PROJET REGIONAL MEDITERRANEEN DE DEVELOPPEMENT DE L'AQUACULTURE MEDITERRANEAN REGIONAL AQUACULTURE PROJECT

TECHNICS USED FOR INTENSIVE REARING AND ALIMENTATION OF FISH AND SHELLFISH Villanova di Motta di Livenza - ITALIA - Vol. II COASTAL FISHCULTURE IN THE UPPER ADRIATIC Mr G. RAVAGNAN I. INTRODUCTION Coastal fishculture in the Upper Adriatic means especially "Venitian Valliculture" and before treating specifically this form of fishculture, I think it advisable to expose some fundamental considerations at first, which enhance this form among other aquaculture systems or indicate the reasons of its present technical orientations. I. 1. Rearing methods In general, the practice of fishculture is grouped into three fundamental methods: extensive, intensive and semi-intensive. These methods differ from one another by the different energetic derivation of their feed diet and, on this presumption, a rearing is counted as extensive when the ambiant environment provides the total feed requirements; intensive when, on the contrary, the feed requirements come entirely from outside; semi-intensive when, the environment along with feed from outside is employed to cover the feed requirements (RAVAGNAN, 1978). I.2. Classification of extensive rearings Extensive rearings cover the most varied technical aspects which are more or less perfected, depending on the socio-economical context where they are found. They can be briefly classified as following: – "Primordial" extensive rearings which differ from the simple water level fishing by the presence of facilities which are not very numerous, primitive and often precarious. – "Structured" extensive rearings fitted with the static and rational. facilities. – "Equipped" extensive rearings which are not only "structured" but are also furnished with the facilities and equipment which give them a good functional feature. I.3. Form of energy Every aquaculture system is always designed to transform a certain quantity of energy into a useful product for man and concerning the different production methods directly connected with aquaculture, three forms of energy employed can be remarked: – "Primary" energy is incident solar radiation; – "Subsidiary" energy is that part of the eco-system (as in rearing) obtained from the adjacent eco-system (tides, winds, temperatures, nutritive salts, etc... ); – "Auxiliary" energy is derived from circuits which are activated by man. 2. VALLICULTURE Vailiculture is a typical example of brackish water extensive fishculture, and therefore carried out in coastal, lagoonal or estuary zones. In the VENICE region, since the beginning of the 16th century, it has not been a primordial or approximative production activity but a well advanced and highly organized one. It originated from lagoonal fishing along with the capacity of observation by the operators of long ago. It derives its name from the latin word "vallim". which means defense, protection, enclosure, and which in our case means the man-made enclosures of water beds which are used for fishculture and they are marked as "valli" in the Venitian documents of the beginning of the 14th century (See BULLO, 1940). 3. THE "VALLE" 3.1. Structures and characteristics The "valle" is a portion of the aquatic eco-system, located on the coast and isolated by man for the practice of fishculture. Long ago "valli" were surrounded by reed, rush or net fencings. These were precarious installations incapable, in any case, of ensuring the independant management of the waters or a satisfactory biological control of the enclosures. Later on, the fencing structures employed were dykes or embankments, water tight structures ensuring complete isolation, to which devices, allowing communication with the outside, were attached. Man proceeded with his work, guided by nature at the beginning, in limiting himself to capturing the natural seed without altering the characteristics of the places, later on, by hindering the movement of the fish and finally, by inventing a satellite eco-system, capable of optimizing its own ecological efficiency, as well as obtaining energy supplies from the basic eco-system. Apart from the enclosure structure and controlled communication devices, a "valle" is equipped with a sea water and fresh water supply (the latter when possible) filter dams, crawls, (colaùri) fishing and selection tanks (lavorieri), Wintering tanks (peschiere), fertilizing systems, and different pumping devices. The surface area of a "valle" can range from tens to thousands of hectares. In the VENICE region, the average size reaches around 300 to 400 hectares, the minimum surface area is 10 to 20 Ha and maximum 1 600 Ha. In this region there is an overall surface area of 18 000 hectares divided up into 47 "valli", nearly all of them are of the "equipped" type, very few of "structured" type and none of "primordial " type. 3.1.1. Ambient conditions The type of soil generally found in the "valli" is of sandy or clayey nature. The depth of the rearing ponds is around 80 cm while for fishing or wintering ponds much greater depths can be found. The typical "valle" environment is that found on coasts or in estuaries; environments having sudden and great variations in salinity and temperature. The coastline where the Venitian "valli" lies has seven important rivers flowing into it, one of which, is the river PO; Nearly all of the "valli" have an indirect communication with the sea, via, more or less important lagoonal water beds which have a very inferior thermic capacity than that of the sea. Consequently, throughout the seasons or in coincidence with particular events, such as the swelling of rivers or sea storms, the salinity conditions are subject to variations of between 5 and 32 ‰ and the temperature is also influenced by the seasons, varying from a minimum 1 - 2° C in Winter, reaching a maximum of 30 - 32° C in Summer. With such conditions, it is very comprehensible that valliculture can only permit the rearing of euryaline and eurytherme species. Fish species generally reared in the "valli" and duration of the rearing season In the Venitian "valli" the following species are most commonly found: Dicentrarchus labrax European sea-bass Sparus aurata Gilthead sea-bream Liza aurata Golden grey mullet Crenimugil labrosus Thicklip grey mullet Mugil cephalus Flathead grey mullet Liza saliens Leaping grey mullet Liza ramada Thinlip grey mullet Anguilla anguilla Eel Atherina boyeri Atherine Along with these species mentioned here above other species can be reared in valliculture such as those given here following: Gobius ophiocephalus Snakehead goby Solea solea Sole Pleronectes flesus European flounder Only the Atherina boyeri and Gobius ophiocephalus reproduce in "valli" ponds while the other species mentioned here reproduce at sea. We have some rearing examples of fresh water species in "valli"; in particular salmo gairdneri and Acipenserides. P. labrax, S. auratus, Mugilides, A. boyeri, Gobius ophiocephalus , A. anguilla show good resistance even in the high temperatures (28 - 30° C) usually reached in the "valli" water in Summer. The minimum lethal temperatures for these species, range between 3 an 5° C (with the exception of A. anguilla which resists even at 0° C). In Winter, the temperatures in the "valli" ponds nearly always drop to 2 - 3° C and sometimes even lower. This explains the necessity of equipping for the wintering of the fish. The rearing operational temperatures range from 10 - 12° C to 28 - 30° C, optimal temperatures are 20 - 26° C. The rearing season starts in the middle of March and continues until the end of October: this covers a duration of about 7 months, with an optimal performance period of four months. Management The management of the "valle" even though entailing various operations, has three fundamental phases which are: seeding, fattening, harvesting or in other words the placing of the products into the rearing, their stay in the ponds, and their capture when they reach commercial size. Seeding Seeding can be carried out following the different methods, which are employed individually or all together. These methods consist in: a) to encourage the "rise" or in other words the migration (anadromous movement) of the young fish from the sea towards the trophic environments,such as those found in the "valli" and to hinder their "return" which is their migration backways (catadromous movement); b) to capture the fry at sea and place them into the rearing ponds; c) to activate the artificial reproduction process, employing more or less complex technics. The "rise" was the only seeding method employed in valliculture long ago; however, soon afterwards, the capture of fry in coastal zones was introduced and when means of transport and advanced techniques were available, they were caught in far off places. At the present time, many artificial reproduction facilities are employed, and it is considered that their popularization and the perfectioning of technologies are essential, not only for the development of sea fish capture but also for the survival of valliculture itself. Artificial or better controlled reproduction has been carried out for all Mugilidae for D. labrax, S. auratus, S. solea and others up to present. The only species where massive reproduction was carried out, was with D. labrax and S. auratus in commercial hatcheries. These species are the most valued, while being at the same time the most difficult to find. The quantity of fry placed into the "valle" every year varies according to surface area available, the trophic characteristics of each "valle" and the distribution of seed depending on the different species. The average amount per hectare/year can be estimated at around four to five thousand fry, 85 % of which is represented by five species of mugilidae and 15 % by S. auratus and D. labrax. The seed requirements for the Venitian valliculture can be roughly estimated at 80 million fry/year. The rate of seed distributed can vary, depending on the year or the species. This is explained by the fact that the marine resources can vary from year to year, something that the valliculture can neither schedule nor control. The survival rate of fry, when examined at commercial size, is around a minimum of 10 - 20 % and a maximum of 30 - 50 %, depending on the species reared and the rearing technics employed. "Valli" having great amounts of D. labrax present, see the production return drop. The seeding of eel was only started in the Venitian "valli" (apart from some exceptions), in the late fifties. The amount of elvers coming from the sea generally sufficed for the standard production of each modal of "valle" employed (sandy or clay), judged in an optimal manner of the "quantity-size" relation and varying on average from 30 to 60 Kg/Ha/year. From 1957 onwards, the Argulus giordanii, crustacean ectoparasites of the Branchiuri group, infested at first, the VENICE lagoon "valli" and afterwards all the "valli" in the region. These destroyed a great amount of the production and so upset all equilibrium which would have permitted a constant and rich production return. From this on, is spite of the persistant presence of Argulus, extra seed of elvers were added, especially "ragani" which are young eel having a body weight of between 15 and 30 gr. The results are not up to expectation and the productions dropped to a third or half of what they were. There are no major difficulties in finding eel seed, especially concerning elvers. Seeding of the "valli" is normally carried out during the months of March, April and May and sometimes in the Autumn months (October - November) for A. anguilla and M. cephalus. The fry known as "novellame" are placed at first into small ponds (a few hectares of surface area) and afterwards they are set free into big ponds known as the "laghi di valle". Fattening The weight increase of the population in the "valle" depends solely on the feed resources found in the rearing ponds, which is the biogenic capacity of the environment. By biogenic capacity is meant, the capacity of the environment to transform into a product of commercial interest a more or less important part of its own natural productivity. In general, the breeder does not intervene directly to increase the natural productivity, but restricts himself to ensuring that there is a good exchange of water. There is no need for a continuous water renewal, in the "valle" but an adequate quantity and punctual intervention is required.In other words, a flow of a sufficient quantity of water, at the appropriate time, in a given phase and circumstance. The hydrous management of the "valle" concerns the defence and enrichment interventions (the conservation and improvement of the ecological effectiveness of the ponds) and the coordination of the rearing operations (seeding, fattening, harvesting, etc... ) In can be calculated that three complete water renewals of the "valle" are required for one year; one at the beginning of Spring, one during the fattening season, and one at the end of Autumn or the beginning of Winter which corresponds to the harvesting phase. Harvesting If the seeding of the "valli" is obtained by the "rise" which is the tendancy that certain fish have of coming into the lagoons or "valli" from the sea, in the search of feed, the harvesting of fish is obtained when these populations return to the sea, urged by the instinct to reproduce or by the thermic conditions. With the exception of A.anguilla the whole population, although sexually and commercially immature , tend to swim back into the sea every year, as they flee above all the low Winter temperature of the ponds. A. anguilla on the contrary can resist to cold, and only heads towards the sea when its sexual maturity puts an end to its trophic period. Therefore, this fish stays in the "pasture" areas from when it enters into the "valle" at elver stage to the age of eight or twelve years, depending on whether it's male or female. The harvesting is performed by the double action of draining (which is a great decrease of the water level inside the "valle") and of "refilling" which is the introduction into the "valle of a hotter and nearly always more saltly flow of water. Therefore by exploiting the rheotactism of the fish as well as the difference of temperature and salinity, the movement of the fish towards the harvesting ponds is obtained. A well designed "valle" will have the possibility of directing all the fish towards one harvesting pond. A "valle" can have many ponds spread out or having a very particular structure. The harvesting pond is located directly behind the control structure for communication with the outside and it must be deep enough to 'enable a depth of 150 m of water when the ponds must be linked up with the fattening zones by a system of sub lagoonals canals, which have a slightly sloped bottom, so that at the harvesting period the fish are either pushed by the draining action and guided by the refelling action. On the inside and edges of the harvesting ponds are found many important structures. The principal ones, being the structure for the introduction and that for the capture. The former is located at the intersection point between the "pasture" ponds and the harvesting ponds, this is a dam, having many openings equipped with sluice gates and devices which enable the directing of the fish into the pond; the latter is a dam enabling the capture of the fish within the the pond itself and capable of stopping both their escape towards the sea or their return into the harvesting ponds, and also of separating eel and other fish species or selecting different size eel themselves. At a certain distance (on the inside) from the main capture structure (which is that placed on the sea-rearing "border") a second one is nearly always found for the capture of the product inside the harvesting pond when the current flows towards the sea and not towards the rearing ponds. On the edges of the ponds there are structures permitting the selection of the immature fish (from a commercial point of view) directing them towards the wintering ponds, and of mature fish ready for market. Wintering the fish In Venitian valliculture language "wintering" means, all the operations which are required to preserve the good health of the fish, by protecting it against frost and bad weather. It is not a typical phase of production but rather the defensive organization of the rearing during the Winter stasis. However, the importance that such a period has on Venitian valliculture (due to cold Winters), impels us to give great consideration to this. Wintering begins in November by the advancement or transfer of the fish into special ponds known under the name of "peschiere" and finishes at the end of March with the "opening" of the "peschiere" and the introduction of the fish into the "pasture" areas. The difficulty of Wintering varies from species to species. The A. anguillas is the most resistant and for which no wintering pond is scheduled, as during Winter, it resists by plunging down deep into the bottom of the pasture ponds. The other species having in order less resistance are as following: D labrax, M. saliens, M. cephalus, Liza ramada, Crenimugil labrosue, Liza aurata, S. auratus. This latter is put to Winter separately, into special ponds where it stays at low densities and where the temperature must be maintened at least at 6° C. During the Winter, the fish lose more or less a lot of weight, depending on the climatic conditions of the season. In the most unfavourable conditions there can be a loss of 10 % or more. The optimal temperature for wintering is that which corresponds to the metabolic equilibrium of the fish, based on the minimum requirement of energy. It varies according to the species in relation to the thermic level of the feeding stimulus. The wintering "peschiere", characteristic of valliculture, are composed of a series of canals of 3 to 6 meters in width and of variable lengths (from 30 to 100 meters), all connecting with one another (even if they are separable by means of sluice gates or grills) and positioned in such a way so as to give shelter to the fish from winds blowing from any direction. Between one canal and another, a more or less wide patch of elevated land is found (about 1 m above the water level) where the cultivation of evergreen plants is carried out and which serve as windbreak hedges. The sandy bottom "peschiere" having a fresh and salt water supply are the more reliable. The canals of a "peschiere" are all the more efficient when they are deep. A depth of 4 - 5 m or more, having a width of not more than about 6 m, ensures the protection against winds, an important thermic capacity and an abundant reserve of oxygen in frost. If the "peschiere" are located near rivers or sand hills which exert a pressure on them, there will be a slow filtration through the sand layers capable of maintaining a certain renewal and of fournishing heat to the environment. This filtration is a very efficient natural heating system. To winter a quantity of ten tons of fish peschiera ponds of 40 000 m2 in surface area must be available. A "valle" of 400 ha must have at least 300 000 m2 of "peschiere". Some years ago, the valliculturer realized that, what limited production, was the wintering capacity rather than the biogenic capacity of the "valle" environment, and devoted his technological research to making the traditional ponds more efficient and experimenting on less costly systems, which were more reliable, when put into use. Many "peschiere" had stand-by heating installations put in, the most valued products were wintered in cement tanks which had a heating system and were sheltered and finally good results were obtained with heated sub-marine tanks. Productive potentiality of the "valli" The productive capacities of the Venitian "valli" reach 150 Kg/ha/year, 50 % of which are Mugilidae, 25 % of D. labrax and S. auratus and 25 % of eel. Taking into account the climatic conditions in which the work is carried out, this level of production could be considered as quite satisfactory but it is certain that a much higher production could be obtained with Venitian valliculture. Rearing cycles Each species reared has its own individual growth index, in relation to which, the commercial size of the product will be defined. For example: S. auratus has a rearing cycle of about 18 months, it reaches commercial size from the 7 - 8 month and the optimal weight is 400 g at the end of the cycle; D. labrax requires a rearing cycle of 37 months to reach a weight of 400 - 500 g; M. cephalus and C labrosus reach their optimal weight of 800 - 1 000 g in about 53 months; L . aurata, saliens and ramada which have an inferior growth index, reach optimal weight of 300 g over a period of 21, 36 months; A. anguilla has a much longer rearing cycle, and it is only captured when it is silvery, in other words when it reaches sexual maturity. At this stage, it can vary in size, depending on it's sex and the climatic conditions found in the "valle". In general the Venitian "valli" produce eels of weights from 200 to 600 g. The present evolution of valliculture For more than ten years, Venitian valliculture, so as to increase its capacity of production, has for aim to overstep the limit imposed by the natural biogenic capacity (to which the productivity of extensive rearing is linked precisely) and directed research towards techniques which would be capable of obtaining good productions in small surface areas as well as increasing the environmental productivity. Many problems were encountered and can be regrouped as following: on one hand, those concerning the adoption of artificial reproduction, intensive and semi-intensiverearing techniques of certain fish species reared, and on the other hand those concerning the adoption of feed chains, on which extensive rearing is based together with operational interventions capable of potentiating them. From the experiments carried out on these two subjects originates the "integrate - valliculture" system, based on the penetration, from an energetic and functional point of view, between the intensive and the extensive.The structured "valli", when this system is employed, has some intensive rearing sectors and others extensive. The former produce independantly and serve as well as first stage for the extensive. From this relation originates a production method of semi-intensive type, where estensive ponds make good use of their natural biogenic capacity and benefit from the energetic supply from the intensive sectors which are as following , semi-reared fishculture material, water current and rearing catabolites.

2) Plan of the energy flow in an integrated valliculture system. It can be noticed that the intensive sectors are supplied with auxiliary energy and artificial feed. On the contrary, the extensive ponds have a supply of primary energy and natural feed. The subsidiary energy could help especially the extensive and building ponds by having fry caught at sea placed into them. By integrating the system a lot of energy passes from the intensive to the extensive in the form of; Hydric flow, fish and organic matter. At the bottom can be seen the productive outlets, some directly coming from the intensives others from the extensives. Intensive rearing All fish species reared in integrated valliculture (with the exception of Atherina boyeri and sometimes part of Anguilla anguilla), start their cycle in the intensive rearing sectors. Some stay there until they reach commercial size (D. labrax and A. anguilla) on the contrary to others (S. auratus, Mugilidae and part of A. anguilla, that stay for more or less long periods, as to complete their cycle, pass into the extensive sectors. The structure of the intensive sectors vary from species to species, as does the rearing density which ranges from a maximum of 25 - 30 Kg/mé 2 for D. labrax to a minimum of 0,5 - 1 Kg/mé for Mugilidae. Extensive rearing The extensive ponds, receiving their fish from the intensive, along with the organic waste matter supply, must have many culture operations carried out principally concerning the good management of the water (by means of pumps) and the draging of the soil bottoms (carried out with the appropriate devices). In this way, valliculture greatly reduces the duration of the rearing cycles and increases, by 300 Kg and more/Ha/year, its production potential of the extensive ponds. Production potential of integrated valliculture An integrated "valliculture unit has the aim of obtaining the highest possible productions with a minimum input of energy coming from outside, therefore it aims at developing to a maximum the natural productivity. The dimensions of the intensive rearing sectors are calculated, not only in relation to the commercial criterions but principally with respect to the energetic - productive benefit that the extensive sectors can take from their integration with the intensive sectors. The production potentiality of integrated valliculture, on an equilibrium basis, therefore combines that of the intensive sectors, reaching in this way remarkably high levels. Experimental facilities located in VENEZIA have shown that it is possible to obtain productions of around 1 000 Kg/Ha/year, 60 - 70 % of which coming from the intensive sectors and 30 - 40 % from the extensives. Experience has shown the benefit of multiplying the production lines, by operating in the extensive ponds rational polyculture, so that the available trophic structure may be used to the full. Consequently, great importance must be given to the choice of species to be reared, to the size relation between the different fish populations, to the period and method concerning their introduction into the environment, to the succession of types of culture and operational interventions linked with increasing the production speeds. It is rightly affirmed that the lower the trophic level to which an organism is introduced the higher the energetic return of the production process will be However it must be remarked that certain fish or shellfish species, even when placed at a quite higher trophic level (3° or 4°) feed off small benthonic or planktonic organisms which are not directly of interest in the humain diet if not introduced into a certain level of the feed chain. In other words, energy is spared, avoiding, this way a decrease in the production speed.

3.) Concerning this plan, the fundamental trophic circuits and the principal productive "outlets" of integrated valliculture system can be remarked Here also can be noticed how the intensive rearing sectors give a supply of energy into the natural circuits, in the form of organic matter.

4.) The waste matter circuits in an integrated valliculture system are underlined here. The plan represents a complete system from artificial reproduction stage to the finished product. The waste matter is an integral part of the productive system. A well managed complex should and could give out through the outlet less nutrients than it receives from the outer ecosystem flow. The practice possibilities of the Venitian "valliculture" model. There where traditional valliculture is found or can be carried out (in other words extensive fishculture), integrated valliculture can also take place. This is perhaps applicable for all the Mediterranean region, and can be applied for the lagoons, estuaries, ponds, marshes or coastal zones which are liable to inundation and give satisfactory results when the climatic conditions are suitable. Principal problems linked with development programmes To conclude, let us point out the principal difficulties linked with valliculture development and therefore sea and brackish water fishculture. Although not being pessimistic, we believe that a serious programme must be based on a realistic view of the real possibilities together with the difficulties which are none the less important. Let us not take into consideration those problems arising with the training of personnel, the commercialization of the products, the financing of the different initiatives, etc... , not because we consider them of less importance but we think that the first to be considered must be the following: a) fry collection. b) ecological defence c) allotment of regions a) Fry collection To seed with marine fish species, fishing at sea is still employed (apart from some species to some degree) which favours the rearing in zones which have an abundant production of fry (Meridional Mediterranean zones) and encourages the breeders working in less favourable zones (Northern Mediterranean zones) to fish new material further away and to transport them to their facilities. As salt and brackish water fishculture only employs marine reproduction species, the continuation and development of this activity are linked inevitably to the corresponding availability of fry. This is the reason why it is necessary to apply suited capture, transport and rearing technologies, to ensure the best possible use of the actual resources (which already offers the possibility of tripling on average the survival rates and therefore the productive potential) and to supply, at industrial level, artificial reproduction techniques. One wonders whether the capture of fry at sea has not the effect of weakening the general productive resources. Normally, the reply can be but reasuring if the poor survival rates in marine environments are taken into consideration. In practice, the more fry transported from the sea into rearings the more satisfactory the results, from a production point of view. The questions arising, when this method is employed are, in our opinion, as following: a) How many fry can the sea furnish ? b) What can be done to maintain (and if possible increase) this valuable potential ? It is difficult to give an answer to the first, without carrying out minute investigations and covering a very vast geographical zone. For the present, we can not have recourse to the very approximative estimations which can however deduce that the present commercial production of Mediterranean rearing facilities come from a few hundred million of fry and that the quantity of available fry could be doubled by developing for this purpose the lagoonal and estuary zones which are neither used for fishing nor rearing. In answer to the second question, it must be remarked that the capture of nearly all the fry born at sea would perhaps bring about eventually a decrease in the number of "free" breed stock (a greater decrease as the rearings become more sophisticated) and eventually a scarcity of fry. It appears that this aspect should be carefully examined, in taking into consideration that it could prove perhaps helpful, not to stop altogether the capture of fry, but to give back a certain number of mature breeders into the sea. However, the fact that only artificial reproduction can ensure the development of coastal fishculture must be kept in mind; This alone indeed permits the control of the reproduction cycle without which fishculture could never be an independant and complete bioculture. b) Ecological defence and allotment of regions The ecological defence of the region (in this case the coasts, the sea, water beds, lagoons, estuaries, rivers, etc... ) is essential not only for the promotion of new fishculture enterprises but also to ensure the very survival of the enterprises already existing. There are still numerous and vast zones, in good ecological condition, available, but it can not be denied that many others are no longer in good condition and that others are more or less in danger. Normally, fishculture is not incompatible with other activities; on the contrary, there are some of industrial type, with which it can fully harmonize; it can also benefit from their presence sometimes when it uses warm affluents. However, it would be absurd to believe that there would be neither damage nor conflict if it is not decided that in certain zones to make fishculture the more privileged activity in relation to all others. INTENSIVE REARING OF SEA-BASS (Dicentrarchus labrax) AND GILTHEAD SEA-BREAM (Sparus aurata) IN RACEWAYS BIOLOGICAL, TECHNOLOGICAL ASPECTS OF FATTENING Mr. H. HELLIN I. GENERALITIES 1.1. Warm waters - Cold waters The growth of Mediterranean marine species by which we are concerned depends greatly on the water temperature in the rearing and is divided into three categories: – Optimal at temperatures around 25° C – Reduced at temperatures below 18° C – blocked at temperatures below 12° C Risks of mortality appear at below 10° C while at 30° C pathological and oxygen consumption problems are remarked. The great influence that the temperature has on the rearing, leads to two types of farm. – The following farms with heated waters: The sea water supplying these farms must be maintained at a temperature of 18 - 20° C the whole year. The continuous optimal growth of the fish is obtained this way and the fish reach commercial size in 18 months. – The fattening farms with sea-water at natural temperature: This solution is only suitable in sites where the temperature of the water never drops below 14° C in Winter time. It is characterized by two growth interruptions in Winter per rearing cycle and the fish reach commercial size after 2 to 3 years. 1.2. The three phases in rearing 1.2.1. The nursery Fattening starts when the weaned fry are transferred from the hatchery. They then undergo a lot of handling before being placed into a special unit, known as the nursery. This unit contains small volume tanks (25 to 50 m3 ) characterized by: – a very good accessibility (better control of the rearing, reduction of manual labour charges) – -the possibility of programming removable partitions while not interfering with the hydraulic system of the tanks. The fry remain in this part for 6 months, until they reach juvenile stage with an average weight of around 20 - 25 g (in a rearing which has controlled water temperatures). 1.2.2. First fattening - fattening When taken out of the nursery the juveniles have reached a sufficient size to be transferred into tanks of more than 60 to 100 m3 in volume. If all the rearing tanks have a unitary volume of more or less 100 m3, no distinction is made between first fattening and fattening. If this is not so, the average size tanks are left for first fattening while the others (100 - 300 m3) are kept for fattening. Juveniles having received first fattening reach around 70 g. before being placed into fattening. The fish leave the farm at a optimal weight for commercialization between 300 and 500 g. 1.3. The different types of tanks I.3.1. Design The tanks form an enclosed space for the fish and their rearing environment: a) Hydraulic characteristics It must be always possible to drain the tanks completely in a short time (half an hour to one hour). They must be designed in such a way so that they can create internal currents leaving no dead zones and maintaining a homogenous rearing environment. Their outlets must be equipped with fish trap systems which do not clog up easily. They must also be located at S.M. concentration zone levels. The water level must be adjustable and constant no matter what the water renewal supply flow may be. The design of the tanks must permit easy access and the use of all the equipment necessary to carry out the rearing. The suitable depth is below 1,20 m so to permit an easy and efficient supervision of the tanks. b) Accessibility The outside circumference around the tanks must permit, easy access to all the important parts, so as to permit the performance of all the rearing operations properly and the ensurance of efficient supervision. The partitions must be smooth and non-porous allowing sterilization and quick cleaning. The partitions are generally rigid panels (concrete - masonry -polyester). 1.3.2. Raceways tanks These tanks are characterized by a length/width ratio of more than 5. They have an upstream water supply and a downstream outlet flow, this permits maintaining a quick continuous longitudinal water current supply on the slant. These tanks make use of the water supply very efficiently and can be cleaned out automatically, the S.M. is transported with the current). They also permit to have high ratios of "water surface/farm land". They are very well suited to adjoining tanks which have reoxygenation between each. 1.3.3. FOSTER-LUCAS tanks These tanks don't differ from the raceways apart from the fact that they have a longitudinal partition and rounded ends. This solution allows the optimal use of the surface while having very rapid interior currents as seen with circular tanks. These currents are produced by the arrival of the renewal water and the air lift systems. These tanks are automatically cleaned out and are characterized by a more homogenous rearing environment than that of raceways 1.3.4. Other types of tanks The circular or earth tanks are not often employed in fattening for work reasons: – Complexity of the rearing operations – Bad use of the surface available On the contrary to the above, it is very advantageous to carry out the fattening phase in floating cages located in the sea or in lagoons when the conditions of the site at right permit so. 2. REARING 2.1. Rearing programme - Stock management 2.1.1. Nursery The weaned fry of 1 to 5 g on arrival at the hatchery farms, are too delicat and are at a too rapid growth stage to be placed directly into the final fattening structures. They are therefore transferred into a special unit, known as the nursery for the reasons here following: a) Frequence of manipulations The nursery must allow quick and easy fishing and grading operations while not causing any stress. These operations are indeed often necessary so to avoid the problem of cannibalism. The great growth speed of the fry causes remarkable difference in size quite quickly. As the sea-bass attacks fry of the same size or inferior to his own size, frequent grading is necessary. This reduces the mortality rate while maintaining a more regular distribution of the population. The manipulations are therefore carried out every 2 to 4 weeks. b) Extreme sensitiveness to the quality and regularity of the physico-chemical parameters of the rearing environment. The principal parameter to have under control is the temperature. It must be as stable as possible (variations ≤ 2 to 4° C per day) and kept at around 25° C. For this, it is advisable to have two water supplies, permitting to increase the temperature of each tank when necessary. The other parameters to be checked carefully are the oxygen ammonia tenor, pH and turbidity. During their stay in the nursery the fry are observed very carefully. They are stocked at optimal loads of around 10 kg/m3 . This phase lasts around 5 months (average temperature between 20 and 25° C) and juveniles of 20 to 25 g. on average are produced. 2.1.2. First fattening - Fattening When the fry are taken out of the nursery they are transferred into enclosures of greater volume. If the optimal loads of 12 kg/m3 were maintained there would be overcrowding and this should entail: – Management problems of the loads (heterogeneity) – more rearing risks should be incurred (fishing, accidents) – manipulations should be more complicated (stress) After a duration of 5 (in heated water) to 10 months (after Wintering) pre-fattened fry of a unitary average weight of 70 g. are obtained and these are then transferred into the final fattening tanks. During this phase the manipulations are reduced to every 4 to 8 weeks because of the decrease in the speed of growth and the loads being more homogenous. Loads at the end of fattening: 15 kg/m3 Average weight: 300 to 500 g. 2.2. Water management The determination of the water supply level depends on the principal parameters here following: 2.2.1. Oxygen consumption in the tanks Taking into account the loads admitted in intensive fattening (15 kg/m3) the oxygen consumption in the rearing tanks can be very high. This depends principally on: – The age of the fish. The younger they are the more oxygen they consume per kg of live weight. – The stress activity rate – The feed rate – The temperature: the higher the temperature the greater the metabolic activity and therefore an increase in the oxygen consumption is remarked. The oxygen supplied through the water renewal must be equal to the difference between the consumption in the tank and the supply by the aerators, this itself depending on: – The temperature and the salinity. The two factors are inversably proportional to the oxygen tenor of the water at saturation. – The supply flow There must be a sufficient supply of oxygen so as to maintain a minimum rate of 4 mg/l of oxygen at the outlet of the tank. 2.2.2. The gassy ammonia rate When most of the oxygen supply is ensured by the aerator systems, the factor determining the minimal renewal rate of water can be the gassy ammonia rate. This rate itself depends on: – The metabolic activity rate of the population. The more intensive it is (stress, feeding period) the greater the production of ammonia. – Tank loads - Feed rate – Other physico-chemical factors Ammonia is discharged in three forms in the rearing environment: – Solid nitrogenous products – Ionized ammonia (ions NH4 +): Ammonium ion – Non-ionized ammonia (NH3 ): dissolved gas Only the third form is highly toxic for fish. The toxicity threshold can however vary depending on the age of the fish (proportional) or the temperature and the level of stress (inversably proportional). Ammonia in rearing environments tends to evolute from the first form to the two other forms (solid solubility) and the report between the two other forms is balanced at a variable level by the following transformation:

The tendency of this balance varies depending on the temperature and the pH (Fig. below)

