, Use in Aquaculture. Universa Press, Wetteren (Belgium).
Bruggeman, E. et al. 1979 Improvements in the decapsulation of Artemia cysts. Pages 309–315 in E. Styczynska-Hurewics et al., eds. Cultivation of fish fry and its live food. EMS Spec. Publ. No. 4. Institute for Marine Scientific Research, Bredene (Belgium).
Dees, L.T. Brine shrimp. 1961 U.S. Department of Interior. U.S. Fish and Wildlife Service. Bureau of Commercial Fisheries. Fishery Leaflet 527: 1.5
Sorgeloos, P. 1977 Occurrence of Artemia in nature and its morphological development from nauplius to adult. Pages 1–7 in E. Jaspers, ed. Fundamental and Applied Research on the Brine Shrimp Artemia salina (L.) in Belgium. Special Publication No. 2., European Mariculture Society.
Sorgeloos, P. 1978 The culture and use of brine shrimp, Artemia salina, as food for hatchery-raised larval prawns, shrimps and fish in Southeast Asia. FAO Report THA/75/008/WP/3: 50pp.
Sorgeloos, P. 1980 The use of the brine shrimp Artemia in aquaculture. Pages 22–46 in G. Persoone et al., eds. The brine shrimp Artemia. Vol. 3. Ecology, Culturing, Use in Aquaculture. Universa Press, Wetteren (Belgium)
Sorgeloos, P., 1977 et al. Decapsulation of Artemia cysts: a simple technique for the improvement of the use of brine shrimp in aquaculture. Aquaculture 12(4): 311–316 pp.
Sorgeloos, P., 1978 et al. The use of Artemia cysts in aquaculture: The concept of “hatching efficiency” and description of a new method for cyst processing. Pages 715–721 in J.W. Avault, Jr., ed. Proc. 9th Ann. Meeting WMS. Louisiana State University, Baton Rouge (LA, U.S.A.)
Vanhaecke, P. and P. Sorgeloos. 1981 International study on Artemia. XIX. Hatching data on 10 commercial sources of brine shrimp cysts and re-evaluation of the “hatching efficiency” concept. Paper presented at the World Mariculture Society - Technical Sessions, Seattle (WA - U.S.A.), 8–10 March 1981
Vos, J. 1979 Brine shrimp (Artemia salina) inoculation in tropical salt ponds: A preliminary guide for use in Thailand. Working paper of the National Freshwater Prawn Research and Training Centre, Freshwater Fisheries Division, Department of Fisheries. FAO/UNDP: THA/75/008: 14p.
Vos, J. and N. de la Rosa. 1980 Manual on Artemia production in salt ponds in the Philippines. FAO/UNDP - BFAR Brackishwater Aquaculture Demonstration and Training Project (PHI/75/005): 44p.
Vos, J. and A. Tansutapanit. 1979 Detailed report on Artemia cyst inoculation in Bangpakong, Chachoengsao Province. FAO/UNDP Field Document. THA/75/008: 54p.
YEASTS AS FOOD ORGANISMS IN AQUACULTURE
C. T. Villegas1
Yeasts, members of Class Ascomycetes, are well distributed over the surface of the earth. They are particularly abundant in substrata which contain sugars. They are also found in the soil, in animal excreta, in milk, on vegetative parts of plants and other habitats, i.e., sand, rivers, estuaries and oceans.
They are particularly noted for their ability to ferment carbohydrates. The fermenting activities of certain yeasts are the basis of the baking and brewing industries. The universal employment of yeast in baking and brewing has resulted in the establishment of another industry, the commercial preparation of yeast cake. By pressing into cakes of great numbers of yeast cells together with inert material such as starch, dry yeast so useful in industry and home, is prepared.
The pelagic zone of the ocean or even a large lake represents a limitless food supply for the individual planktonic animal but this source of energy may be so thinly dispersed and getting enough to eat is still a problem. Aquaculturists have seen the need for substitute food source and yeasts could be the answer.
