Fish Disease Diagnosis and Control
Mediterranean Marine Aquaculture: an Overview
Lecture notes by Dr. Panos Varvarigos presented at the
The combined roles of the:
1. Aquaculture Systems (pivotal role)
Author: Dr. Panos Varvarigos
Freelance Veterinarian – Fish Pathologist, Athens, Greece.
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I. About the presentation
1.1. Structures, implements and materials: ● Cage type, size and layout ● Netting, net washers, antifoulants (mesh size; fouling; effluents) ● Night lights (necessary for sea bass) ● Equipment / measuring devices (tested, calibrated).
1.2. Environmental sensitivities: ● Sea depth, currents, nature of bottom ● Seasonal fluctuations of water parameters ● Inflow of contaminants from agricultural land ● Natural biota (plankton, parasites, wild fish shoals) ● Presence of predators (seals, dolphins, sea-birds, etc.).
1.4. Health and production management: ● Fry transportation and offloading ● Initial stocking of fry per cage ● Stocking density by fish size and net space ● Feed quality, storage and feeding pattern ● Vaccination and grading ● Disinfection ● Man-made diseases (overfeeding, trauma).
1.5. Data collection: ● Fish history of stocking, feeding, growth performance ● Fish movements (traceability) ● History of handling (grading, net change, movements, vaccination) ● Accidents at handling (net folding, suffocation, trauma) ● Adverse weather (storms, strong winds, lightning) ● Water parameters (temperature, dissolved oxygen, plankton blooms) ● Recent pathologies and treatments ● Epizootiological archives.
1.6. On site inspection and testing: ● Shoaling behaviour, reactions to stimuli and feeding ● Water conditions (temperature, dissolved oxygen, clarity, plankton) ● Presence of dead and moribund fish ● External lesions (ulcers, exophthalmus, reddening, fin erosion) ● Recent history (onset, empirical treatments, handling, bad weather) ● Past epizootiological archives (related to season and species) ● On site necropsy (skin, fins, gills, internal organs) ● Rapid diagnostic testing (rapid "elisa" kits) ● Sampling for laboratory examination (whole fish, organs).
2.1. Design, structures and equipment: ● Water source (pump-ashore, borehole, recirculation) ● Site characteristics (climate, landscape) ● Water treatment (filtration, degassing, UV-ozone, biofiltration) ● Ergonomy (interdepartmental flow of people and inputs) ● Easy maintenance and disinfection (modular disassembling).
2.2. Environmental sensitivities: ● Water quality and bacterial load (naturally filtered) ● Water dissolved gas pressure (gas bubble, nephrocalcinosis) ● Temperature control (ambient, water) ● Light intensity and photoperiod (controlled indoors) ● Salinity control (availability of fresh water) ● Disinfection measures (foot baths, implements, tanks, pipes).
2.3. Health / production management
2.3.1. Larvae juveniles and fry: ● Age and stocking number (per tank or raceway) ● Control of algae and live prey production ● Feeding pattern of live and inert feeds; quality ● Observing fish behaviour and feeding adequacy ● Water clarity (surface skimmers, siphoning) ● Bacterial load monitor in water, algae, live prey cultures ● Fish sampling and microscopic watch of development ● Vaccination and grading (size variation, anatomic disorders) ● Establishment of emergency procedures.
2.3.2. Brood-fish and fertilized ova: ● Age, number and sex ratio per unit (tank, basin) ● Feed quality and feeding pattern ● Behaviour ● Vaccination ● Antiparasitic treatments ● Spawning management (photoperiod, hormonal induction) ● Fertilised ova collection system ● Egg inspection, disinfection, incubation, hatching rate ● Quarantine and conditioning of incoming brood-fish.
2.4. Epizootiology: ● Vertical transmission of pathogens ● Pumped in pathogens ● Inadequate hygiene ● Temperature controlled diseases (bacterial and viral) ● Salinity dependent pathogens (usually parasites) ● Stamping out, temporary shut off.
2.5. Data collection: ● Egg hatching rate and larvae survival ● Water parameters (verification of regulated conditions) ● Handling and medical treatments (transfers, grading, vaccination) ● Fish feeding and growth rate ● Daily records of phyto- and zoo- plankton production ● Brood-stock health, treatments, fecundity, egg quality ● Fish movements (traceability) ● Routine disinfection checks.
2.6. On site clinical inspection, microscopy, testing: ● Presence of dead and/or moribund fish ● External lesions and symptoms (ulcers, shining heads) ● Dispersion in the water and swimming behaviour ● Response to feeding and to external stimuli ● Evaluation of feeding status; necropsy; microscopy (e.g. calculi) ● Rapid diagnostic tests; microbiology ● Water and live prey bacterial and parasitic loads ● Vitality and enrichment status of live prey.
I. About the presentation
Fish health management (disease prevention, monitoring, control and treatment) comprises a vital economic element in any aquaculture enterprise
This course aims to provide "front line" information of recent "aqua-health" research, but also knowledge not to be found in textbooks, which is based on the day-to-day practical experience of field working professionals.