Effects of pH and temperature on the distribution of ammonia and ammoniumion in water. Data from LIAO et al. (1972) 2.2.3. Other physico-chemical parameters 2.2.3.1. Temperature: Rearing technique employed in heated water: We have observed in 1.1. that the temperature is an essential parameter for the management and competitivity of a rearing. In the case of rearing with controlled temperature, a minimum supply flow must be maintained in such a way so that the thermic loss can be counterbalanced in the tanks. The rearing water supplied to the farms may be heated by means of: a. Industrial thermic systems: With this option which is wide spead, the water flows more or less directly from the coolong circuits of industrial plants (petrochemical, power stations). This solution which is on general rather complex leads to heavy investments. THe complexity is caused by the quantitative and qualitative irregularity of the water (temperature, physico-chemical characteristics) . This water may be employed directly when it has been mixed with "natural" sea- water or when treated., or as warm water in the heat exchangers if the physico-chemical characteristics are not suitable for use in the rearing. b. Geothermal drilling: This solution, limited to very precise geographical areas, is very advantageous when carried out in shallow water with high temperatures (above 25° C) as the drilling in this case is not very expensive and there is a constant hot water supply. The water is obtained from artesian wells or by means of pumps and it is employed as a hot fluid in the fresh water-sea water heat exchangers. This solution usually leads to an increase of investment concerning: The limited choice for the site which could lead to overexpenditure for land or the development of sites which are not very suitable (too narrow, too elevated unsuitable edaphic characteristics}. Hydraulic systems are more complex (double or triple circuits) involving treatments, recycling, pumping stations and the facilities must be covered so as to limit the loss of heat in regions which have bad climates. 2.2.3.2. Suspended matter tenor A too high tenor of S.M. is harmful as it can cause asphixia (clogging of the gills) and feeding problems (decrease in the viability) 2.2.3.3. pH. This is a very important parameter, as it conditions the balance between toxic ammonia and non toxic ammonia. It is advisable to maintain the pH above a level of 7,5 and below a level of 8,5. In the rearing tanks, the pH varies inversably to the tank load. If the pH should vary, it will be necessary to raise the renewal rates. 2.2.4. Hydraulic aspects The renewal rates in the tanks which are calculated depending on the loads in the tanks and the limiting physico-chemical parameters (NH4 - 02 S.M. tenor). To define the limit design of the hydraulic structures it is necessary to take into account: – The average and maximum hourly renewal rates (maximum needs) – The minimum time - for filling and draining the tanks – The balance coefficient of the needs for all the tanks. Finally, the design of hydraulic systems in rearing farms must taken into account, so as to enable the rational management of the water: – Possibility to increase or decrease the renewal flows to maximum or minimum without causing any perturbation. – An isolation as complete as possible between the systems of the different rearing units so as to limit the contamination risks and to adapt the water qualities to each rearing phase. 2.2.5. Usual values The average value of the renewal rates practised with intensive rearing is as following: – 1 to 15 renewals per hour – Drainage of the tank within 15 to 30 minutes – Filling of the tank in 30 minutes 2.3. Feeding Feeding is the most important element in the running of a fattening unit. 2.3.1. Generalities The principal points to be taken into account for the management of feed in a rearing unit are as following: 2.3.1.1. Calculation of the food rations The food rations distributed daily must be calculated very precisely, for each category to ensure optimal growth, while applying managerial plans for the rearing. These calculations are carried out while taking into account: – the size of the fish – the type of food – the t° of the water – the control and follow up of the rearing. The daily food ration varies on general between 1 and 3 % of the total biomass for sea-bass and gilthead sea-bream in fattening. To optimalize the distribution of this daily ration, 4 or 5 feeds will be programmed during the day, divided out evenly from sunrise to sunset. The distribution of the feeds is one of the principal jobs of the manual labourers. Therefore it is advisable to employ automatic distributers although they may demand heavy investments. 2.3.1.2. The food formulation balance - Appetency Special attention must be given to the food formulations and the stocking of this food so as to obtain a perfectly "balanced diet. A food unbalance, even slight, can have heavy consequences, deteriorating the sanitary state of the population, causing risks of epidemic and a transformation coefficient increase of the food. On parallel, the appetency of the food must be checked, and this principally when dry feed is distributed. Unappetizing food leads to a decrease in consumption, losses and the rapid pollution of the tanks. 2.3.2. Dry and wet food Presently two principal types of food can be distinguished: 2.3.2.1. Dry feed: At the present time this is the most renown formula employed for the fattening of sea-bass and gilt-head sea-bream. The food is regularly distributed in the form of dry pellets which is supplied by food manufacturers. This solution, although the simpliest and the more easily mechanized one has two major disadvantages: – high costing (average transformation coefficient, high cost per kilo of feed). – Still average adaptation of the food distributed to the needs of the fish. (appetency problems, deficiencies, osmotic exchange. – It has nevertheless the advantage of lowering the manual labour (distribution) and stocking costs. 2.3.2.2. Wet food Wet food is still not widely employed and it is badly controlled in the rearing of sea-bass and gilthead sea-bream. It is however quite advantageous as it brings a remedy to the disadvantages of the dry food described here above. It is manufactured in wet food pellet form in the farm from minced waste fish combined with "premix" type of meal which is supplied by the food manufacturers. The development of this food is blocked by: – problems of technological order in the mechanization of its manifacture and its distribution. – the definition of formulations depending on the components available and their biochemical composition. – higher investment costs. 3. THE FARM IN GRAVELINES 3.1. Introduction The first rearing unit in GRAVELINES by which this report is concerned, was the first industrial marine aquaculture unit to be set up in France and the first aquaculture centre unit. Different fattening techniques have been developed over these past years on the Mediterranean coasts and many difficulties have been encountered, the principal one being of economic order. The length of time that the rearing cycle demands, which for sea-bass reared in natural conditions is 30 months seriously limits the profitability of the operations. The valorization of the thermic effluents was the principal reason for the implementation of this farm on this site, located in the immediate vicinity of the power plant and the SEPIA International, this was sanctioned following the international meeting organized in 1982 by SERAG (Syndicat for the study of an aquaculture network system in GRAVELINES) to define the aquaculture design of this farm. The principal aim of this implementation was to demonstrate,on one hand, the fiability of the rearing techniques and on the other hand, the profitability of these marine fish rearings. 3.2. Design of the pilot farm The passage of the water in the cooling circuits of the power plant raises the temperature of the sea-water by 13° C which during the year evolues between 15° C and 29/30° C. So as to ensure these temperatures in the rearing environments, even when the existing temperature is much lower, it has been decided to cover the entire farm (3 000 m2) with agricultural green houses. This cover prevents evaporation caused by wind, cooling caused by the difference of the temperature at the water/air interface, the decrease in temperature due to snowfalls or even the decrease in salinity caused by heavy rainfall. The farm has also been designed to limit exterior agressions (pollution from the inner port in DUNKERQUE, dredgings, chlorination in the cooling circuits of the power plants). A recycling circuit permits the isolation of the farm from the outside for about 8 days in "protected" rearing conditions. The circuit comprises of: For the nursery: – a special discharge circuit – a decantation unit – a pump cover and a pumping station – a purification unit (biofiltration of the waste water) For the fattening: This system is simpler as it doesn't need a real depuration unit. The temperature of the rearing environment can be optimized by mixing a heated water supply (450 l/s) and a sea water supply at natural temperature. Thus the highest possible temperatures can be maintained in Winter (using only heated water) while in Summer, the highest temperature in the rearing environment is limited to 25 - 26° C which seems ideal for sea-bass and gilthead sea-bream. The water management complex allows this manipulation. It must be remarked that the possibility of two independant circuits with different temperature exists: – one for the nursery – the other for the fattening units The nursery and fattening tanks have been designed while taking into account the following points: – Working facilities (flow regulation - observation - feeding the fish - drainage) – Automatic cleaning. The outlet is placed at the bottom of the tank near the end and the water level is regulated by an overflow. The water discharged is channeled at a normal flow rate through the evacuation pipe or flushed through this channel when the tank is being cleaned out. 3.3. Description of the pilot farm 3.3.1. The water circuits The farm has at disposal three types of water supply: – heated water – "natural" sea water – recycled water These different sources of water are mixed proportionally depending on what has been decided at water management level, so as to ensure the best conditions possible to the fish, from a temperature viewpoint as well as a physico-chemical one The water supply overflows are evacuated by means of drain sluices. Heated sea-water This water is pumped downstream from 3 structures of the EDF power plant at a maximum flow of 400 l/s. Its temperature varies between 15 and 32° C. This heated water is used entirely as soon as its temperature reaches 25° C. In Summer it is mixed with cold sea-water. Cold sea-water This water is pumped up stream from the EDF power plant into a sea-water supply canal which joins up with the inner port of DUNKERQUE. It permits the temperature of the rearing water to be lowered during Summer. Recycled sea-water This recycling permits, the recovery of water which has already been used in the rearing, making it compatible once again by means of successive treatments, for the rearing of fish and then it is distributed throughout all the tanks. This method not only permits a complete isolation of the structure from the outside environment (power plant stoppage, pollution), but it also ensures a greater overall flow in the tanks when necessary (temporary overflow of the structure). The quality of the water is verified daily by means of physico-chemical and the steady flow, in the tanks is optimized according to the fish load contained in the latter. 3.3.2. The rearing units They are two in number: the nursery and fattening. The nursery It has FOSTER LUCAS type of tanks of around 35 m3 each in volume. Each tank has a water supply by means of either two circuits (the model employed can be regulated from 5 to 30 l/s). – the nursery circuit – the fattening circuit A low-pressure aerator, type air lift, permits to reinforce the speed of the current and to increase the supply of dissolved oxygen in the rearing environment. An outlet located at the bottom, equipped with an anti-vortex system, permits the elimination of wastes (faeces - uneaten food) deposited on the bottom of the tank. Automatic pressure feeders completes the equipment of the tanks. The fattening unit It has two series of 14 adjoining tanks, having a unitary volume of around 65 m3 THese tanks are identical, apart from their size to those in the nursery. All the rearing units get their water supply by means of reinforced concrete channels which are placed crosswise in the tanks and are located on the lateral walls of the latter. A series of platforms situated on top of the walls of the tanks permit the usual work (feeding, observation, flow regulation, treatments, etc... ). 3.3.3. Annex equipment Along with the rearing unit, the farm has the following facilities: – a wet food manufacture workshop with a cold room. – a technical room which is divided into two parts:

• a boiler room

• a room for the TGBT – a generating plant – componants (decanters - pumping station - biofilters of the recycling circuits 3.4. The running of the farm - First results 3.4.1. General characteristics The general characteristics of this rearing are as following: – The annual production when in full operation is 65 to 70 t. (around 35 t. in 1985). – The production staff comprises 4 people: 2 technicians and 2 workers one of which a fisherman. – The number of fry placed into rearing each year is around 300 000 divided up between sea-bass and gilthead sea-bream. – The global surface area of the tanks is around 2 200 m2 for a total surface area of 3 200 m2 (60 % ratio). – The warm water flow is 1 600 m3 /hour, the cool water flow is 800 m3/h. 3.4.3. First results The management of this farm was placed in the charge of the Aquanord Maritime Cooperative which regroups specialists in aquaculture, professional fishermen, and experts on the commercialization of sea-products. After a first trial carried out in the provisonary facilities in 1983 and whose objective was the appreciation of certain characteristics of the site, the production was started in June of 1984 with the introduction of 200 000 fry: 170 000 sea-bass fry 20 000 gilthead sea-bream fry 10 000 sole fry (trial) The growth rate remarked was on the whole comparable to the initial previsions. The survival rate however did not come up to expectations, this is quite normal when we realize that this operation began before the works had been completed. After a year's work, the first sales took place, which confirmed on one hand the technical hypothesis and on the other hand the heavy commercial demand and the high sale prices. The quality of the fish was unanimously appreciated by french and foreign experts. Since the first trials, demands have increased and the demands exceed our production possibilities (5 to 6 ton per week). 3.5. Future production This pilot farm is the first element of the aquaculture site in GRAVE-LINES. A hatchery will be built in the coming months which will give autonomy to the site and a rapid possibility of expansion. This "aquaculture zone" created as a traditional development zone will regroup finally: – around 22 production farms (22 "lot")

– common circuits:•.supply - heated water - cool water

• evacuation

• VRD – Annex service units

• Hatcheries

• Service cooperatives (food sales)..

• Research and training units

• Veterinary unit If this realization is a success, France will pass from experimental and small scale production stage which it is at present doing, to large scale production stage of marine fish, GRAVELINES will finally permit more than 1 200 t/year. 1. GENERALITIES 1.1 Warm waters - Cold waters 1.2 THe three phases in rearing 1.3 The different types of tanks 2. REARING 2.1 Rearing programme - Stock management 2.1.1. Nursery 2.1.2. First fattening - Fattening 2.2 Water management - Temperature 2.2.1. The oxygen consumption in the tanks 2.2.2. The gassy ammonia rate 2.2.3. Other physico-chemical parameters 2.2.4. Hydraulic aspects 2.2.5. Usual values 2.3. Feed 2.3.1. Generalities 2.3.1.1. Ration calculation 2.3.1.2. Formulation balance - Appetency 2.3.2. Dry or wet food 2.3.2.1. Dry food 2.3.2.2. Wet food 3. THE REARING FARM IN GRAVELINES 3.1. Introduction 3.2. Design of the farm 3.3. Description of the farm 3.3.1. The water circuits 3.3.2. The rearing units 3.3.3. Annex equipment 3.4. Running of the farm - First results 3.4.1. General characteristics 3.4.2. First results 3.5. Future perspectives INTENSIVE FISHCULTURE - ITTICA UGENTE S.P.A TORRE MOZZA DI UGENTO - LECCE Mr. L. CORBARI The fishculture is located in the Pouilles (Southern Italy) on the Ionian coastline; it employs sea water and it combines, sectors for the production of eel and white fish (gilthead sea-bream and sea-bass) along with a controlled reproduction centre. It began functioning in 1981. Characteristics of the facilities The fishculture is located near a series of tanks, the water level of which varies according to the tides, and these are used for intensive rearing. The total surface area consists of 25 hectares of land and 50 hectares of tanks and canals which communicate with the sea. The intensive rearing production structure consists of the following: An eel rearing sector, which has 24 tanks, each having a surface area of 1 250 m2 , concrete walls, supply and outlet systems, and a pre-tank which is employed for capture, selection and fish prophylactic treatment purposes. Eighteen tanks have concrete bottoms, while six others have compact gravel ones. 2 000 quintal of both commercial size eel (200 gr) and semi-grown products (weaned eels) are reared in this sector, every year. White fish rearing sector (gilthead sea-bream and sea-bass); Allows the growth of fish from fry size to commercial size (more than 300 g); It has the following structures: Pre-fattening: 20 tanks with concrete bottoms and walls and having a surface area of 25 m2 each; they are sheltered and used to rear fry throughout the first rearing months (January - April). The tanks are divided out in 4 rows; they are equipped with water supply and outlet systems, a liquid oxygen and compressed air central device for the supplementary aeration of water and a water thermoregulation apparatus. The first stage: has for purpose, the rearing of white fish during the first year. It has two series of 18 rectangular concrete tanks, each having 50 m2 in surface area and adjoining long walls. The supply canal runs along the short walls of the tanks while the outlet canal runs along the ones opposite. This disposition permits having a near constant water flow in the whole tank. The second stage: has the purpose of rearing white fish until commercial size is. reached. It has two series of 11 rectangular concrete tanks of 240 m2 each and has adjoining long walls. The supply and outlet systems are identical to these found in the first stage. This sector permits the production of 800 quintal of commercial size fish. The artificial reproduction of white fish sector permits the production of gilthead sea-bream and sea-bass fry. It is equipped with a phyto-zooplankton section of 24 cylindro-conical tanks for larval rearing, and reproduction tanks. The facilities are completed by special and normal structures, such as the pumping station, the electric press, offices, a cantine, a machine workshop, a warehouse, a cold room, a room for the preparation of feed stuff, etc... Sales 1984 1 300 quintal of eel: average rearing time: 18 months, average density: 2 6 kg/m2; conversion rate: 1:3. 100 quintal of gilthead sea-bream and sea-bass coming from artificial reproduction. Average rearing time: 24 months for gilthead-sea-bream and 36 months for sea-bass. Average density: 5 kg/m2; conversion rate: 1: 2,5. Personnel: The fishculture employs 20 people: 1 biologist, 1 administrative employee, 4 specialized technicians and 14 manual labourers. Research: On parallel with production, ITTICA UGENTO, has many applied research activities, such as ichtyo-pathology prophylaxis and the improvement of rearing techniques along with the automatization of the management.

Intensive rearing facility Legend: 1. First fattening 2. Offices 3. Larval rearing 4. Power station 5. Generator set 6. Pumping station 7. White fish rearing tank: stage 1 8. White fish rearing: Stage 2 9. Eel rearing tank 10. Supply canalization 11. Outlet canalization 12. Sluice gate 13. Semi-intensive tank

Valli 10 000 ha Reproduction Sevrage Intensif Mollusques Intensif 6 ha artificielle civelles Lagune 1973 1974 1975 1975 1977

Valli 10 000 ha Artificial Weaning Intensif Molluscs Intensif 6 ha Reproduction Elvers Lagoon 1973 1974 1975 1975 1977

ITTICA UGENTO SPA - Les Ventes - Sale N° Functions 1 Biologist in charge of Production and Research 1 Technician in charge of Production and Organization 1 Administrative employee 3 Specialized technicians 14 Workers

ITTICA UGENTO SPA PRSONNEL

Thermic tolerance of the different brackish water species reared Inferior or superior critical intervals Feeding interval Optimal growth interval

Average temperature of the water (Puglies) and rearing period

EEL Standard Production cycle

1977 1984 ∆% Costs (Lit. 77) (Lit. 84) Elvers (kg) 1 600 3 900 5 500 + 41,0 Feed (kg) 500 1 200 1 100 - 8,3 Annual manual labour cost 6 000 000 15 000 000 22 000 000 + 46,6 Energy (Kwh) 25 60 110 + 83,3 Profit Price of eel 4 500 10 900 9 800 - 10,1

EELS: Evolution of the cost of the different production sectors and of the sale price Actualization while using the ISTAS (2.4161)

EELS Production costs variations in % compared to profit considered equal to 100.

EELS Production cycles

EELS Average time period variation in production costs depending on the profile considered at 100. Possibilities for improvement in efficiency – Increase in the multiplication factor • Decrease in mortalities (Pathological study) • Optimization of feeding • Rearing techniques and conditions – Decrease in the work factor • Mechanization • Automatization – Energy saving • Improvement in • Technological transfer • Use of renewable energy Actions on fixed prices – Increase in rearing 1977 1984 % Costs (Lit 77) (Lit 84)* Sea bass fry 150 360 300 - 20 Sea bream fry. 400 1 000 1 000 - Food 400 1 000 1 050 + 5 Work 6 000 000 16 000 000 22 000 000 + 46,6 Energy 25 60 110 + 83,3 Profit Cost of white fish per kg 6 800 16 500 25 000 + 51,5

Evolution of the costs of the different production levels and the sale price Actualizating while employing the ISTAS coefficient (2.4161) SEA BASS SEA BREAM Reproduction Verified and reproducible Verified Larval rearing Acceptable inputs Uncertain and poor input Fry rearing Important mortalities Excellent results Rearing fattening Not optimal but acceptable Must be perfected, but good

WHITE FISH Schematic situation of intensive rearing

THEMES SPECIES Eel Sea bream Sea bass Pathology and prophylaxis * ** Reproduction ** Feeding ** * Rearing method * * **

Research themes and their priorities INTENSIVE REARING OF EEL Mr. G. ARLATI 1. PRELIMINARY VIEWS The following arguments must be taken into consideration: – Rearing methods, stagnant or running water – Choice of an ideal spot for the implantation of the eel fishculture. 1.1. Rearing methods: stagnant or running water It is possible to schematize eel rearing from the following fundamental characteristics: a) quantity and quality of the water; b) type of "seed" (elver or small eel), ("ragano"); c) feed; d) pathology; e) market. 1.1.1. Water To develop and live, eels have an urgent requirement of pure water and if possible abundantly. There must always be a good amount of oxygen present; when the density per surface area is increased, it must be possible to guarantee the oxygenation by means of auxiliary equipment. 1.1.2. "Seed" By eel rearing is meant, the rearing of very small eel or eel, at elver stage, which are taken from the sea, to commercial size. It is not yet possible to reproduce the complete biological cycle of eel in a laboratory: although the artificial insemination problem has been solved, it is still impossible to ensure the survival of the leptocephalus at larval stage. 1.1.3. Feed Every type of intensive zootechnical rearing is based on the distribution of special food which is suited especially to the product reared. With eel culture, it is not sufficient to employ a well balanced diet to ensure the result: the growth of the product depends, in fact, on the water temperature, on which depends the quantity of food that must be distributed, and the appetency of the food itself. It is also necessary that this latter, as it is very expensive, guarantees a good conversion index. 1.1.4. Pathology Parasites and bacterial diseases must be foreseen, by respecting the quarantine period for the fish brought from outside and the scheduled prophylaxis treatment of the environment, and if these should appear , quick intervention is demanded by employing adequate therapeutic treatments. 1.1.5. Market A breeder knowing how to sell his eel will obtain a better return than one having no commercial experience. 1.1.6. Eels are reared in earth or concrete tanks, in greater densities than that, which could be tolerated by the quantity of physical oxygen available in water. For this reason, two different fundamental methods of oxygen supply are employed, which in turn give rise to two rearing models: rearing in stagnant water and rearing in running water. With the stagnant water model, which favours the growth of the phyto-plankton green algae in the water of tank, the oxygen is furnished by the photosynthesis. A dense photosynthesis (20 cm optimal transparence) can not increase unless the water flow is very slow or inexistant. In fact, for these tanks, which have more or less stagnant water, the water exchange per day is around 5 % of the total volume. With the running water model, the oxygen requirement of the eels is furnished by a constant new water flow, as seen with the culture of trout. A continuous renewal of abundant running water is therefore required for this type of tank. The temperature and the quantity of water at disposal bring about the definition of the choice between one model or the other: – When there is a limited amount of water at disposal, it is possible to have stagnant water tanks or at the most, recycled water tanks having more or less running water. – When there are great quantities of water at disposal, it is possible to have either stagnant water tanks and if the mild climate permits so, running water tanks. The temperature of the water is a fundamental point: 19 - 28° C for the european eel. From an economic point of view, the rearing of eel is advantageous when sizes of 120 - 160 grammes are reached within two years or less, when starting off with elvers (0,2 - 0,3 g.); this rapid growth is only possible in optimal temperatures (21 - 26° C). The european eel does not eat in temperatures of under 8 - 9°C, but better results are obtained at temperatures of more than 17° C (22° C optimum). 1.2. Choice of the ideal spot for the implantation of an eel culture The spot will be all the more suitable for rearing when the specific conditions here following are present: 1) Availability of a good water supply, channeled or pumped from a canal, or better still, coming from underground sources or wells. For instance, around 400 - 450 m3 is required per day to rear 40 tons of eel per year. 2) The water must not be contaminated (by insecticides, phytomedicaments, etc... ) Alkaline or neutral water is the best, while acid water (below 6,5 pH) is not very recommendable for rearing. The eventual presence of wild eel in the water which is to be employed indicates clearly that the water is of good quality and suitable for rearing. 3) The rearing site must not receive sea inflows or be flooded. 4) The water exchange in the tanks must preferably take place by means of downflows. In other words, the energetic costs must be as limited as possible, the use of pumps for the distribution of water must be avoided when possible. 5) The soil must not be porous so as no water leakage is possible from the bottom or sides of the tanks. Clay mixed with sand is the most advisable. 6) The site must receive sunshine so as to favour the algae bloom, which by the chlorophyllian synthesis produces oxygen. 7) the site must be cleared so permiting the oxygenetion of the surface water by the wind. 8) Good roads and electricity must be available 9) The availability of important markets in proximity must be ensured. 2. INTENSIVE REARING The technics employed for the intensive rearing of eel are based on three essential conditions: favourable temperatures, sufficient oxygen, artificial food. Favourable temperatures for european eel rearing vary from 8 to 28° C; The optimal temperature for the introduction and weaning of small wild eels varies from 18 to 24° C; for the weaning and growth of elvers, it is situated around 22 to 26° C. The further the temperature is away from the optimal values, the longer the biological cycle of rearing will be. The dissolved oxygen content required for the survival of the eel is from 2-3 mg/l for elvers and 0,5 mg/l for adult eels. In rearing, these oxygen requirements are greater and vary depending on the temperature (and the salinity), the size of the eels and their metabolic activity. Generally, saturation values of 80 - 90 % are acceptable up to 20° C, and a value around saturation for temperatures of more than 20° C. Normally, artificial feed for eels is made in the form of moistened paste balls, mixtures of meals, oils, water and having or not a certain quantity of minced fresh fish content. 1.1. Intensive rearing cycle At present, intensive rearing can be subdivided into three parts: – Elver weaning and rearing – Small wild eel ("ragano") weaning – Eel rearing The breeder must establish his own production with fish coming from outside: elver or "ragani" caught in sea or brackish waters, or with small eel which have grown from weaned elver or "ragani" size, and coming from other rearings. This method is employed so as to avoid the appearance of the "closed cycle" at biological level, and it gives diversified results to the breeder. Only the description of the different phases in the rearing cycle, and the most renown methods and technics shall be taken into account here. 2.1.1. Elver weaning and rearing Elver weaning and rearing is carried out in both brackish and fresh water, in special tanks, built especially for this purpose. Generally concrete or polyester resine tanks are employed, of different shape (rectangular, square, or round) and of variable size (from 8 to 20 m2) and having the required equipment from a technological point of view at disposal, (independant hot and cold water circuits, premixed 02supply, air supply by means of different types of diffusers: blowers, venturi, etc... an outlet circuit having partial or total feedback of the hot water and when required the possibility of discharging non reusable water, etc... ) These tanks are located in enclosed green houses or sheds. Elver rearing is known as the 1st stage and it is economically advantageous when there are higher temperatures than 21-22° C up to 25° C available. Elvers, coming from the sea, can be of variable weight (0,20 gr. to 0,40 gr) depending on the season and the area where caught, available from November to February the italian eel is smaller, while those found on the French or English Atlantic coast are bigger and are available in the Spring, from March to May. The "seed" density varies depending on the rearing method employed: if stagnant water tanks, having an exchange of 6 - 7 l/minute, are employed, the density is 0,4 kg per m2; if running water tanks, having an exchange of 40 - 45 1/ minute about are employed , it is 3 kg. per m2 . At this point weaning takes place, by employing a minced mixture of worms and anchovy- and/or sardines, blended with feed in powder form, or with a binder. Little by little, a meal of more or less compound food is added to the paste. It is a complete and self sufficient food which becomes less and less appetizing (anchovy, live worms, etc... ).Within 4-5 weeks, if the product is healthy and well adapted to the environment and has not suffered any stress for the latter, the importance of adapting the elvers gradually, within 24 - 36 hours, which have come from temperatures of 9 - 12° C, to ambiant rearing temperatures must be remarked; it is weaned at 22 - 25° C; it is necessary to control the hygienic and sanitary aspects and carry out the required treatments when necessary. At this stage, a first selection is made, as a uniform and homogeneous size facilitates growth speed: the smaller eels will be placed back into the weaning tanks, the others, when weighed, will be fed a moistened paste to fatten them and another selection will be made 3 or 4 weeks later. At this rythme, within 8 months the elvers reach the size of small eels ("raganello") of 15 - 16 gr. A typical cycle can be resumed schematically in the following way: Individual average weight rearing days at 22° C Number of selections carried out 0,25 4-5 weeks 1(weaning) 0,5 2 months and 1 week 3 1 3½ months 5 2 4 months and 3 weeks 7 4 5 months and 4 weeks 10 8 8 - 9 months 13 16 10 - 12 months 16 In stagnant water tanks, it is possible to have a maximum density of 1,6 - 1,8 kg/m2 about, while in running water tanks, 5 kg/m2 is obtained. In any case, the densities, apart from depending on the water exchange and the size, also depend on the oxygen supply technic employed. Finally, if all the operations are to be profitable, it is necessary to obtain 20 - 25 kg. of small eels within 9 months from 1 kg of elvers, and to reach 30 - 35 kg within a year. The above data is purely indicative, as the minimum percentage, less than 15 %, will reach 15 gr within 5-6 months; the greater part , 45 to 65 %,will reach a pay size in the scheduled time; and the rest, known as the "tails" will take longer to fatten. The present cost of elvers is 27 - 30 000 lira/kg; the cost of the finished product of this 1st rearing stage, which is but a rough estimate, as there exists no real market, is around 46 - 52 000 lira/kg. Italy's future for eel culture lies in the rearing of elvers, but up to the present day, very few 1st stage rearings exist, which would provide a guaranteed production. This is explained by the fact that 5 or 6 years ago, breeders were encouraged to start off experimental elver rearings at. industrial scale, but a more conclusive evaluation on the costs/benefits showed that this would only be of profit if low costing hot water was at disposal. The secret of a good first stage rearing result is based on the selection, the feed and the sanitary operation programme. 2.1.2. Small wild eel weaning ("ragano") It is not necessary to have water of more than 22° C at disposal for small wild eels, good performances are also obtained at 18° C. Some breeders even employ water from sources having temperatures of 13° - 14° C, but this is only carried out when a small number of animals are to be weaned for rearing purposes, as the lenght of the weaning cycle is rather long (4-5 months) and not always conclusive. It must be remarked that the weaning of elvers as well as small eels must try to "domesticate" a good amount of animals in the shortest time possible. A good environment, efficient technical management of the facilities and a correct weaning method would ensure success. The concrete or polyester resine tanks, located indoors, facilitate the weaning and sanitary controls; these tanks can be of variable size and form (from 24 to 200 m2 ). As most of the small eels are taken from the Mediterranean French coast, it is necessary to carry out an antiparasite treatment on arrival, employing formaline and green malachite for two days before starting to wean them and keeping them under observation (quarantine) for at least 2-3 weeks before sorting them out; taking these precautions, it is possible to intervene immediately if a serious and contagious disease should occur among them. It is better to eliminate immediatly a certain number of contaminated elements than to run the risk of the whole rearing catching the disease: the consequences would be all the greater and costly in the end. After this stage, weaning starts by distributing whole sardines threaded on to a wire through the head or a moistened paste mixed with fish (40 %) or with red worm (10 %). During the weaning period, it is recommendable to carry out two antiparasite treatments each week as means of prevention, preferably each morning and in any case the fish must always be fasting. After about one month, a first selection may be carried out: the smaller ones being kept in weaning while the others will receive less and less feed attractants in the paste. After about two weeks, a second selection is carried out followed by another, two or three weeks later on, at the third sorting out the product will be big enough to go into open air tanks for fattening purposes or 2nd stage growth. It is presumed when employing this method, that concrete tanks and the required structures, so to facilitate fishing and selection, be at disposal. Elver tanks are quite commonly employed to wean small eel during certain periods. In all cases, if there are similar conditions present, the density of the product may be freely multiplied by three compared to that for elvers. Most of the Italian eel cultures are second stage rearings employing concrete fisheries (RAVAGNAN model). This area, which is easily controlled and isolated, takes up about 1/10 of the total surface of the tank and it is possible to carry out, along with the management operations (collection and fishing of the fish), the weaning of small wild eels. However, this is only feasible under one condition; that during this period, the rest of the earth tank is not employed for rearing purposes, as this operation would be very risky and could have an effect on the sanitary situation in the tank, resulting in serious consequences. Indeed, the more knowledageable breeders, employ the fishery to wean small wild eels, who have been accostumed to fattening feed, and set them free, after selection, into the other part of the tank. In this case, the "seed" density is 1,5 to 2 kg/m2 when the whole rearing tank is taken into consideration. It is also very important not to recollect the outlet water, but to let it discharge into a small decantation lake (3 000 - 8 000 m2 ) before finally flowing out of the rearing. During the weaning period, the average mortality varies from 10 to 30 %, depending on whether, enclosed structures or open air rearing, are employed by the production centres. Small wild eel of 15 - 20 gr average size cost 5 000 to 5 500 lira/kg, (unfortunatly this price will rise, as there are fewer and fewer small wild eel available on the market today) and are sold when weaned at 2 - 3 times their original price. 2.1.3 Eel rearing The fattening cycles of eel takes place outside, in either fresh water - which is preferable, as it quickens growth, or in brackish water in earth tanks, of square or rectangular shape and having variable surfaces (400 to 1 600 m2). It is not Obligatory to have at disposal "hot" water 22° C, but temperatures of below 14 - 15° C do not give good results. In the North of Italy, where the possibility of obtaining water at 12 -14° C from wells exists, the most commonly employed method, is that of using semi-current water, which has a constant and very slow flow (6-8 1/sec. for a surface of 1 000 m2, which roughty corresponds to the same number of cubic meters). All solutions are possible as long as the collection and fishing of the product is feasible, as the management is the most important operation. Tanks have been especially built so to permit the automatic selection of the product. The average period for the rearing cycle is linked with the climatic situation and the seasons; in the North, to reach commercial size, when beginning at small eel size (25 g), takes 8-9 months; in the South, the same result is obtained in 5 - 6 months. The fish already sorted out and weaned, are placed into the fattening tanks, in the Spring. This way, part of the product of commercial size will be available for Christmas, a time when there is a great demand for this product. The "seed" densities can vary from 1 to 3 - 4 kg/m2 , depending on the method employed; the maximum density reached, is three time more. During this period, mortality reaches 15 - 30 % and for fresh water rearing it is necessary to carry out sanitary treatments against ichthyophtyriase, protozoan parasites which are very contageous and the cause of drastic economical consequences: these diseases are the cause of absense of growth and even death if widely spread throughout the product, when the proper controls are not carried out. Here below is a production plan for a 2nd stage rearing, carried out in semi- current water flows having an average annual temperature of 18 - 19° C (Centre and North of Italy). Individual average size Rearing days N° of progressive Selecting apparatus (gr) selections (diameter mm) 15 - 20 Seeds 0 8 25 - 30 1-2 months 1 10

