|30 March||Departure from Ghent to Rome|
|31 March||Meeting at FAO Fisheries Department with Messrs. Pillay, Mistakidis, De Wouytch, Kvaran, Henderson and Pedini|
|1 April||Departure from Rome|
|2 April||Arrival in Bangkok|
|3 April||Meeting at FAO Regional Office with Mrs. Delmendo|
Meeting at the Department of Fisheries with Messrs. Chertchai Amatyakul, Ariya Sidthimunka, Umpol Pongsuwana, Kasemsant Chaloyandeja and Renu Hutasingh
|4 April||Visit of private salt ponds on the way to Rayong (accompanied by Mr. Kasemsant Chaloyandeja and Ms. Renu Hutasingh)|
|Visit Rayong Fisheries Station (Komes Chareonpranis, Director and Mr. Uthai Patanoporin, Chief Brackishwater Aquaculture Division)|
|- Slide lecture and discussion with the local officers on “Optimization and economy on the use of Artemia in aquaculture”|
|5 April||Initiation of practical work with Artemia at Rayong Fisheries Station:|
|- demonstration of cyst decapsulation|
- layout for the construction of Artemia air-water-lift-raceway
- demonstration of ricebran preparation as food for Artemia
- layout for the construction of flow-through culturing system of Artemia using filtered pond water
|6 April||Visit Chachoengsao Macrobrachium Research Center (Mr. Somsuk Singholka, Director):|
|- slide lecture and discussion with local officers on “Optimization and economy on the use of Artemia in the aquaculture
- demonstration of cyst decapsulation
|7 April||Initiation of practical work with Artemia at the Chachoengsao Fisheries Station:|
|- decapsulation of cysts|
- construction and practical set-up for Artemia air-water-lift raceway
- preparation of ricebran as food for Artemia
|8 April||Visit Samut Sakhorn Fisheries Station and private salt ponds (accompanied by Messrs. U. Pongsuwana and K. Chaloyandeja)|
|- discussion with local officers on pond design and management for Artemia cyst production in salt ponds|
|9 April||Visit Songkhla Fisheries Station (main station + 3 substations) (accompanied by Dr. Pairoj Bhromanonda and Mr. Swasdi Wongsomnuk)|
|10 April||Lecture and discussion with local officers on the use of Artemia in the aquaculture hatcheries and on the planning for Artemia cyst production in salt ponds to be constructed at the Shrimp Farm of the Songkhla Fisheries Station|
|11 April||Return to Rayong Fisheries Station (accompanied by Mr. K. Chaloyandeja)|
|- inspection of Artemia work initiated since last visit:|
• raceway culturing of larvae (1 m3 raceway) with ricebran suspension
• flow-through set up for culturing of Artemia in pond water
- discussion of working plan for the future
|12 April||Return to Chachoengsao Macrobrachium Research Center (accompanied by Mr. Kasemsant Chaloyandeja and Ms. Renu Hutasingh)|
- inspection of Artemia work initiated since last visit:
• raceway culturing of larvae
• decapsulation of cst and hatching of the naupli
- discussion of working plan for the future
|14 April||Meeting at UNDP Regional Office with the Deputy Regional Representative and his Assistant, Dr. N.S. Subbaraman and Mr. A.R. Patten, respectively.|
|Meeting at the Department of Fisheries with Director-General Chertchai Amatyakul, Mr. Ariya Sidthimunka and Mr. Vanich Varikul|
|Slide lecture and discussion on “The use of Artemia in aquaculture and the practical possibilities for optimization in Thailand” at the Faculty of Fisheries, Kasetsart University, NIFI, Bangkhen|
|Meeting with FAO officers Mrs. M.N. Delmendo, Dr. L.B. Marcelo and Mr. D.D. Tapiador|
|Departure from Thailand|
|15 April||Arrival in Belgium|
|22 October||Arrival in Bangkok|
|23 October||Meeting at UNDP Regional Office with Messrs. Nixey, England, Ratana and Vos|
Discussion on working programme of Mr. Jan Vos, FAO Associate Expert
|24 October||Meeting at the Department of Fisheries with Mrssrs. Ariya Sidthimunka, Somsuk Singholka, Anand Tunsutapanich and Jan Vos. Visit Chachoengsao Macrobrachium Research Centre; discussion on Artemia programme|
|25 October||Visit Rayong Fisheries Station|
- discussion on working programme of Ms. Renu Hutasingh and Mr. Jan Vos
- Preparation of the layout for the pond and canal construction + working plan for Artemia inoculation and production in Samut Sakhorn with Ms. Renu Hutasingh, Mr. A. Tunsutapanich and Mr. Jan Vos