1 Researcher, Tigbauan Research Station, SEAFDEC Aquaculture Department, Tigbauan, Iloilo, Philippines.
2. YEASTS IN AQUACULTURE
Alon (1974) utilized various readily available food materials such as Torula yeast, food yeast (Saccharomyces), baking yeast and soya powder to simplify the mass culture of the rotifer Brachionus plicatilis. For the California strain of Brachionus, the rotifer grew nearly exponentially on all diets with various yeast and soya powder giving much better growth than the control diet, Chlorella. The Torula yeast diets yielded the highest population, final rotifer concentration was 70 percent higher than the control. The other diets of food yeast, soya powder and baking yeast gave 45 percent, 40 percent and 20 percent higher rotifer density than the control after 15 days. The food yeast resulted in 178 rotifers per ml while soya powder and baking yeast gave 170 and 140 individuals per ml while the Chlorella control yielded 123 individuals per ml. Relatively slow growth was observed for the Japanese strain. A better response in terms of population growth was noted for baking yeast, food yeast and Torula yeast compared with the control. The baking yeast gave the best growth with a final rotifer concentration of 106 individuals per ml, about 90 percent higher than the control with only 55 rotifers per ml. He concluded that powdered yeast products, readily and cheaply available on the commercial market may facilitate rotifer mass culture for utilization in the larval culture of marine animals.
Similar conclusions have been reached by some Japanese workers. Hirata and Mori (1967) reported the use of Chlorella and baker's yeast in the culture of the rotifer, B. plicatilis. Hirayama and Watanabe (1973) tested caked yeast Rhodotorula sp. and dried marine yeast (AS Y-4011) as food. Caked yeast (still living) Rhodotorula sp. has less nutritional effect than the marine Chlorella. Dried marine yeast added to diluted Chlorella suspension was very effective. He concluded that yeast may be most effective as supplemental food when enough phytoplankton cannot be supplied. Furukawa and Hidaka (1973) have reported successful mass culture of the rotifer, B. plicatilis with final densities ranging from 100–1 000 individual per ml within experimental periods of 27–37 days using freeze dried marine yeast Turulopsis candida var. marinae as food material for the rotifer employing a method of water exchange and periodic thinning.
In Japan, the most practical technique for the production of a copepod Tigriopus japonicus is a combination culture with a rotifer, Brachionus plicatilis fed with baker's yeast or W-yeast (Fukusho et al., 1978; Fukusho, 1980). Chlorella minitissima were first cultured in 200-ton concrete tanks A and B (10 × 15 m × 1.4 m depth) which were fertilized in advance and seawater was added gradually to full amount with aeration until the density of 97 × 105 cells per ml in Tank A and 145 * 105 cells per ml in B were attained. Tigriopus were seeded with the rotifers at density of 15/L in A and 29/L in B. Then W-yeast and baker's yeast were supplied in Tanks A and B at the rate of 1 g/1 million rotifer/day. Total amounts harvested during 89 days were 168.0 kg and 81.5 kg with maximum densities of 22 048 ind/L and 10 000 ind/L for Tank A and B, respectively. The maximum densities for rotifers were 169/ml and 46/ml.
Tigriopus japonicus fed with W-yeast and baker's yeast as food for the larvae of mud dab, Limanda yokohamae (30 days old, 10.30±0.51 mm in T.L.) for 23 days improved survival rate and the growth of fish. T. japonicus cultured with W-yeast were better than those fed with baker's yeast (97.4%– 97.0% survival; 14.3-14.0 vs. 12.5-12.0 times increase in body weight). Both groups showed excellent survival rate and growth. These results indicate that T. japonicus is a suitable food organism for mud dab and that its nutritional quality is successfully improved by culturing it with W-yeast.
Furukawa (1972) and Hirata (1975) had some success in small tank culture feeding of marine yeast diet and baking yeast to Penaeus japonicus. Furukawa (1972) enumerated a few points that have to be taken into account in the cultivation of fry by means of marine yeast as follows: (i) the temperature of the water should be kept slightly lower (less than 27°C) although this causes delay in growth; direct sunlight should be avoided in order to check the propagation of diatoms and prevent pH from rising; (ii) when the density of marine yeast is maintained at about 1×106 cells/ml the pH can be maintained at about 8.1 At density of 1×108 or more, however, the pH value will become higher than adequate; (iii) the rate of zoea feeding on yeast is approximately 50 percent at the maximum while the rate of growth per day is between 1×106 - 1×107 per ml at the maximum. He stated that the ecosystem of living creatures comprising a combination of the marine yeast and zoea or a combination of the marine yeast, rotifer and zoea, provides a milder and more favourable environment for the growth of zoea than the ecosystem of phytoplankton and zoea.