It is not obligatory to carry out all the selections recommended, but it is necessary to do three at least (the 1st, 3rd and 5th) so as to obtain an average size male of 120 - 180 gr. and a female of 300 - 350 gr. These selections are carried out in the aim of having fish of uniform and homogeneous size, this permits a shorter rearing period. The selection may be carried out manually, it is more difficult but more efficient also than the automatic option which is quicker but not as exact. At adult stage, the eel transforms sexually. When adult stage is almost reached, the eel is not profitable to rear because its feed conversion decreases greatly, becoming practically non-existant.. This is why, breeders principally aim at producing as many females as possible: these being the only ones reaching a weight of more than 250 - 300 gr. as their maturity takes longer; on the contrary, males reach maturity from a sexual point of view when they reach a size of 120 - 150 gr. and must be commercialized (fig. 1). Good breeders can obtain a percentage of 30 - 35 % of females which require 6- 8 months rearing to reach this size. Present market prices are 11 000 - 12 000 lira/kg (three fish weighing 300 - 350 grammes each) while for males of 120 - 150 gr. (7-8 fish per Kilo), the prices range from 7 800 - 8 000 lira per kilo. 3. FEEDING 3.1. Feed requirements The eel is a carnivorous and predatory fish having a tendancy for cannibalism, it has a stomach. It preferes moistened feed. It's feed requirements have been thoroughly studied by the japanese researchers, but only concerning the species reared on site. The essential amino-acid requirements can be seen here following, after NOSE, ARAI and HASHIMOTO (1972). Essential amino-acid requirements for 100 gr. of proteins Arginine 3,9 Metionine + Cistine 4,5 Istidine 1,9 Phenylalanine + Tirosine 5,2 Isoleucine 3,6 Triptophane 1,0 Leucine 4,1 Treonine 3,6 Lisine 4,8 Valine 3,6

Also certain data concerning the composition of feed on sale in Japan can be found in table 1. The biological value of the protein, which means it's essential amino-acid content, is of particular importance in the artificial diets for eel. It must not differ too much from the concentration relations existing in the protein of the organism of the fish. Here finally, can be seen the nutrient requirement of warmwater fishes from the National Academy of Science - WASHINGTON, a mixture for the mineral integration: Minerals gr/100 gr of food

Ca Co3 0,750 and the composition of the vitamin mixture: Vitamins For each kg of food Vitamin A 5 500 IU Vitamin D3 1 000 IU Vitamin E 50 IU Vitamin K 10 mg Colin 550 mg Niacin 100 mg Riboflavin 20 mg Piridoxyne 20 mg Tiamin 20 mg Calcium pantothenate 50 mg Biotin 0,1 mg Folacine 5 mg Vitamin B 12 20 mg Ascorbic acid 30 - 100 mg Inositol 100 mg Two types of food exist in Italy, the first one, known as the starter, has high protein content (50 -55 %),.and poor lipid content (5 %); it is given to elvers and during the weaning of small wild eels, it costs around 1 100 lira/kg The second one has a lower protein content and a greater lipid one, it is especially suited for growth and fattening, till the eels reach commercial size, it costs around 1 000 lira/kg. 3.2. Feed technics A moistened paste is prepared just before being distributed to the eels, as shown in tables 2 and 3. This paste is prepared with equal quantities of meal and water (the water being the same as that found in the tank). Some breeders add a fish oil percentage which can vary from 2 to 9 % depending on the water temperature and the size of the fish, otherwise a percentage of minced blue fish (5 - 10 %) can be added instead . Some breeders add a vitamin mixture to the diet every two months (1 - 2 % of dry matter) during 5-6 days so as to avoid deficiencies, due to lack of vitamins, occuring (Tab. 4). The food is distributed daily by means of feeders, which are placed on the surface of the water, 4-6 times per day for elvers, 3-4 times for small eels in weaning and twice for eel in growth, respecting the quantities indicated in the daily food table. When the work must be carried out in lower temperatures, the paste is given out only once per day. A real and practical estimate, to verify the quality of the food, is made by watching how well the food is accepted and its conversion index; to enable this, the breeder must mark down the food consumption for each tank. For elvers (0,2 - 12.gr), the conversion index is 1,4 - 1,8; for small eels (12 - 25 gr) it is 1,8 - 2,2; while for those in fattening (25 -180 gr) it can vary from 2,3 to 3,2. The percentages are given in dry matter by calculating the meal at + 1,7 of the amount of fish given,and they are valid for optimal rearing temperatures (22 - 23° C). Fats must not be added when the temperature falls below 15° C. 4. GENERAL PROGRAMME PLAN When a rearing is started up, the principal aspects of the technical management must be taken into account. For this, let us examine a programme model, concerning the rearing cycle in a typical fresh water facility in the North of Italy which has water supplies from wells 15 - 16° C at disposal and when required an additional water supply from a source of non polluted water. Although the optimal conditions of temperature are not found here (Tab. 5) it is still feasible today, to produce eels, starting with small eels and not elvers, when a temperature of 18 - 20° C is present for 10 months of the year as shown in the theoretic plan (Table 6). The fundamental problems, which shall be briefly confronted here, concern: a) the possibilities of finding small eels; b) food; c) sorting out; d) an adequate programme of the environmental prophylaxis, carried out while respecting the elementary sanitary norms. 4.1. Finding small eel ("ragano") The possibility of finding small eel is essential for the smooth running and success of the entreprise. The small eel should be lively, strong and healthy. Most of the small wild eel come from abroad at the present day (5 000 lira/kg, 50 - 60 fish per kg). Weaning will be all. the easier when they are healthy. Above all, a good breeder must get the wild eel to adapt to artificial food, or rather to get the greatest amount of fish to "adjust" as quickly as possible. By employing the more recent and sophiscated methods, 85 to 90 % of the fish can be weaned in 3 - 4 weeks. For this phase, indoor concrete tanks, having a constant water exchange, water from sources or wells, must be employed, in preference. 4.2. Feed Apart from the weaning period during which special substances bringing on a appetite, are added to the paste, the food diet must be autosufficient and well balanced. Indeed, a good breeder usually adds 5 to 10 % of blue fish, which is minced just before the distribution of the feed takes place, special "integrators" in the appropriate quantities, depending on the requirement of the rearing, are added. The nutrient function of the eel is linked with the temperature of the water and the dissolved oxygen content present in the tank (Fig. 2) and this latter depends on the weight of the eel (Fig. 3 and table 7). The food conversion indexes of the plan are those obtained with the food there presented, and integrated in the appropriate way to the above mentioned temperature: They are at optimum in these conditions, which must n't stop improvements being made in the near future. 4.3. Selection Selections are very important in modern eel rearing. The aim of these, is to obtain a tank of fish of uniform and homogeneous size, therefore the greatest possible number is fed and a quick weight increase is obtained. In fact the rearing cycle is reduced and consequently the biological risk. The selections shown in the plan, concern the principal selections, which means, those which are really necessary in any environment and which must take place immediatly after weaning and when commercial size is reached, for both males and females (big eels). Two additional selections are necessary for each fattening cycle, they are obligatory within the programmed management, as the equipment and methods of today are at disposal and permit not only the collection and fishing of the product more quickly (transfer by means of a siphon) but the real sorting out (semi-automatic sorter). 4.4. Prevention The basis of sanitary protection, for every rearing, is prevention rather than treatment; this is why it is necessary to intervene adequatly, employing the precise programmed operations of prophylaxis on the environment, the parasites and the bacteria. This sanitary protection programme, includes the intervention methods, linked with the rearing environment and the particular situation in which the work is carried out. Some prevention normes, exist consisting very simply of keeping optimal conditions in the tanks. In this way, the pathological problem will not be as important, as there will be fewer occasions for "stress". Therefore it is very important to follow certain methods of management: – ensure that the rearing tanks have a sufficient supply of non-contaminated water every day. – respect the quarantine period and disinfect adequately the environment every second day, at least over 2-3 weeks, (depending on the temperature) for those fish coming from outside before placing them into fattening tanks. – dry the tank completely by removing organic residue at the end of each rearing cycle at least and by leaving it in the sun for 7-10 days: a better productivity will be obtained this way. – distribute a well balanced and adequate food diet. – Carry out good anti-bacterial prophylaxis, followed by polyvitamined treatments, at least 2-3 times per year, at the most critical moments. – eliminate dead eels from the tank immediately and incinerate them. 5. CONCLUSION AND ECONOMIC CALCULATION It must be understood at once, that the total use of the production possibilities of a rearing, is not possible before 2-3 years after it has been commenced. Let us now analyse quickly some data on production, as seen in the plan. Even in the non optimal conditions of the example given (18 - 20° C), productivity is quite good and still profitable, from an economic point of view: indeed, in 11 - 12 months, 60 % of the fish reached adult male stage (120 - 150 gr; 6-8 fish/kg) 12 - 13 months after their arrival at small eel stage; the remaining 40 % reached adult eel size after 22 - 23 months. Until male stage is reached, mortality is from 22 - 28 % (average 27 %) and 35 % for the complete biennial cycle of the big eel; taking into consideration the mortality of the different rearing phases and the male/female relation of , the small eels bought, it is possible to obtain 42 adult males (5-6 kg), and 23 b female eel (11,5 - 13,5 kg) when starting off with 100 small eel. These results are feasible for every good breeder who is prepared and capable of dealing with the problems in. the proper way which may arise in the course of the daily work. Finally, taking as an example a rearing which has a water surface area of 15 000 m2 and a annual production capacity of at least 40 tons of marketable heavy product, the yearly economic calculation (table 8) where the operational result (gain) is given. TABLE 1 SOME DATA CONCERNING THE COMPOSITION OF JAPONESE COMMERCIAL FOOD (from H. AOYE, reported by TOMIYAMA & HIBIYA, 1977) Elver 1 Elver 2 Ragani 3 Adult 4 Proteins min 49,0 min 47,5 min 46,0 min 45,0 Lipids min 3,0 min 3,0 min 3,0 min 3,0 Fibre max 1,0 max 1,0 max 1,0 max 1,0 Ash max 17,0 max 17,0 max 17,0 max 17,0 Calcium min 2,5 min 2,5 min 2,5 min 2,5 Phosphorus min 1,3 min 1,3 min 1,3 min 1,2 Diet composition 1 Animal substance 69 69 68 65 Cereals 19 22 21 22 Others 12 9 11 8 Oil cake - - - 5

1) In miscellanea: fish meal, yeast meal, liver meal, linseed meal, soyabean meal. Vitamins: Vit. A oil, calciferol, tocopherol acetate, thiamine nitrate, pyridoxine hydrochloride nicotinic acid amine, calcium panthotenate, folic acid, cyanocobolamin, vit. K, riboflavin, biotine, inositol, chloride, ascorbic acid. Minerals: Calcium carbonate, ferric acid, fumaric acid, potassium chloride, magnesium sulphate, calcium phosphate, manganese sulphate, cupric sulphate, zinc sulphate, calcium iodate, cobalt chloride. TABLE 2 DAILY FOOD TABLE Elvers Conditioned eels small eels (0,2 - 0,4 gr) (4 - 12 gr) Food in % b/w 10 4-6 3 - 5 % of water in the food 140 - 160 140 - 160 130 - 140 % of fats in the food 0 0 3 - 5

Table 3 DAILY FEEDING SCHEDULE FOR ADULT EELS (25 - 120 - 300 gr.) Water temperature % Feeding Feed % oil Water 4-8 small quantities 100 0 100 - 120 8-10 0,25 - 0,50 100 0 100 - 120 10 - 12 0,50 - 0,75 100 0 100 - 120 12 - 15 0,75 - 1,25 100 0 100 - 120 15 - 18 1,25 - 1,50 100 3-4 100 - 130 18 - 21 1,50 - 2,00 100 5-6 100 - 130 21 - 23 2,00-2,50 100 7-8 100 - 130 23 - 26. 3,00 100 10 100 - 130 27 - 30 1,50 - 2,00 100 10 100 - 130 Healthy eels are voracious animals and consume the food given to them quickly. Consequently, it is important to study how eels eat and to only give them the quantitiy that they would eat. Table 4 DEFICIENCY SYNDROMES OF WATER SOLUBLE VITAMINS IN A. japonica (by HASHIMOTO, 1974) Vitamins Deficiency signs Thiamine Anorexia, poor growth, ataxia, fin haemorrhage, dark colouration Riboflavin Anorexia, poor growth, fin haemorrage, dermatitis, photophobia, lethargy Pyridoxine Anorexia, poor growth, nervous disorders, epileptic fits, convulsions Pantothenic acid Anorexia, poor growth, ataxia, mortality, shin haemorrhage, skin lesions, dermatitis Inositol Anorexia, poor growth, grey-white intestine Biotin Anorexia, poor growth, ataxia Folic acid Anorexia, poor growth, dark colouration Choline Anorexia, ataxia, grey-white intestine Nicotinic acid Anorexia, poor growth, ataxia, anemia, skin haemorrhage, skin lesions Vitamin B 12 Anorexia, poor growth Para-aminobenzoic acid Nothing to report Ascorbic acid Anorexia, poor growth, fin - head - shin haemorrhage, lower jaw lesions Table 5 INITIAL CONDITIONS OF THE WATER TO BE EMPLOYED FOR EEL REARING Elements Values (optimum) PH 6,0 - 9,0 (7,0 - 7,8) Dissolved oxygen 60 - 100 % (more than 80 %) Salinity 0,3 – 5 % (0-0,2%) Alkalinity 0 - 3 mEq/l (1,5 - 2,0)

N - NH3 0 - 2 ppm (0)

H2S 0, 1 ppm (0) Iron 0 - 1, 0 ppm (0 - 0,05) Temperature 13 - 30° C (20 - 25° C) by OGAMI, 1974; translated by HOSHINA, 1978 Table 6 ROGRAMME PLAN (18 - 20° C) Phase Duration of Final Rearing cycle Mortality Density Water Food Average food the phase individual % (Kg/m2) exchanges daily % Number of conversion indexes weight (gr) (number of times/day times per day) 1 st 20-40 days 16-18 Weaning 12-18 10-15 6 - 8 ad libitum 3 - 4 - ° 2 nd 3 months 45-50 fattening 4 from 1-2 1 - 2 2 - 2,4 1 - 2 1,8 - 2 * to 3 - 4 3 rd 4 months 80-90 Fattening 4 from 2 0,5 - 1 2 1 - 2 2 - 2,3 * to 4-5 4 th 3 months 120 - 150 final fattening 2 From 3 1 - 2 2 1 - 2 2,4 - 2,6 of males to 6 – 8 ° 5 th 4 months 240 - 260 Fattening of 2 from 3 0,5 - 1 2 1 2,5 - 2,8 big eels to 8 * 6 th 4 months 360 - 390 Fattening of 3 from 3 0,5 - 1 1,8 - 2 1 2,3 - 2,6 big eels to 8 * 7 th 3 months 500 - 600 Final fattening 2 from 3 1 - 2 1,8 - 2 1 2,6 - 2,9 of big eels to 8 °

° = Necessary selection - * = Advisable selection TABLE 7 - Eel: Individual and per kg oxygen consumption of body weight Weight of eels (gr) 5 10 20 50 100 150 200 300

O2 consumed (mg/h) 1,1 1,7 2,9 5,5 8,9 11,9 14,9 19,3

O2 consumed (mg/kg/h) 214 171 143 108 89 78 73 64

02 consumed (cc/kg/h) 150 120 100 76 62 55 51 45

TABLE 8 Annual Economic Calculation of a 15 000 m2 of Water Rearing Gains (average price eels L 8 - 9 OOO/Kg (*) % Lira 100 320 000 000 Management costs 1. Feed 33 105 000 000 2. Seeds 18 57 000 000 3. Salaries and emoluments 12 40 000 000 4. General cost of functioning 10 31 000 000 5. Depreciation 8 25 000 000 Total Costs 81 258 000 000 Results of the Operation 19 62 000 000

(*) NB: eels: L 7 300 - 7 600/Kg - Free rearing; Big eels: L 10 000 - 15 000/kg - Free rearing.

Fig. 1. Course of growth of Anguilla anguilla; female (0); male (+). From Bulletin of the Japanese Society of Scientific Fisheries.

Fig. 2. The feeding activity for eels and the dissolved oxygen content depending on the temperature. (Mx: maximum; A: average; L: low; N: nil. Here following as the subdivision of the temperature values concerning the feeding activity (L: mortality , Sub O: suboperative; OP: operative; OT: optimal; CR: critical; Semi O: semi operative (by RAVAGNAN )

Fig. 3. The oxygen consumption for eel depending on weight. (by RAVAGNAN) INTENSIVE REARING OF MULLET Mr. F. GHION It is only in the last few years that the rearing of mullet is being carried out while employing reliable methods. This practice was derived from the standards required in "valliculture". With the intensive rearing of fish the first year, a means to increase the steady rate was remarked with this practice. In addition to this, the integrated valliculture technique was employed, in which, mullet play a principal role. In turn, and following the availability of food, suitable for this species, the rearing of mullet in fresh water became developed, while limited to the 2 species, which in such an environment show a more rapid growth rate: Mugil cephalus and Liza ramada. This slow development was backed by a considerable series of experimental tests, carried out for the formulation of a diet, especially suited to mullet, which up to this had not existed. This text gives the history of this evolution along with the synthesis of the research activity which has been carried out over the past ten years. MULLET IN TRADITIONAL VENITIAN VALLICULTURE I assume that you already have a knowledge of the rearing practice involved by valliculture techniques which programme more or less similar interventions for all the species reared and so those applicable for mullet. Thus there exists for mullet, seeding in the Spring (the fry is either bought or obtained from the stock), growth in Summer in the "valli" tanks, direction back and capture in the estuary which is followed by the distribution into the Wintering tanks. THis cycle is repeated each year until commercial size is reached. In fig. 1, can be found the 5 species of mullet deviated into our "valli". Beside the name of the fish, the size and the commercial value are indicated, and as can be remarked these vary depending on the fish species. Figures 2 and 3 show the growth variations in intensive valliculture. These graphs represent the general values. The growth can vary from one valli to another and also from one season to another. It is however evident that 3 years of growth is necessary for a species sold at the smallest acceptable commercial size (ex. 300 g. L. aurata) and a longer time for the others. As already remarked here above, intensive rearing of mullet was introduced in the aim of obtaining a superior steady rate later on. Indeed, we thought, that the rearing of fry in small tanks for several months would permit the distribution in the extensive tanks,of fish of greater size, which would be capable of defending themselves and escaping carnivorous fish (fig. 4). A diet especially suited to mullet was unfortunatly not available, so the food given normally to trout and other fish was employed. By simply taking into consideration the natural food of mullet, it is clear that this food if not incorrect was, at least,inadequate. In the food chain, mullet is placed indeed at a quite low trophic level. In examining fig. 5, which represents the components of the aquatic ecosystem, it can be said that mullet is located at primary consumer level while invading somewhat, all the same, the secondary consumer zone (carnivorous). Litterature indeed recalls that mullet is principally detritivorous and phyto planktonphageous but occasionally, it can ingest small organisms. From an energetic view-point while admitting that for each ring of the food chain only 10 % of the energy used is restored, it seems evident that mullet, when compared with the other fish in the "valli", makes better use of the primary energy produced by the real ecosystem. This better use can be estimated at around 10 to 100 times more than that of the other fish. The natural diet of the mullet which also comprises a great quantity of inert material, thus appears as a poor diet. In general, it would seem correct to believe that artificial food should follow the same pattern. The most important point in a diet from a commercial view-point is represented by the protein rate. A series of research tests have been carried out so as to define the minimum protein rate of the food for mullet, and the summary of these is given here following: Figure 6 shows the growth variations of fry (M. cephalus), submitted to diets of variable protein content (from 20 to 80 %). The results obtained after 20 weeks of rearing in an aquarium are shown in table 2. The comparison is easier in fig. 5. The minimum content is found around 40 %; diets containing 20 to 30 % shows an advantage obtained at growth level while diets with high protein content show their complete futility. Other tests have been done with C. labrosus , L. aurata and L. ramada, while using 2 different diets (see fig. 7). The essential difference between A. and B diets is the carbohydrate rate content, the lipid content is nearly always constant and that of proteins varys from 10 to 50 % (see fig, 8). Type A diet which presents a lack in carbohydrates did not permit to establish a minimum protein level (M.P.L) while for type B diet (with more sugar content) the M.P.L was reached for each species, at around 20 % (fig. 9). Other tests of the same type, trying to establish the influence of the lipid content or the origin of proteins, have given to conclude that a diet for mullet must contain a rather low protein rate (20 to 30 %) , a high carbohydrate rate (60 %), and a lipid rate of between 5 to 10 %. It is clear that such values can be reached if plant life diets are employed. Intensive rearing of mullet is thus developed by following these instructions . The preparation of a food can not however be limited to its simple formulation. In the special case of mullet, the importance of the granulometry of the natural food is known. It is also known that mullet can adopt very diverse food techniques by adapting themselves to the characteristics of the substratum. Thus, mullet feed by sucking and filtering the bottom layer, by scraping the patina covering the submersed material, and by sucking up the tiny fragments of floating organisms. Figure 10 shows the results obtained in a test carried out to reveal the quantity of food eliminated by the mullet, depending on the size of the particles offered. With this method it is noticed that 40 % of the food commercialized on the market is eliminated. It must be remarked that some particles of this food are bigger than 1 000 urn. There is less waste depending on the number of big particles eliminated. Furthermore, the chemical analysis carried out on this food eliminated, shows that the use of the protein part is at it's best, when the size of the particles is less than 250 urn. This can be also due to the fact that the protein part of the food is generally represented by fish meals or finely minced meat. However, while analysing all the results obtained by both tests, it seems clear that a finely minced food causes less wastes and is thus finally more interesting. The following figure shows the results of an experiment carried out so as to verify the efficiency of the filtration systems of mullet. For this test, 5 diets of different granulometries were prepared (from under 50 to 1 000 microns) together with 5 lots of inert material (sand). Each separate diet was mixed with one lot of inert material in every possible combination, while using around 30 % of food and 70 % of inert material. The organic substance content of the diets obtained was then defined. After the food had been given to the fish (one diet at a time) and it was ensured that this food had been ingested, the fish were sacrified and the analysis of the organic substances found in the stomach was established. The variation between the organic substance content of the food and that of the stomach was chosen as indication of the ingestion capacity. From the results of fig. 11 and 12, it appears when too fine material is presented, the mullet is incapable of selecting out the organic material, while it is quite capable of doing so when the particles are of- between 100 to 250 urn. Thus it can be concluded that these are the ideal dimensions for mullet food. As for the distribution of food, a test was carried out in concrete tanks where the food was distributed in the following manner: 1. Whole food distributed automatically over the water surface. 2. Food mixed with sand distributed automatically over the water surface. 3. The same but with adult fish. 4. Food mixed with water and placed on the bottom. 5. Food mixed with water and sand, placed on the bottom with adult fish. 6. Food mixed with the mud of the "valle" and placed at the bottom. For this experiment, juveniles weighing on average 10 g. for all species were employed. The results are found in fig. 13. It can be remarked that the best growth is obtained with the first system which also gave the best indication of conversion. The use of sand seems to have little influence on growth apart from in the case where adult fish are mixed with juveniles. No advantage is obtained when the mud from the valle is mixed with the food and this confirms the observation made here above on how mullet are uncapable of selectiong out very fine material (like the clay employed). These observations permit us to arrive at a new conclusion concerning the feeding of mullet. It seems interesting to distribute whole food over the water surface. From a practical view-point, this technique is very simple. By applying these results, the intensive rearing of mullet can become widespead. It is limited during the first and second year in valliculture and widespead in fresh water rearing stations until commercial size is reached, this is obtainable in two years. To recall the observations made here above on the natural feeding habits of mullet and the ecologic value (energetic saving) that this species represents, it can be said that our final objective can not only limit itself to producing an artificial food but it seems more important to do research work which would increase the production of mullet while making better use of the natural environment. It is evident that in this sector, the problems appear to be more complex as they concern the natural procedures which we would like to channel in a predifined direction. The idea of wanting to increase the primary production through fertilization, work, etc... is valid and applied correctly in fresh water aquaculture. This appears more difficult in a sea-water environment and especially complex when we refer it to valliculture. Let us examine fig. 14 which shows the frequence of occurance of organisms found in the stomach of 40 mullet of different species captured in a pilot facility of integrated valliculture. This figure brings nothing new to the feeding habits of mullet which are already well known. The recent presence of all the animal species already described, abundance of organic and inorganic detritus, phytoplankton and plant life. Fig. 15 shows the situation of one species: L. ramada captured in sea-water. The selection of small material seems quite evident- The same species captured in fresh water shows a real preference for plant organisms along with rotifers which were evidently in full growth at the time of capture. This confirms on one hand the adaptability of the species to the most varied conditions of the environment from a Trophic viewpoint and on the other hand it also shows how mullet can take an immediate advantage from the flow back of an intensive facility thus also anticipating the fertilizing effect. It can also be affirmed that in this case, the mullet play a depuration role for the "water by eliminating the suspended matter. As for the ploughing machine used in valliculture, it seems that this has already been referred to. I would only like to add, that experimentally, 1 t/ha of yearold mullet was reared in a pond of around 4 ha, by fertilizing the environment with 18 t of organic matter for the whole season and an intervention by means of the machine was employed every fortnight. Apart from the high concentration of fish observed (estimated) it was verified that the intervention of the machine helped get rid of sulphuret and phosphorus contained in the mud at the bottom. This ploughing permits on one hand better hygiens in the bottom and the water and on the. other hand recirculates a great quantity of fertilizer which induces a better phytoplanktonic production (increase of the chrolophyl in the water). The establishment of the exact technique of management for the ploughing machine and the possibility of introducing organic matter into the environment remains a problem to be looked into. The careful use of the suggestions dictated by common sense together with the knowledge of the feeding habits of mullet introduce the development of the semi-intensive rearing of mullet which gives good hope for the future. It is also clear that the industrial production of this species can not be performed until sure and cheap methods for Wintering can be ensured. LIST OF FIGURES Fig. 1.Specific characteristics of the different species of mullet. Fig. 2.Growth curves of mullet in extensive valliculture. Fig. 3.Curves obtained for the different species of mullet depending on whether the semi-intensive or intensive method was employed. Fig. 4.Comparison between the survival percentages obtained in intensive and extensive rearing (first year) of the different species reared in valliculture. Fig. 5."Pond" ecosystem diagram; the principal factors Fig. 6.Ponderal increase of M. cephalus mullet reared while diets of different proteinic content were employed. Fig. 7.Ponderal increase of O+ fed mullet while diets of different pro-teinic content were employed. Fig. 8.Comparison between the ponderal increases of the different species of mullet (fry) reared while diets of different proteinic and glucidic content were employed. Fig. 9.Composition of the different diets employed in the experiment described in fig. 8. Fig. 10.Graph attesting the waste percentage of the food given to mullet depending on the size of the particles (above) and the percentage of use of proteins also depending on the size of the particles of food (below). Fig. 11.Experimental data of a trial in the aim of verifying the efficiency of the filtration system of mullet. Fig. 12.Table of the results (average value) of the tests illustrated in fig. 11. Fig. 13.Data of a trial in the aim of verifying the influence of the systems of the food distribution on the increase and conversion index. Fig. 14.Distribution of prey frequency for mullet depending on their size. Fig. 15.Idem fig. 14 but for L. ramada captured in sea water environments. Fig. 16.Idem fig. 14 but for L. ramada captured in fresh water environments. Fig. 17.Idem fig. 14 but for L. ramada captured at the outlet of an intensive facility for sea-bass. Fig. 1. Specific characteristics of the different species of mullet adults)

Fig. 2. Growth curves of mullet in extensive valliculture

Growth curves classed as "good" for the extensives of Venitian "valle". In the different "valli" the rearing cycle must be prolonged so as to obtain these results. (Seeding is carried out in March - April using fry of 2 - 4 cm. These graphs do not permit the definition of the decrease in weight during Winter.) Fig. 3. Growth curves obtained for different species of mullet while employing the intensive, semi-intensive method.

Semi-intensive rearing period.

Fig. 4. Comparison between the survival percentages obtained in intensive and extensive rearing (first year) of the different species reared in valliculture.

Fig. 5. Drawing of a "pond" ecosystem. The principal factors are: I. Organic and inorganic abiotic substance II. A. Generators - Vegetation with roots II. B. Generators - Phytoplankton III. 1 A. Primary consumers (Herbivora) - On the bottom III. 1 B. Primary consumers (Herbivora) – Zooplankton III. 2. Secondary consumers (Carnivora) III. 3. Tertiary consumers (Secondary carnivora)

Fig. 6. Ponderal growth of M. cephalus fry reared while employing diets of different proteinic content

Fig. 7. Ponderal increase of 0+ mullet fed for 20 weeks diets of different proteinic content

Fig. 8. Comparison between the ponderal increases of the different species of mullet (fry) reared while employing diets of different glucidic and proteinic content.

PROT. LIP. EST. INAZ. A B A B A B 10% 9.88 - 10.66 10.21 - 9.87 6.83 - 76.07 20% 20.13 22.28 11.06 9.64 11.87 64.68 30% 30.39 32.67 10.46 9.19 17.16 51.25 40% 43.00 44.15 10,24 10.06 19.85 38.56 50% 54.00 55.27 10.20 10.78 26.04 29.70 Fig. 9. Composition of the different diets employed in the experiment shown in fig. 8.

Fig. 10. A graph showing the percentage of waste from the food given to the mullet, depending on the size of the particles (above) and the percentage of use in proteins, also depending on the size of the particles of food (below). Size of the inert particles (μ) < 50 50 - 100 100-250 250-500 500-1 000 - 14.6 - 27.7 - 67.9 + 114.0 + 60.1 - 28.7 - 29.7 - 42.4 - 56.2 - 6.6 - 61.6 - 2.0 - 48.5 - 26.6 - 58.1 + 90 0 - 21.8 - 10.8 - 64.1 - 54.1 - 64.4 - 61.8 + 87.5 + 16.9

< 50 - 14.8 - 15.3 - 57.4 - 63.7 - 48.9 + 56.5 + 10.4 - 23.5 - 23.3 - 18.3 - 38.2 - 39.6 - 27.3 + 40.2 + 26.4 - 29.2 + 22.7 + 115.5 + 7.1 71.6 + 72.6 + 37.1 + 34.9 - 32.2 + 0.4 + 57.9 +100.4 193.1 +213.9 + 28.6 + 36.8 - 25.8 + 003.9 + 1652 + 43.4 182.5 +175.1 + 31.7 + 31.9 - 9.0 + 35.7 382.3 50 - 100 + 22.7 + 59.9 + 62.8 - 11.4 - 6.5 + 70.2 + 9.8 + 53.3 + 722 +135.7 + 76.3 + 96.6 + 113.7 - 17.3 - 22.7 + 163.7 + 174.9 + 123.8 +150.7 +134.4 +118.9 + 95.1 + 67.1 - 41.4 - 42.4 + 32.1 + 67.8 + 200.5 + 10.7 + 97.0 +210.2 + 121.8 + 0.4 - 30.8 - 23.5 + 68.9 + 111.8 + 163.4 +186.4 + 1763 +117.8 + 239.4 + 4.3 - 9.0 +139.3 100-250 + 43.8 + 184.5 + 73.3 + 40.9 + 46.4 - 56.4 - 50.4 + 87.1 + 66.6 + 38.6 + 169.6 + 45.7 + 54.7 + 45.4 - 44.5 - 0.8 + 188.9 - 28.0 + 44.9 + 95,9 +123.1 + 21.2 - 16.0 - 42.8 + 55.3 + 61.3 + 79.6 + 241.7 +253.2 + 70.9 - 49.3 - 27.0 + 20.6 + 41.3 + 128.5 + 2532 250-500 + 24.5 - 773 - 47.5 + 64.6 + 46.4 + 68.4 - 32.6 4.0 + 155.6 + 83.4 + .12.3 - 49.5 - 16.0 - 12.0 - 63.3 31.1 + 83.8 - 15.9 + 71.7 + 2.3 + 45.4 + 64.0 - 14.6 + 13.5 - 11.5 +154.9 +110.0 - 18.5 - 49.3 - 53.2 - 55.0 + 8.7 + 126.2 +139.3 +201.1 500-1 000 + 43.4 63.8 + 108.2 +161.5 +138.7 Size of the particles of the food Fig. 11. Experimental data of a test carried out in the aim of verifying the efficiency of the filtration system of mullet. Fig. 12Tableoftheresults(averagevalue)fromtestsshowninfig.4 Size of the particles of food (μ )

500-2000 250-500 100-250 50 –100 < 50 < 50 + + + + - 42.4 76.3 33.4 213 3.2 50 -100 - - - - - 18.3 24.5 24.1 62.7 6.1 Size oftheinertparticles( 100-250 + + + + - 22.6 75.2 95.7 40.7 16.1

250-500 + + + + - 120.1 39.0 μ 51.1 81.6 915 ) 500-2000 + + + + + 100.7 116.6 119.7 184.3 53

Average weight Average weight Conversion increase (%) Index Feeding system Init. Final Dry food on the surface 9.8 24.1 145.9 2.42 - automatic feeder

The same as above + 10.4 32.7 214.4 2.73 sand 72.9 180.4 149.2

The same as above 12.8 29.8 132.8 2.82 with small and big fish

Paste on the bottom 9.7 19.5 101.0 3.32

Paste with sand on the 9.9 20.0 102.0 3.72 bottom with small and 59.0 128.3 117.4 big fish

Paste on the bottom + 13.3 27.8 109.3 4.25 mud

Fig. 13. Data from a test carried out in the aim of verifying the influence of the feeding systems on the growth and conversion index.