|26 October||Visit Chachoengsao Macrobrachium Research Centre; discussion on future Artemia working plan for Messrs. Vos and Tunsutapanich. Meeting with Dr. Arporna Sribhidadh, Deputy Minister of Agriculture and Cooperatives|
|27 October||Meeting at the UNDP Regional Office with Messrs. England and Vos Meeting at the Faculty of Fisheries, Kasetsart University, NIFI, with Messrs. P. Suraswadi, Suchit Bhimjoying and Somsuk Singholka|
|28 October||Visit Samut Sakhorn Fisheries Station; discussion of the layout for the construction of ponds and canals for Artemia inoculation and cyst production with Messrs. Poonsak Khawanukul, Anant Saraya, Sagha Wattanachi, Chairaj Phibalpenratana, Renu Hutasingh and J. Vos|
|29 October||Visit Chachoengsao Macrobrachium Center; discussion on practical aspects of the Artemia working plan with Messrs. Somsuk Singholka, Anand Tunsutapanich and Jan Vos|
|30 October||Departure for Manila|
|Writing of report and discussion with the Acting Project Manager, Dr. Herminio R. Rabanal, in Manila, Philippines|
|4 November||Visit of the Artemia ponds of Mr. Ceferino de los Santos, Jr. in Barotac Nuevo, Iloilo, Philippines|
|Discussion Leader||: Dr. Patrick Sorgeloos (Belgium)|
|Rapporteur||: Dr. Richard A. Neal (U.S.A.)|
Whereas there is a present and potential future shortage of good quality Artemia cysts, and considering the widespread need of Artemia cysts in aquaculture research and development in many countries of the world, the following recommendations are proposed:
Research on the biology and ecology of Artemia should be encouraged to provide needed background data for more efficient production of Artemia.
More efficient procedures for hatching and utilization of Artemia should be encouraged based upon available information.
Research on automated production systems for the mass production of Artemia cysts should be encouraged.
Investigation should be conducted to develop substitutes for Artemia.
A committee should be appointed by FAO to collect and disseminate statistics on:
projected future demand for Artemia cysts
This committee should send a questionnaire to likely countries to establish information on under-utilized sources of Artemia cysts.
In literature, the hatchability of a given sample of cysts mostly has been expressed as hatching percentage, namely the number of live nauplii hatching out of 100 cysts. This criterion, however, does not take into account the degree of purity of the product; in other words it does not consider the quantity of debris included in the batch of cysts. In this regard, the concept of hatching percentage is misleading since a figure of 90% hatching may indeed be correct, despite the fact that the product may carry a significant quantity of debris.
Inasmuch as Artemia cysts are always sold on a weight basis, the criterion of first importance to the customer is of course the number of live nauplii which he will get from the total quantity of product purchased.
The following simple techniques can be applied to determine the weight of product, which under standard conditions of incubation, will yield 1 million nauplii:
3 samples of 250 mg are taken at random from the batch of cysts to be analyzed
the cysts are hydrated in 100 ml graduated cylinders in 80 ml natural seawater at 30°C and are kept in suspension by a continuous aeration from the bottom of the tubes
after 1 hour, the water volumes are adjusted to 100 ml with seawater and 5 subsamples of 0.250 ml each are taken from each tube with a glass pipette or preferentially with an automatic micropipette
each subsample is pipetted in a small plastic tube (5 ml content), the pipet is cleaned with seawater in the same tube and the water level in the tube is finally brought to 4 ml with seawater
since at the end of the hatching period a total count will be performed on each tube, the volume of seawater in the tubes may vary in function of the type of tube used (minimum 4 ml)
the tubes are closed with a cap and clamped into a rotating axle at 5 rpm; the whole set up is incubated at 30°C in continuous light conditions
after 48 hours, the content of each tube is fixed by addition of a few drops of Lugol's solution1 (stains the nauplii dark) or with another fixative.
