All rights reserved. Reproduction and dissemination of materials contained in this manual, in paper or electronic form, for educational and non-commercial purposes, are not authorized without prior written consent from the copyright holder provided the source of information

is fully acknowledged. Reproduction of material contained in this manual for resale or commercial purposes is prohibited without permission of the copyright holder. Applications for such permission should be addressed to the Director General of Fisheries Research, Ministry

of Agriculture &Fisheries Wealth, P.O. Box 427, Muscat 100. Sultanate of Oman.First edition March, 2012.

Suggested citation format: Al Rashdi K.M., Eeckhaut I. and Claereboudt M. R. 2012. A manual on Hatchery of Sea

cucumber Holothuria scabra in the Sultanate of Oman. Ministry of Agriculture and Fisheries Wealth, Aquaculture Centre, Muscat, Sultanate of Oman. 27 pp.

Ministry of Agriculture and Fisheries WealthDirectorate General of Fisheries Research

Aquaculture Center

A Manual on Hatchery of Sea Cucumber

Holothuria scabrain the Sultanate of Oman

Khalfan M. Al Rashdi Igor Eeckhaut

Michel R. Claereboudt

FOREWORD

The Sultanate of Oman is richly endowed with marine resources that provide livelihood to fishermen in the coastal communities. Sea cucumber is just one of these resources providing source of income to Mahout Bay fishers (men and women). The species Holothuria scabra is one of the most sought-after beche-de-mer products being sold to international markets.The Ministry of Agriculture and Fisheries Wealth through the Agriculture and Fisheries Development Fund has created a project titled Feasibility study of sea cucumber aquaculture in Oman and provided the much-needed financial support to be able to conduct research studies aimed at developing techniques in the hatchery of the sea cucumber Holothuria scabra. Because of the high demand for beche-de-mer products, fishers put so much fishing pressures on this resource causing decreasing fishery production to the degree of overfishing. Therefore, there is an urgent need to improve and intensify the production of hatchery-bred sea cucumber juveniles for grow-out culture and for stock enhancement.This manual shall serve as a guide to aquaculture technicians, students, entrepreneurs and marine enthusiasts, who shall endeavor to multiply the efforts of the authors.

Saoud H. Al-Habsi, PhDDirector General of Fisheries Research

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ACKNOWLEDGMENT

We thank the Agriculture and Fisheries Development Fund for providing the necessary fund for the conduct of various research activities and final

printing of this manual. Special thanks are due to His Excellency Dr. Fuad Al Sagwani, Minister of Agriculture and Fisheries Wealth and His Excellency Dr. Hamed Al Oufi, Undersecretary

of Fisheries Wealth for their support and encouragement to write this manual. Also we thank Dr. Saoud Al Habsi, Director General of Fisheries Research and Dr. Fahad Ibrahim, Director of Aquaculture Center for their unwavering support in sea cucumber aquaculture project and their sincere encouragement to finally see the

printing of this manual. We thank Madagascar Holothurie Company (Madagascar) and University of Mons

(Belgium) for taking their time off

to help us in the hatchery trials and without their technical support this manual will not come out.

Khalfan M. Al Rashdi Igor EeckhautMichel R. Claereboudt

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FOREWORD

ACKNOWLEDGMENT

INTRODUCTION

Diversity of sea cucumbers in Oman

Status of Sea Cucumber Fishery in Oman

Biology of Holothuria scabra

Habitat and Environment

Feeding and Growth

Sex and Reproduction

Maturation and Development

Artificial Propagation Techniques

Source of broodstock

Preparation for Spawning

Hatchery Techniques

Hormone-induced oocyte maturation and fertilization

Induced spawning

Pre-spawning and spawning behavior

Egg collection, counting, incubation and hatching

Post-spawning maintenance

Larval rearing

Food and feeding

Preparation of settlement substrates

Larval settlement and metamorphosis

Some problems in larval rearing

References

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CONTENTS

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INTRODUCTIONThe sea cucumber fishery in the Sultanate of Oman is centered on one species, the Holothuria scabra. The H. scabra fishery is limited on the eastern side of Mahout Bay area in Al-Wusta region, which is characterized by seagrass beds with fine sand in sheltered flats and lagoons. H. scabra is considered one of the most commercially valuable species for beche-de-mer production and have been widely fished in the tropics. The A-grade classification of beche-de-mer from sandfish H. scabra commands one of the highest prices on the international market. China has become the largest producer of sea cucumber worldwide.