Fig. 14. Frequences of the distribution of prey for mullet depending on their size.

Fig. 15. L. ramada Sea water

Fig. 16. L. ramada: Fresh water

Fig. 17. Sea bass intensive rearing discharges TROPHIC COMMUNITY CORRELATIONS IN FERTILIZED PONDS FOR THE COMMERCIAL CULTURE OF THE KURUMA PRAWN Penaeus japonicus Bate IN THE NORTH ADRIATIC SEA (North-East Coast of Italy) LUMARE F., ANDREOLI C., GUGLIELMO L., MASELLI M.A., PISCITELLI G., SCOVACRICCHI T., TOLOMIO C. INTRODUCTION The Penaeus japonicus, a penaeid of Asian origin which was introduced into Italy and acclimatized in 1979 (LUMARE and PALMEGIANO, 1980), is now considered to be one of the most interesting species for Italian and Mediterranean aquaculture. Italy's environmental features and advanced bio technological knowhow make it a favourable environment in which to develop prawn culture using this species (LUMARE, 1983; LUMARE, 1983 a). One of the most promising approaches to prawn culture is through extensive breeding in fertilized ponds to increase the final production levels. An initial experiment has been conducted along these lines in a small area of a "valle da pesca" (Italian traditional extensive fish farm) in the Venetian lagoon (1 ha), which gave a final yield of 294,2 Kg after 5 months' management (LUMARE et al. , 1984). This paper is a tentative initial interpretation of the main correlations at various levels in the trophic chain, in which the final link is prawn production, and is intended to provide preliminary indications for production purposes. MATERIALS AND METHODS The research was conducted in two "valli da pesca" in the Venetian Lagoon (fig. 1). The first, Valle Sparesera, covered 11 ha, and was divided into smaller ponds (fig. 2) in which the bed sloped at a gradient of about 0,40 %. The low water replacement rate was due to the weak effects of the tide (ponds A, B, C) and, in the case of ponds D end E, it was partly due to the fact the pump was only brought into service at the end of June, to ensure an exchange rate of 1 % of the whole volume par day. The bottom of the ponds were very high in sand content (45 ù) and low in organic matter (0,9 - 1,1 %). In the second area, in Valle FOSSE, a 1 ha square pond was dug, shored up with earth and fitted with grilles for water replacement. The bed has a high lime content (70 %) and a high organic matter content (4,3 %). A pump provided a total water replacement rate of 1,5 % A considerable time before sowing, the Valle SPARESERA ponds were completely drained to eliminate predators; since the pond in the Valle FOSSE could not be completely drained, sodium hypochlorite was added to remaining pools (0,015 litres/m2 ). About a month before the Penaeus japonicus post-larvae sowing operation (table 1), the fertilization programme was started, following the schedule and using the quantities given in Fig. 3. Initial organic fertilization was effected in very shallow water (10 - 20 mm), while the subsequent phases were effected on the full ponds (0,60 - 1,30 m). During fertilization, water samples were taken at regular intervals (every 1-4 hours for 1,5 - 3 consecutive days) at a preselected station on each pond, to measure the phosphate, nitrate, nitrite, ammoniacal nitrogen, 02 , pH and salinity levels and the temperature. Every 15 days, in addition to the physical and chemical readings, phyto and zoo- plankton samples were taken from each pond. The phyto-plankton was collected at a depth of about 20 cm, and fixed in 4 % neutralized formalin. The quantitative and qualitative study was conducted using the UTERMOL method, to calculate the cellular volume (ANDREOLI and FRICANO, 1983) and the wet biomass. The zooplankton was collected by skimming the surface with a 200 urn mesh net, and was fixed in 4 % neutralized formalin; Subsanples were analized to identify and count the species using the "DOLLFUF curve". The dry weight and wet weight were established using the technique described by LOVEGROVE (1966). Phyto and 'zoo- plankton turbidity levels were periodically measured using a SECCHI disk, to form the partial basis for the fertilization programme. Monthly measurements of the biotic communities on the pond bottom were taken on the basis of three samples taken from a station on each pond using a 0,04 m2 VAN VEEN grab. The samples were passed through a serie of sieves with a minimum mesh of 0,5 mm. The organisms collected were fixed in 4 % neutralized formalin and then classified, counted and measured in terms of dry and wet biomass. At various stages in the breeding process, penaeid samples were taken and immediately fixed in 4 % formalin; Their stomach contents were examined and a qualitative analysis of their prey and percentage frequency was effected. Penaeid samples were periodically taken for the biometric analysis of sexual maturity levels, impregnation rates and sex ratios. Periodic water temperature measurements were also made to establish the effect of temperature changes as a conditioning factor on the growth of the penaeids. The prawns were harvested using netted traps and an electric fishing system beginning early September in ponds D and F, and in October in the others, through to the end of November. RESULTS Fig. 4 gives the ammoniacal nitrogen, nitrite, nitrate and orthophosphate levels in the water of pond D in Valle SPARESERA. This pond was also considered representative of the other ponds (A, B, C, E) of which it reflected the general patterns, with slight variations. Fig. 5 gives the readings for these parameters for pond F in Valle FOSSE, where fertilization was only effected at the beginning of the breeding phase. Fig. 6 gives the temperature readings in pond D. But this situation may also be considered indicative of the other ponds in the two "valli da pesca". The same applies to the pH readings, which varied between 7,9 and 9,4 averaging 8,4, while salinity ranged from 26,6 to 38,5 %. Fig. 7 gives the quantitative variations in the wet weights of the phyto and 200- plankton and the corresponding chlorophyll a quantities in pond D in Valle SPARESERA; the same parameters are considered in fig. 8 in relation to pond F in Valle FOSSE. Periodic water transparency readings were taken on the ponds when conditions were calm, to ensure that the reading expressed the quantities of phyto and zoo- plankton. A SECCHI disk was used to provide an empirical assessment of the state of eutrophication of the ponds, to relate this to prawn production and to see if it might in future be used by prawn farmers. Fig. 9 gives the water transparency reading for ponds D and F during the experimental period. Fig 10 gives the histograms for the biomasses of the biotic communities on the pond bottoms, expressed in wet weight, allowing for the metabolized inert substances (molluscs shells and valves). Account was also taken of the edible fraction of these alone, namely the soft parts of Hydrobia sp and Cerastoderma glaucum (Brug.). With regard to the latter, only sizes below 3 mm were considered, being the size which the penaeid could attack because of their soft valves and because they are only found in the Valle SPARESERA up to the end of May. In the following months, although C. glaucum were plentiful in the macrobenthonic communities in Valle SPARESERA, they did not form part of the P. japonicus diet because they were generally larger than 3 mm . In Valle FOSSE, Hydrobia sp constituted the main component of the fraction of mollusc utilized. Table II gives the percentage composition of the prey found in the stomachs of the P. japonicus. It shows the different natural diets, which reflects the different structures of the macrobenthos in the two "valli". Fig. 11 shows the growth pattern of the penaeid populations in the two ponds considered (D and F). One relevant finding is the fact that the size in terms of weight of the pond D population dropped during the final breeding phase, which is striking proof of the scarcity of trophic resources in the environment. This situation is corroborated by the final weights given in table III, which shows the production of penaeid in the ponds (A, B, C, D, E, F) in both "valli da pesca". The sex ratio of the different populations of P.japonicus was around 1 (table III), with a slight predominance of females; the-impregnation rate (ie. the presence of spermatophore in the thelycum) was generally low in comparison with the 98 % found under normal breeding conditions (LUMARE, unpublished data). The percentage of females at an advanced stage of ovarian maturation (stages III - IV) was also very low indeed (0 - 8,6 %) . DISCUSSION Water fertilization is practised in many forms of fresh-water fish culture (MOAV. R. et al., 1977; WAHBY, 1974; BISHARA, 1978; BISHARA, 1979) and sea water fish culture (CHEN, 1972). It is also practised in many forms of prawn farming; for example, the farming of Penaeus vannamei Boone and P. styli rostris Stimpson in Central America, and P. monodon Fabricius in polyculture in the Philippines (ELDANI and PRIMAVERA, 1981). For penaeid production, KITTAKA (1975) emphasizes the value of this practice, coupled with predator control and improving the pond bed conditions. In Italy, water fertilization was successfully adopted for the first culture experiments with P. japonicus (LUMARE end al., 1984). It is vital to gain a better understanding of the relationship between the features of the pond bed, fertilization, and productivity at the various trophic levels, and final production, if this production approach is to be properly planned. The two areas used for the experiment were therefore chosen in terms of the differing features of the pond beds, and different fertilization programmes were designed accordingly. THe effect of this diversity was measured more directly throught the torbidity of the phyto and zooplankton. Yet this diversity was not matched by the expected diversity in the sizes of the phyto and zoo-plankton populations, because pond D was surprisingly found to have a larger biomasses than pond F. Probably one of the main reasons for this was the part played by the microbenthonic populations that were not measured in the experiment, although they will have to be taken into account in future. The macrobenthonic biomasses also appeared to be larger in pond D, yet for the purposes of counting the biomass relevant to the trophism of the penaeids, the curve superimposed on the histogram in Fig. 10 shows that the trophic base was larger in pond F (the average size of the biomass, calculated on the basis of the last three readings, was 113,6 Kg/ha, compared with 41,2 Kg/ha in pond D). To calculate this figure, the sizes of the zoological groups which were wholly utilizable were taken into account (mainly annelids, crustaceans, insects) and only the soft parts of the attackable molluscs (Hydrobia sp and others). No account was taken of the soft parts of the percentage of Cerastoderma glaucum which were too large (over 3mm) to be used by the penaeids. A comparison of the stomach contents of the penaeid populations of the two ponds (D and F) also revealed different diets correlated to the differing biocetonic composition of the pond bottoms, probably due to the specific pedological features of the soil (high sand content in pond D, mainly lime in pond F, and different percentages of organic matter). Table II shows that Chironomidae and Capepoda make up 81,5 % of the penaeids'diet in Valle SPARESERA, while Hydrobia sp and Corophiidae account for 82,4 % of the diet in Valle FOSSE. The differing quantities of macrobenthonic biomasses in the two environments are also reflected in the final production in each case In this connection, it should be noted that the final yields (Kg/ha) in the two intercommunicating ponds D and E were different. This might be due to the fact that harvesting began on 3 September in pond D, while in pond E it began about one month later. Harvesting early in the first pond thinned out the population and provided a better distribution of the trophic resources, and hence reduced cannibalism which was much higher in pond E, and in nearly all the others. One may assume that the pattern of the environmental parameters of the water and the related trophic situation ensured sufficient nutritional levels in terms of the size of the penaeids, almost until September in pond F, and only until the end of July in pond D. In the following periods, the macrobenthonic populations were no longer able to meet the penaeids' feeding requirements (the daily requirements is about 20 % of their body weight, at the size and temperature levels considered here), thus causing cannibalism. This was even more evident in ponds which had a higher sowing density. In pond F, growing out trials with a lower sowing density (1,8 P 23/m2) had yielded a recovery rate of 48 % the year before (LUMARE and al., 1984) Proof of the difficult dietary situation in Valle SPARESERA in the final breeding phase was the softening of the penaeids'carapax, attributable to the low content of mineral salts. More evident proof of this was the unusually low impregnation percentages, except in pond F, which was further evidenced by the extremely low ovarian maturation rate (stages III - IV) in all the ponds, whereas in a previous experiment, pond F has produced an 84,6 % rate (LUMARE and al., 1984). ACKNOWLEDGEMENTS The authors wish to thank the Department of Ecology, the Environment and Fisheries of the VENICE Provincial Government for having placed "Valle SPARESERA" and the "Valle FOSSE" ponds at their disposal, and for their invaluable help in the management and harvesting operations. They also thank the Venetia Fisheries and Aquaculture Development Consortium CoSPAV), CHIOGGIA, for its logistical support, and Mr. G CASOLINO, of "Istituto per lo Sfruttamento Biologico delle Lagune - CNR", LESINA, for his help with the graphics. Literature Cited ANDREOLI C. e G. FRICANO - 1983 - Il Po: Ulteriori osservazioni di densità e di biomassa del fitoplancton nel tratto di fiume prossimo alla centrale termo-nucleare. di CAORSO (Piacenza). Un ciclo annuale (settembre 1980 - agosto 1981). Riv. Idrobiol. (in press). BISHARA, N.F. - 1978 - Fertilizing fish ponds. II - Growth of Mugil cephalus in Egypt by pond fertilization and feeding. Aquaculture, 13; 361 - 367. BISHARA, N.F. - 1979 - Fertilizing fish ponds. III - Growth of Mugil cephalus in Egypt by pond fertilization and feeding. Aquaculture, 16; 47 - 45. CHEN. T. P. - 1972 - Fertilization and feeding in coastal fish farms in Taiwan. In "Coastal aquaculture in Indo-Pacific Region". PILLAY T.V.R. (Ed.); 410 - 416 ELDANI A. and J.H. PRIMAVERA - 1981 - Effect of different stocking combinations on growth production and survival of milkfish (Chanos chanos (Forskäl) and prawn (Penaeus monodon Fabricius) in polyculture in brackishwater ponds. Aquaculture, 23, 59 - 72. KITTAKA J. - 1975 - Food and growth of Penaeid shrimp. Proceedings of the First International Conference on Aquaculture Nutrition, October, 1975; 249 - 285. LOVEGROVE T. - 1966 - The determination of the dry weight of plankton and the effect of various factors on the values obtained. Some contemporary studies in marine science. Ed. H. Borners, LONDON. LUMARE F. - 1983 - Italy farms kuruma prawn; Fish farming International, 10 (3); 10-11 LUMARE F. - 1983 a - Italian valliculture and its future development. Rapp. Comm. int. Mer Medit.., 28 (6); 85 - 89. LUMARE F. e G.B. PALMEGIANO - 1980 - Acclimatazione di Penaeus japonicus Bate nella laguna di LESINA (Italia Sud-Orientale). Riv. It. Piscic. Ittiop. A XV, 2; 53-58. LUMARE F. , T. SCOVACRICCHI , G. PISCITELLI, G. GRASSO - 1984 – Prime esperienze di allevamento commerciale a gestione controllata del peneide Penaeus japonicus Bate in una valle da pesca nella laguna di VENEZIA; XVI Congr. SOc. It. Biol. Mar., 25 - 30 settembre 1984, LECCE; 1-18. MOAV R., G.WOHLFARTH, G.L. SCHROEDER, G. HULATA and H.BARASH - 1977 – Intensive polyculture of fish in freshwater ponds. I. Substitution of expensive feeds by liquid cow manure. Aquaculture, 10; 25-43. CAPTIONS Fig. 1. Map of the Venetian Lagoon, showing the locations of Valle SPARESERA (I) and Valle FOSSE (2). Fig. 2 Planimetric map of Valle SPARESERA showing the ponds used for the production experiment. The arrows indicate the direction of water flow caused by the tide and pumping system. Fig. 3 Schedule for the pond fertilization program (ponds A, B, C, D,E, F) throughout the whole experiment, from pond preparation to just before harvesting. Fig. 4 Nutrient content measurements at the moment pond D was fertilized in Valle SPARESERA. The arrows indicate the dates on which the fertilizations were carried out. Fig. 5 Nutrient content measurements in Valle FOSSE. The arrows, left, indicate the sole date on which fertilization was effected (organic + inorganic) Fig. 6 Water temperature readings in pond D in Valle SPARESERA. Fig. 7 Histogram of the phyto and zoo-planktonic.biomasses given in wet weight, with corresponding chlorophyll a curve in Valle SPARESERA. It also shows the qualitative composition of the most representative species of the community. Fig. 8 Histogram of the phyto and zoo-planktonic biomasses given in wet weight, with corresponding chlorophyll a curve in Valle FOSSE. It also shows the qualitative composition of the most representative species of the community. Fig. 9 Transparency curves of the water in pond D in Valle SPARESERA and pond F in Valle FOSSE using the SECCHI disk. The arrows indicate the dates of fertilization; the last arrow, right, (coloured white), shows the final fertilization, which was not applied to pond D. Fig. 10 Histograms of the macrobenthonic biomasses given in wet weight in pond D, Valle SPARESERA, and pond F, Valle FOSSE; The qualitative composition of the main zoological groups are given for these biocenoses. The curves express the fractions of the biomasses actually used by the penaeids, namely, annelids, crustaceans and insects taken together, and the soft parts of the mollusks that they are able to attack (Hydrobia sp and others), and the Cerastoderma glaucum under 3 mm in size. Fig. 11 Curves showing the weight increases of the populations of Penaeus japonicus in pond D and F in the Valle SPARESERA and Valle FOSSE, respectively

- FIG. 1 -

- FIG. 2 -

- FIG. 3 -

- FIG. 4 -

- FIG. 5 -

- FIG. 7 -

- FIG. 8 -

- FIG. 9 -

- FIG. 10 -

Fig. 11

Pond marks A B C D E F Surface (ha) 0,67 0,64 0,65 4,50 4,96 1,00 7.5.84 7.5.84 Sowing data 12.6.84 12.6.84 12.6.84 12.6.84 11.6.84 11.6.84 Total number 7,060 13,546 19,835 117,080 130,280 25,205 post- larvae Post larvae Density (sp/m2 ) 1.0 2,1 3,0 2,6 2,6 2,5

Tab. I - Sowing programme of Penaeus japonicus post larvae (P23 27) in Valle SPARESERA (A,B,C,D,E ponds) and in valle FOSSE (F pond).

Species Valle SPARESERA (D pond) Valle FOSSE (F pond) FORAMINIFERA 9,88 0,30 ANNELIDA 6,36 4,80 Spionidae 6,30 Nereidae 0,06 4,80 MOLLUSCA 0,60 45,1 Hydrobia sp 0,40 41,80 Cerastoderma glaucum 0,20 0,80 Abra sp 1,10 Unidentified 1,40 CRUSTACEA 20,40 47,20 Ostracoda 0,40 0,20 Copepoda 18,80 Mysidacea 0,30 2,90 Amphipoda Corophiidae 0,30 40,60 Gammaridae 2,40 Isopoda Idoteidae 0,50 Decapoda Palaemonidae 0,20 0,20 Unidentified 0,40 0,40 INSECTA 62,70 2,20 Chironomidae 62,70 2,20 PISCES 0,06 0,40 Gobidae 0,06 0,40 Tab. II - Percent frequency of preys in stomach contents of Penaeus iaponicus stocks in Valle SPARESERA (D pond) and in Valle FOSSE (F pond) reared. Different qualitative pattern of preys are related to macrobenthonic biocenosis differentiation in the two "valli da pesca".

Pond marks A B C D E F weight (g) 0,037 0,037 0,037 0,040 0,040 0,043 Initial size lenght (cm) 1,73 ± 0,36 1,73 ± 0,36 1,73 ± 0,36 1,71 ± 1,57 1,71 ± 1,57 1,71 ± 1,63 Initial number specimens 7,060 13,546 19,835 117,080 130,280 25,205 Rearing period (days) (1) 111 111 111 108 135 84 Final number specimens 1,805 2,619 3,203 46,134 25,376 9,696 Catch rate {%) 25,5 19,3 16,7 39,4 19,5 38,5 weight (g) 23,28 ± 2,76 11,47 ± 1,69 15,00 ± 2,04 24,13 ± 3,74 22,57 ± 3,92 35,58 ± 6,64 Final size lenght (cm) 14,57 ± 0,54 11,69 ± 0,49 12,80 ± 0,56 14,90 ± 0,58 14,58 ± 0,77 16,33 ± 0,89 Final production (kg/ha) 61,90 46,90 75,10 257,90 118,30 345,00 Actual Final production per pond (kg) 41,50 30,10 48,80 1 160,80 586,40 345,00 Sex ratio (♀♀:) 1,3: 1 1,0: 1 1,0: 1 0,98: 1 1,0: 1 1,0: 1 Impregnation rate (%) (99) (87) (87) (117) (131) (104) 68,7 0 2,3 67,5 65,6 85,6 Ovarian maturation rate (%) (III - IV stage) 0 0 0 2,6 0 8,6

Table III - Biometric, biological and production data on the Penaeus japonicus bred in Valle SPARESERA (ponds A,B,C,D,E) and Valle FOSSE (pond F). The impregnation rate and ovarian maturation rate have been calculated as the average of the last four samples (September - October). The number in brackets indicate the numerical size of the samples. (1) The breeding period in days, from sowing date to the date the harvesting of the penaeids began. NEW REARING TECHNIQUES AND LARVAE QUALITY IMPROVEMENTS IN SEA-BASS (Dicentrarchus labrax) HATCHERIES MEREA: Equipe Méditerranéenne de Recherche Aquacole (Mediterranean Aquaculture Research Team) Mr. D. COVES 1. Introduction From 1974 to 1982, large scale sea-bass rearing has permitted the production of several hundreds of thousands of fry per year in French hatcheries. However, the rearing results were incertain, irregular and poor, with survival rates ranging between 0 and 40 % after two months. Therefore, producers were forced to use big rearing volumes and great quantities of larvae to ensure sufficient production. Since 1983, the PALAVAS experimental centre has perfected rearing techniques and so larval behaviour, survival and animal, productions are better. As this technique proved to be reliable at semi-intensive and intensive pilot scale it is now possible to propose technological norms and estimate production costs for this phase of sea-bass rearing. 2. Review of rearing conditions and block points with the "green water" technique: 2.1. General conditions of rearing The feeble density larval rearing technique known as the semi-intensive "green water" type has been described by BEDIER (1979). It is distinguished by the principal parameters here following: a) Initially low concentration in rearing with about 20 larvae per liter in a black tank of 10 m3. b) The rearing environment is stagnant with the phyto-planktonic bloom for the first 20 days. The water renewal is carried out progressively little by little. c) The artificial rearing, with which mercury or fluorescent lighting is employed gives intensities of 2 000 to 5 000 lux at the surface of the tanks when hatching starts. 2.2. Anomalies encountered Two principal anomalies occur frequently at this point: a) An anomalie in behaviour is remarked from the 20th to 30th day in rearing (temperatures of 18 to 20° C). It is distinguished by fits of giddiness together with loss of appetite and the production of white faeces. In some cases these symptomes are followed by the death of the whole stock. b) Anatomical anomalies. On one hand, some rearings present a high percentage of individuals having a malformation of the mandibular arch (prognathic, bilateral or simple operculata); on the other hand, a skeletal lordosis type malformation appears in 80 to 90 % of the animals when they reach 20 cm in lenght (CHATAIN in process) So many weak fish, greatly reduce rearing preformances and survival rates at weaning stage are very average (CHATAIN in process). In this last case there is a correlation between the presence of a lordosis and the absence of a functional gassy bladder (CHATAIN in process) 3. Solutions to block points 3.1. A pragmatic approach. From 1984 onwards, the overall improvement of the quality of the rearing environment has been elaborated. A pragmatic approach was first employed and this technique led to the reproduction of larvae in natural living conditions (COVES in process, COULET, 1985) In consequences, the "green water" was abolished and in place there is a continuous water renewal from the beginning of rearing. The artificial light (neon) was replaced by a natural light of more feeble intensity (maximum 1 000 to 1 500 lux from November to March) and so discontinuous (9 to 12 hours per day) during the rearing season. On the principle of this method, two rearing techniques are employed simultaneously. One of these techniques known as the intensive, is carried out in tanks of 2 m3 with great quantities of larvae at the beginning (50-100 larvae per liter) the other technique known as the semi-intensive, in tanks of 10 m3 with small quantities of larvae (10-25 larvae per liter) (tab. n° 1, page 6) The results obtained at pilot scale for both techniques are very significant right away. a) The fits of giddiness stop completely and mandibular and opercular anomalies disappear. b) Survival rate progresses from 15 % to 40-50 % on average (fig. n° 1, 2, 3, 4 and tab. n° 2 page 7, 8, 9, 10, 11). c) The results obtained on all the tests show that this method is reliable when the purely technological problems are under control (pumping, heating). d) The average percentage of animals having a functional air bladder is greater but results vary a lot still. (fig. n° 1 and 3 and page 7 and 9, tab. n° 2 page 11). 3.2. Experimental approach: On parallel with the pragmatic attempts carried out at pilot scale, many tests have been tried out at experimental level in small pay load cylinder-conical units of 500 liters. In 1985, the results described here following have been transferred to intensive pilot scale so a new and extremely reliable rearing technique could be started. 3. 2. 1. Influence of colour of the tanks The comparaison is carried out between white walled and black walled tanks. The results obtained are clear. Walls painted in black encourage better behaviour, predation, growth and survival (RONZANI in process). 3.2.2. Influence of the incident light parameter The different tests carried out in black walled tanks having incandescent artificial lighting showed the important part played by the light parameter on the quality of the rearing. a) The photoperiod Constant lighting handicaps the inflation of the gassy bladder and doesn't permit good consumption of prey during the interval which corresponds to the natural night period ( RONZANI in completion). b) Light intensity During a first experiment the 70 lux-1 800 lux scale was tested for the first 40 days of rearing. The intensity tested was constant for 9 hours out of 24 hours. Only the 1 800 lux doublet shows abnormality in behaviour and giddiness from the 20th to the 30th day (WEPPES, JOASSARD in process). During a second series of experiments, the 50-5 000 lux scale was tested during the first 20 days of rearing. The correlation was found between the light intensity and the rate of the gassy bladder inflation (JOASSARD in process). 3.2.3. Influence of the environment and the feed sequence Recent experiments have permitted to define a new method of rearing by controlling the environment and the feed sequence, during the first fortnight of rearing. This technique finally gives the possibility of obtaining good rates of the gassy bladder inflation from the 12th day onwards, ensuring a good quality fingerling production without any malformation. Tested at intensive pilot scale, this method gave reliable results from a quantity point of view (survival between 35 and 55 %) and a quality point , of view (normal fingerling rates between 75 and 95 %) (fig. n° 3, page 9). 4. Technico-economical norms The numerous tests at experimental level and the reliability of the results obtained when transferred to pilot level have permitted the estimation of the technico- economical norms. These are all the more realistic as these calculations concern the rearing of about 1 million larvae of 45 to 50 days of age produced in 12 tanks of 2 m3 each, carried out in 3 cycles during the 1985 season (fig. n° 4, page 10). These norms form a tool of efficient programmation more than the economic balance of a production technique would. These figures show clearly that the cost of production, not counting the depreciation of a 45 - 50 day old larvae is relatively low, 0,12 FF; and that only a drop in the cost of feed (which is 90 % per price of cysts) could permit a real reduction in the cost of this phase of sea-bass rearing (tab. n° 3, page 12 - 13). 5. Conclusion Two years of zootechnical research work has permitted to raise the important block points which has hindered sea-bass rearing. This break through now permits every potential producer, to organize clearly the management of his enterprise. The next step to be taken is the productivity returns which will stay relatively low as long as Artemia is not replaced by artificial feed. Adress: Chemin de Maguelone 34 250 - PALAVAS LES FLOTS FRANCE TABLE N° 1: REARING METHOD. 1984 INTENSIVE SEMI -INTENSIVE - volume (m3 ) 2 10 - 15 - shape cyllndro-conical circular TANK - colour black black ( whitebottom ) ONE DAY OLD LARVAE PER LITER 5 0 - 100 10 - 25 - period ( h) 9 - 12 LIGHTING [natural] - maximal intensity 1000 - 1500 (lux ) quality opened circulating system -renewing rate 5 30 – 50 WATER (% total volume.h-1) (D1) (D45) - temperature(°C) 14 18- 22 ( D1) (D45) AERATION (liter. mn-1m3) 0.4 3.0 (D1) (D45) - Brachionus plicatllis D 6 - D 12 (led on yeast and algae) -Artemia nauplii D 9 - D 20 PREYS ( San Fransisco ) - Artemla metanaupIIi D 16 - D 45 (led on dry compounded powder )

Time (years) Figure N° 1:- Low density sea-bass larval rearing, Dicentrarchus labrax Evolution of survival and of normal air bladders rates after the first 50 days during the 1976 - 1984 period.

Figure N° 2: - Low density sea-bass larval rearing, Dicentrarchus labrax Evolution of yulds and productions obtained after the first 50 days during the 1976 - 1984 period.