The total number of nauplii hatched in each tube is determined by filtering the suspension on a small gauze filter, placing the filter in a petri dish and counting the nauplii under a dissection microscope. The average number of nauplii produced per gram of cyst product is then calculated. This number can also be expressed more practically as the quantity of product that has to be incubated to produce 1 million nauplii.
250 mg product in 100 ml seawater
5 sub-samples of 0.250 ml giving n1 n2 n3 n4 n5 nauplii or average n nauplii in 0.250 ml
number of larvae per gram product: n × 4 × 100 × 4
weight of product needed for the production of 1 million nauplii
1 Preparation of Lugol's solution:
The reasons for applying the cyst decapsulation technique in the aquaculture hatcheries are discussed in detail in the SEAFDEC consultancy reports (see above) and in the following papers: Sorgeloos, P. et al. (1977b), Bruggeman, E. et al. (1977), and Sorgeloos and Bruggeman (1978).
In view of the local price difference between the two forms of hypochlorite, the decapsulation of Artemia cysts in Thailand should preferentially be done with technical bleaching power (e.g. calcium hypochlorite, for example, Nielon-60-P power containing 60% active product imported from Nissin Denka Co., Ltd., Japan. Local price: ± 38 Baht per kg).
Ratio cysts/active product: 2–3 g cysts for 1 g active product (take into account the percentage of inert ingredients when calculating the quantity of product needed).
Ratio cysts/sodium carbonate: 7 g technical Na2CO3 per 10 g cysts. For economy reasons, Mr. Anand Tunsutapanich (Chachoengsao Macrobrachium Research Center) is using technical calcium oxide instead of sodium carbonate at a rate of 3 g CaO per 10 g cysts. The use of CaO instead of Na2CO3 does not affect the efficiency of decapsulation, however, no information is available yet of its impact on the hatching efficiency.
Total volume of decapsulation solution (seawater + bleaching powder + sodium carbonate): 200 ml per g cysts. Preparation of the decapsulation solution - first dissolve the bleaching power (e.g. 10 minutes of aeration) then add the sodium carbonate. If a small particulate precipitate is formed, let it sediment (eventually overnight) and only use the supernatant.
Ratio desactivation solution/cysts: minimum of 0.5 ml of a 1% solution of technical sodium thiosulphate (Na2S2O3.5H2O) or sodium sulphite (Na2SO3) per 10 g cysts.
Hydrate the Artemia cysts in seawater or tapwater (the aeration should keep all the cysts in suspension).
After 1 hour filter off the cysts on a 150–200 micron screen and wash them with seawater or tapwater.
Resuspend the cysts in a bucket with the decapsulation solution which has been cooled to a temperature of 15–20°C (eventually add ice).
Stirr the suspension continuously with a stick in order to keep all the cysts in suspension. Check the temperature and eventually add ice to avoid a temperature increase above 40°C.
Depending on the temperature, it takes from 5–10 minutes before the decapsulation process is finished (orange-like colour of the suspension). Filter off the decapsulated cysts on a 150–200 micron screen and thoroughly wash them with seawater or tapwater until there is no more smell of chlorine.
Resuspend the decapsulated cysts in seawater or freshwater (1 kg of cysts in about 5–10 liters water). Add the desactivation solution and agitate the suspension during a few minutes.
After sedimentation of the decapsulated cysts to the bottom of the container, remove the floating materials (non-decapsulated cysts, transparent membranes, plumes, etc.). If a large portion of non-decapsulated cysts (which means that the products used do not have the exact activity or that the procedure has not been followed properly): harvest the floating non-decapsulated cysts and resuspend in saturated brine solution (300 g technical salt per liter), change the brine solution after 2 hours and again after 4–12 hours. These cysts can now be stored until a new batch of cysts will be decapsulated: filter off the brine and add the cysts to the new batch of cysts which will be decapsulated - see working procedure 1 and onwards.