DIVERSITY OF SEA CUCUMBERS IN OMANAlthough there are a growing number of studies focusing on echinoderms and holothuroids worldwide, the Arabian Sea has not received much attention. There have been a few publications on echinoderms in the region, but with the exception of recent studies on the fisheries and stock assessment of Holothuria scabra (Al-Rashdi et al. 2007a; Al-Rashdi et al. 2007b), there are no first hand records of sea cucumbers in the Sultanate of Oman except a recent study done by Michel & Khalfan (2011). The Sultanate of Omans coastline extends over than 3,500 km in three connected bodies of water: the Arabian Gulf, the Sea of Oman and the Arabian Sea. The country has always depended on the sea, however human pressure on its natural resources is steadily increasing from fishing, extensive infrastructure development, tourism and industry.

The marine climates in each of the three bodies of water are very distinct. The Arabian Sea coastal ecosystems withstand the full force of a seasonal coastal upwelling during the summer monsoon (JuneSeptember) with sea surface temperatures often dropping well below 20C. The cool upwelled water is accompanied by a steady influx of nutrients that feeds the growth of extensive beds of benthic algae interspersed with rich coral communities. In the Arabian Gulf and the Sea of Oman, sea surface temperatures in the same summer period often exceed 32C, whereas in the winter, water temperatures drop to below 22C. Despite these extremes, coral communities flourish along the hard-substrate shores and support a rich echinoderm fauna. There is only one marine protected area (the Daimaniyat Nature Reserve) in the Sea of Oman, and it covers about 20 km2 and encompasses a string of nine

However, the worldwide supply of beche-de-mer could hardly meet the Asian market demand. The world beche-de-mer market is largely controlled by Chinese traders and Hongkong SAR is still the major world market followed by Singapore which is rather more stable. From the nutrition and medicinal point of view, sea cucumber is an ideal tonic food. It has higher in protein and lower in fat than most other sea foods. It has been a source of chondroitin sulphate which is well-known for reducing arthritic pain. Currently, there are an increasing number of commercial products containing sea cucumber and its extracts for nutritional and medicinal purposes.

Fig. 1. An adult sandfish, Holothuria scabra

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Holothuroidea

Holothuria leucospilota

Holothuria atra

Holothuria scabra

Holothuria nobilis

Fig. 2. Some sea cucumbers species found in Omani waters

small islands. In total, 17 species of Aspidochirotida, 2 species of Apoda and 2 species of Dendrochirotida were observed during the survey. Additional species of Dendrochirotida were observed in the south of Oman. Although eight species were found in all or nearly all locations, many others were only found in four locations or fewer. Holothuria scabra, for instance, was restricted to two locations on the Arabian Sea near the Island of Masirah, and Holothuria arenacava was only found in a few sandy embayments near Muscat. The overall number of species decreased from the Arabian Sea (19 species) to the Sea of Oman (13 species) to the Arabian Gulf (8 species). Several species were recorded for the first time during the survey: Holothuria arenacava, H. nobilis, a common yet unidentified species of Holothuria, H. cinerescens, and a mottled pink Actinopyga, these last three were from Dhofar (southern Oman). Holothuria hilla and Holothuria impatiens were also observed. From a community standpoint, both the non-metric multidimensional scaling analysis and the cluster analysis, identified a major split in the structure of sea cucumber communities between the Arabian Sea and the Sea of Oman. This first subdivision is mainly due to six species that were restricted to the Arabian Sea coast of the Sultanate: Holothuria scabra, H. nobilis, Actinopyga miliaris, Actinopyga sp. (unidentified species), H. cinerescens and an unidentified species of Holothuria sp. Secondary subdivisions of communities distinguish the northern part of the Arabian Sea coast from its most southern part (Dhofar). In the Sea of Oman, sea cucumber communities from the central part of the Gulf (Muscat) are separated from those in the northern and southern regions. The holothuroid community found near Mahout Island appeared to be relatively distinct from both that of the Arabian Sea and the Sea of Oman.