Figure N° 3: High density sea-bass larval rearing, Dicentrarchus labrax Evolution of survival and normal air bladders rates after the first 50 days during the 1981 - 1985 period. * Result of the first cycle 1985 ** Result of the second cycle 1985 *** Result of the third cycle 1985

Figure N° 4: High density sea-bass larval rearing, Dicentrarchus labrax Evolution of yulds and productions obtained after the first fifty days during the 1981 - 1985 period. Table N° 2 AVERAGE PILOT PRODUCTION RESULTS, 1984

3 METHOD rearing quality totaI survival rate D45 swimbladder inflation total fry number per m total normal fry3 number (%) rate D45 (%) D45 per m D 45 SEMI 1 57.0 48.5 10270 4980 - INTENSIVE 2 57.0 48.5 10270 4980 1 22.5 28.0 21100 5890 INTENSIVE 2 44.5 23.0 44300 10080

* with an inflated swimbladder 1 means obtained from all attempts 2 means obtained from attempts realised without any trouble Table N° 3 SEA-BASS LARVAE REARING TECHNICO ECONOMIC RESULTS, 1984

INTENSIVE SEMI-INTENSIVE REARING METHOD AND RESULTS SUMMARY

* TANKS VOLUME (M3) 2 10 * D1 LARAVAE A LITER 100 18 * SURVIVAL RATE (%) 44 57 * TEMPERATURE 18 18 * RENEWING RATE (%) 5 → 50 3 → 30 .-1 -3 * AERATION (l.MN .M ) 0.4 → 3.0 0.4 → 3.0 * PREYS ROTIFERS ► ARTEMIA ROTIFERS ► ARTEMIA

10 000 D45-50 LARVAE COST % % COST PRODUCTION COST (FF) (FF)

D1 LARVAE 39.1 2.4 1.9 30.9

* Rotifers 19.6 174.1 PREYS * Nauplii 25.0 56.3 77.8 3.7 * Metanauplii 866.8 1069.8

* Heated water 28.6 66.7 FLUIDS * Normal water 0.9 1.9 4.3 1.8 * Air 0.1 0.3

* General 205.0 198.0 LABOUR 39.5 16.0 * Counting 435.0 57.3

* TOTAL PRODUCTION COST 1620 1603 Table N° 4: Technico-economical norms of intensive sea-bass larval rearing 1985 1. TECHNOLOGY and RESULTS - Rearing enclosure = black cylindro-conical tanks, pay load: 2 m3 - Initial load = 100 larvae per liter - Survival rates = 40 % - Hourly renewal rate = 5 to 60 % - Aeration = 0.4 to 3,0 1. mm-1 m-3 - Feed = Rotifers then artemia

2. PRODUCTION COSTS Cost of rearing unit tank of Production cost of the whole 2 m3 stock: 100 000 J 45 Tax free francs Francs H.I% Manual labour 1 850 2 056 18 Animals 122 136 1 1 Feed and Processing 8 203 9 114 96 79 Fluids 266 296 3 2 Excluding Manual labour 8 591 9 546 100 Total general 10 441 11 602 100

BIBLIOGRAPHY BEDIER, E. , 1979 - Production à l'échelle pilote d'alevins de loup (Dicentrarchus labrax L. ) . Symposium on the early life - History of fish -Woodshole, USA, 2-5 Avril 1979. COULET, J.L., 1985 - Synthèse des données acquises sur l'élevage larvaire du loup (Dicentrarchus labrax). Technique semi-intensive et intensive. Rapport interne IFREMER, équipe MEREA - 27 pages. POST LARVAE PRODUCTION OF P. JAPONICUS INCREASE IN PRODUCTIVITY BY MODIFYING THE FEED SEQUENCE TEAM MEREA Mr G. Le MOULAC The reproduction of Penaeus japonicus has been studied for years. The Mediterranean climatic conditions in France favours the rearing of this species. The production of post-larvae in hatcheries is easy. The pathological problems (black branchiae disease) caused by maintaining brood-stock in captivity, have been resolved. Survival rates for broodstook are good at the end of a nine month period (60 %). Maturation is obtained at 18° C. Spawning takes place when the temperature is raised abruptly to 25° C. Larvae rearing is carried out at 26° C while supplying mono specific algae cultivated in algae rooms and Artemia nauplii when these reach Z3 stage. The final density is around 100 P3 per liter. Pre-fattening is carried out at 23° C. The feed comprises of Artemia nauplii and deep frozen feed. The initial density is 10 P3 per liter and there is a survival rate of 70 % at P23 stage. These established facts permit the proposition of an intensive production model in hatcheries and the technico-economic study relative to this. The definition of the productive cost gives the opportunity of having an efficiant programmation tool at disposal, to increase productivity. A 20 % increase on the production cost of post-larvae has been obtained, by modifying the feed sequence. N/Ref: GLM/MPD N° 85.05.306 INTRODUCTION The production of P, japonicus post-larvae is no longer a hindering factor to the development of shrimp culture in France. The technico-economic study shows clearly that this production is profitable. The techniques employed show good results. The progress envisaged now is at the increase level of productivity which should cut down the production costs of post-larvae. The first productivity increases quickly obtained concerned feed, by optimizing the output and by employing low cost feed. DESCRIPTION OF THE PRODUCTION CHANNEL 1) Stocking in tanks Brood fish stocking during Winter demands the abidance of a certain number of rules. 1.1. The origin of the brood fish (lagoons): To have at disposal fish which do not need to be fattened during Winter. 1.2. The temperature to be kept is 18° C which limits the growth of the animals. 1.3. Survival depends on the regular drainage of the tanks. 1.4. Fresh feed: mussels, crab, calmar. 1.5. Dividing up of animal stock: 2 females for 1 spawning with a ratio of 7 females for 3 males and a density of 15 animals/m2. 2) Spawing Mature animals at 18° C are placed into a tank at 25° C without having the peduncle ablated. – Sand bottom, important renewal, 180 000 nauplii spawned per female. – Spawning all year round. 3) Larvae rearing At high density, starting 220 nauplii per liter which show a 70 % survival rate at P3 stage. 3.1. The feed consists of unicellular algae: – Chaetoceros calcitrans – Phaeotachylum tricornitum – Monochrisys lutherii, through out the whole period of rearing and Artemia nauplii from Z3 stage onwards. 3.2. Feeble renewals of 10 to 20 % per day until P3 stage is reached. 3.3. A constant 1 000 lux light on the cylindro-conical tank of 2 m3 by means of a 150 w spot light. 3.4. Anti-fungic control - Constant drip of treflan. Anti-bacterial control - Furazolidone from Z1 stage, every 48 hours until Mysis 1 stage is reached. 4) Nursery - Prefattening 4..1. Having 10 P3 per liter during 20 days, survival is 70 %. 4.2. The feed consist of Artemia nauplii until P10 stage and from then deep frozen adult Artemia. 4.3. A 100 % renewal per day 4.4. A temperature of 23-24° C. 5) Technico-economic study of the channel Brood fish Nauplii P3 P23 Animals 11 % 75 % 36 % 26,5 % Heating 27,5 % 10 % 6,5 % 11,8 % Pumping 2,7 % 0,6 % / 1,2 % Air 0,3 % / / 0,6 % Feed 5,1 % / 41 % 56,5 % Manual labour 53,2 % 15,5 % 16,4 % 3,5 % Unitary Price Tax free 120,9 F/0 1,62/1 000 N 9/1 000 P3 48/1 000 P23

Table 1 - Analysis of the production cost (1) The facts established permit the economic study of the reproduction channel. The most important points in feeding are at larvae rearing and pre-fattening stages. Increases should be quickly remarked at these points. 6) Productivity increases 6.1. Prefattening In substituting deep frozen Artemia by Daphnie which costs 11F/kg while the former costs 48 F/kg. 20 % gain on the cost of P23 Brood Fish Nauplii P 3 P 23 Animals 11 % 75 % 36 % 34 % Heating 27,5 % 10 % 6,5 % 15,2 % Pumping 2,1 % 6 % / 1,5 % Air 0,3 % / / 0,7 % Feed 5,1 % / 41% 43,9 % Manual labour 53,2 % 15,5 % 16,4 % 4,5 % Unitary price Tax free 120,9 F/0 1,62/1 000 N 9/1 000 P3 38/1 000 P23

Table 2 - Analysis of the production costs productivity improvements in prefattening 6.2. Larvae rearing Survival of 60 % - 70 % (mixed algae in larvae rearing.) 30 % gain on the cost of a P3 10 % gain on the cost of a P23 Brood Fish Nauplii P 3 P 23 Animals 11% 75 % 36 % 25,3 % Heating 27,5 % 10 % 6,5 % 16,8 % Pumping 2,7 % 0,6 % / 1,8 % ir 0,3 % / / 0,9 % Feed 5,1 % / 41* % 50,3* ½ Manual labour 53,2 % 15,5 % 16,4 % 5 % Unitary Price Tax free 120,9 F/O 1,62/1 000 N 6/1 000 P3 34/1 000 P23

Table 3 - Analysis of production costs improvement of productivity in larvae rearing 6.3. Prefattening Compound dry feed. Conversion index = 3 unitary price = 30 F/kg 40 % gain on the production cost

Brood Fish Nauplii P 3 P 23 Animals 11 % 75 % 36 % 46,1 % Heating 27, 5 % 10 % 6,5 % 30,8 % Pumping 2,7 % 0,6 % / 3 % Air 0,3 % / / 1,5 % Feed 5,1 % 41 % 9,2 % Manual labour 53,2 % 15,5 % 16,4 % 9,2 % Unitary Price Tax free 120,9 F/0 1,62/1 000 N 6/1 000 P3 20/1 000 P23

Table 4 - Analysis of production costs improvement of productivity in prefattening 6.4. Evolution of gains on the production costs of post larvae. A gain of 20 % is remarked on the production cost of P23 post-larvae when deep frozen Artemia are subtituded by Dapnies. A 10 % gain follows with the use of an algae cocktail in larvae rearing. A 40 % gain can be scheduled in prefattening when granulars are employed. 1 2 3 4 Animals 26,5 34 25,3 46,1 Heating 14,8 15,2 16,8 30,8 Pumping 1,2 1,5 1,8 3 Air 0,6 0,7 0,9 1,5 Food 56,5 43,9 50,3 9,2 Manual labour 3,5 4,5 5 9,2 Unitary Price T.F. 1 000 38 38 34 20 P 23 Production cost % profit 20 % 10 % 40 %

Table 5 - Analysis of evolution of gains in productivity when the feed sequence is modified CONCLUSION From this point on, the gains on production costs are obtained with brood fish by recovering and cryopreservating fertilized eggs or Nauplii,so allowing the use of broodstock the whole year round. On larvae rearing and investiment (algae room) by the use of micro-particles INDUSTRIAL MANUFACTURE OF COMPOUND FOOD FOR MARINE SPECIES Mr. J. J. SABAUT The aim of industrial manufacture of compound food is to feed animals in rearing, while ensuring their maximum growth at the lowest production cost and preserving optimal hygiene. The perfectioning of compound food for aquaculture was started in the fifties and became widely used in the seventies with the development of salmonculture in fresh and marine water. Food is an essential element for all animal production. It can represent 40 to 60 % of the production costs, depending on the methods employed and the species reared. The constraints defining which food is to be manufactured and what industrial structures are to be built so as to ensure the coverage of the rearing needs, are as following: – knowledge of the species reared and of their food needs, – availability of raw materials: nature, price, stockage, renewals, – importance and divisions of the rearings, – variations in the seasonal needs concerning the quantity and quality of food. 1. Knowledge of the species reared and their nutritive requirements On the contrary to land animals, fish have great requirements of proteins. These are carnivorous species mostly, which require quality proteins, rich in the essential amino-acids. The proteinic requirements are shown in table 1 and 2. These species do not make good use of carbohydrate as a source of energy. Important imputs of fatty matter, rich in unsaturated fatty acids, ensures this requirement while at the same time giving the necessary fatty acids. Depending on the species, the fatty matter imputs vary from 8 to 15 % of the ration which comprehends 40 to 50 % of proteins. All compound food manufacturers, depending on their competance and especially on the technological capacities of their facilities, define the food ranges having specified characteristics. In table 3, the sea-bass food composition, commercialized by the Aqualim Company is shown, along with the table for the daily distribution of food, expressed in live weight percentages of fish. 2. Availability of raw materials Essential raw materials, employed in compound food for aquaculture have high proteinic content. For fish meal, along with the local production available in numerous countries, meal found on the world market is employed. This comes from big exporting countries: Norway, Danemark, Island, Chili, Peru. In any case, it is very important to define the type of fish meal, as their characteristics can vary considerably from one product to another. It is also necessary to ensure the non-oxydation of the constituent fatty matters, as when not verified, these products would be inadequate and even toxic for the animals being reared. Other proteinic matter of animal origin is also employed, such as meat meal, blood meal, hydrolysates, lactoserums, yeast, etc... Among the vegetable proteins, the most commonly employed, although in smaller amounts, are soya meal and whole cooked soya beans, distillery solubles, wheat or corn glutens, lucerns, cereals and wheat by-products. The diverse characteristics of these products are shown in table 4, 5, 6 and 7. For the manufacturer, the choice between all these products depends on the formulation objectives, the analytic characteristics, the price and supply possibilities. The calculations are carried out by means of a computer and optimization programmes which take all these constraints into account. All the products and in particular animal meal should be stocked properly, so to ensure perenniality of their nutritive qualities. It should be advisable to ensure a proper rotation of the stocks thus avoiding that they become stale or oxydate. this depends on the buying facilities of the local products and on importation, in relation with the food qualities to be manufactured for a given period. Avoid stocking over many months especially in high temperatures. 3. Technological aspects At it is necessary to cover the food requirements of the species reared, it is also essential to distribute standard size food particles, suitable to the size of the mouth of the fish during the production cycle. The particle sizes vary from a hundred microns to ten millimeters. This presentation aspect is of prime importance. The manufacture of compound food is based on the caking of dry meal by compressing it through circular die holes having different diameters (See figure 1). This technic, commonly employed for the production of compound food for land animals has been adapted, which led to the construction of very specialized units for the manufacture of compound food in aquaculture. Certain constaints have been remarked. They result principally in the following points: – As little dust as possible and good caking (pelleter) of the particles is required so as to limit losses and pollution of the rearing waters. – The proteinic products which are the base of the formulation are difficult products to manipulate, mince or cake and require properly adapted technical solutions. – Specific and costly investments are required to manufacture the food particles of less than 2 mm in diameter. – Finally, due to the improvement in knowledge on the nutritive requirements of fish, it becomes imperative to incorporate more and more quantities of fatty materials, which leads to the requirement of the appropriate technical solutions. Indeed, from a standard diagramme, such as that given in figure 2, it was necessary to modify and innovate on it, which means a unit for the manufacture of fish food is quite original when compared to that for land animal food. The animal food industry is a heavy industry which requires high investments for a feeble return. The raw materials make up more than 80 % of the cost price of the food manufactured. To define the production capacity of one unit, the rearing divisions which would buy the food must be well known, along with their capacities and their production cycles. There are indeed important seasonal variations concerning quantity and quality. Each manufacture has its own particularities which must be studied according to the market requirements and production perspectives over the next 10 years. The raw material supply conditions must be taken into account (in bulk or in sacks) together with the availability of a specialized manual labour and the desired automatization level of the circuits and of the whole unit. TABLE N° 1 ESTIMATE OF THE PROTEINIC REQUIREMENTS OF CERTAIN FISH (1) Species Crude proteinic requirement in the food ensuring optimal growth (g/Kg) Rainbow trout (Salmo gairdneri) 400 - 450 Carp (Cyprinus carpio) 380 Chinook salmon (Oncorhynchus tshawytscha) 400 Japanese ell (Anguilla japonica) 445 Plaice (Pleuronectes platessa) 500 Royal sea-bream (Chrysophrys aurata) 400 Herbivorous carp (ctenopharyngodon idella) 410 - 430 Japanese red sea-bream (Chrysophrys major) 550 Seriola (Seriola quinqueradista) 550

(1) by C.B. COWEY, 1978 TABLEAU N° 2 The essential amino acid requirements of six species of fish Chimooka Japaneseb Rainbowc Channele Salmon Eel Carpb Carpc Trout Tilapiad Catfish Arginino 2.4 (6.0/40) 1.7 (4.0/42 1.6 (4.3/38.5) 1.52 (3.8/40) 1.40 (3.6/40) <1.59 (4.0/40) 1.03 (4.29/24) Histidine 0.7 (1.8/40) 0.8 (1.9/42) 0.8 (2.1/38.5) 0.50 (1,4/40) 0.64 (1.6/40) ND 0.37 (1.54/24) . Isoleucine 0.8 (2.2/41) 1.5 (3.6/42) 0.9 (2.5/38.5) 0.02 (2.3/40) 0.96 (2.4/40) ND 0.62 (2.58/24) beucine 1.0i (3.9/41) 2.0 (4.8/42) 1.3 (3.3/38.5) 1.64 (4.1/40) 1.76 (4.4/40) ND 0.84 (3.50/24) Lysine 2.0 (5.0/40) 2.0 (4.8/42) 2.2 (5.7/38.5) 2.12 (5.3/40) 2.12 (5.3/40) 1.62 (4.1/40) 1.50 (5.00/30) Methionine 0.6 (1.5/40) 1.2 (2.9/42) 1.2 (3.1/38.5) 0.64 (1.6/40) 0.72 (1.8/40) <0.53 (1.33/40) 0.56 (2.34/24) Cys - 1% Cys - 0% Cys = 0% Cys = + Cys = + Cys = 0.74% 60% replaceable by Cys 0.9 (2.1/42) 0.8 (2.1/38.5) Cys - 1% Cys - 2% Phenylalanine (4.1/41) 2.2 (5.2/42) 2.5 (6.5/38.5) 1.16 (2.9/40) 1.24 (3.1/40) ND 1.20 (5.0/24) Tyr - 0.4% Tyr - 0% Tyr = 0% Tyr = + Tyr = + Total Tyr + Phen

1.2 (2.9/42) 1.3 (3.4/38.5) 50% Phen Tyr - 2% Tyr = 1% Threonine O.8 (2.2/40) 1.5 (3.6/42) 1.5 (3.9/38.5) 1.32 (3.3/40) 1.36 (3.4/40) ND 0.53 (2.21/24) Tryptophan 0.2 (0.5/40) 0.4 (1.0/42) 0.3 (0.8/38.5) 0.24 (0.6/40) 0.20 (0.5/40) ND 0.12 (0.5/24) Valine 1.3 (3.29/40) 1.5 (3.6/42 1.4 (3.6/38.5) 1.16 (2.9/40) 1.24 (3.1/40) ND 0.71 (2.96/24) Total 12.3 (30.49) 14.8 (35.4) 13.7 (35.8) 11.28 (28.2) 11.64 (29.1) 7.48 (30.92) a Data for chlnook salmon and rat from Mertx (1960) b Data for Japanese eel and carp from Mose (1979) c Data for Rainbow trout and carp from Ogino (1980) d Data for tilapia from Jackson & Capper (1982) e Data for channel catfish from Wilson et al, (1977, 1978, 1980); Harding et al, ( 1977); Robinson et al. (1978, 1980a, b, 1981) Values are expressed as grams per 100g of dry diet. In parenthesis the numerators are grams per 100g of protein and the denominators are the percent total dietary protein TABLE N° 3. RATION CHART - WHOLE NUTRIMENT FOR SEA-BASS Weight-of sea- TEMPERATURE PATTERN GRANULOMETRIES bass/ing below 15 °C 15° to 19° to 23° to 26 26° to 28 ° 18 °C 23 °C °C 28°C and + 1st stage Inf.to 0,5 unlimited distribution depending on the appetite of the fish 2nd stage 0,5 to 1 Feed 4 4,2 4,5 4 3 3rd stage 1 to 3 according to 3,2 4,2 4,5 4 3 the fish Crumbs 3 to 8 appetite 2,8 3,0 3,3 3,0 2,9 1,5 mm 8 to 15 2,5 2,7 3,0 2,7 2,5 2 mm 15 to35 1,9 2,3 2,6 2,3 2,0 3,2 mm 35 to 100 1,3 1,6 2,0 1 ,8 1,5 4,5 mm 100 to 500 1,1 1,4 1,8 1,6 1,3 6 mm Sup.to 500 1,1 1,4 1,8 1,6 1,3

This chart indicates the quantity of feed in Kg for 100 kg of fish to be distributed daily according to the type of nutriment and the rearing water temperature. Remarks: The ration bases must be adapted to the particular conditions of each rearing.Control the dissolved oxygen content. ANALYTIC GUARANTEES: WHOLE NUTRIMENT FOR SEA BASS Humidity % gross % Fats % Cellulose % Minerales % Granulometries proteins 1st stage small grain 10 56 12 2 13 2nd “ “ “ 10 56 12 2 13 3rd “ “ “ 10 56 12 2 13 Crumbs 4th stage 10 52 12 2 13 Granulars 1,5 mm 10 52 12 2 13 “ 2 mm 10 49 15 3 13 “ 3,2mm 11 46 15 3 13 “ 4 , 5 mm 11 46 15 3 13 “ 6 mm 11 46 15 3 13

TABLE N0 4 - ANIMAL MEAL

Meat Fish 45 50 55 60 Fish Anchovy Sardin Smelt herring soluble 60 65 65 68 70 Centesimal Composition Dry material 93 93 93 93 92 92 92 92 92 50 Crude proteins 45 50 55 60 60 65 65 68 72 30 Fatty materials 10 10 9 8 10 7 5.50 10 9 4 Cellulose ------Nitrogen Fece extract 3 3 4 3 3 5 6.50 2 0 6 Minerals 35 30 25 22 19 15 15 12 11 10 Calcium 12 10 8 7 5 4 4,50 4 3 0.10 Total phosphore 5 4,50 3.50 3.30 3 2,50 2.70 2.50 2 0.50 Assim. phosphore 5 4,50 3.50 3.30 3 2,50 2,70 2.50 2 0.50 D.N.M- Ruminants 36,00 40,00 46,80 51.00 51,00 55,30 55,30 58,50 61,90 26.40 Energy Raw Kcal/kg Poultry 3613 3903 4136 4287 4473 4561 4483 4888 4941 `2328 Metabolisable “ 1870 2070 2270 2400 2950 2880 2800 3100 3200 -430 Pork: Digestible “ 2930 3160 3320 3480 3540 3540 3430 3910 4410 1900 Metabolisable " 2550 2750 2870 2990 3050 3030 2920 3360 3780 1640 Net" “ 1790 1910 1940 1930 2070 1960 1850 2250 2490 1090 Net" “ 0.83 0,89 0.90 0.92 0,96 0,91 0.86 1.05 1,16 0.51 Ruminants Digestible TDFp 100 61 55 68 70 77 77 75 83 83 42 Net UF/Kg 0.70 0,77 0.83 0.86 1.01 1,01 0.98 1,14 1.14 0.56 Net UA/Kg 0.56 0,62 0.67 0.68 0,73 0,69 0.86 0.80 0,82 0.33 Amino-acids percent of Raw products LYSINE 2.16 2,53 2.78 3.12 4.57 5,08 5.12 4.84 5,06 1.38 METHIONINE 0.60 0,68 0.74 0.80 1.57 1.81 1.83 1.93 1.85 0.42 METHIONINE + 0.99 1,10 1.20 1.55 2,14 2,38 2.41 2.55 2,44 0.60 CYSTINE THRE0NINE 1.36 1,54 1.68 2.06 2.44 2,72 2.75 2 94 2 89 0,70 TRYPTOPHANE 0.18 0,20 0.22 0.36 0.62 0,69 0.50 0.70 0 77 0.11 ARGlNINE 3.10 3,39 3.72 3 81 3,52 3,76 3.71 3 57 3.67 1 28 GLYONE 6 48 7,54 8.29 7.24 4,33 3,93 4.C2 2 64 4 12 2,57 GLYONE + SERINE 8 23 9,51 10.45 9.83 6.79 6,43 6.58 6 51 6,84 3 27 HISTONE 0 66 0,81 0 89 1.05 1 26 1,61 1.57 1.29 1 38 1 09 ISOLE DONE. 1 31 1 44 1 58 1 85 2.59 3,08 3 06 3 17 3 30 0 90 LEUCNE 2 52 2,84 3.12 3 79 4.19 4,84 4.80 5, 24 5 16 1.37 PHENYLALANINE 1 43 1 56 1 72 2 02 2 27 2 71 2.72 2 65' 2 75 0.70 PHENYLALNINE+TYR 2 27 2.62 2.88 3.38 4.16 4,85 4 91 4.78 4,67 1.16 OSINE VAUNE 2.04 2,14 2 35 2.78 2.95 3,47 3 53 3.76 3,80 1.00 Digestible nitrogenous materials D.N.M Total digestible food T.D.F Forage Unit F.U. Starch Unit S.U TABLE 5 Animal by-products Dairy products fats

Animal by-products Dairy products Fats Feather Slaughter. Blood Skimmed Lacto Butter Soya Corn meal by meal milk serum milk Lard Fat oil oil products Centesimal composition Dry material 90 90 90 95 95 95 99 99 99 99 Crude proteins 83,50 60 85 35 13 32 - - - - Fatty materials 2.50 14 1 1 1 5 99 99 99 99 Cellulose 1,50 2,50 ------Nitrogen Feed extract - - - 51 72 48 - - - - Minerals - 2, 50 13,50 4 8 9 10 - - - - - Calcium 0.20 3,80 0,30 1,30 0,90 1,30 - - - - - Total Phosphore 0,75 2,20 0.25 1 0,70 1 - - - - - Assim.phosphore 2.75 2,20 0,25 1 0?70 1 - - - - D.N.M Ruminants 65,80 48,00 68,00 33,30 11,70 28.80 - - - - Energy Raw Kcal/Kg 5254 4825 4938 4179 3786 4257 9207 9207 9207 9207

Poultry Kcal/Kg 2340 3000 2760 2770 2170 2900 8800 7110 9020 8800 Metabolisable Pork Digestible Kcal/Kg - 3930 3650 3870 3500 4060 8650 8850 8650 8650 Metabolisable Kcal/Kg - 3440 3060 3570 3380 3780 8370 8370 8370 8370 Net Kcal/Kg - 2470 1800 2300 2270 2580 8370 8370 8370 8370 Net UF/Kg - 1.15 0.84 1,07 1,06 1.20 3.89 3,89 3.89 3.39 Ruminants Digestible TDF p - - 80 85 85 85 218 218 218 2.18 100 Net UF / Kg - - 1.05 1,16 1.16 1,16 3.70 3,70 3.70 3.70 Net UA / Kg - - 0,71 0,70 0,70 0.72 2.40 2,40 2,40 2.40 Amino acids percent of raw products LYSINE 1.87 2,23 8.05 2,91 1,05 2.27 - - - - MSTHIONINE 0.43 1.17 1,02 0,79 0,20 0.64 - - - - METHIONINE + 4 .27 2,10 1.76 1.10 0,47 0.99 - - - - CYSTINE THREONINE 3.66 1,98 3.74- 1,52 0,77 1.50 - - - - TRYPTOPHANE 0.40 0,45 1.04 0,45 0,14 0.45 - - - - APGININE 6.58 3,72 3.68 1.21 0,29 1.18 - - - - GLYCINE 6.54 2,84 3.81 0,66 0,25 0.73 - - - - GLYOINE + SERINE 6.34 6,21 8,03 2,59 0,85 2.45 - - - - HISTONE 6.54 0,75 5 40 0,99 0,22 0.89 - - - - ISOLEUONE 2.20 2,26 0.91 1,94 0,75 1.76 - - - - LEUCINE 6.91 4,03 11.07 3,37 1.19 2.91 - - - - PHENYLALANINE 4.11 1,74 6.12 1.70 0.38 1.53 - - - - PHENYLALANINE + 6.5 2,23 8.58 3,45 0,71 '2.97 - - - - TYROSINE VAUNE 7.5 2,86 7,90 2,30 0,72 2.04 - - - -

Digestible nitrogenous materials D.N.M These indicative energetic values are given for fresh chickens. There seems to Total digestible food T.D.F be differences for laying hens and turkeys Forage Unite F. U. Starch Unit S.U. TABLE 6 Cereal by-products

Soft wheat Hard wheat White Second Fine Germs wheat Bran Bran wheat Bran Bran Bran wheat Centesimal composition Dry material 83 88 88 88 83 88 88 88 88 15 15 16 15 26 16 16 15 12 Crude proteins Fatty 3 4 4,50 4,50 8 3.50 4,50 4 12 materials Cellulose. Nitrogen 1 3,50 7 10 3 1.50 9 10.50 5 Feed extract 67 63 56 53 46.50 64,50 54,50 54 51 Minerals 2 2.50 4,50 5,50 4.50 2.50, 4 4,50 8 - Calcium 0,05 0.08 0,10 0,15 0.07 0,10 0,15 0,15 0.06 - Total Phosphore 0.30 0.60 0,90 1,20 1 0.90 0,90 1 1.40 - Assim. phosphore 0.10 0.20 0,30 0,40 0.33 0,30 0,30 0,33 0.47 D.N.M Ruminants 13,50 12.80 13,10 11,70 23.90 14,40 13,10 11,70 8,50 Energy Raw Kcal/Kg 3922 3954 3914 3857 4255 3944 3934 3872 4096 Poultry Kcal/kg 3050 2750 2100 1400 3030 2000 1200 31 00 Metabolisable Pork Digestible Kcal/Kg 3630 3240 2920 2600 3630 3630 2890 2600 3280 Metabolisable Kcal/Kg 3480 3100 2780 2470 3390 3470 2750 2470 3120 Net Kcal/Kg 2500 2220 1980 1760 2453 2490 1950 1760 2450 Net UF Kg 1,16 1.03 0,92 0,82 1.14 1.16 0,91 0.82 1.14 Ruminants Digestible TD.Fp.100 80 79 70 59 82 80 70 61 75 Net UF / Kg 1.10 1,08 0,90 0,70 1.13 1.10 0,90 0,72 1,00 Net UA / Kg 0,79 0,75 0,63 0,50 0.81 0,79 0,63 0,50 0,76 Amino acids percent of raw products LYSINE 0,47 0,53 0,72 0,59 1,45 0,45 0,67 0,53 0,59 METHIONINE 0,24 0,24 0,25 0,21 0,43 0,25 0,24 0,22 0,25 METHIONINE + CYSTINE 0,54 0,54 0,56 0,51 0,80 0,57 0,55 0,53 0,52 THREONINE 0 42 0,45 0,53 0,48 0,89 0,46 0,52 0 47 0,44 TRYPTOPHANE 0 20 0,20 0,21 0,20 0,24 0,21 0,21 0,20 0,10 AFGININE 0 81 0296 1,11 1,04 1 84 0,82 1,13 0,95 0,95 GLYCNE 0 60 0,59 0,85 0,81 1,36 0,53 0,79 0,58 0,61 GLYCNE - SERINE 1,26 1,22 1,52 1,44 2,36 1,29 1,47 1,35 1,14 HISTONE 0 35 0 26 0,40 0,39 0,53 0,36 0,41 0,36 0,32 ISOLUCAE 0 54 0 52 0,54 0,49 0,84 0,59 0,55 0,54 0,45 LEUCNE 0 07 0 93 0,96 0,89 1,52 1,03 0,69 0 95 0,81 PHENYLALANINE 0 64 0 60 0,59 0,57 0,89 0,72 0,62 0,63 0,52 PHENYLALANINE + 1,05 1,01 1,03 0,98 1,56 1,20 1,08 1,03 0,92 TYROSINE VALINE 0,69 0,71 0,78 0,72 1,25 0,71 0,76 0,72 0,69 Digestible nitrogenous materials D.N.M Total digestible food T.D.F Forage Unit F.U. Starch Unit S.U.

TABLE 7 CEREALS

Hulled Naked Soft Forage Hard Paddy Oats Corn Barley Rye Millet Oats Oats Wheat Wheat Wheat Rice Centesimal composition Dry material 87 67 87 87 87 87 87 87 87 87 87 Crude proteins 10,50 14 16 11 11 13.50 9 10 8 10 10 Fatty materials 4,30 5 5 2 2 2 4 2 2 1.50 3 Cellulose 10 2 2 2,50 2,50 2.50 2,50 5 9 2.50 2,50 Nitrogen Feed extract 59,20 64 62 70 70 67.20 70,20 67,50 63.50 71 70 Minerals 3 2 2 1,50 1,50 1.80 1,30 2,50 4.50 2 1,50 - Calcium 0,10 0,10 0.10 0,05 0,05 0.05 0,01 0,06 0,05 0.07 0,04 - Total Phosphore 0,35 0.35 0,35 0,35 0,35 0,35 0,30 0,35 0.30 0.30 0,30 - Assim. phosphore 0,12 0,12 0.12 0,12 0,12 0,12 0,10 0,12 0,10 0.10 0,10 D.N.M Ruminants 8,30 - - 8,80 8.90 11,10 6,60 7,40 6,50 7.90 6,60 Energy Raw Kcal/Kg 3836 3969 4001 3786 3786 3813 3866 3729 3615 3723 3822 Poultry Kcal/kg 2550 3190 3190 3080 3080 3050 . 3370 2700 2650 2800 3250 Metabolisable Pork Digestible Kcal/Kg 2680 3830 3430 3430 3440 3500 3130 2720 3310 3490 Metabolisable 2580 3650 - 3330 3330 3320 3410 3050 2660 322C 3400 Kcal/Kg Net Kcal/Kg 1890 2680 - 2380 2380 2360 2490 2180 1920 2290 2450 Net UF/Kg 0.88 1.25 1,11 1.11 1.10 1,16 1.01 0.89 1,05 1.14 Ruminants Digestible TD.F p 100 64 - - 79 79 79 83 75 74 77 72 Net UF / Kg 0,79 - 1,08 1,08 1,08 1.15 1,00 0.98 1.05 0,35 Net UA / Kg 0.60 - - 0,76 0,76 0,75 0,81 0,67 0.66 0.75 0,76 Amino acids percent of raw products LYSINE 0,42 0,55 0,62 0,31 0,31 0,36 0,25 0,37 0,28 0,40 0,23 METHIONINE 0,17 0,23 0,25 0,17 0,17 0,22 0,19 0,17 0,18 0,18 0,16 METHIONINE + CYSTINE 0,47 0,62 0,71 0,42 0,42 0,54 0,39 0,40 0,36 0,45 0,34 THREONINE 0,34 0,45 O,49 0,31 0,31 0,38 0,32 0,34 0,28 0,34 0,33 TRYPTOPHANE 0,15 0,20 0,23 0,12 0,12 0,16 0,09 0,12 0,10 0,11 0,10 ARGNINE 0,65 0,85 1,03 0,53 0,51 0,62 0,40 0,51 0,61 0,55 0,39 GLYCNE 0,50 0,66 0,77 0,43 0,42 0,53 0,33 0,42 0,36 0,46 0,33 GLOINE + SERINE 0,98 1,30 1,50 0,93 0,92 1,14 0,77 0,84 0,74 O,89 0,79 HISTIDNE 0,21 0 28 0,33 0,24 0,24 0,29 0,25 0,22 0,17 0,23 0,22 ISOLEUCINE 0,41 0 55 0,65 0,40 0,40 0,50 0,34 0,38 0,32 0,35 0,44 LEUONE 0,74 0,98 1,17 0,72 0,72 0,89 1,17 0,70 0,59 0,63 1,38 PHENYLALANNE 0,51 0,63 0 85 0,48 0,51 0,62 0,45 0,51 0,39 0,44 0,53 PHENYLALANINE + 0,86 1,15 1,45 0,78 0,84 1,02 0,81 0,83 0,74 0 73 0,96 TYROSINE VALINE 0,55 0 73 0,88 0,49 0,47 0,60 0,46 0,53 0 48 0, 51 0,55 Digestible nitrogenous materials D.N.M Total digestible food T.D.F Forage Unit F.U. Starch Unit S.U.