The further treatement of the decapsulated cysts which settled out in the desactivation solution can be either one of the following methods:
direct distribution to the cultured species after washing out the sodium-thiosulphate or sulphite.
incubation for hatching:
upon completion of the hatching process, harvest of the nauplii on a 150–200 micron screen; thorough washing (removal of all dissolved organics, mainly glycerol); distribution to the cultured species.
dehydration in brine and storage until later use:
aerate the suspension for another 2 hours (eventually overnight) store in fresh brine until use; see above (a) and (b)
Do not expose the decapsulated cysts (hydrated or dehydrated) to direct sunlight (UV-rays kill the embryos).
Whereas for years the demand for Artemia cysts has exceeded by far the offer resulting in a continuous increase of the price of this product, and creating a serious bottleneck for the further expansion of the aquaculture hatcheries (Sorgeloos, 1976), this situation was reversed recently by the commercial exploitation of several new natural resources.
At the recent FAO/EIFAC Symposium on Finfish Nutrition and Feed Technology (Hamburg, Germany, June 20–23, 1978), it appeared however that there is still an important lack of information on the commercial sources of Artemia cysts.
We hope to alleviate this gap by the list given below. We would like to emphasize that the latter list has been worked out without any consideration on the potential quality of the respective cyst-products (Sorgeloos et al., 1978).
Informations gained from various sources indicate that the present commercial supply of cysts averages about 100 metric tons per year.
|Brazil||Macau strain (i.e. San Francisco Bay Artemia inoculated in the natural salt ponds of Macau - RN)|
|CIRNE - Rio de Janeiro|
|Av. Presidente Vargas|
|417 - A - 21° andar|
|Rio de Janeiro|
|Compagnie des Salins du Midi et de l'Est|
|51, rue d'Anjou|
|Paris - 8|
|Spain||Various strains in the Cadiz area|
|Edificio Casa del Mar, 5° planta|
|U.S.A.||Argentina strain (Buenos Aires)|
|P.O. Box 1303|
|Homestead, Florida 33030|
|Australian strain (Shark Bay)|
|270 Howard Avenue|
|New Haven, Connecticut 06519|
|Australian strain (Western region|
|Middle East strains (multiple sources)|
|P.O. Box 91272|
|Los Angeles, California 90009|
|Canadian strain (Chaplin Lake)|
|Pennsylvania Pet Products|
|P.O. Box 191|
|Spring City, Pennsylvania 19475|
|Great Salt Lake (Utah) strain|
|Sanders' Brine Shrimp Co.|
|1255 W. 4600 South|
|Ogden, Utah 84401|
|San Diego (California) strain|
|San Diego Brands|
|American Salt Company|
|San Diego Airport|
|San Diego, California|
|East San Francisco Bay (California) strain|
|D.C. Park and Associates|
|874 Carlston Avenue|
|Oakland, California 94610|
|North and South San Francisco Bay (California) strains|
|San Francisco Bay Brand, Inc.|
|8239 Enterprise Drive|
|Newark, California 94560|
Meeting with Mr. Somsuk Singholka (Director, Chachoengsao Macrobrachium Research Center), Mr. Sombhong Suwannatous and Mr. Anand Tunsutapanich (Officers, Chachoengsao Macrobrachium Research Center) and Mr. Jan Vos (FAO Associate Expert - Growing Food Organisms, THA/75/008)
|Date||:||26 October 1978|
|Subject||:||Working plan of the Artemia activities of Mr. Anand Tunsutapanich and Mr. Jan Vos|
|Results||:||After a thorough discussion of the detailed plan of operations with regard to the further Artemia activities of Mr. Anand Tunsutapanich and Mr. Jan Vos at the Chachoengsao Macrobrachium Research Center, it was worked out and agreed upon. This programme is outlined in detail in paragraph 2.3.5 of the present report. Furthermore, it was suggested for effective implementation of the Artemia work at the Chachoengsao Station that the following arrangements should be carried out:|
Mr. Jan Vos, as necessary, to provide with a full-time technical aid for better execution of his Artemia activities.
Mr. Anand Tunsutapanich and Mr. Jan Vos will alternately have weekend responsibility with regard to the supervision of the running Artemia activities. Their mutual arrangements will be communicated to the Station Director.
Short weekly meetings will be called by the Station Director with Mr. Anand Tunsutapanich and Mr. Jan Vos for reporting on their respective progress and problems with regard to Artemia matters in order to stimulate an efficient exchange of information.