Epifanio (1979) studied the value of torulan yeast, Candida utilis as food for four species of bivalve molluscs. They were fed diets consisting of varying proportions of the yeast and the diatom Thalassiosira pseudonana. Juvenile Argopecten irradians, Mercenaria mercenaria and Mytilus edulis grew as fast or faster than with the control (100% algae) when fed diets containing as much as 50 percent yeast. Growth of soft tissue in Crassostrea virginica was decreased with the amount of yeast in the diet. His results showed that the yeast, Candida utilis, is suitable as a replacement for up to 50 percent of the algae in the diets of three of the four species tested. He stated that the apparent inability of C. virginica to utilize yeast suggests some differences in the digestive physiology of this species.
The use of yeast, as feed in aquaculture can be summarized as follows:
It has been used successfully as feed during the early larval stages (up to zoea 2) of Penaeus japonicus.
Yeast and powdered yeast products have facilitated the mass culture of the rotifer, Brachionus plicatilis for utilization in the larval culture of marine animals.
Some species of yeast (W-yeast) have improved the nutritional quality of the copepod Tigriopus japonicus, a suitable food for the mud dab, Limanda yokohamae.
It has been found most effective as supplemental food when enough phytoplankton cannot be supplied.
Alon, Noel C. 1974 Simplified culture of the rotifer, Brachionus plicatilis Muller using yeast diets and EDTA. (Handout)
Epifanio, C.E. 1979 Comparison of yeast and algal diets for bivalve molluscs. Aquaculture 16(3): 187–192
Fukusho, K. 1980 Mass production of a copepod, Tigriopus japonicus in combination culture with a rotifer, Brachionus plicatilis fed W-yeast as a food source. Bull. Jap. Soc. Sci. Fish. 46(5): 625–629. (In Japanese with English Abstract)
Fukusho, K. et al. 1978 Effect of initial supply of Chlorella sp. on the copepod Tigriopus japonicus production in combination with the rotifer, Brachionus plicatilis, feeding baker's yeast. Bull. Nakasaki Prefectural Institute of Fisheries No. 4, pp. 47–56
Fukusho, K. et al. 1980 Food value of a copepod, Tigriopus japonicus, cultured with W-yeast for larvae and juveniles of mud dab Limanda yokohamae. Bull. Jap. Soc. Sci. Fish. 46(4): 499–503. (In Japanese with English Abstract)
Furukawa, I. 1972 Rearing methods of prawn, Penaeus japonicus Bate fed marine yeast. Yooshoku 9(9): 38–42
Furukawa, I. and K. Hidaka. 1973 Technical problems encountered in the mass culture of the rotifer using marine yeast as food organism. Bull. Plankt. Soc. Japan 20(1): 61–71. (In Japanese, English Summary)
Hirata, H. 1975 An introduction to the rearing methods of prawn, Penaeus japonicus Bate in Japan. Mem. Fac. Fish., Kagoshima University 24: 7–12
Hirata, H. and Y. Mori. 1967 Mass culture of marine rotifer, Brachionus plicatilis fed baking yeast. Saibai Gyogyo 5: 36–40
Hirayama, K. and K. Watanabe. 1973 Fundamental studies on physiology of rotifer for its mass culture - IV Nutritional effect of yeast on population growth of rotifer. Bull. Jap. Soc. Sci. Fish. 39(11): 1129–1113
Villegas, C.T. et al. 1980 The effects of feeds and feeding levels on the survival of a Prawn, Penaeus monodon larvae. Memoirs of the Kagoshima University Research Center for the South Pacific 1(1): 51–55
CULTURE OF INVERTEBRATES AS FOOD ORGANISMS
FOR VILLAGE-LEVEL FISH HATCHERIES
Rafael D. Guerrero III1
Small-scale fish hatcheries, particularly for fishes and the giant prawn, Macrobrachium rosenbergii, are highly viable enterprises in developing countries such as the Philippines and Thailand. With simple village-level technologies, Asian fish farmers have demonstrated that many forms of food organisms can be economically produced without the need for sophisticated equipment.
This paper is a review of some practical methods that may be applied by fish farmers in producing food organisms for fish and prawn hatcheries. The organisms include Moina, chironomid larvae mosquito wrigglers and earthworms.