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Fig. 3. Women of Al Wusta province, collect sandfish

during low tide

STATUS OF SEA CUCUMBER FISHERY IN OMAN

There have been very few studies conducted

on the fishery of sea cucumber in the Sultanate

of Oman. Harvesting of sea cucumber mainly

Holothuria scabra, constitutes a minor fishery

in Oman which takes place in Mahout Bay.

Due to the increased demand for beche-de-

mer in the international markets, a revived sea

cucumber fishery was noted in 2003. The fishery

is usually linked with the shrimp fishing season

that runs from September to March. However,

sea cucumber harvesting is done only when the

number of shrimp landings decreased usually

during November.

Despite the lack of regulations in the harvesting

of sea cucumbers, some general fishery

management rules are being practiced in the

region such as the restriction on the use of

SCUBA for harvesting any marine resource. Sea

cucumbers are only collected by hand during low-

tide or by skin-diving in deeper areas. The peak

months for harvesting sea cucumber are from

September to January, despite the lack of specific

closed season. Studies showed evidences of

rapid overfishing of sea cucumbers in Oman.

Recently, the average harvest per fisher in 2007

was only less than 20 per fishing trip compared to

about 100 sea cucumbers per fisher per fishing

trip in 2005. In 2005, 50% of the fishers were

Women, and because of the stock depletion their

percentage has dropped in 2008 down to 15%.

Processed sea cucumbers showed a significant

number of very small individuals of less than 6 cm (dried form) which correspond to about 12 cm of

live specimens.

Fig. 4. Small caught H. scabra which reflects overfishing

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At present, the government through the Ministry of Agriculture and Fisheries Wealth has already initiated some measures to monitor the sea cucumber fishery with the objective of providing regulatory framework that will ensure the sustainability of the resource.Classification and Morphology of Holothuria scabraSea cucumbers are holothurids which are related to sea stars, brittle stars, and sea urchins. The taxonomy of sandfish H. scabra is:

Phylum: Echinodermata, Class: Holothuridae (with tube feet), Family: Holothuriidae (with circular body and single gonad), Genus: Holothuria (Metriatyla) Rowe, 1969, and Species: H. scabra Jaeger 1833.

Sea cucumbers have elongated and cylindrical body. The body wall is the edible part and accounts for more than 50% of the total animal weight. The body surface is relatively smooth with small papillae or sensory tube feet, with black dots and dark transverse wrinkles. The mouth is on the ventral surface at the anterior end of the body while the anus is located dorsally at the posterior end of the body. Sea cucumbers have tiny calcareous plates called spicules in their skin (Fig. 4). Microscopic examination of spicules can be used for species identification.

Fig. 4. Sea cucumber

morphology

Fig. 5. Natural habitat of Holothuria scabra

Biology of H. scabraHabitat and EnvironmentSea cucumbers are an abundant and diverse group of worm-like and usually soft-bodied echinoderms. They are found in nearly every marine environment, but are most diverse on tropical shallow-water coral reefs. They range from the intertidal, where they may be exposed briefly at low tide, to the floor of the deepest oceanic trenches. They are found in sandy-muddy areas with lots of sea grasses; mostly in shallow waters of 5-10 meters deep. The more preferred areas have high levels of nutrients. They can tolerate lower salinity up to 20 ppt for short period of time.

Feeding and Growth Juveniles and adults of sea cucumbers feed on detritus or decaying organic matter. However, under captive or culture conditions they can feed on dry seaweeds and artificial feeds. Sea cucumbers grow to a size of 300-500 g (15-16 inches long) over one year which is equivalent to a monthly growth rate of about 15-25 g month.