1. Separation of the heavy debris (sand, skelets, etc.) in a saturated brine solution: poor the cysts in brine, either agitate (aerate) continuously or discontinuously in order to detach all clumps of cysts and have all dirt settled on the bottom of the container. There is no time limitation for this process.
2. Harvest the floating cysts from the surface of the brine solution; wash the cysts in freshwater or seawater (the cooler the better-cf. start cyst metabolism)
3. Pour the cysts in freshwater or seawater and let them settle at the bottom: the light debris (empty shells, plumes, etc.) will float at the surface (for some batches agitation of the surface layer might be needed to detach the cyst-clumps. This process should last not longer than 15 minutes.
4. Further processing can be done following various ways:
4c. Transfer the cysts to a cloth sack and remove the excess water by gently squeezing the sack and cyst-mass.
5c. 5c. Distribute the cysts on a drying surface consisting of a table or trays
made of wire screen covered with cotton muslin (tight woven grade; will
absorb water from the cysts).
In order to spread the cysts in thin layers (maximum a few mm thick)
use a woven wire basket (5 mm mesh).
By properly shaking this basket containing the cysts, the cysts will be sifted through the mesh and be evenly distributed over the drying surface.
6c. Drying should take place in a housing designed as a drying room. Temperatures as high as 40°C will not harm the cysts. Air exchange is required in order to maintain a minimal humidity (drying the cysts in direct sunlight might result in a decrease of the viability - due to heat-absorption the temperature inside the cyst shell might exceed 40°C which is lethal for hydrated cysts).
7c. Stop the drying process once the cysts are dry (the water content should be below 9%; preferably between 2 and 5%). Cool the drying room in order to eliminate or reduce the static charges that can be generated when hot dry cysts are brushed off the drying tables. Screen the cysts through a 300 to 500 micron screen.
8c. The viability of the dried cysts is guaranteed when packed under
vacuum or under nitrogen atmosphere (away from oxygen and otherwise
free radicals can be produced and these irreversibly damage the
The nitrogen packaging process has to be performed in 2 steps:
• flushing of the cysts with nitrogen; overnight storage under nitrogen (description of oxygen from the alveolar layer);
• flushing of the cysts with nitrogen followed by packaging under nitrogen atmosphere.
LITERATURES RELATED TO THIS SUBJECT
Benijts, F., G. Vandeputte, P. Sorgeloos, 1977 Energetic aspects of the metabolism of hydrated Artemia cysts: 79–87. In Fundamental and Applied Research on the Brine Shrimp, Artemia salina (L.) in Belgium, Special Publication No. 2, European Mariculture Society, Ed. Jaspers, E., Institute for Marine Scientific Research, Bredene, Belgium: 110p.
Clegg, J.S., 1976 Interrelationships between water and metabolism in Artemia cysts. II. Carbohydrates. Comp. Biochem. Physiol., 53A: 83–87
Clegg, J.S., J. Cavagnaro, 1976 Interrelationships between water and cellular metabolism in Artemia cysts. IV. Adenosine 5'-triphosphate and cyst hydration. J. Cell. Physiol., 88: 159–166
Crowe, J.H., 1971 Anhydrobiosis: an unsolved problem. Amer. Naturalist, 105: 563–574
Crowe, J.H., J.S. Clegg, 1973 Anhydrobiosis. Dowden, Hutchinson and Ross, Inc. Stroudsburg: 477p.
Hellfrich, P., 1973 The feasibility of brine shrimp production on Christmas Island. Sea Grant Technical Report UNIHI-SEA GRANT-TR-73-02: 173p.
Rakowicz, M., 1975 Notes on Artemia salina. Manuxcript: 13p.
Sorgeloos, P., 1976 The brine shrimp Artemia salina L.: a bottleneck in mariculture? FAO Technical Conference on Aquaculture, Kyoto, Japan, 26 May - 2 June 1976, Experience Paper 77: 12p.
Sorgeloos, P., 1977 Automatic washing and sieving of Artemia cysts. Manuscript: 3p.
Sorgeloos, P., 1977 et al., The use of brine shrimp, Artemia salina in aquaculture Proceedings of the 3rd Meeting of the ICES Working Group on Mariculture, Brest, France, May 10–13, 1977. Actes de Colloques du C.N.E.X.O., 4: 21–25