2.1 Culture of Moina sp.
The Moina is a cladoceran which is fairly common in freshwater habitats. This zooplankter is a filter-feeder of bacteria thriving in manure solutions.
The female Moina can breed either sexually or parthenogenitically. Each female can produce 7–24 young. The average life span of M. macrocopa is 8.72 days (Bellosillo, 1937).
Ventura and Enderez (1980) reported on the production of Moina sp. in concrete tanks using hydrated chicken manure applied in sacks. The Moina population was maintained at 20 000–40 000/0.5 m2 with abundant Anabaena sp., Scenedesmus sp., Chlorella sp., Schroederia sp. and Staurastrum sp. Chicken manure was applied at the rate of 20 g/m2, while water depth was maintained at 40–50 cm.
1 Consultant, SEAFDEC Aquaculture Department, Tigbauan, Iloilo, Philippines
In a preliminary study, Mane (personal communication) applied approximately 210 kg of dried chicken manure in a 0.1-ha freshwater pond 50–60 cm deep. After a week, Moina sp. was collected in abundance using a scoop net of bolting silk cloth. No record of the production is available.
Moina is usually fed live to fish fry and Macrobrachium larvae. It may also be frozen for preservation and fed to the fry while thawing.
2.2 Culture of chironomid larvae
Chironomid larvae and pupae are among the desirable and practical food items used in feeding carnivorous fish fry. Chironomus sp. feeds on algae and has high reproductive capacity. Each female lays about 2 300 eggs in one batch which hatch in about 3 days at high temperatures. The larvae attain a size suitable for feeding purposes in 16–20 days (Lutz et al., 1959).
In Hong Kong, chironomid larvae are farmed in 700–1 000 m2 vegetable fields with mud 10–13 cm deep. In a 1 000-m2 field, 1 180–1 480 kg of dried chicken manure are applied as fertilizer. The yield at 15–30°C is about 140 kg in a 675-m2 field every 50 days (Shaw and Mark, 1979).
The larvae may be kept alive for hours and days for transport in containers under cool and moist conditions.
2.3 Culture of mosquito wrigglers
Mosquito larvae or wrigglers are very suitable food organisms for all kinds of fish. These aquatic insects feed on vegetable matter and may be cultured in old jars filled with water fertilized with manure (Jocher, 1969).
In a preliminary study, Guerrero (unpublished data) produced Culex sp. larvae in an open drum with an almost pure culture of Kirchneviellalunaris, a colonial green alga. The water in the drum was fertilized with human urine at 10 parts per thousand.
Mosquito larvae are collected by means of fine-mesh scoop nets. They can be kept alive for several hours in a damp cloth.
Dried and ground wrigglers mixed with other feed ingredients as yeast and ricebran can serve as excellent food for fry.
2.4 Culture of earthworms
Earthworms are beneficial soil annelids. They improve soil condition and produce humus. Aside from these, they are also valuable as a cheap source of protein for fish feeds.
Tropical countries have an ideal climate for the culture of earthworms. There is also an abundance of agricultural wastes such as animal manures that can be utilized efficiently and economically for earthworm production.
Earthworms can be mass-produced by breeding them in worm beds, boxes or tanks indoors or outdoors. They grow best in an organic matter substrate containing 10–12 percent protein (Swingle, 1961).
In culture studies on Perionyx excavatus, Guerrero (1980) found circular concrete tanks as suitable culture units. Substrates tested to be satisfactory for P. excavatus production consisted of Murrah buffalo manure alone for breeding, 75 percent manure and 25 percent Leucaena leucocephala (ipil-ipil) leafmeal for fattening and a combination of 50 percent manure, 25 percent ipil-ipil leafmeal and 25 percent sawdust for growing.
Earthworms are harvested by hand or by means of mechanical harvesters. Use of selective screens has also been employed successfully. Young worms or juveniles are usually fed live to fish fry and/or shrimp post-larvae. Processing of earthworm meal is simply done by drying and grinding the worms.
Proximate analysis of dried P. excavatus showed it to contain 64.7 percent crude protein, 14 percent fat, 13.9 percent NFE, 4.2 percent moisture and 3.2 percent ash. Some essential minerals and vitamins (per 100 g) analyzed in earthworm meal were calcium - 286 mg, phosphorus - 10 004 mg, vitamin A - 733 I.U., thiamine - 0.12 mg, riboflavin - 2.86 mg and niacin -5.0 mg.
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