Sex and Reproduction

By external features, it is not possible to determine the male from the female sea cucumber. Sex can only be determined by biopsy or dissection of animal to obtain

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Fig. 5. The life cycle of sandfish, Holothuria scabra

gonad sample. However several studies showed that animals are ripe at most times of the year, with one or two seasonal peaks.

Sea cucumbers can reproduce both sexually and asexually, although most species have defined sexes. During asexual reproduction, the sea cucumber will divide itself down the median line, also known as fission, to create two separate organisms, which will complete their development by regenerating the missing body parts. Some can only complete this process if certain parts are present.

In contrast, sexual reproduction or spontaneous spawning involves the external fertilization of eggs and the subsequent fertilization by sperm within

the water column. The fertilized eggs will hatch and the larvae will live a planktonic or free floating existence, swimming along with the aid of tiny cilia for mobility, eating and growing until they have reached the metamorphosis stage.

Metamorphosis and Development

Sea cucumbers undergo a metamorphic phase that transforms from larva to adult (Fig. 5). During metamorphosis, larvae undergo a complex development and regeneration of internal organs. The newly emerged juveniles sink to the bottom in order to complete their development and take on the adult form, but will continue to grow until they reach the full-adult size.

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ARTIFICIAL PROPAGATION TECHNIQUESIn order to determine their potentials for commercial culture or for restocking of depleted marine areas it is necessary to develop mass seed production techniques for the sea cucumber.

Source of Broodstock Wild sea cucumber broodstock are usually used for artificial propagation. Animals are collected during their reproductive season so that they are ready for immediate spawning. Ideally, broodstock must be around 250 to 500 g in weight (Fig. 3). Furthermore, animals with no bodily damage or lesions due to collection procedures, have smooth and shiny skin with transparent mucous layer, and have not eviscerated should be selected.

Sea cucumbers are collected by hand

during low tide. While at sea, collected animals should be kept in insulated containers with aeration using portable battery-operated aerator if holding time is more than two hours. It is better if the animals defecate while in the holding containers before packing in transport bags. Although sea cucumbers can tolerate low dissolved oxygen levels and high temperature (up to 30oC) for a long period of time without eviscerating, they must be provided optimal holding conditions i.e. water temperature of 27-30 oC to prevent stress which can cause premature spawning.

Prior to packing, animals must be cleaned by gently washing the body with seawater and should be packed at low density or better yet individually if using smaller plastic bags filled with one liter of sea water and inflated with oxygen.

Fig. 6. Newly-caught sea cucumber H. scabra broodstock from Mahout Bay

Fig. 7. Selected broodstock prior spawning.

Preparation for Spawning

Upon arrival in the hatchery, the animals are acclimatized while in the transport bags with ambient seawater. Their bodies are gently cleaned before placing in the tank with flow-through sea water and aeration. Sediments or faeces are siphoned at the tank bottom immediately prior to induction of spawning. The animals are induced to spawn after about 30 minutes to 1 hr (Fig. 7).

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HATCHERY TECHNIQUESHormone-Induced Oocyte Maturation and Fertilization

Broodstock are dissected to obtain their gonads for the in-vitro fertilization (Fig

7-Part 1). A cut was made using a pair of scissors at the lower ventral side of its body. Body fluid is drained and the gonad

is taken out using forceps and placed in a beaker. Gonads are then cut into small pieces using a pair of scissors, and then sieved in 60 m plankton net with filtered

sea water in order to get the oocytes or the spermatozoa in the case of males. Then, the oocytes are induced to mature within 2hr by adding the artificial maturation

hormone solution prior to adding the milt to fertilize. The fertilized eggs are kept in a small basin without aeration until hatching.