Figure N° 1

1 - Emptying, of raw materials 16 - Mixing 4 - Weighing FIGURE N° 2 18 - Caking press 6 - Premixing 24 Vertical cooling system 9 - Grinding Base Unit 31 - Reserve and sacking

FEEDING PRINCIPLES FOR MARINE FISH UNDER INTENSIVE CULTURE Mr. A. G. J. TACON INTRODUCTION In contrast to extensive and semi-intensive farming systems where fish derive all or a substantial part of their nutrient needs from naturally available food organisms, fish maintained under intensive culture conditions are totally dependent on the external provision of a nutritionally "complete" diet throughout their culture cycle. Traditionally, "complete" diets have taken the form of a dry or moist pelleted feed consisting of a combination of different feed ingredients, the overall nutrient profile of which, as near as possible, approximates to the. nutrient requirements of the fish species in question under conditions of "maximal" growth. Alternatively, complete diets may consist of a single food item of high nutrient value (i. e. trash fish, live food organisms - Artemia), or a combination of both. For the majority of marine farmed fish, there is scant information on basic nutrient requirements. This is in sharp contrast to the intensive poultry or trout industry, where basic dietary requirements for protein, essential amino-acids , essential fatty acids , vitamins and minerals are well established. At present, these shortcomings are overcome by using fresh or frozen fish/dry compound feed combinations with a high inherent nutrient safety factor; the use of which being economically justified with good management by the high market value of the farmed species (i. e. sea bass, gilthead bream, turbot, sole, eel). FEEDING PRINCIPLES AND GUIDELINES From a nutritional viewpoint, the principles governing the feeding of fish under intensive culture can be viewed as follows: 1 - That the food given is palatable and consumed completely with minimum effort and wastage. 2 - That the nutrient profile of the food given approximates as far as possible to the known dietary nutrient requirements of the fish species in question. 3 - That the food is digestible, keeping the fish healthy and growing normally. 4 - Ideally, that the food is efficiently converted into new body tissue in the form of fish growth (i. e. optimal feed conversion efficiency). In practice, however, from a commercial farming viewpoint, the choice of a particular feeding strategy is based on an assessment of three basic criteria (for intensive culture): 1 - Feed availability and handling. 2 - Feed performance. 3 - Food and feeding cost/unit of production/unit time and the rate of return on capital. For example, four basic hatchery feeding strategies are currently available for the mass rearing of marine fish larvae from first feeding, through metamorphosis, to the post-larval stage. These include: 1) The exclusive use of a succession of live planktonic food organisms (i. e. algae, diatoms, flagellates, yeasts, rotifers and Artemia salina nauplii). 2) Use of selected live and/or frozen plankton in conjunction with "fresh" and/or frozen fish, mollusc or crustacean tissue. 3) Use of selected live and/or frozen plankton in conjunction with dry feed materials or formulated diets. 4) Exclusive use of microencapsulated or microparticulate larval diets. Using the guideliness listed above, each hatchery feeding strategy can be assessed as follows: FEED AVAILABILITY AND HANDLING A. - LIVE PLANKTONIC FOOD PRODUCTION 1. Source of culture organisms a) Local 1. Species available 2. seasonality 3. personnel requirement for collection b) Imported 1. Dependability of supplier 2. minimum quantity of order 3. lead time on orders 4. variability in performance (hatchability - Artemia cysts) 5. import restrictions - licence/tax/country 2. Maintenance and production of culture organisms a) Stock cultures 1. Space requirement .- laboratory/phytolaboratory 2. personnel requirement - specialized training 3. service requirements - power/light/gas/air/UV/aircon - backup 4. culture media requirement - inorganic salts/trace elements/vitamins/chelating agents/antibiotics 5. equipment requirement - autoclave/filters/glassware/microscope/cell counter/plankton culture vessels b) Production cultures 1. Extensive continuous culture (green water system) requirements - fertilizers/personnel/air supply/space/tanks - backup 2. Intensive batch culture system requirements - space/air and CO 2/supply/personnel/fertilization/UV and light/plankton culture vessels/phytolaboratory. c) Survival/stability 1. Survival/shelf life of stock cultures 2. survival of production culture - frequency of culture failures 3. necessity to keep stock cultures all year round B. FISH, CLAM OR CRUSTACEAN TISSUE FEEDING OPTION 1. Sources a) Fisherman b) Processors c) Farm staff/hired labour 2. Quality a) Species available b) Forms available 1. whole 2. heads, tails, skin, bone 3. offal c) Handling at source 1. icing 2. freezing 3. boxing 4. salting 5. storage d) Handling/processing on site 1. Service requirements - electricity/gas/air/water - backup 2. equipment requirement - mincing/blending/sieving/boiler 3. personnel requirement 4. storage requirement - space/freezer/refrigerator 5. depandability of services - power supply/backup e) Nutrient content 1. proximate composition 2. particle size requirements 3. seasonal variations in nutrient content 4. possible contaminants/anti-nutritional factors 5. spoilage characteristics/storage life f) Larval feeding requirements 1. manual feeding - feeding regime 2. automatic feeders 3. service requirements - electricity/air 4. personnel requirements for feed preparation/feeding - man/hours/day 3. Quantities available a) Daily basis b) Weekly basis c) Seasonality d) Dependability of supplier e) Alternative species available f) Transport requirements for feed delivery to hatchery g) Minimum quantity of order IN HOUSE PRODUCTION OF A DRY/MOIST DIET 1. Feed ingredient availability a) Local sources 1. nutrient content 2. variability in composition 3. seasonality 4. particle size 5. possible contaminants/antinutritional factors 6. processing/cooking requirements 7. spoilage characteristics/storage life 8. storage requirements - space/refrigeration 9. dependability of supplier 10. minimum quantity of order b) Imported ingredients 1. nutrient content 2. variability in composition 3. seasonality 4. particle size 5. possible contaminants/anti-nutritional factors 6. processing/cooking requirements 7. spoilage characteristics/storage life 8. storage requirements - space/refrigeration 9. dependability of supplier 10. minimum quantity of order 11. lead time on orders 12. import restrictions 2. Feed preparation and handling a) Power requirement 1. electricity/gas/petroleum/steam 2. dependability of power supply - backup b) Water requirement c) Personnel requirement - man hours/day d) Feed preparation space requirement e) Feed processing equipment requirement - grinder/hammermill/blender/mixer/extruder/pelleter/freeze drier/solar drier/air drier/oven f) Larval feeding requirements 1. manual feeding - feeding regime' 2. automatic feeders 3. service requirements - electricity/air 3. Feed storage a) Shelf life/stability b) Freezing/refrigeration requirement c) Packing d) Maximum storage capacity e) Minimum production batch D. IMPORTATION OF MICROENCAPSULATED/MICROPARTICULATE LARVAL DIET 1. Sources Japan, Taiwan, S.,E., Asia, U.S. Europe 2. Quantities available a) Dependability of supplier b) Minimum quantity of order c) Lead time on orders 3. Quality a. Nutrient content 1. proximate composition 2. particle size distribution 3. stability in water 4. variability in composition 5. spoilage characteristics b. Larval feeding requirements 1. manual feeding - feeding regime 2. automatic feeders 3. personnel requirement - man hours/day 4. Storage a) Storage space requirement b) Power supply requirement - electricity/gas/petrol 1. refrigerator 2. freezer 3. dependability of power supply - backup c) Packing and shelf life of feed FEED PERFORMANCE 1. Behaviour of feed in water a) Buoyancy b) Aggregation/dispersion characteristics c) Stability/leaching characteristics 1. rate of nutrient loss 2. effect on water quality 3. inducement of algal/bacterial growth/contamination d) Swimming behaviour of planktonic food organisms 2. Feed requirement for each critical growth phase a. mg/larvae/day b. food density/larvae/day c. particle size requirement 3. Larval development a) Feed days required for each growth phase 4. Feeding behaviour of larvae a) Distribution within culture tank 1. even distribution 2. clumping/aggregation 3. surface/mid-water/bottom b) ability to capture prey/feed particles c) Feeding incidence of larval population (%) d) Visibility of faecal strands e) Feed palatability/attack response f) Abnormal swimming behaviour g) Incidence of cannibalism 5. Larval survival a. Mean survival rate (%) for each critical growth phase b. Frequency of mass larval mortalities - batch failures c. Incidence of bacterial/parasitic infections related to poor water quality resulting from poor diet stability d. Incidence of larval deformities 1. scoliosis/lordosis 2. Short body dwarfism 3. Tissue wasting disease 4. moulting capacity (shrimp) 5. frequency size distribution 6. Additional hatchery service requirements related to feeding option used a) Water exchange (%) frequency b) Aeration c) Lighting d) Tank cleaning e) Water sterilization f) Use of antibiotics FOOD AND FEEDING COST/UNIT OF PRODUCTION/UNIT TIME 1. Capital (fixed costs related to feeding option)1 a) Land - total hatchery area devoted to live food production, feed preparation and storage b) Structures - shed, store, laboratory, tanks, etc... devoted entirely to live food production, feed preparation and storage. c) Machinery/equipement - pelletizer, grinder, feed dispensers, blender, boiler, silos, oven; freeze drier, autoclave, refrigerator, freezer, pumps, filters, microscope, air conditioner, mincer, sieves, etc... directly associated with feeding option. 1. Although the capital items listed represent the total fixed capital investment for a particular feeding regime, for economic evaluation only the total fixed or capital outlay will be considered. This outlay is primarily in the form of a sinking fund of contribution, which covers the deperciation (amortization period) and loan interest payments on the cost of the land, structures and machinery or equipment over a pre-determined period. For complete financial analysis (since many capital items listed have dual functions, i. e. not necessary restricted to the feeding option alone), all aspects of the hatchery operation must be considered (i. e. investment requirements such as hatchery building, larval rearing tanks and accessories, installation cost, electrical facilities including backup services, air supply, water supply, plumbing, filter systems, utilities, laboratory equipment,; vehicles, etc... ) together with insurance costs, business permit/license fees and land taxes where applicable. 2. Operating (variable) costs related to feeding option a. Personnel - (manpower requirement, including level of technical skill required) b. Energy - (electricity, fuel, oil) c. Feed procurement, handling (delivery), storage and processing cost. Additional factors which may also have to be taken into consideration include import duties/taxes, minimum quantity of order, availability of foreign exchange/credit facilities. A crude estimate can be made of feed costs/unit of fish produced using these values. d. Maintenance/spares e. Fertilizers and chemicals f. General supplies and materials g. Miscellaneous 3. Market value of fish and revenue from sales/year 4. Total cash outlay of hatchery/year (includes hatchery operating costs, sinking fund and insurance, etc... ) 5. Cash outlay/106 larvae produced/unit time 6. Net income (before taxes, 3 - 4) 7. Income over total outlay (%) SPECIFIC DIFFICULTIES ASSOCIATED WITH THE FEEDING OF MARINE FISH 1. Live food requirement during first feeding The majority of marine farmed fish have small diameter eggs (1-2 mm) producing small larvae (0,5 - 1,5 mg wet weight) with poor yolk sac reserves. For some species, the hatchlings are so poorly developed that their mouth is still closed and gastro- intestinal tract non-functional (i.e. gilthead bream). In addition, after a short yolk-sac abdorption period, larvae are often incapable of consuming feed particles 50 m. Bearing these factors in mind, therefore, and the problems associated with diet stability and nutrient leaching, it is perhaps not surprising that most commercial hatchery rely on the use of live food organisms (commonly Brachionus plicatilis and Artemia salina nauplii) for the first feeding larvae until metamorphis is complete. Despite the economic efficacy of a well managed marine fish hatchery using a live food feeding regime, there are numerous disadvantages associated with a live food feeding strategy, including: 1) High initial capital investment costs - fabrication of expensive and sophisticated live food production facilities, including laboratory with high energy service requirements. 2) Land/space requirement - valuable hatchery space, which may otherwise be used for larval production, is devoted to live food production. 3) Stock culture maintenance requirement - feeding regimes involving the use of pure diatom/algal species and specific rotifer strains necessitate the maintenance of stock cultures on a yearly basis; usually requiring the construction of an air conditioned laboratory for this purpose. 4) Labour requirement - The maintenance and production of live food organisms necissates a high labour (skilled) requirement of live food production units does not favour the development of small scale hatcheries by the traditional farmer with limited cash funds. 5) Small scale hatchery development - the current high capital investment costs and high labour (skilled) requirement of live food production units does not favour the development of small scale hatcheries by the traditional farmer with limited cash funds. 6) Weather effect - The production of live food organisms in outdoor tanks is affected by the climatic conditions, resulting in variable larval survival, depending on the season. 7) Variable quality and nutritive value - The quality and nutritive value of live food organisms is variable depending on strain, source and culture method used (WATANABE et al., 1983). 8) Availability and cost - On the basis of culture techniques used at the Centre Océanographique de Bretagne (France), the dry weight cost of Artemia nauplii and rotifer Brachionus spp has been estimated to be US $ 220/ kg and US, $ 2 000/ Kg respectively (GIRIN, 1977). Similarly, in many developing countries, the importation of Artemia cysts necessitates import clearances, taxes and the availability of foreign exchange facilities. Bearing these factors in mind, it is essential that a simple and inexpensive artificial feeding package be developed if intensive marine farming systems are to be realized by the traditional fish farmer. 2. Feeding behaviour and diet stability Maximum benefit from feeding can only be achieved if the food provided is ingested by the fish. An understanding of the feeding behaviour of the fish is, therefore, essential. The diet presented must have the correct texture, particle size, density (buoyancy) and attractiveness to elicit an optimal feed response. Marine fish, and especially their larvae, appear to be particularly exacting in this respect. For example, although marine fish held in captivity generally rely on sight to locate their food, they also rely on chemoreceptors located in the mouth or externally on appendages such as lips, barbels and fins. Consequently, with many marine fish species food particles are carefully sensed before being taken into the mouth; the presence of feeding attractants within the food acting as ingestion stimulants. Feed palatability and feeding attractants: At its lowest level the often poor palatability of a hard dry pelleted diet can be improved simply by adding 10 - 20 % water so as to give a soft pellet texture (i. e. flat fish). For many marine fish species specific dietary feeding attractants have been found to elicit a feeding response under captive conditions, including the nucleoside-inosine and inosine-5-monophosphate (for turbot, MACKIE and ADRON, 1978; PERSON Le RUYET et al., 1983) and the quaternary amine-betaïne, either alone (for 50 g. sole, MACKIE et al., 1980) or in combination with free L.amino acids and inosine (sole; MACKIE et al., 1980, CADENA ROA et al., 1982; METAILLER et al., 1983). The practical importance of feed attractants and diet palatability is particularly critical during the weaning of marine fish larvae from a live to a non-living diet. Similarly, as attempts are made to replace the fishmeal component of practical fish feed with unconventional protein sources of an alien nature to the fish (for example, soybean meal), the problem of diet texture and palatability will become even greater. Finally, by improving feed palatability, the period of time the feed remains in water can be reduced, thus minimizing nutrient leaching. Feed stability and nutrient leaching: Since many marine fish species have a slow feeding behaviour, in that they "sence" their food prior to ingestion, diet stability and consequent leaching of water soluble nutrients poses a major hazard. Nowhere is nutrient leaching more of a hazard than in diets for larval fish where mouth size necessitates the use of food particles with a very high surface area/volume ratio. For example, GRABNER et al., 1981, reported the loss through leaching of almost all the free, and about one-third of the free plus protein bound amino acids from frozen or freeze-dried zooplankton (Artemia salina and moina sp.) after a 10 minutes water immersion period at 9° C. SLINGER et al., 1979, reports the loss through leaching of up to 50 - 70 % in vitamin C, 5 - 20 % loss in pantothenic acid, 0 - 27 % loss in folic acid, 0 - 17 % loss in thiamine and a 3 - 13 % loss in pyridoxine activity through leaching, after a 10 second water immersion period (1,18 - 2,36 mm diameter trout pellet). Similar water stability tests with complete diets for penaeid shrimps report water soluble vitamin losses of 97 % (thiamine), 94 % (pantothenic acid), 93 % (pyridoxine), 90 % (vitamin C), 86 % (riboflavine), 50 % (inositol), and 45 % (choline) after a one hour immersion period in sea water (CUZON et al., 1982). Although to a certain extent these effects can be minimised by using dietary feeding attractants and short feeding intervals (i. e. regular feeding), various microencapsulation and micro-binding stabilization techniques have recently been introduced for the manufacture of artificial diets to overcome these problems. Although it is not the intention of this discussion paper to review these techniques here, the use of extrusion cooking techniques to produce expanded and rehydratable water stable feeds seems particularly promision (MELCION et al., 1983; CADENA ROA et al., 1982). The combination of extrusion cooking techniques with the subsequent application of a lipid-vitamin emulsion onto the outside of the expanded feed may be particularly profitable avenue for research. The advantages and disadvantages of producing an extruded diet are shown in Table 1. 3. Dietary protein requirement The majority of marine fish species examined to date are carnivorous in feeding habit, and consequently have a high dietary requirement for protein (minimum of 40 - 50 % on a dry weight basis) and a low tolerance for dietary carbohydrates (TACON and COWEY, 1985). At present, high quality fish meals supply the major proportion of the protein component within complete diets for marine fish, with levels of up to 70 % (of the total diet) being used within some starter rations. In view of the high market cost of good quality fish meals, it is not surprising that feed costs may account for 40 - 50 % of the total operating cost of the farming operation. Apart from being an expensive feed commodity, and of uncertain supply within the next decade, the utilization of high quality fish meals for fish feeding is also inefficient in terms of our utilization of sea fishery stocks. Clearly, alternative and ideally less expensive sources of good quality protein must be found (for review see TACON and JACKSON, 1984). Table 1 - Disadvantages and Advantages of Extrusion Cooking DISADVANTAGES 1. Expansion processing requires more expensive equipment than straight pelleting (including steam pelleting). 2. Process requires higher pressure, steam addition and mixture temperatures. 3. Reduced production rates, despite a very high power (energy/electricity) requirement. 4. Resulting pellets require further drying to reduce moisture content. 5. Higher vitamin supplement (destruction of heat labile vitamins). 6. Altered feed ingredients - particularly the use of feedstuffs with a high starch content. 7. Risk of over expansion - excessive bulkiness. 8. Reduced voluntary feed intake by fish. 9. Added cost of above to feed (10 - 20 % of total value). 10. 10. Possibility of maillard-type,reaction occuring and reducing the biological availability of specific amino-acids. ADVANTAGES 1. Allows feeder to observe fish - particularly under conditions of poor visibility (if an expanded "floating" feed is produced). 2. Ingredients are cooked which gelatinize the starch, resulting in strong intermolecular bonding. 3. Pellets are more durable and have superior water stability (reduced leaching of water soluble nutrients). 4. Increased biovailability and disgestion of carbohydrates (higher digestible energy content). 5. Better feed conversion efficiency. 6. Delayed gastric evacuation. 7. Faeces are coarse and lumpy (as compared to fine and watery with steam pelleted diets). 8. Less dust with expanded feeds. 9. Reduced feed wastage - incorrectly adjusted feeders - no over-feeding. 10. Lower feeding rate. 11. Floating property - allows determination of food consumption 12. Facilitates water and/or oil absorption so as to produce a rehydratable semi- moist diet (larval applications) or a high lipid diet (for use at low water temperatures, or with carnivorous fish species with a low carbohydrate tolerance). REFERENCES CADENA ROA, M., C. HUELVAN, Y. LE BORGNE and R. METAILLER. 1982 Use of rehydratable extruded pellets and attractive substances for the weaning of sole (Solea vulgaris). J. World Maricult, Soc, 13; 246 - 253 CUZON G., M. HEW and D.COGNIE. Time lag effects of feeding on growth of juvenile 1982 shrimp Penaeus japonicus (Bate). Aquaculture, 29; 33 - 44 GRABNER M., W. WIESER and R. LACKNER. The suitability of frozen and freeze dried 1981, zooplankton as food for fish larvae: biochemical test program.Aquaculture, 26; 85 - 94 MACKIE A.M. and J.W. ADRON. Identification of inosine and inosine -5' – monophosphate as the gustatory feeding stimulants for the turbot, Scophthalmus maximus. Comp. Biochem. Physiol., A60; 79 - 83 MACKIE A.M., J.W. ADRON and P.T. GRANT. Chemical nature of feeding stimulants for 1980 the juvenile Dover sole Solea solea (L). J. Fish. Biol., 16; 701 - 708 METAILLER J.P., J. GUILLAUME, J. MEHU, R. METAILLER and G. CUZON. 1983 preparation by extrusion cooking of improved feeds for marine animals. Proceedings "Cost 91" Extrusion Cooking, Athens, 14 - 18 November 1983 (In.press). PERSON - LE RUYET J, B. MENU, M.CADENA ROA and R. METAILLER. 1983 Use of expanded pellets supplemented with attractive chemical substances for the weaning of turbot (Scophthalmus maximus). J. World Maricult. Soc. 14; 676 - 678 SLINGER A.G.J., A. RAZZAQUE and C.Y. CHO. Effect of feed processing and leaching 1979 on the loss of certain vitamins in fish diets. In Finfish nutrition and fish feed technology edited by J.E.HALVER and K.TIEWS. Schr. Bundesforschungsanst. Fisch. Hamb.; (14/15) Vol. 2; 425 - 434 TACON A.G.J. and C.B. COWEY. Protein and amino acid requirements. In Fish Energetics – 1985 new perspectives (P. Calow and P. Tytler, eds). Croom Helm Press Ltd. London, pp. 155 - 183 TACON A.G.J. and A.J. JACKSON. Utilization of conventational and unconventional protein sources in practical fish feeds - A review. international Symposium on "Nutrition and Feeding in Fish" 10 - 13 July 1984, Aberdeen, Academic Press, London (In press). WATANABE T., C. KITAJIMA and S. FUJITA. Nutritional value of live organisms used 1983 in Japan for mass propagation of fish. A review.Aquaculture, 34; 115 - 143. MARINE FISH FEED Mediterranean species needs Mr J. GUILLAUME INTRODUCTION Those species found in the Mediterranean can not be clearly defined. The species reared for commercialization have no particular characteristics either. As the climatic conditions have little influence on. the nutritional requirements, a bibliographical review is uncalled for. Therefore, it appears preferable to limit this report to two typical families of Mediterranean aquaculture; sparidae and serranidae. As there is little information available on these families; when referring to nutrition, it is that for teleoster which is generally taken into account while applying the classical works (HALVER 1972; HOAR and RANDALL 1969-979; Anonyme 1981). It often happens that the studies carried out on the best known family have been taken as reference: salmonidae. But, before going into details, let us recall the excellent review by ALLIOT and PASTOUREAUD, 1984 also. Before giving a brief account on larvae broodstock needs and the different facters liable to modify requirements, juveniles shall be the first case to be examined, while reviewing the essential energy and food needs. Finally, the nutritional particularities of the fish by which we are concerned, shall be taken into account. I - JUVENILE FEED REQUIREMENT 1.1. Energy requirements 1.1.1. The importance of this requirement The first requirement remarked, when fish or superior vertebrates fast, is their energy requirement. Experiments carried out by ROZIN and MAYER, 1961, point out clearly that when the energetic density of a diet is changed, the gold fish can adopt its consumption in such a way so that its energetic consumption is nearly always constant. Numerous nutritional and non-nutritional factors can modify this balance (FLETCHER 1984) but it can be admitted that if the fish is fed ad libitum, it can make up for its needs, as long as the diet given is sufficiently appetizing and not greatly diluted with non- nutritional feed. This principle, now greatly accepted by land vertebrates, furnishes a very simple and good way of expressing nutritional needs. Indeed, for a given animal in a given condition, if the needs or standards employed by the food manufacturers are expressed in percentages of the energetic level, the animal fed ad libitum, will have satisfied its requirements in amino-acids, vitamins and other nutriments along with its energetic requirements. Theoretically, this would justify the systematic measurement of the energy consumed each time the requirements in any nutriment are studied. It would also justify a thorough study of the different factors liable of . modifying the energetic consumption and so the energetic requirements of fish. Many works have been certainly devoted to this subject and just taking the more recent reviews as example, let us mention the synthesis by FISHER,1979; BRETT and GROVES, 1979.The great relation of the energetic consumption evolution are well known: proportionality between the "basal" or routine consumption (standard metabolism) and the body weight, revised to a power while not taking into account the temperature, size nor species (around 0,8 power); the relative effect of temperature and salinity; the physical activity of the fish etc..Nevertheless, there are yet many problems to settle – The effect of temperature has been hardly ever studied in a zone of sufficient size, so that, for a given species the range of "proportionalite", (existance of a constant Q10) the compensation zone, where respiration reaches its maximum and the highest temperature limit is compatible with life, cannot be defined. The relation, between growth rates and temperature for different sizes or ages (BRETT 1979) has not been thoroughly examined either at rearing level. – Few global models have been proposed and it is rarely found that the same study comprehends a way to plan the effect of size, feed rates or temperature.Particular attention should be paid to the works carried out by HOGENDOORN and al, 1982 and HOGENDOORN, 1983 on the african cat fish Clarias lasera on this subject. – The ration tables are more or less of empiric origine. Those by HOGENDOORN, 1983, are somewhat an exception. 1.1.2. Energetic consumption of sea-bass and of sea-bream There are few studies dealing with consumption for those fish by which we are concerned. For records, the works carried out by PIONETTI, 1984, on the egg and embryo of sea-bass, that by ELDRIGE and al, 1982 and by CECH, 1981, on the embryo and juvenile of the american striped bass Morone saxatilis, could be taken into account. However, to our knowledge there is but one thorough study on the energetic consumption of the european sea-bass (Dicentrarchus labrax),that by BICAL, 1979. This author, points out that the relation between oxygen consumption (thus energetic consumption) and body weight is very close to the general interspecific relation by WINBERG, 1956, and also that found for the american striped bass, Morone saxatilis by KRUGER and BROKSEN, 1978. The energetic consumption increases regularly when the temperature rises from 8° to 25° c, although the maximum Q10 is between 10° and 15° c, reaching more than 20° c. Therefore, the sea-bass seems to be a relative hot water fish, which is not very unlikely. Salinity has little influence on the energetic consumption, even when it varies around 2,5 to 45 %. This unexpected result goes to show that the osmoregulation consumption is overestimated and could be interpreted as a consequence of the euryhalinity of the species. With sea-bream Chrysophrys major, a different result can be achieved, while using the classical theories: this fish can live longer at a salinity level of 12%, which means, in conditions where osmozeregulation consumption is at it's lowest level (WOO and MURAT, 1981). Finally, it can be said that the sea-bass is a relatively classical fish from an energetic point of view. BOHAC, 1981, has already proposed ration tables in function of the body weight and the temperature of the water. However, it seems that additional work in this field is necessary, especially so that the optimum temperature and the "routine margin for the activity" (by FRY, 1957) may be defined precisely. Our knowledge being limited, it also seems difficult to define the consumptions linked with physical activity, which in turn is itself linked with climatic factors and rearing conditions. Empiric works are still necessary in this sphere. As for sea-bream Sparus aurata, there is hardly any data available on the energetic metabolism. However, the preliminary works by DOSDAT, 1984, carried out in rearing tanks and not in a laboratory, shows that is must be assumed that a fish having a relatively high metabolic consumption, even greater than that of sea-bass, could be partly due to the quicker growth speed of sea-bream at the age when examined. 1.1.3. Use of the energy from the diet and coverage of sea-bream requirements For the past century zootechnicians who deal with land animals have proposed partition methods of the chemistry energy of food , in faeces energy, nitrogenous wastes energy and energy used by the animal (metabolisable, digestible or net energy) The latter fraction being used to cover the consumption of the standard metabolism, muscular consumption.anabolism consumption etc... This method has been transposed to fish by the authors (ex RUMSEY, 1977; CHO, SLINGER and BAYLEY, 1982). The only theoretical particularity with fish is the persisting doubt on the linearity relations between losses (ex: faeces and food injected(HOGENDOORN, 1983) Theoretically, it is therefore possible to estimate for a fish, the capacity of any food to cover the energetic requirements above mentioned, or at least for fish of moderate consumption; if sufficient measures, were employed, energetic values, digestible or metabolisable tables, could be made out for the principal food ingredients so to formulate diets of known energetic level. In practice, this work is more difficult with fish than with land animals, due to the fact that it is very hard, to collect excrements in water and especially to measure branchial and urinary nitrogenous wastes. To our knowledge, no data on the digestible or metabolisable energetic value is available on the sea-bass or the sea-bream, only a few measures on proteinic digestibility have been carried out by ALLIOT, 1982. Salmonidae, which are the nearest species, were employed. to take these measures (RUMSEY,1977; CHO and al, 1982). 1.1.4. The role of proteins, glucides and lipids as sources of energy. Technology being as it is, it is impossible to say precisely what quantity of respective energy is furnished by proteins, glucides and lipids in the food distributed out to sparidae and serranidae, as their digestibility is still unknown. By the food distributed and their estimated digestibility, it seems clear however, that the balance of these three main categories of food play a principal part not only for growth (this is evident for proteins, lipids, another molecule source is also known) but also for the good use of food energy itself. This balance influences the part of energy employed for the thermic consumptions and tissue synthesis. But because of the double part played by proteins and lipids, which supply unsynthetizable molecule though organism and energy sources, it is very complex to determine the best balance between these three categories of food. This has only been tackeled very briefly: for instance, research work has been carried out on the optimal proteinic rate and the best calory-nitrogene relation, etc... ) 1.1.4.1. Optimal proteinic rates for the sea-bass as for the sea-bream have been often studied, the results of which are not very convergent; the differences remarked may be attributed either to variations in requirements themselves (differences in species, in size, growth rates special environmental conditions) or to the study methods employed and especially to the type of diet (digestibility of the various elements, nature and percentage of glucides and lipids, digestibility and biological value of the proteins, etc...) The same applies for the different species of sea-bream as the "need" estimates start at 40 % SABAUT and LUQUET, 1973 (Sparatus aurata) reaching 55 % after YONE, 1976 (Chrysophrys major). The estimates are more homogenous with sea-bass as they go from 50 % (ALLIOT and al, 1974) to more than 53 %, if the works carried out by METAILLER and al,1981, are taken into account. The latter shows that there is improvement of growth weight when the proteinic tenor of the diet passes from 53 to 63 %.But even with higher rates, proteins are well digested by sea-bass (BRIGAUDEAU, 1981; PERES, 1981; ALLIOT, 1982) and are always an excellent source of energy. 1.1.4.2. The energy/protein relation (often known as calory-nitrogen relation) gives a more homogenous criterion than proteinic rates do, so the energetic level is taken into account , even if it cannot be thoroughly estimated. A thorough study has been carried out on juvenile sea-bass by ALLIOT and al, 1979 and by ALLIOT, 1982, the optimal value of this relation is situated around 7 to 8 Kcal of metabolisable energy per g.of proteins. Greater values, corresponding to a certain proteinic deficiency, are explained by the decrease in the conversion rate which doesn't even show an improvement in the proteinic efficiency coefficient, often remarked with land vertebrates. However this relation does not take into account the two possible origins of non proteinic energy. 1.1.4.3. The optimal rate of lipids has been studied with sea-bass as with sea- bream. Once the essential fatty acid needs have been satisfied, it seems,lipids are the most appropriate source of energy, that spares proteins. By the fact, like most teleoster, these fish make good use of low fusion point lipids at digestive as well as at metabolic level, but a limit above which growth rates decreases has been remarked by many authors. It is around 12 % for sea-bass (ALLIOT and al, 1974) and 9 % for Sparus aurata (MARAIS and KISSIL, 1979) and 10 % for Chrysophrys major (YONE and al, 1971-1975). The differences are too slight for them to be imputed to specific requirments. It can also be remarked that the maximum dose tolerated is feeble when compared, with that supported by salmonidae, where 15-16 % rates are usually found in certain marketable feed. It is not to be taken for granted that this superior limit has been studied adequately and so requires further study. 1.1.4.4. Glucides are a more rare source of energy in the natural feed of fish by which we are concerned. Indeed it can be estimated that with prey fish, one or more percent of glycogen is found in the dry matter. However, it should be remarked that up to 10 % of glycogen dry matter basis can be found in the lung tissues of molluscs. It is well known that teleoster have little or no enzymatic store concerning glucydasis (PERES, 1979) A second limit for the use of glucides is found at the metabolic level, the fish being prone to diabetes (SHIMENO and al, 1979). By this fact, if some herbivorous fish are excluded and especially sole, which consumes mollusc rich in glycogen, a great increase of glucides is quickly remarked, through the bad regulation of the glycemia, hypertrophy of the liver, linked with the storage of glycogen, slow growth, etc... There are certainly big differences between the various types of glucides. Glucides of feeble molecular weight digested by land vertebrates are also digestable by fish which evidently doesn!t stop these same constituants from causing the metabolic disorders which we have spoken about. Starches, which very well assimilated by earth vertebrates are not so by fish, moreover as high rates are incorporated. This phenomenon especially remarked with salmonidae, has been also found with sea-bass and sea-bream. Thus the bad effect that the glucose has on growth and hepatomegaly has been described for japonese sea-bream by FURUICHI and al, 1971, above a 10 % ratio, and for sea-bass by ALLIOT and al, 1979, who employed a much greater amount (around 20 %). It must also be remarked the negative effect that glucose has on the absorption of proteins,stressed by the first authors. Thus it is quite clear, that glucides can but spare a small quantity of proteins, the highest limit being difficult to define, is around 15 % which is less than for salmonidae. It is surprising to remark that the lowest limit of the glucide rates has been very briefly studied; It could be supposed that sea-bass and sea-bream can completely do without glucides as there are no essential needs of these and that the gluconeogenesis is very effective with carnivorous fish of this type (BEVER and al, 1981). We shall however mention the preliminary results obtained by SPYRIDAKIS and METAILLER, 1985, which show that growth is delayed if the diet only includes small amounts of glucides, 6 % therefore quite sufficient. 1.1.4.5. In. conclusion, the energetic needs for sea-bass and for sea-bream have not yet been studied precisely. The energetic value of the food given, is concluded from the experiments carried out on other species: The routine measures of digestable energy being at its beginning for sea-bass, to our knowledge. In spite of these blanks, there is much more information available on the energy furnished by proteinic, lipidic and glucidic food. these studies seem to show that there is little hope of feeding carnivorous fish rich glucidic diets, a part from the more predator species of molluscs which calls for a more detailed study. 1.2.Protein needs 1.2.1. The qualitative needs in essential amino-acids The essential amino-acids required by the sea-bass have been defined by METAILLER and al, 1973. From this study, at least nine amino-acids are unsynthetizable by sea-bass, there are still a few doubts about cystine and tryptophan which have not been studied while tyrosine could be classed among the semi- indispensable amino-acids, as the synthesis from phenylalamine seems possible. However, the qualitative needs of the sea-bass are very similar if not identical to those of other teleosteans and land vertebrates. For the japonese sea-bream, SAKAMOTO and YONE, 1972, employing the most classical technique of purified diets without amino-acids, showed clearly the essential and non-essential amino-acids for these species; cystine and tyrosine are not essential. No information in this sphere on Sparus aurata is known. 1.2.2 Quantitative needs of essential amino-acids The amount of essential amino-acids could be defined in many ways, by employing diets, lacking the essential amines-acids and by adding progressively this substance or by employing more or less mixed proteins. Both methods don't necessarly give the same results, as pure amino-acids are not always properly employed by fish (THEBAULT, 1983). Although the real needs are the quantities expressed per day for an animal of given weight and of physiological characteristics, the "needs" are often in fact, the food percentages permitting the maximum growth of the animal. By using the growth speed as criterion , LUQUET and SABAUT, 1973, have been able to define as following for sea-bream the lysine, sulphur amino acids and tryptophan needs. The authors find no better performances when the diet contains more than 1,5 % of arginine. This could signify that there is little need of this essential amino- acid or that its degree is perturbed by the particular part that arginine plays in the urea cycle, in relation with osmoregulation. It is a common known fact that with trout, the arginine need decreases when the fish is changed from fresh water to salt water (KAUSHIK, 1979). These essential amino-acid needs are very similar to those found for the quinnat salmon (Oncorhynchus tshawytscha) apart from arginine, which seems to indicate that the similarity between the needs, which is evident from a qualitative point of view is also applicable from a quantitative point of view. Sea-bass require fewer essential amino-acids, but the study carried out by THEBAULT, 1983, from a different criterion must be remarked: the plasma tenor and free amino-acid tissues. This author shows, that with a diet containing barm and purified soya protein, the need is 1,2 to 1,3 % of the food which is 18 to 20 mg per day and per 100 g of live weight. The other essential amino-acids must be studied individually: In default of this work, it should be better to adopt the balance of the essential amino-acids of the ideal protein, obtained from a hen's egg or that of the "average- protein" of the species considered; both methods furnish debatable results. 1.2.3. Proteinic quantitative needs With superior vertebrates which are reared at industrial scales, the diet formula is usually estimated, by taking as basis, the essential amino-acid need expressed per unit of metabolisable or digestible energy. This method however assumes that there be a sufficient amount of normal amino-acids so that the proteinic anabolism is not limited. To estimate the "true" (daily necessary supply) total protein needs, the maintainance needs and growth needs could be measured. The first measure was carried out by ALLIOT (1982) employing sea-bass and the need would be from 390 mg. per 100 g.-1 of the metabolic weight. However, the proteinic need of growth expressed per g. of proteinic gain has not yet been established It is very difficult to define, because of the double role played by proteins which are at the same time a nitrogenous and an energy source. 1.2.4. In conclusion, the quantitative "need" of serranidae and sparidae in essential amino-acids, is only roughly known. They have yet to be expressed in function of the food energy while the age of the animal must be taken into account, as is applied with superior vertebrates. From a practical point of view, as long as great quantities of biological high value proteins, such as animal meal, are employed, the essential amino- acid need is more or less covered when the proteinic need is adequate. In other words, all proteins become limiting more quickly than amino-acids when the total nitrogenous matter level is lowered. The need of one of these elements, independantly of the others, can be determined however and numerous studies shall have to be reexamined if proteins and amino-acids are to be cut to a minimum. 1.3. Fatty acid needs Since the works carried put by CASTELL (1972) which show how necessary serie w-3 fatty acids are, (the linolenic acid family) numerous studies have been carried out on this problem, sea-bass and sea-bream included (see review by COWEY and SARGENT, 1977; CASTELL, 1979). It is only possible to cover the essential amino-acid needs of this serie, which are mostly poly-unsaturated long clain fatty acids 20: 5 w-3 et 22: 6 w-3, by supplying directly these same fatty acids (YONE and FUJII, 1975, a and b, FUJII and YONE, 1976). Indeed, on the contrary of that remarked with trout for example, the desaturation elongation mecanisms permitting the synthesis of these fatty acids from linolenic acid (18: 3 ,w-3) are quite insufficient (YONE, 1978; WATANABE, 1982). Excess of linolenic acid has visibly a bad effect on the lipidic metabolism and consequently on the growth of fish. From a quantitative point of view the japonese sea-bream does not have a great need of serie w-3 essential fatty acids. YONE, 1978, estimates them at 0,5 p in the diet, the latter containing 2,5 % of cod liver oil or 5 to 6 % of fish oil about. The role played by serie w-6 fatty acids has not yet been clarified for the fish by which we are concerned. It has been proven that these fatty acids which are the principal essential fatty acids of land vertebrates, play a role somewhat similar for fish, they can be the origin of the synthesis of certain prostaglandines for gold fish (HERMAN and al, 1984). Generally, feeble amounts are incorporated into triglycerids and also into phospholipids. However, if there are great quantities present in the food and if serie w-3 essential fatty acids are lacking especially, growth is delayed and the lipid composition is totally modified. It is unknown if sparidae and serranidae have a need of these fatty acids and precise data, on the tolerable maximum, is also lacking. From a practical point of view with serranidae and sparidae the w-3 and w-6 series (if necessary) of essential fatty acid needs are always covered if the food contains an amount of oil corresponding to the optimum which is empirically established, as long as oil of marine origin is employed. It seems feasible to use vegetable oil rich in serie w- 6 fatty acids once the needs of polyunsaturated long chain fatty acids have been satisfied (YONE and al, 1971) but the consequences on growth and body composition have yet to be established. Another suject calls for reflexion: what happens about the needs of the sea-bass when it is reared in low salinity as the w-6/w-3 relation increases generally with euryhalin fish when they are changed from sea water to fresh water (see CASTELL, 1979) ? 1.4. Vitamins The role and the need of vitamins by fish have been thoroughly studied, especially for salmonidae (HALVER, 1972) Fish have the same vitamin requirements as land animal with the possibility of three exceptions: Vitamin D, vitamin K and paraminobenzoic acid for which a direct role in the physiological metabolism of the fish is still unsure. In the other cases, the only particularities of fish, concern the symptoms of deficiency which not only differ from those known for land animals but seem to vary from one species to another. Other evident incertitudes are found concerning the quantitative needs: The present values published are often overestimated, which means rather the normes or recommandations than the true needs. The work carried out by GODELUCK, 1983, on vitamins for sea-bass is very complete. Employing simplified diets (not purified) containing or not the vitamin in question, this author was unable to deprive the sea-bass juvenile of ribor-flavine, pyridoxin and even vitamin A and E, which seems to indicate that there is overstimation of the needs or an underestimation of the ingredient supply. With vitamin C it is different It is known that the ingredients employed in the usual composition of food do not contain vitamin C and moreover that pure ascorbic acid added to this food is always destroyed partially by the normal technological procedures. By this fact, it is difficult to determine the true need of sea-bass. To allow for losses during manufacture and stocking, the same supply as that employed for salmonidae is recommended: 500 mg/kg of food. For japonese sea-bream, research work has been started by the japonese author (YONE, 1976). After studying the effects that a semi-purified diet lacking in a vitamin each time, these authors point out the essential role of group B vitamins sensu stricto such as thiamine, riboflavine, pyridoxin, pantothenic acid and vitamin B 12 together with choline, inositol and ascorbic acid. The results obtained on para aminobenzoic acids, folic acid and biotine are inconclusive; but for the two last vitamins mentioned it is known that small quantities are required and maybe the experimental period was too short (102 days) to induse a net deficiency. The diet employed by YONE (op.cit.) containing fish oil, make it impossible to carry out studies on group A vitamins. To our knowledge no study of this type has been carried out on sea-bream (Sparus aurata) However the description given by PAPERNA and al, 1980, on the systemic granuloma which has been since linked with the deficiency in vitamin C for turbot, (TIXERANT and al, 1984) must be taken into account. There are few measures of quantitative needs available. For sea-bass they are practically inexistant: for sea-bream they are limited to inositol and pyridoxine. The inositol requirement for growing fish is around 0,05 to 0,1 % of the diet (YONE, FURUICHI and SHITANDA, 1971) while the pyridoxin requirement is around 0,2 - 0,5 or 0,5 - 0,6 mg/kg-1 depending on what criterion is taken, the weight gain or the hepatic transaminasis. Activity (TAKEDA and YONE, 1971) It must be remarked that these values are only slightly inferior to those obtained for the same vitamin B 6 with Sparus aurata by KISSIL (1979). Finally it can be said that little is known concerning the quantitative requirements in vitamins for serranidae and sparidae. The works carried out by GODELUCK (op.cit.) accredit the thesis by which the sea-bass has fewer requirements than was supposed, even fewer than those for salmonidae. Assuming this, it should be better to aim all research work on vitamin E which has the antioxydizing role of intervening in the food itself (where it is possible, avoid using the synthetic antioxydizing agent) and at tissue level, together with vitamin C, which in artificial food is often deficient. However, the exact importance of the requirements in vitamins A and B must be defined prudently until precise measures have been taken. 