Hatchery operation: culture of larvae basic methodology, feeding and nutrition, factors influencing growth and survival, and settlement and metamorphosis
5.1 BASIC METHODOLOGY
The hatchery culture of bivalves is as much an art as it is a science and the old adage applies that "there are many ways to skin a cat". In a similar vein, the success of a hatchery is related more to the skill and intuitive "feel" for the work of the manager and technicians than it is to the location, the scale and quality of the physical structure and the sophistication of the available equipment. Every hatchery is different in the way it is managed and in the nuances of the manner in which the various aspects of culture are approached and the work done. There is no standard methodology as such but there are common denominators that relate to the need to fulfill the biological requirements of the different bivalve species through their early developmental stages.
This section of the manual synthesizes the various approaches and the methods used in the culture of larvae from the fertilized egg to settlement with emphasis on some of the more commonly cultured species.
5.1.2 Methods for embryo development
22.214.171.124 Tanks for embryos and larvae
Fertilized eggs are permitted to develop to the fully-shelled "D" veliger stage in tanks of the type shown in Figures 50 and 52. This early veliger stage is known as the D-larva stage because of the characteristic "D" shape of the shell valves (Figure 51). D-larvae of the various, commercially cultured bivalves are similar in appearance.
A wide range of circular or semisquare (square with rounded corners) tanks can be used for embryo development and also for larval rearing (Figure 52). They should be constructed from preferably either "virgin" (new, non-recycled) polyethylene or fibreglass (alternatively known as GRP - glass reinforced plastic or fibreglass). Previously unused vessels should be filled with seawater and allowed to soak with weekly changes of water for 2 to 4 months before use. Soaking removes toxic substances that leach from the surface of new plastics which may be harmful to larvae.Steam curing of fibreglass tanks substantially reduces the period the tanks need to be seawater soaked.
Figure 50: Fertilized eggs can be incubated in various types of tanks in filtered seawater for a period of 2 to 3 days, depending on species and temperature.
Flat-bottomed or steeply tapering conical tanks (i.e. almost flat bottomed) are most commonly used for embryo development (Figure 52). Shallowly tapering conical tanks (shaped like an ice cream cone) are less satisfactory because the early embryos are immobile and will tend to aggregate together at the bottom of the cone.
Figure 51: Photomicrograph of Crassostrea gigas D-larvae (48-h after fertilization). Mean size is 75 µm shell length.
Surface area of the tank base rather than water depth is more important. Aeration during this early stage is not recommended. The mechanical effects of the disturbance it creates can lead to abnormal development.
Figure 52: Suitable rearing vessels for embryo (and larval) development. A - 200 l steeply tapering conical fibreglass tank with bottom drain; B - 125 l polyethylene flat-bottomed tank; C - 1 000 l insulated polyethylene, square tank with rounded corners.
126.96.36.199 Water treatment
Culture tanks are filled with seawater filtered to 1 to 2 µm particle size (Figure 53A) and heated to the required temperature (usually 18 to 24ºC; cooler for cold water species). Some hatcheries disinfect the water following fine filtration by passing it through an ultra-violet light (UV) unit (Figure 53B), the value of which is questionable unless it is used properly and with discretion.
UV units should be maintained according to manufacturer’s recommendations and a record kept of hours of lamp usage. Lamps must be replaced as they reach the specified hours of use at which time the quartz silica sheath that separates the lamp from the water flow needs to be cleaned with a soft cloth soaked in alcohol. Moreover, these units are designed to disinfect freshwater and are not as efficient in killing or immobilizing marine bacteria and other micro-organisms.
As a rule of thumb - if UV disinfection is considered necessary - it is best to pass water through two or three similar units, connected in series, at half the flow-rate recommended for a single unit (Figure 53). It should be remembered that limiting the diversity of bacteria in a culture of embryos or larvae may reduce competition and thereby permit potentially harmful bacteria to predominate. Modern thinking is that the probiotic approach is the better option. This approach entails controlling the density of larvae carefully, feeding them properly with only the best cultured algae available and, paying attention to the hygienic operation of both the cultures and equipment.
Figure 53: Examples of suitable equipment for water treatment. The multiple bag filtration unit (A) is arranged for fine water filtration. One bank of 3 filters is in use while the second bank is serviced and readied for use. These filtration units contain bags that progressively remove particulate matter from 10 µm down to 2 µin three stages. The uv disinfection unit (B) consists of lamp units arranged in series and is designed to treat a continuous flow of previously filtered seawater. This is the recommended arrangement in seawater treatment rather than relying on the water being treated by a single lamp unit.
It is sometimes beneficial to filter the water and to fill the culture tanks 24 hours before they are needed. This is more applicable in hatcheries located adjacent to estuaries contaminated by industrial or domestic wastes, or by the leachings from richly metaliferous geological strata (and mine workings) in the catchment area, which may contain elevated quantities of heavy metals. The water is then treated by adding 1 mg per l of EDTA (sodium salt - as used in the preparation of algal culture medium) and 20 mg per of sodium metasilicate and is vigorously aerated for 24 hours. Pre-treatment helps to complex heavy metals and render them non-toxic to the particularly vulnerable early stages in development of bivalve larvae. The water does not need to be re-filtered after pre-treatment but aeration is switched-off during embryo development.
188.8.131.52 Culture of embryos
Embryos are stocked in the culture tanks about 2 hours after fertilization and at the appropriate density. Fully developed D-larvae are recovered 24 to 48 hours later, depending on species and water temperature (Figure 54). Either no or very low aeration is used during embryo development.
Embryo stocking densities for many of the commonly cultured oviparous oysters and clams can be as high as 50 000 to 80 000 per l of culture, although 20 000 per l is more generally considered to be the safe upper limit (Table 10). In contrast, similarly high initial embryo densities of many of the scallop species leads to abnormal development and numbers are usually restricted to 10 000 to 15 000 fertilized eggs per l of culture tank volume in warmer water species. Egg densities are more commonly based on the surface area of the tanks rather than on tank volume in cold water scallop species, where maximum density should not exceed 1 000 per cm2 (Table 10).
Figure 54: Development of embryos from the early trochophore (A) to the fully shelled D-larva stage (D). The ciliated swimming feeding organ (velum) can be seen in B and early shell valve formation in C. Fertilized eggs will develop to fully formed D-larvae in less than 2 days in many warm water species but the entire developmental process can take 4 or more days in cold water species.
Table 10: Summary data of typical embryo densities (thousands per l), initial D-larva size (shell length, µm), densities of D-larvae (thousands per ml) and culture conditions in terms of suitable temperature (+ 2ºC) and salinity (+ 5 PSU) for the culture of embryos and early larvae of a number of bivalves. Notes: N/A - not applicable: embryo development takes place within the mantle cavity in Ostrea edulis. * Embryo densities in cold water scallops are calculated as embryos per unit area of the base of tanks rather than per unit volume. Maximum density should not exceed 1 000 fertilized eggs/embryos per cm2.