Fig. 8-1. Part 1 of induced oocyte maturation in sea cucumber H. scabra

1. Dissect

3. Cut gonad into pieces

4.Wash and filter5. Add hormone to oocytes

2. Remove gonad

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Fig. 8-2 Part two of induced oocyte maturation in sea cucumber H. scabra

Fig. 9 Monitoring of water temperature in the spawning tank

Induced Spawning

1) Temperature shock- water temperature is increased by 3-5oC for one hour, either by adding sun-warmed seawater into the spawning tank or by using aquarium heater (Fig. 8). Water temperature is kept within 28-32oC with constant stirring to maintain temperature uniformly within the tank. In some cases, water temperature is cooled down by 5oC below ambient for one hour and then raise the temperature again above ambient by 3-5oC. After treatment, replace water with fresh seawater at ambient temperature and wait until the animals spawn.

2) Drying and water

pressure application- animals are placed in the spawning tank without water for half an hour before subjecting them to powerful jets of seawater for several minutes. Seawater is replaced at ambient temperature.

3) Sperm-induced technique- one to two ripe males (previously identified) are dissected to obtain the gonads. Gonads are cut into small pieces and added to the spawning tank at ambient water temperature.

4) Spirulina bath- dried alga Spirulina is added at a concentration of 30 g per 300-500 L of seawater. Algamac 2000, a commercial preparation of spray-dried cells of Schizochytrium algae may also be used at 0.1 g/liter. Mix

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Fig. 10. Spawning behavior of male H. scabra

well by stirring the water and leave for one hour before replacing with fresh seawater at ambient temperature.

5) Treatment combinations may also be used:

a) Dry treatment + Cold shock treatment + Hot shock treatment

b) Hot Shock treatment + Spirulina bath

c) Dry treatment + Hot shock treatment + Spirulina bath

Pre-Spawning and Spawning Behavior Broodstock display signs or behavior indicating that spawning is imminent, such as: 1) rolling movements, 2) lifting the front-end of the body and swaying, 3) gliding or climbing the tank wall.

Sea cucumber H. scabra are broadcast spawners. Spawning usually occurs in the afternoon, or at night following collection of the broodstock. Spawning has been observed just prior to the full-moon and new-moon phases. Males usually spawn first, releasing a continuous stream of milt (or the hydrated suspension of spermatozoa) for several minutes or even hours. Female usually shows bulging of the front-end of the body followed by

intermittent egg release, sometimes at one hour after the first male releases milt. Fertilization takes place in the water.

Females can spawn 2-3 times over a period of one hour but may stop when disturbed. On the average, a 500-g female can spawn at a range of least 1 to 3 million eggs. During spawning, it is advisable to remove the males from the tank once they

Fig. 11. A: male gonads containing sperms and B: female gonads with mature oocytes.

started releasing sperm in order to prevent poly-spermy or a condition of excessive amount of sperm that can damage the eggs.

Egg Collection, Counting, Incubation and HatchingWhen spawning is complete, breeders are removed from the spawning tank. Eggs are siphoned into an 80m wet sieve and washed several times with UV-treated

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Fig. 12. Embryonic and larval development of sandfish H. scabra at water temperature of 25-29oC.

seawater to remove excess sperm and accumulated dirt during spawning. Newly-spawned eggs are white, spherical and visible to the naked eye. Sea cucumber eggs measure 80 to 200 m.

To estimate the number of eggs, place all eggs in a bucket with known volume, e.g. 10L. Stir the water gently to disperse the eggs in the water column. Take five 1-ml subsample, then count number of eggs in each subsample using the Sedgewick-Rafter chamber under a compound microscope. Get the average of the 5 subsamples and multiply with the total volume of the water in the container. Estimate also the fertilization rate by dividing the number of fertilized eggs by the total number of eggs counted.

Eggs are stocked in the hatching-cum-larval rearing tanks at a stocking density of 0.5-1 per ml for incubation, hatching and subsequent rearing the newly-hatched larvae. Hatching occurs at 36 hrs after fertilization at water temperature of 25-29 oC (Fig. 7).