1.5. The minerals Although a marine fish has the same inorganic element needs as a land animal or a fresh water fish, it is difficult to say the food requirement of these elements, as for most of these, the result obtained from its organism depends at first, on the branchial and intestinal absorption of water drank, on the renal and intestinal excretions, and only slightly on the intestinal absorption of food (LALL, 1979). The principal exception is phosphorus which although absorbable from water, is found in insufficient quantities to satisfy the needs of the organism. It is due to the research work carried out by YONE (1976) that the nutritional role of phosphorus with japanese sea-bream was proven and the first estimates of needs were given. The works show that the deficiency delays growth and transformation rates, decreases the serum's mineral phosphorus level and bone calcification and alters the body composition. The requirement is 0,6 and 1,38 % of the diet which is a rather high level. It can be remarked that YONE (1976) associates (without insisting) this need to a phosphocalcic ratio (Ca/P) of 0,5 about. As there is a correct calcium contribution by sea-water even if there is no food supply, the signification of this ratio should be taken prudently; which doesn't otherwise signify that there are no interactions between Ca and P in the diet on the phosphorocalcic metabolism. It can be remarked that the works carried out by BACLE (1980), although not as thorough as those by the japonese team, on sea-bream, show however the preponderant role of phosphorus with this species. The only other relatively rare mineral element found in the sea water is iron, which intervenes, as known, in the synthesis of hemoglobin. The comparaison of purified food with or without additives has been carried out by YONE (1976) with the japonese sea-bream. No significant: difference of performance has been remarked, which shows clearly that even when this oligoelement is absent, no severe deficiency is remarked. However with dosages of hemoglobin, the authors show clearly a typical anemia of iron deficiency. No study is known on other minerals, such as manganese, copper or zinc. A deficiency in these elements seems most improbable, but it must be remarked, even with a great food supply, excess in calcium can inhibit the absorption of the oligo elements and so cause deficiencies. The phenomenon, well known with land vertebrates is also liable to happen with fish (LALL, 1979). KOENIG (1984) draws attention to cases found with fresh water fish, fed diets containing fish meal rich in ash. 1.6. Other requirements The different types of needs, accepted by animals have been reviewed. However, fish may not have strickly the same needs as superior vertebrates. Many tests have been carried out where fish fed fresh fish or other living feed had a quicker growth speed than their homologues fed manufactured artificial feed . It has been assumed that other organic molecules (unknown growth factors) could be responsible for these differences. But in many cases natural food is more advantageous due to the distinct property that it contains, not mentioning its nutritional value: appetence. At present it is known that the different attitudes that fish has towards food depends greatly on the small organic and hydrosoluble molecules, such as organic basis, amino-acids, nucleotides; this, when the fish start taking and ingesting feed.Numerous studies on the effects of these compounds have been carried out on japonese sea-bream. (GOTH and TAMURA, 1980, FUKE and al, 1981). In practice these works are still uncertain but it must recalled that the appetence or inappetence must be taken into account. There is little information, on the above for sea-bream Sparus aurata but the first research work carried out by TANDLER and al, 1983, however could be taken into account. 1 7. In conclusion, although disimilar, the studies presented point out a certain number of particularities differentiating the salmonidae from the Mediterranean marine fish, at nutritional levels: there is a much smaller capacity of desaturation-elongation of the linolenic acid; there are greater global proteinic requirements, and it is more difficult to replace the proteinic nutriments by glucids and lipids. However these particularities are still often very vague; no data has been calculated. II - VARIATIONS IN REQUIREMENTS 2.1- The effect of age: the case of larvae The vertebrate juveniles are characterized by a growth rate which decreases regularly with age. By this fact, the relative importance of the maintenance needs rise quickly at the expense of the growth need. With vertebrates, this is clearly remarked by the regular decrease of the optimal amino acid rates as the animal grows. The vitamin and mineral supplies are often increased for young animals, as the energetic level is modified inversely. Salmonidae have the same proteinic requirements (HALVER, 1969-1970) and it is believed that the same applies for sparidae and serranidae. Therefore ALLIOT and Pastoureaud, 1984, estimate that the optimal proteinic rate drops from 60 % with 1 g sea-bass fingerlings to 45-50 % for 20 g juveniles. There is little information available on the increase depending on age, of the essential fatty acid requirements. The works carried out by LE MILINAIRE and al, 1982 and GATESOUPE and al, 1984, show however with turbot that the w-3 serie of polyunsaturated fatty acid requirements decrease considerably from young larvae to juvenile stage, droping from 1,5 to 0,5 % about in the diet. With sea-bass juvenile, GATESOUPE and al, 1984, obtained different survival and growth rates by modifying the feed itself or by enriching it with live prey Brachionus plicatilis and Artemia salina employed to feed larvae. When these experiments were tried out, the effects, of the essential fatty acid rates obtained, were not as good as those obtained with turbot. The authors conclude that the sea-bass larvae has feeble requirements and so they are difficult to define. 2.2.Broodstock There is very little information available on broodstock fish. In numerous rearings these animals are fed fresh fish or other "living feed". When, on rare occasions, artificial feed was employed, vitamins and essential fatty acid were added, from which the egg benefited directly as no precise data was available. Since then, some information is available through the experiments carried out by the WATANABE team. This author pointed out that the japanese sea-bream had feeble requirements in proteins. He pointed out the beneficial effect of w-3 serie polyunsatured long chain fatty acids, phosphorus, carote-noid (carotene or canthaxantine), the negative effect of serie w-6 polyunsaturated long chain fatty acid excesses. He also showed that better results are obtainable (greater amounts of eggs and higher percentage of floating eggs) by adding deep freeze seiche or krill meal to the feed (WATANABE and al,a,b,c,d). 2.3 The effect of the environmental factors The temperature has great influence on the quantitative requirements; but less, as already seem on qualitative requirements. To begin with, it would seem reasonable to neglect the eventual variation of these requirements during the year, apart from the Winter season perhaps , where by adding the appropriate feed, the risks of juvenile mortality are perhaps reduced, although the cause of mortality is not of nutritional origin. There is a great lack of information also concerning the relation between proteinic requirements and salinity. The reinforcement of the tendency of ammoniotelism while lowering salinity levels should promote a decrease of nitrogenous needs with see- bass in brackish water. So that, some small differences were remarked in this direction with salmonidae (ZEITOUN and al, 1973). But it is still risky to apply these with sea-bass or sea-bream. It has been remarked that the eventual consequential effect of salinity on the amino acids and essential fatty acids are still not very well known. 2.4. As long as further studies have not been carried out on the effect of the environmental factors and on age, it will be difficult to really adapt this feed to the type of fish (fingerling or juvenile) to the season or to salinity. There is only one rearing phase which needs for sure a distinct food reproduction. Unfortunatly it is the most ignored physiological stage from a nutritional point of view. III - CONCLUSION The data recorded for the synthesis is quite dissimilar and neither one of the two species by which we are concerned can be considered well known from a nutritional point of view. In many cases other neighbouring species were employed: striped sea- bass and japonese sea-bream. The data recorded on these species seems more applicable for the sea-bass and sea-bream than that recorded on salmonidae. The data recorded on one species must not be automatically applied for another species even if both belong to the same family. The example of salmonidae and certain land animals especially show that the requirement can differ for two very closely related zoological species. It is difficult to say whether the strict carnivorous and non amphibiotic character (although the sea-bass is euryhalin) of both Mediterranean species is responsible for all the particularities which differenciates them from salmonidae (glucids not well tolerated, greater requirement of phosphorus and especially the incapacity to synthetize w-3 serie poly unsaturated long chain fatty acids from linolenic acid) It can be confirmed however that these differences are not to be disregarded and that feed permitting the satisfactory growth of trout need not give the same results with sea-bass or sea-bream, although the opposite may not be true. As these great differences are known, it should be advisable to give precise quantities so to permit the feed manufacturers to make up suitable diets for each species, and this at low cost. Unfortunatly the data recorded is still insufficient and often concerns other neighbouring species. 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Nutrition azotée et croissance chez la daurade et la truite. In Actes de Colloques n° 1, Colloque sur l'Aquaculture, Brest, Oct. 1973, 243-253. METAILLER R., ALDRIN J.P., MESSAGER J.L., MEVEL G., STEPHAN G., 1981. Feeding of european sea-bass Dicentrarchus labrax: role of protein level and energy source. J. World Maricul. Soc. 12, 117-118. METAILLER R., FEBVRE A., ALLIOT E., 1973. Note préliminaire sur les acides aminés essentiels du loup ou bar, Dicentrarchus labrax Linné. Etud. rev. C.G.P.M 52, 91-96. PAPERNA I., HARRISSON J.G., KISSIL G.W., 1980. Pathology and hispathology of a systemic granuloma in Sparus aurata L. Cultivated in the gulf of Aqaba J. Fish dis. 3, 213-221. PERES G., 1981. In nutrition des poissons (op. cit.) Enzymologie digestive. I Les prothéases, l'amylase, les enzymes chitinolytiques, les laminari-nases, 55-67. PIONETTI J.M., 1984. Exigences environnementales et indicateurs physiologiques et biochimiques au cours de l'ontogenèse de poissons marins en Aquacul- ture. In BARNABE G. et BILLARD R. Ed. L'Aquaculture du bar et des sparidés. INRA Publ. PARIS, 1984, 127-137. ROZIN P., MAYER J., 1961. Regulation of food intake in the gold fish. Am. J, Physiol. 231, 968-974. RUMSEY G.L., 1977. Fish nutrition. Recent advances. Proc. Int. Symp. Diseases of cultured salmonids. Seattle WH? Washington 4-6 april, 16-40. SABAUT J.J., LUQUET P., 1973. Nutritional requirement of the gilthead bream, Chrysophrys aurata, quantitative protein requirements. Mar. Biol. 18, 50- 54. SAKAMOTO S., YONE Y., 1972. Amino acid requirements of red sea-bream, cité par YONE, 1976. SHIMENDO S., HOSOKAWA H., TAKEDA M., 1979. The importance of carbohydrate in the diet of a carnivorous fish. In Proc. World Symp. on Finfish nutrition and Fishfeed Technol. Hamburg 20-23 juin 1978. Berlin 1979. I -127-143. SPYRIDAKIS P. et METAILLER R., 1985. Communication personnelle. TAKEDA T., YONE Y., 1971. Studies on nutrition of red sea-bream. II - Comparaison of vitamin B6 requirement level between fish fed a synthetic diet and fish fed beef liver during prefeeding period. Rept. Fish Res. Lab. KYUSHU Univ. n° 1 - 37-47 cité par YONE, 1976. TANDLER A., BERG B.A, KISSIL G.W., MACKIE A.M., 1982. Effect of food attractants on appetite and growth of gilthead bream Sparus aurata. J. Fish Biol. 20: 673-681. THEBAULT H., 1983. Etude du besoin en methionine chez le loup, Dicentrarchus labrax en milieu contrôlé. Thèse de doctorat 3ème cycle en 0céanologie. Université d'Aix-Marseille II - 101 pp. TIXERANT G., ALDRIN J.F., BAUDIN LAURENCIN F., MESSAGER J.L., 1984. Syndrome granulomateux et perturbation du métabolisme de la tyrosine chez le turbot (Scophthalmus maximus). Bull. Acad. Vet. de France - 57, 75-85. GOH Y., TAMURA T., 1980. Alfactory and gustatory responses to amino acids in two marine telosts. Red sea bream and mullet. Camp. Biochem. Physiol. 66 C, 217-224. YONE Y., 1976. Nutritional studies of red sea bream. In Proc. First Intern Conf Aquaculture Nutrition. Newmark, Delaware, University of Delaware 39-64. YONE Y., FUJII M., 1975. a. Studies on the nutrition of red sea-bream. XI - Effect of 3 fatty acid supplements in corn oil diet on growth rate and feed efficiency. Bull. Jpn Soc. Fish 41. 73-82. YONE Y., FUJII M., 1975. b. Studies on nutrition of red sea bream. XIII -Effect of 3 fatty acid supplement in a corn oil diet on fatty acid composition of fish. Nippon Suisan Gakkaiishi 41, 79-86. YONE Y., FURUICHIM., SAKAMOTO S., 1971. a. III - Nutritive value and optimum content of lipid in diet. Rep. Fish. Res. Lab. KYUSHU Univ. I, 49-60. YONE Y., FURUICHI M., et SHITANDA K., 1971. b. Vitamin requirements of red sea bream. I - Relationship between inositol requirements and glucose levels in the diet. Bull. Jap. Soc. Sci. Fish. 37, 149-155, cite par YONE, 1976. WATANABE T., 1982. Lipid nutrition in fish. Comp. Biochem. 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N° 194. 191 pp. WOO N.Y.S., MURAT J.C., 1981. Studies on the biology of red sea bream. Chrysophys major. III - Metabolic response to starvation in different salinities. Mar. Biol. 61, 255-260. ZEITOUN T.H., TACK P.I., HALVER J.E., ULLREX D.E., 1973. Influence of salinity on protein requirement of rainbow trout (Salmo gairdneri) fingerlings. J. Fish Res. Board Can. 30, 1867-1873. Anonyme, 1981. Nutrition des poissons. Actes du colloque C.N.E.R.M.A Paris, Mai 1979. Editions du CNRS, Paris. QUALITY CONTROL OF FEED IN PISCICULTURE Ms M. AMERIO INTRODUCTION In intensive pisciculture, feeding, although dating back centuries in the economic achievement of fish rearing for commercial purposes, is more or less a recent science when speaking in terms of: definition and covering of nutritional requirements, distinction of the feed particularities, preparation and distribution techniques of feed. The cost of feeding now represents, as far as rearing prize species is concerned (sea-bass, trout) 60 % of the total cost of management. KLENTZ G. W. (Verone, 1982) (1) shows clearly an example of the high cost attributed directly to feeding: the weight growth of a group of fish having a consumption index (= gain in body weight/dry feed injested) of 1,9 kg of feed for 1 kg of fish, is more expensive ( in terms of feed costs) than a same group of fish having a consumption index equal to 1,5. There are numerous reasons for these high costs: high mortality rates during the different rearing phases, the high technology applied for the preparation of the feed, the use of raw materials of animal origin, having a high unitary cost (special fish meal and oils). Fresh and marine water fish, intensively reared, can but take a partial benefit from the natural feed of the environment, consequently they mostly depend on artificial feed. Besides, fish are a "fantastic biological machine" as they have the great capacity of converting feed into flesh: they are capable of converting 1,5 to 2 kg of food into 1 kg of flesh, whilst for the same production, chickens require 2,3 to 2,4 kg and bovines 5 to 7 kg (2). This great conversion capacity, well explained by the fact that as fish are heterothermic, they do not use up energy to keep a constant body temperature, and also because of their specific weight, not much effort is demanded of them for movement. There is also a smaller demand of energy for the energetic metabolism of excretion for fish, when compared to other species: the product from the nitrogene excretion is ammoniac instead of urea. For the formulation of low costing compound feed, the feed requirements of the animals (concerning proteins amino-acids, lipids, etc... ) must be known, along with the characteristics and nutritive content/of the feed, and the price of each ingredient. It is still unknown exactly, what use each species makes of each of the different feed components, and so quite often, the feed is calculated on the presumption that the food value of a substance is the same for fish as that for monogastric animals, or that an ingredient can be replaced by another, when only taking into account the nutritive content. It is therefore the quality of the feed which essentially plays the determinant part. Feed must be taken in a larger sense: as carrier of nutritive principles and substances which have an oligodynamic action, but the possibility of feed having eventual toxic actions or more precisely, which have an antinutrional action must be taken into consideration also. From this point of view the feed not only becomes a carrier of indispensable substances for body growth and maintenance but also a modulator of the organic defences and a way of immunity. PROTEINS The feed naturally ingested by fish (fish, crustacae, mussels, etc... ) contains high concentrations of proteinic substances (from 50 % to 70 % of the dry matter) The artificial feed scheduled for carnivorous fish is also manufactured in such a way that it contains at least 40 % of raw protides (N x 6,25). Therefore fish have a great proteinic requirement when compared with other monogastric animals (chickens, pigs). The great proteinic requirement can be justified very synthetically, by the three following factors: – slow proteinic speed at muscle level (which make up 60 % of the whole body) when compared with other animal species. 3 g of proteins /g of R.N.A (ribonucleic acid) are synthetized per day. There is a much greater proteinic synthesis speed (10 to 20 times more) at liver and gill level. – the high rate of amino-acids which are directly utilized by the oxidation processus for the energy supply and which can consequently become the limiting factors at proteinic synthesis level. – the limited capacity of the use of carbohydrates at digestion level. LUQUET (1975) (3) draws attention to the, fact that the great doses of proteins, when feeding trout, can be a great waste. It must be considered that out of 100 g of proteins distributed to trout, 10 to 15 g are eliminated through excrements, 30 to 40 g through the gills and the urine, under the form of ammoniac. This evident bad use of proteins could be caused by a bad proteinic balance, but also by the fact that a great part of the proteins are used for energetic reasons. Lipides and carbohydrates also, can have an economic effect on proteins, but it must be taken into account the fact that lipides are expensive and that over a long period their use could create problems in the agglomeration preparation not forgetting the damage of hepatic character. At least 50 % of proteins in fish feed is of animal origin generally (fish meal, blood meal, meat meal, lacto-serum) having a good biological value. The biological value of a protein (THOMAS, 1909), (4) is represented by the percentage relation between the nitrogene contained in the organism and that really absorbed). The formulation of fish feed also comprehends proteins of vegetable origin (soya meal, cereal by products, cotton meal). It is widely known that carnivorous fish make better use of proteins of animal rather than of vegetable origin, and as this has been remarked as far as trout are concerned, who are unable to use non proteinic nitrogen, urea, ammonium citrate, this last product can even be toxic. The digestibility of proteinic fish feed is influenced by numerous factors (directly concerning the fish or the feed - species, age, water temperature, ingestion, the treatment the feed received), in any case it is greater for proteins of animal origin that for those of vegetable origin (biological value feed table). The nutritive value of proteins, especially those of vegetable origin, can be greatly improved by thermo-mechanical treatments, such as compressing, flaking, extrusion, etc. The compressing treatment consists in forming cubes which can receive a more or less long thermic treatment (dry or steam from 1 to 10 minutes); The flaking thermic treatment consists in steaming under high pressure (1,5 to 7 athmospheric pressure) over a very short time (a few minutes) which is then followed by a compressing treatment. The extrusion treatment is the preparation of the feed (grinding and adding water) followed by the cooking phase and setting of the product, (a homogeneous paste is then obtained and pressed through die holes. All the physical chemical phenomena which are verified during the different phases of treatment -gelatinization or dextrinization of the starches, denaturation of proteins, inactivation of the anti-nutritional factors- are closely linked to each of the variable factors concerned: moisture, temperature, pressure, duration of treatment. The thermic treatments cause the denaturation of proteins, so facilitating the hydrolytic action of pepsine and tripsine (proteolytic enzymes) and quickening the digestion of the protein. To obtain the correct estimation of a protein, the amino acid compositions must be known along with their availability when the feed reaches the mouth of the animal, after the necessary manipulations have been carried out (drying, grinding, cubing, extrusion, etc... ) For this, it seems necessary to carry out a correct analysis of the proteinic amino acid composition (a) Tryptophan is destroyed by the hydrolyse acid of proteins, this is why a specific hydrolyse is carried out in an alkalin environment. Methionine itself is affected by the hydrolyse acid treatment, as it oxidizes and changes into sulphonated methionine. Great quantities of sulphonated methionine can be found in the protein, if this latter has undergone, during food manufacture transformation, an oxidizing or thermic treatment; so a measurement of methionine as sulphonated methionine should be made. COWEY C.B. (1978) (5) precises that the oxidized form of methionine could be utilized by fish, but this is not certain. The thermic treatments, if carried out in certain humid concentrations and on raw materials which have a good sugar content, bring about the formation of the MAILLARD products (between amino group of lysine and a radical glycoxydic). Following this chemical reaction, the lysine present in the feed is not completely available biologically. An exact estimate of the qualitative and quantitative composition of the food protein permits the verification whether or not the protein can satisfy the amino-acid needs of the species. The necessary amino-acids for salmonidae and for sea-bass are as following, arginine, histidine, lysine, isoleucine, leucine, methionine, cystine, phenylamine, tyrosine, threonine, tryptophan, valine. For fish, the isoleucine/leucine relation is important, as it must be always inferior to 1: 3. The knowledge of the amino-acid composition permits the application of certain calculation methods: the chemical rate, the index of essential amino acids by OSER; the index of the proteinic balance by ARNOULD, which give the global estimate of the protein, without carrying out biological research, which is very difficult and costly when concerning fish. It must be remarked that the different methods of calculation are restricted to taking into consideration the limiting amino-acids, evaluating the proteins by respecting a minimum, or evaluating the proteins by comparing between their essential amino-acid patrimony and that of whole egg protein (taken as the protein of comparaison) Therefore this is not only a problem of covering the need in the different amino- acids but a problem of the amino-acid balance, between the essential and unessential amino-acid rates. The synthesis of a protein is a action carried out continually at the different organ levels and the speed of which depends on the concentration of the monopeptides which react well and are proportional to the product concentrations. From one organ to another, varies the speed of the synthesis and that of a turn over change.according to the specific characteristics of the organ, for instance it is greater at liver level than at muscle level, ARNOULD (1971) (6) following a series of experimental controls, proposes an evaluation system of proteins, based on the calculation of the identified needs. ARNOULD's amino-acid balance index is obtained in fact by the product of the relations between the different essential amino-acids of a given protein (egg protein) and the amino acid protein examined. In the calculation, the unessential amino-acids are taken globally into consideration. The more ARNOULD's index approaches 1, the greater, from a biochemical point of view, the speed of the global synthesis of proteins and the less the waste caused by imbalances. The amino- acids causing the great imbalance, are shown clearly by their relation (the further they are from 1, the greater the imbalance). Normally, the fact that the whole rate of amino acids contained in feed is only employed partially is not taken into account; the concept of amino acid availability is identified with the concept of digestibility, in the sense that this is normally an integrant part of the proteinic structure, it becomes available at absorption level only when the proteins have been digested. However there is no perfect superposition between the concept of digestibility and that of availability for the interferences in the digestive system, but with fish, on the contrary to what happens with pigs, there is little interference in the intestinal microflora on the amino acid rates and the transit time of food is rather quick. Finally it must not be forgotten that the qualitative control of food, rich in proteins of animal origin must schedule the measurement of the amines-biogenes (histamine, putrescine, cadaverine, etc... ) in other words, the composites which can form after the processus of decarboxylation of the amino-acids. This parameter which also furnishes a measure of the quality of food conservation, is important. The amino-by products can create a toxic or pharmacological effect on the animals. ANTINUTRITIONAL EFFECTS INTRINSIC ANTINUTRITIONAL FACTORS These are factors linked with the specific characteristics of a vegetable, varying in the same species from "cultivar to cultivar" and under genetic control. Some of these are thermolable and the appropriate treatment could reduce them remarkably. EXTRINSIC ANTINUTRITIONAL EFFECTS These are molecules of natural origin and have a synthesis which can contaminate food more or less accidentally. This accidental contamination is partly due to the technological evolution; what is meant by this, is the residue and metabolic waste from pesticides, and partly the metabolic waste from micro-organisms which can develop in foodstuff. The ingestion of substances which have an antinutritional effect, causes, apart from lower performances, a phenomenon linked with acute toxicity: hemorragies and death, or a phenomen of chronic toxicity: alteration in tissues, abnormalities of the vital organs, such as the liver and kidneys. a) EXTRINSIC ANTINUTRITIONAL EFFECTS: – Heavey metals – Pesticide residue – Mycotoxins Heavy metals The risks of contamination of raw materials and of ready made foodstuff, caused by heavy metals must not be neglected above all, if the high contamination of the area is taken into account, which for some, lead especially, can cause great damage. It is widely known that the average level of lead has greatly increased, and unfortunatly, it can be confirmed that in industrial countries, plants or areas having a "natural" level of lead are inexistant. Amongst the heavy metals which can be of risk to fish, let us state, apart from lead, arsenic, cadmium and copper. Selenium, when in high concentrations can also have a toxic effect. It is difficult to say precisely the specific effect of each metal. They have without doubt, specific consequences on the immunity system and on the liver and kidney tissues. As for arsenic found in trout, doses of 7,5 mg/100 g of dry matter over a period of 12 months, cause hepatomes. (7) Following ingestion or contact with certain doses of arsenic, alterations in the gills and kidneys may be noticed. The ingestion of hexavalent chrome has a direct consequence on the intestine. The consequences of copper excess are: congestion of the gills, hyper excitability, an acute reaction to the infectious effect of Gram + bacteria; a 1 mg /g dose of the dry matter in the food diet causes a delay in the normal pigmentation of the trout. Pesticides A lot has been written on this subject, which should be taken into account. It seems advisable to recall that with pesticides, as with other toxic substances a remarkable noxious effect on young animals is noticed. The abnormalities remarked, lead from the simple loss of appetite to nervous disorders and sterility, etc... Mycotoxin Mycotoxins are metabolites, produced by fung uses. The toxic substances of mycelion origin, differing from those of bacterial or vegetable origin, do not give rise to the production of antibodies and consequently immunity. They have various effects on the organism of animals (neurotoxic, nephrotoxic, hepatotoxic, hemotoxic, dermotoxic, enterotoxic, osteotoxic and immuno-suppression). There are more than one hundred mycotoxins, and syndromes of unknown etiology can be attributed to them. Those studied concerning fish are aflatoxins. By this term is meant a group of metabolites, produced by Aspergillus flavus and Aspergillus parasiticus stocks, during their growth on the foodstuff. Following research work carried out in the states,. foodstuffs containing more often aflatoxins are: corn cotton grains, flour extracted from cotton, pea-nut flour, rice, soya beans, etc... The tonic effect that aflatoxin B1 has on trout was given special attention. JACKSON (1963) (8) showed the hepatocancrogenic effect of aflatoxin B1, given out in doses of 0,1 to 0,5 pp over a time laps of 4 to 6 months. Acute effects: hemorragies and hepatic necrosis are remarked within 3 to 10 days, when 0,5 mg/kg of body weight is given. Aspergillus flavus develops quite easily in pea nut and cotton grain flour (especially when these foodstuffs are stored in depots of holds of ships). b) INTRINSIC ANTINUTRITIONAL FACTORS Following their chemical make-up, the antinutritional factors of intrinsic origin can reduce the digestibility of proteins and of polysaccharides specifically or not inhibit the digestive enzymes, necessary for the decomposition of the macromolecules and/or inferior to the absorption processus level of nutritive substances. Tannins There are aromatic substances of phenolic type. They are found concentrated in the external layers of certain cereal caryopsises (sorghum, barley). Consumption of food rich in tannin causes the reduction of the body growth and the use of proteinic nitrogen. The effect is caused by the capacity that tannin has, of linking itself in a non-specific way to proteins, by forming resistant complexes to the protease actions of the digestive system. They can have a cancrogenic effect: HALVER (9) shows that doses of 7,5 mg /100 g of dry matter in the food diet causes hepatic histological alterations which bring about an hepatic tumer. Phytic acid or inositol hexaphosphoric acid and its salts Phytates are widespread in cereals and can make up 35 to 97 % of the phosphoric content, which in this form, can not be assimilated either by animal or man. Phytates play an antinutritional part, as they can interfere with the absorption of certain mettalic ions such as calcium, iron, magnesium, zinc, in forming insoluble complexes which are evacuated in the excrements. They can also link themselves with proteins as shown by SPINELLI J (1983) (10).With trout fry, fed diets which have a 0,5 % phytic acid level, a reduction of 66 % in the digestion of proteins was remarked, together with decreases of around 10 % in growth and the inconsumption ratio of the feed. Lectins Also known as hemoagglutins or phytoagglutins: These are macromolecules, which, depending on the type, agglutinate different types of isolated cells. Studies carried out on leguminous lectins have proven the capacity that these substances have of interfering in the absorption of nutritative substances, following the interaction with the epithelial cells of the intestinal mucous membrane. Resorchinols By this term is meant, the different alchilic by-products of resorcinol, containing uneven numbers of 15 to 23 carbon atoms in the alchilic chain. These componants, partially thermolable are found present in variable quantities in rye, wheat and tritical. It was found by certain scientists that resorchinols play a possible antinutritional role, and have been stated as responsible for the loss in appetite and the decrease in growth, remarked in numerous animal species when given diets which have a high rye content. Gossipole The cotton grain meal contains a variable 0,03 to 0,2 % of gossipole: phenolic type pigment, responsible for the toxic phenomenon remarked in monogastric animals. This pigment apart from being directly toxic, can link itself to lysine and consequently decrease the possibility of using proteins. It has been remarked that trout show a toxicity phenomenon caused by gossipole: anorexia and ciroide fluid in the liver. Amylasic inhibitors In wheat, barley and rye, are found great quantities of proteinic molecules, capable of inhibiting the amylasis (enzyme responsible for the destruction of the polysaccharides of the food diet). In wheat caryopsis, the amylastic inhibitors make up about 2/3 of the albumin content and around 1 % of the proteinic content of wheat meal. They are found in the endosperm of the caryopsis. From a nutritional point of view, the characteristic resistance of these inhibitors to thermic treatments and to the trypsine effect is interesting; these properties probably depend on the remarkable capacity of the proteinic molecule stabilized by the numerous dissulphuric bridges; Research carried out on chickens, fed diets enriched in antiamy-lastic factors of wheat proved in the decrease of growth and the hypertrophy of the pancreas. The amylasis activity of trout and the reports with the anti-amylasic factor of cereals, is being investigated at present at my research institute within the frame of the finalized programmes of the C.N.R. (Centre National de Recherches). Proteasic inhibitors The most commonly known, are those of soya. It can be said that they are found in all leguminous, even if in variable amounts. Raw soya meal contains 1,4 % of the KUNITZ trypsic inhibitor and 0,6 % of the BOWMAN-BIRK chymotrypic inhibitor: the former is thermolable, the second (great inhibitor of chymotrypsin) is relatively thermostable. They cause the hypertrophy of the pancreas and a decrease in the growth rate, probably as there is a second deficiency in essential amino-acids (sulphurated amino-acids especially) caused by the hypersecretion of the pancreatic enzymes. Saponins These are complex glycoxydes of triterperoid alcohols, found in soya at a rate of 0,5 % about of the dry matter. By their polarity, they are insoluable in dissolvents such as hexane and consequently they remain present in soya extract. Saponins cause the bitter taste found in soya and have a hemolitic activity. In normal conditions they are not absorbed at intestinal level, while they are hydrolised by the bacterial enzymes in the sacrum and colon. Glucosinolates They are chemically made up of glycoxydes which have sulphur atoms in their molecules. They are generally found in the parenchymal tissue of crucifer grains. They have a concentration of around 4 to 10 % in whole grains. From the hydrolysis of glucosinolates, made up by the enzyme known as mirosinasis, present in the grains, toxic compounds can be liberated. The mirosinasis enzyme is also synthetized by part of the bacteria which can live in the symbiosis of the subjects intestine. Industrial treatment adopted in the preparation of the colza meal can destroy this enzyme. Crude fibre Naturally found in foodstuff, it can mean in the case of fish and especially carnivorous fish a factor of antinutritional action, as it is hardly destroyed at all, due to the reduced dimensions of the digestive tube and feeble action of the cellulosolitic microflora there present, it reacts as an inert material, raises the transit speed of the food and reduces the possibility of using the other components found in the food. Otherwise it can have a direct physical effect on the absorption of the digestive enzymes. LIPIDES By the term lipides (commonly known as fats) is meant the numerous substances, of different chemical nature, characterized by common properties, among which is found their insolubility in water, their solubility in organic solvents and their presence in the molecule of radical fatty acids. Lipides have different important functions: 1) The structural components of the membranes 2) The reserve and depositing substances 3) The components of the cellular surface which play a part in the determination of the immunity of the tissues. 4) and last but not least, some of them have essential biological activities. From a biological point of view, lipides can be subdivided into two groups: depositing lipides and cellular lipides. The depositing lipides are found in the differenciated cells, also known as fatty cells (adipocytes); they are a reserve material (they decrease during fasting) or they have a protective function. The cellular lipides extract themselves with more difficulty from cells (as they are often linked to proteins) and they do not diminish greatly during fasting. Lipides comprise of the following: triglycerides, phospholipides, sfingomyelins, sterols and waxes. Fats are the most important source of energy. The energetic reserve, furnished by lipides permit the fish in their natural environment, to answer the needs during certain physiological moments -fasting - reproduction and migration. Before speaking of fats from a nutritional point of view, one of the most important roles that they play for fish must be remarked: phospho-lipides of cellular membranes, are responsible for the cellular exchanges and of the flexibility of the membranes, depending on the temperature and the pressure of the water. To estimate the food value of lipides in the diet, certain factors must be taken into consideration: 1) digestibility 2) The presence of toxic substances 3) Oxidizing level 4) Essential fatty acid content 5) Maximum level tolerated by the animal in question The digestibility of fats by fish is greatly connected to the fusion point of the fat itself and consequently to the degree of saturation. The insaturable fatty acids are more easily digested than saturated ones (NOSE, 1966) (1) Experiments carried out with salmonidae, fed diets containing saturated fats showed clearly, how these are the cause of what is known as the ciroid degeneration of the liver, which was pointed out by GHITTINO (12). The formation of the ciroide in the liver is a complex phenomenon and has multiple effects. Fish species are capable of digesting high quantities in the diets (20 to 30 % of the food) as long as there are adequate quantities of choline, methionine and vitamin E. Recent research work carried out showed that the digestibility of fats is not dependent on the temperature, at least where trout is concerned. (13) Ingested lipides are hydrolyzed in the digestive system by lipases and phospholipases; the fatty acids are thus liberated and afterwards they are metabolized in the liver- At this point, different metabolic processes occur: direct oxidation in the aim of producing energy, conversion into other fatty acids (elongation phenomena or dehydrogenation of the carbon chains). The liver of fish plays an important part for the deposit of fats, which is quite different from with what takes place with other animals. Lipids depositing in the viscera and tissues of fish are characterized by great quantities of carbon long chain fatty acids (20-22) and containing up to. 6 double vonds in the chain. The greater part belongs to the linolenic group (ω 3); some differences are remarked, depending on the fish species and the environmental factors. Fresh water fish lipides contain more ω 6 fatty acids when compared to marine species (ω 6/ ω 3 is equal respectfully to 0,37 and 0,16 for fresh and sea water species. Fish phospholipides are particularly rich in long chain poly-unsaturated fatty acids (PUFA) These are necessary in the food diet, where they increase the food utilization. Concerning the food for trout, at juvenile stage, it should contain 1 % of (ω 3) linolenic acid. It must be remembered that marine species have a greater requirement of 3 serie. Fish have the capacity of elongating the carbon chain of fatty acids, this depending solely on the precursors presence for the ω 6 and ω 3 fatty acid series in the food. This metabolic system seems to be modulated by the concentration of some unessential fatty acids in the diet (oleic acid for example as given by HALVER, 1975) (9). A high percentage of polyunsaturated fatty acids in the food cause difficulties for the stability maintenance of fats, in storage conditions; the reason for which, so as to avoid it becoming rancid, or certain vitamins being degraded and the appearence of toxic phenomena in fish, it is necessary to employ anti-oxidizing substances. The natural antioxidant, and perhaps the most efficient, is alpha-tocopherol- aoetate (vitamin E) which by oxidizing easily, protects the lipide rate from such a risk. The integration moment of the vitamins in the food must be taken into account. The oxidization of fats can be the cause of serious problems when feeding fish: it is a reaction which can be verified quite easily -contact with oxygen- favoured by the presence of certain substances or metals which catalyse this reaction. Along with developing toxic substances, the fact that the fats becomes rancid can reduce the use of the other components in the feed (liposoluble vitamins, proteins). The consequence of injesting oxidized food is all the more serious for trout fry as for sea-bass fry. Recent experiments carried out by GHITTINO, CORBARI and AMERIO (14) demonstrate clearly that after a relatively short period (20 to 30 days) of feeding a rancid fats diet, sea-bass fry have ciroid fluid in the liver, showing anemia symptoms and high death rates. It must be remarked that certain natural oils (ex. cotton oil) toxic substances can be found present, such as cycloprenoid fatty acids (sterculic, malvalic acids). VITAMINS The research work carried out so to define the requirements, activity mechanisms and consequence due to deficiency or excess of vitamins in the diet, dealt especially with salmonidae (15). VITAMIN A: indispensable vitamin, A vitamin A deficiency diet brings about the delay in growth and in bone development; disorders at epithelial cell level are remarked also. Excess of vitamin A cause hyper-vitaminose disorders: Let us recall that the liver of fish is an organ that metabolizes and accumulates fatsoluble vitamins. Vitamin A has a positive action on growth, due to the fact perhaps that it has a protective action on the epithelial cells of the gut, and it allows a good absorption of the nutritive substances and prevents the entrance of pathogenic micro-organisms in the organism. Concerning the stalibility of vitamin A, in the ration it must be remarked that its esters (acetate-palmitate) are more stable when compared to the free form. Vitamin A is sensitive to the presence of peroxides. VITAMIN E: belonging to the tocopherol family, the most important being ∝ - tocopherol. Tocopherols are quite stable towards heat and acids, while they oxidize very quickly in the presence of a nascent oxygen, peroxides or other oxidized products. This is the reason why they play a protective role for fats. This action is found on both the inside and outside of the cell. The vitamin E esters are more stable when compared with free forms. Vitamin E along with selenium and vitamin C, have positive effects on the reproduction of fish specie. VITAMIN B1 (Thiamine): It was the first vitamin to be considered essential for trout. Vitamin B1 deficiency syndrome includes anorexia, neurites, poor growth, increased sensivity to infectious agents. In practice, this deficiency can be found when fish are fed fresh fish: In the fish viscera, there is an antivitamin-thiaminase which destroys the greater part of vitamin B1 present in diet. CHOLINE, INOSITOL: they are essential for the normal metabolism of the fats. VITAMIN C (L-ascorbic acid): Ascorbic acid is readily oxidized in to dehydroascorbic acid. It acts as a biological reducing element in hydrogen transport Vitamin C is involved in different reducing mechanisms: hydroxylation of tryptophan, of tyrosine and of proline. It is necessary for the formation of hydroxy proline which is one of the main constituants of collagen; it plays a synergistic role with vitamin E as an antioxidant; it is necessary in the synthesis of folic acid and increases immunity. Concerning trout, a requirement of 200 mg/kg of food is noticed. When formulating a food diet, so to cover the vitamin requirements correctly, without taking into account the specific needs, the real avaibility of the vitamin factors in foodstuff, the presence of anti-vitamin factors must be taken into consideration. It is also very important to know the vitamin stability present in the food. This is a current and very important problem not only for the particularities of scientific character but also practical. Most vitamins have very feeble molecules which are easily changed in their structure with a consequent loss of certain physical, chemical and biological properties. The main factors effecting the stability of vitamins are as following: Humidity - A great loss in vitamins was noticed in a sample having a greater value of moisture than 8-10 %. Concerning certain vitamins such as vitamine K3, C and B1, humidity can destroy them while increasing the catalytic activity of certain oligo- elements (Fe, Cu, I). In other cases, such as vitamin A, humidity can damage the protective layer and consequently increase their vulnerability to noxious agents. Temperature: Activity losses of vitamins are found proportional to the temperature (increasing with temperatures of 18 to 40 ° c). Other factors: Incompatibility existing between the different vitamins, E.G. vitamin C and B1, between choline and some components of B group vitamins; type of support ( losses are greater with a mineral support or a vegetable by-product support, such as grape skin). Following recent research carried out by Prof. MARCHETTI (1980) (16) in Italy, from a total of 400 samples, it was found that the greater losses concerned vitamin K3 (70 % of that declared), vitamin B1 and vitamin C (50 %). BIBLIOGRAPHY 1) KLONTZ G.W. - Esperienze e nuove acquisizioni nell'allevamento ittico intensivo - Acquacoltura 82 - Verona, 16 ottobre 1982 2) BARBUJANI F. - Convegno sui problemi dell'Acquicoltura in Italia - Chioggia Sottomarina, 9-11 maggio 1983 3) LUQUET P. (1975) - La Pisciculture française - 442 (11a) II trimestre 4) THOMAS - Citato da BORGIOLI E. in nutrizione a Alimentation degli animali domestici - Edagricole 1972 5) COWEY C.B. (1978) in Finfish Nutrition and Fishfeed Technology - Ed. da Halver/Tiew vol. 1, pag. 3-14 6) ARNOULD R. - in Proteines et acides amines en nutrition humaine et animale Ed. De Vyst A. (1972) 7) ASHLEY L.M. in Fish Nutrition - Ed. Halver - Academic Press. (1972) 8) JACKSON (1963) citato da Ashley L.M. in Fish Nutrition - Ed. Halver - Academic Press. (1972) 9) HALVER J.E. - Proc. 8th Int. Cong Nutr. Prague - CZECHOSLAVAKIA 1969 10) SPINELLI J. (1983) - Aquaculture 30, 71-83 11) NOSE T. (1966) EIFAC Fourth Sess., Belgrade, DOC 66/sc II-7 12) GHITTINO P. (1970) - Ittiopatologia, Vol. II - Edagricole 13) AUSTRENG E. (1980) - Aquaculture 19, 93-95 14) GHITTINO L. (1984) - Riv. Ita. Piscic. Ittiop. 19 (3), 95-114 15) PHILLIPS A.M. Jr e BROCKWAY D.R. (1957) - Prog. Fish Cult., 19 (3), 119-123 16) MARCHETTI M. (1980) - Techn. Molit., 31 (3) PATHOLOGICAL ASPECTS OF INTENSIVE REARING Mr G. GIORGETTI Please excuse me for not being able to speak to you in french, this language being more familiar to you. Many thanks for your kind invitation. I will try to resolve this language problem by employing tables and slides. Health, a well accepted factor, desively conditions the economic results of the zootechnical interests; this applies for all intensive rearings, and more specifically fishculture (table 1). It is clearly known that the environment is a fundamental factor for the pathological phenomenon to arise and develop. Environment taken as a stress factor: means the effort made by the animal to adjust to an environmental situation which is not truly appropriate. When one enters into a cattle shed or pig sty one can sense immediatly if something is wrong (too hot, ammonia, etc... ) but in a fish farm, where the environment is water, no abnormalities are remarked although there may exist a bad environmental situation (Tab. 2-3-4-5). The sanitary situation is very important, and will become more important, for at least three reasons, given here under: 1. The objectives of the Management techniques are to increase the amount of products per unit of space. This means a greater concentration of animals; and therefore greater risks of infection; 2. The present environmental situation leads us to believe that the environment is becoming worse rathan than better. 3. At present, genetics in fishculture do not concern health problems but only the conversion and growth rates of the fish, delayed or advanced laying, etc... This caused the fish being weaker and more sensitive to the pathological agents. Sudden mortality, decreasing progressively,is generally linked with environmental situations while mortality which increases progressively is generally linked with disease. To make a diagnoses, the 3 different sources of information here following must be employed: – anamnesis, – inspection, – clinic. Anamnesis A new disease is clearly remarked when the mortality rises and the food consumption decreases. Inspection will permit us to notice that the fish show abnormal behaviour in the tanks, the fish are found on the bottom or edges of the tanks, so avoiding being carried by the stream flow. The fish will show an abnormal reaction to stimuli. Two non-specific lesions are remarked when the clinical test is performed, the fish are suffering and the lesion degree can vary in relation to the disease: exophthalmus and hyperpigmentation. Disease can be caused by: – Virus, – bacteria, – parasite, – Mycetes – Environment. and for each of these catagories the therapy and prophylaxis must be mentioned. Virus (Slides). No therapy, no prophylaxis as active immunisation, only prophylaxis plan possible (Table 6, 7). Bacteria (Slides). The Italian Ministerial Ordonance of 4 August 1969 regulates the use of chemiotherapic and antibiotic drugs to be mixed with feed, against bacterial disease. (Tab. 8 and 9). Today vaccines are employed for prophylaxis (Tab. 10, 11, 12, 13, 14 and 15). Parasites (Slides). The prophylaxis and therapy employed for parasitic disease are based on the appropriate use of disinfectants, as seen in table 10. Mycetes These can be generally considered as negative elements of secondary irruption in a pathological process. Disinfectants shall be employed as therapy. Environment (Slides). It is always responsable for the evolution of pathological events, as already seen, but sometimes it is the sole cause of disease, as in the case of the well known "gas bubble disease" (acute and chronic) and in "gill disease". The only prophylaxis and therapy possible is to eliminate the cause which has defined the pathological phenomenon. TABLE 1 TROUT PRODUCTION IN WESTERN EUROPE (Metric tons) COUNTRY 1978 1979 1980 1981 1982 1983 DENMARK 17 500 17 950 17 500 21 700 20 600 21 000 ITALY 17 840 19 900 19 900 20 635 20 000 19 000 WEST GERMANY 7 500 8 000 8 800 12 000 13 000 13 000 AUSTRIA 1 300 1 400 1 480 2 400 2 300 2 300 U.K 2 770 3 070 4 200 5 200 5 500 5 500 IRELAND (freshwater) 220 400 490 610 600 IRELAND (Seawater) 170 340 600 BELGIUM 300 300 300 300 400 400 FRANCE 18 000 18 000 19 000 24 000 25 000 26 000 NORWAY 2 200 2 690 3 275 4 485 4 500 4 000 FINLAND 3 300 3 600 4 000 5 400 5 700 6 000 SPAIN (Estimates) 7 500 9 000 10 300 11 000 11 000 12 000 Total 78 210 80 130 89 155 107 780 108 850 110 400 Table 2 Interaction of factors necessary at the development of the disease in aquaculture