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Fig. 13. Broodstock used in induced spawning are returned to holding tank with sand bottom to recuperate and remature

Table 1. Important water quality parameters during larval rearing of

H. scabra

Paramater Levels Remarks

Salinity, ppt 32- 36

Temperature. oC 26-30

Rearing water UV-filtered

Illumination, lux 400 Natural light during daytime, artificial light at nighttime

Dissolved oxygen, ppm 6-9 Use fine bubble-producing airstone diffuser

pH 0.07-0.4

Ammonia, ppm 0.07-0.4

Post-Spawning Maintenance

After spawning, animals are put back in sand-bottom tanks lined with flow-through seawater (Fig. 6). The animals are maintained at a stocking density of 20-30 animals per 1-ton tank in static aerated seawater. They can be fed artificial feeds consisting of prawn head waste, soya bean powder, rice bran and sea grass powder, at a feeding rate of 50 g per day. Broodstock held at lower density and fed artificial diets can re-mature and spawn more than once.

Larval RearingNewly-hatched auricularia larvae are maintained in the same tank where they are hatched. Auricularia is the stage when sea cucumber larvae commence feeding on exogenous food sources. Larvae are stocked at 1000/liter of water volume in 1000-L plastic buckets filled with 800L UV-filtered seawater (Fig. 7).

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Fig. 14. Larval rearing of sand fish H. scabra: batch-culture method in 100-L plastic buckets (A-B); larval rearing in flow-through tank (C-D), cotton-filter for copepods entering through water supply.

Food and FeedingThe use of mixed algae is best for sea cucumber larvae however, the best feed combination and appropriate feeding levels have to be looked into. Auricularia larvae can be fed a mixture of different algae such as Chaetoceros muelleri, C. calcitrans, Isochrysis galbana, Nannochloropsis sp., and the temperate alga Phaeodactylum tricornotum. P. tricornotum is cultured at low temperature of 19-24oC in the laboratory with artificial lights (Fig. 12). The sea water medium is 1-m mesh filtered and UV treated, and enriched with chemical fertilizers at the rate of 2 to 3 ml per l of water. Chaetoceros, Isochrysis and Nannochloropsis are all cultured in the laboratory or in outdoor tanks (water temperature: 29-30oC) and fertilized with Guillards F/2 medium (Trademark: Proline).

Feeding of auricularia larvae starts on Day-2 or two days after hatching (Table 2). Initially, algal density used is at 20,000 cells per ml and is maintained until Day-7. From Day-8 until Day-15 when the larvae transformed to mid-auricularia and then late auricularia, algal density is increased to 40,000 cells per ml. From auricularia, larvae transform into doliolaria which is non-feeding. It preceeds the stage when

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Fig. 15. Algal food culture under laboratory

condition

Fig. 16 Corrugated PVC plates

immersed in tank with

flow-through sea water to coat with benthic diatoms

the larvae settle in the tank bottom or on any substrates provided and begin their benthic life as pentactula larvae.

Preparation of Settlement Substrates

Fig. 16 Corrugated PVC plates immersed in tank with flow-through sea water to coat with benthic diatoms

Suitable settlement substrates are provided (similar to abalone hatcheries) such as corrugated PVC plastics, fiberglass plates, mesh screens and rough tiles. Prior to use, the plates are prepared by immersing in shaded tanks filled with seawater that were previously inoculated with various benthic diatom species such as Navicula sp. or Nitzschia sp (Fig. 13). In other hatcheries, the plate surfaces are painted with commercially available Spirulina powder made into paste for coating at 1-2 g powder per m2. It usually takes 4-5 days to coat plate surfaces with diatoms. Then

the Spirulina-coated plates are immersed in tanks with flow-through sea water in a partially shaded area.

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Days post-hatch

Larval stage Feeding Water management

0 (stocking) 32- 36 No No change

2-7 Early to mid auricularia 20,000 cells/ml*100% change every 2 days

8- 15 Mid to late auricularia 40,000 cells/ml* 100% change daily

18-20 Doliolaria Non-feeding stage same

22-29 ppm Pentactula Benthic diatoms & Sargas-sum extract Flow-thru

30-40 Juveniles Benthic diatoms & Sargas-sum extract Flow-thru

*mixed algae: Chaetoceros, Phaeodactylum, Nannochlo-ropsis, Isochrysis

Table 2. Feeding and water management schemes during larval rearing of sea cucumber Holothuria scabra

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Larval Settlement and Metamorphosis

Metamorphosis or the transformation to doliolaria stage begins two weeks after fertilization. However, most larvae can be observed swimming continuously searching for subtrates for settlement. Since doliolaria are phototactic (positively attracted to light), it is best to cover the tank to prevent aggregation of larvae on the water surface.

The early juveniles began to attach at the bottom of the rearing tank on the 3rd or 4th week after fertilization. They appeared as transparent (visible only by using light torch) and their size is estimated to be 0.5 mm. At this stage, they are fed daily with finely ground Sargassum particles (that passed thru 120 m filter) at an amount of 1 ml/100 L tank volume. Not all larvae undergo metamorphosis at the same time; many of them may stay at auricularia or doliolaria stages while others are transformed into juveniles. Hence, a mixture of phytoplankton and Sargassum particles should be added into the culture tank during this period (which lasts about 2 weeks).

Some Problems Encountered During Larval RearingCopepods infestation is one problem that can be encountered during larval rearing of sea cucumber. Copepods attack sea cucumber larvae either directly or by repeated collision causing bodily damage. There is difficulty in eliminating copepods with sea cucumber larvae as their sizes are similar. However the application of trichlorofon (Dipterex) has been recommended. Also, water filtration should be improved to minimize copepod and ciliate infestations.

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REFERENCES

- Agudo, N. 2006. Sandfish Hatchery Techniques. The Worldfish Center, Secretariat of the Pacific Community, and Australian Center for International Agricultural Research (ACIAR), 42pp.

- Al-Rashdi, K.M. & M.R. Claereboudt. 2010. Evidence of rapid overfishing of sea cucumbers in the Sultanate of Oman. SPC Beche-de-mer Infor. Bull., 30: 10-13.

- Al-Rashdi, K.M., S.S. Al-Busaidi & I.H. Al-Rassadi. 2007. Status of the sea cucumber fishery in the Sultanate of Oman. SPC Beche-de-mer Infor. Bull. 25: 17-21.

- Al-Rashdi, K.M., Claereboudt, M.R., and Al-Busaidi, S.S., 2007. Density and size distribution of the sea cucumber, Holothuria scabra (Jaeger, 1935), at six exploited sites in Mahout Bay, Sultanate of Oman. Agriculture and Marine Sciences Journal 12:34-49.

- Asha, P.S. & P. Muthia. 2002. Spawning and larval rearing of sea cucumber Holothuria (Theelothuria) spinifera Theel. SPC Beche-de-mer Infor. Bull., 16: 11-15.

- Conand, C. 2004. Present status of world sea cucumber resources and utilization: An international overview. P.13-23. In: Lovatelli, A. et al. (Eds.) Advances in sea cucumber aquaculture and management. FAO Fisheries Tech. Pap. No. 463., 425pp.

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- Maruyama, Y.K. 1980. Artificial induction of oocyte maturation and development in the sea cucumbers Holothuria leucospilota and Holothuria pardalis. Biol. Bull., 158: 339-348.

- Pitt, R. & N.D.Q. Duy. 2003. Breeding and culture of the sea cucumber Holothuria scabra in Vietnam. Aquaculture Asia, 8(1):36-39.

- Pitt, R. & N.D.Q. Duy. 2004. Breeding and rearing of the sea cucumber Holothuria scabra in Vietnam. In: Lovatelli, A. et al. (Eds.) Advances in sea cucumber aquaculture and management. FAO Fisheries Tech. Pap., 463:33-346.

- Leonet A., Rasolofonirina R., Wattiez R., Jangoux M., Eeckhaut I. (2009). Invertebrate Reproduction and Development 53, 13-21.

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