Table 3

Sometimes an aetiological agent is conditioned by another (erytrodermatytes of the carp condition by the virus) Table 4 Chemical Biological 1, Water chemistry 1. Population Density 2. Pollution 2. Other fish, lateral swimming space requirement 3. Diet Composition 3. Micro-organisms (pathogenic & non pathogenic) 4. Nitrogenous and other metabolic wastes 4. Macro-organisms (ecto- & endo-parasites)

Physical Procedural 1. Temperature 1, Handling 2. Light 2. Hauling 3. Sounds 3. Stocking 4. Dissolved gas 4. Feeding methods a) manual b) automated 5. Disease treatments

Biological, chemical, physical and procedural environmental factors which can adversely impact fish in intensive culture and cause stress. Table 5 A

SAMPLES 82 OUTBREAKS SAMPLES 83 OUTBREAKS Number of Samples: 6 978 Number of Samples: 7 465 Viral disease Viral disease IPN 168 IPN 78 VHS 129 VHS 74 Bacterial Disease Bacterial Disease Aeromonas sp 12 Aeromonas sp 24 Red mouth Red mouth 23 Foruncolosis 10 Foruncolosis 3 Myxobacteriosis 8 Myxobacteriosis 18 Vibriosis 58 Vibriosis 34 Mycotics Disease Mycotics Disease Saprolegnosis 23 Saprolegnosis 13 Parasitic Disease Parasitic Disease Acantocefalosis 9 Acantocefalosis 4 Chilodoniasis 4 Chilodoniasis 1 Costiasis 8 Costiasis 12 Dattilogirosis-girodattilosis 93 Dattilogirosis-girodattilosis 69 Hexamitiasis 11 Hexamitiasis 8 Ichtiophtiriasis 60 Ichtiophtiriasis 34 Lentosporiasis 26 Lentosporiasis 3 Oodiniasis 2 Oodiniasis 7 Tricodoniasis 34 Tricodoniasis 22 Diplostomum voluvens 1 Diplostum voluvens - PKD 38 PKD 28 Table 5 B Samples 1984 Outbreaks Total samples examinated N. 8836 Viral Disease IPN 56 UHS 83 Negative tests 79 Bacterial disease Aeromonas sp. 16 Alcaligenes sp. 4 Red mouth 76 Foruncolosis 10 Myxobacteriosis 31 Vibriosis 16 Negative tests 150 Mycotics disease Branchiomicosis 1 Saprolegnosis 29 Parasitic disease Acantocefalosis 6 Apiosomiasis 2 Argulosis 1 Chilodoniasis 2 Costiasis 1 Dattilogirosis - girodattilosis 112 Hexamitiasis 10 Lentosposporiasis 7 Nematodi 9 Oodiniasis 6 Tricodiniasis 16 Ictioftiriasis 59 Environment-poisoning 26 Liver degeneration 31 Branchial disease 334 Bubble gas disease 46 PKD 26 Traumas 2 Water analysis 325 Various 329 Table 6 VIRAL HAEMORRHAGIC SEPTICEMIA COST Trout production 1983: 18 000 ton VHS mortality 30 % of the total Trout sail price: £ 2 700 kg Total amount product: £ 48 600 000 000 VHS cost in Italian lire each year L 15 000 000 000 VHS killes fries and fingerlings which cost more than market size trout.

Table 7 Trout F. Trout F In Healted Tr. Far. Re WF Tr. 2e Time VHS Free T.F Year Prov. TN. Plane Far. Healted. 1978-79 67 16 8 1 1 8 1980 69 50 8 1 0 15 1981 70 50 7 2 1 21 1982 70 50 4 4 3 24 1983 70 49 0 3 6 27 Total 70 49 27 11 11 27

Official prophylaxis plane against viral haemorrhagic septicemia (VHS) in Province of TRENTO. Table 8 Tab. 2. Test of sensitivness to antibiotics Chloramphenicol +++ Tetracycline +++ Furanose ++ Sulphamides - Neomycin - Table 9 Therapy and Disinfection table DISEASE DISINFECTANT QUANTITY DURATION Viral haemorrhagic sept. Caustic lime (External plants) 20 - 30 q/ha one month Viral haemorrhagic sept. Formalin (Internal plants) 2 % Viral haemorrhagic sept. Iodofori (1,7 % of activity) 30 - 50 p.p.m 5 minutes Bacterial Diseases (1) Tetracycline 75 mg/kg/p.v * 6-7 days - Interruption 30 days Bacterial Diseases Chlortetra 75 mg/kg/p.v 6-7 days - Interruption 30 days Bacterial Diseases Ossitetra 75 mg/kg/p.v 6-7 days - Interruption 30 days Bacterial Diseases Sulphamerazine 120 - 220 mg/kg/p.v 21 days - Interruption 60 days Bacterial Diseases Furazolidone 60 - 70 mg/kg/p.v 10 days - Interruption 30 days Parasitic Diseases Ammonia salt (10 % of activity) 20 p.p.m 20 - 30 minutes Parasitic Diseases Formalin (2) 150 - 250 p.p.m 20 minutes Parasitic Diseases Acriphlavine 5 - 10 p.p.m 60 minutes Argulosis Masoten (in the ponds) 0,2 - 0,3 p.p.m Against Algae Copper sulphate 1/10 000, 2-3 kg/ha Eggs Iodofori (1,7 % of activity) 15 % 10 seconds Eggs Green malachite 1/200 000 10 seconds, 1-2 times a week

(1) Italian Ministerial Ordinance 4 August 1969 (2) Pay attention to the water temperature; The higher is, the lower the need of the disinfectant concentration (till 10 %, 250 p.p.m, between 10 % - 15 %, 200 p.p.m, over 15 %, 150 p.p.m). Table 10 VACCINE ADMINISTRATION METHODS

Table 11 FACTORS WHICH INFLUENCE THE IMMUNITARY LEVEL IN VACCINATED FISH 1 - Administration method of the vaccine 2 - Cell number in vaccinal solution 3 - Water temperature 4 - Size of the subjects to vaccinate

Table 12 VIBRIO VACCINE EXPERIMENT Table 1 (1° Experiment without hyper-osmotic solution) Type of Vaccine Total fry Total dead fry % mortality Formalin 96 8 8,33 Heat 93 17 18,23 Heat + C.V. 86 20 23,26 Commercial 96 28 29,25 Control 87 27 31,03

Table 2 (2° Experiment with hyper-osmotic solution) Type of vaccine Total fry Total dead fry % mortality Formalin 78 3 3,84 Heat 78 6 7,69 Heat + C.V 73 5 6,85 Commercial 81 6 7,40 Control 88 20 22,72

Table 3 (Experiment in field with inactived vaccine with formalin without pre-vaccinal hyper-osmotic bath Fry Dead Fry % mortality Vaccinated 252 989 5 500 2,17 Control 319 480 97 360 30,40 Table 13 MONOSPECIFIC OR BIVALENT VACCINE EFFICACY IN TROUT FARM AFFECTED BY VIBRIOSIS AND REDMOUTH Group Spontaneous mortality % 1. Vibriosis vaccine 12,3 2. Redmouth vaccine 31.9 3. Bivalent vaccine 23,2 4. Group of Control 38,1

Table 14 VIBRIO VACCINE EXPERIMENT ON SEA BASS Tank 1 Tank 2 Tank 3 Medium Vaccinated animals 16 % 12 % 18 % 15,4 % Tank 4 Tank 5 Tank 6 Medium Not vaccinated animals 32 % 32 % 28 % 30,7 %

Table 15 % Pathological events in farmed sea-bass (Dicentrarchus labrax) - Vibrio 60 % - I.P.N. 20 % - Tricodine 10 % - Girodattili 5 %

Vaccination trials of Sea-bass fry Infection test with Vibrio anguillarum (30' in 10 1 of water with 28 x 107 cell/ml) MORTALITY Days Vaccinated Not vaccinated 1 0 0 2 0 4 3 0 0 4 0 7 5 2 1 6 1 2 7 1 2 8 0 0 9 1 0 10 3 0 11 0 0 12 0 0 13 0 0 14 0 0 Total 8 14