The scope of this presentation is to convey up-to-date practical advice of what to do and what to avoid in order to optimise fish health and wellbeing under the structural and environmental constraints, which prevail in any farming system and location.
This presentation covers the aspects/topics with a bearing on fish health, namely, the type of farming system, environment, epizootiology, actions by management, etc. As the opening presentation to the course it introduces the participants in the multidisciplinary subject of marine fish health under commercial intensive conditions.
The main fish species grown in the eastern Mediterranean are sea bass (Dicentrarchus labrax, family Serranidae) and sea bream (Sparus auratus, family Sparidae). Each accounts for about half of total output. [photoarchive]
Intensive production started in Greece about 20 years ago utilising simple wooden structures. Given sufficient market demand for these fish and funding from the E.U., large hatcheries to provide the necessary fry were built within a few years and the cage farm technology intensified. Nowadays several fish feed mills produce the pelleted and extruded diets required, while state institutes are equipped to research fish husbandry, nutrition and pathology. Greek production alone accounts now for close to 100,000 metric tonnes of output, whereas hatcheries provide about 300 million fry to on-growers.
In two decades a small business has been converted to a large industry with all the production economics, marketing and financial parameters of big business attached to it. Industrial scale fish growing resulted to a sharp drop in farmed fish welfare. In addition, pathogens were given a huge chance to flourish and cause damage. Often the initial design of fish farms did not account for expansion beyond a certain level. Production expansion became market driven and at times ignored environmental constraints and the biological limitations of the fish themselves. The result was disease, frequently in the form of catastrophic epizootics.
Soon it was recognised that maintaining the good health of the growing stocks was a key issue for business success. Research advanced and experience was gained as regards the relations among the environment, the pathogens and fish biology and how these could be monitored and regulated by proper husbandry and novel technology in order to alleviate stress and reduce the detrimental effects of the various pathogens. Disease prevention rather than cure has become a must for the industry.
Despite the benefits of a good, timely diagnosis and effective treatment, disease prophylaxis forms the cornerstone of modern fish pathology as served by us veterinarians.
III. New entrants to intensive culture present with new challenges
Market demand for as well as the increase in sea bass and bream output obviated the need to grow additional marine species with high market value. Most new entrants in farming belong to the family Sparidae (breams). The following species have been successfully reared in the eastern Mediterranean to-date:
● Sharp snout sea
bream (Diplodus puntazzo)
● White Bream (Diplodus sargus)
● Red porgy (Pagrus pagrus)
● Striped sea bream (Lithognathus mormyrus)
● Pandora (Pagellus erythrinus)
● Corb or Shi drum (Umbrina Cirossa)
● Meagre (Argyrosomus regius)
● Grey mullet (Mugil cephalus)
● Dover sole (Solea solea)
Hatchery techniques and on-growing know-how is being accumulated gradually and these species are now commercially available. However, their contribution to the total farmed output is still marginal for a list of reasons as follows:
· Their specific nutritional requirements remain poorly researched. Feeding these new farmed species is in the main by means of commercial bass and bream diets.
· Raising these fish on existing installations displaces bass and bream representing a considerable opportunity cost.
· Some species (e.g. pandora and white bream) proved to be slow growers (they need about 3 years to reach market size).
· Some of these species proved to be very sensitive to particular pathogens (e.g. sharp snout sea bream is devastated by the myxosporean parasite Enteromyxum leei; sole suffers greatly from the Nodavirus causing viral nervous necrosis and from Photobacterium damselae subsp. piscicida causing pasteurellosis).
· Sensitivity to a larger spectrum of pathogens is still unknown; hence as yet unknown epizootics are likely to emerge, representing a high production risk.
· Treating diseases is not always feasible, or economically acceptable, nor always environmentally compatible. For example, there are no suitable medications against endo-parasites, whereas knowledge gaps exist in the environmental compatibility of most antiparasitic bath treatments (formalin baths, anti-louse baths with chemical compounds). Medications against gill flukes could be drawn from drugs used on terrestrial farm animals to treat flatworm parasites, but research is needed to establish safety, MRL's, potency and withdrawal periods. On the other hand, the licensed antibiotics against bacteria are only a few. Vaccines against bacterial diseases are also few, whereas vaccines against viral diseases are not yet available.
Thus, management measures, such as routine disinfection, cage-net cleanliness, avoiding fish over-crowding and over-feeding, combined with deep waters with sufficient currents/water exchange, comprise the major options to prevent disease.
IV. Introduction to the presentation
In intensive marine fish culture, diseases play a vital economic role. Veterinarians working in the field as farm consultants not only need to provide a sound diagnosis and to suggest a feasible treatment, but in addition, must have the knowledge and the experience to recognise those elements that may provoke disease or may help to prevent it on a particular installation.
The farming system, which may be open (sea cages) or semi-controlled (pump-ashore), is central to any decision. On any farm, either a cage farm or a land-based farm, fish health/welfare is affected differently by several factors that may be grouped in topics as follows:
· The particular structural design.
· The natural environment.
· Epizootiology in particular regions and/or during certain seasons.
· Data collection, record availability and presentation on site.
· Clinical inspection frequency and presumptive diagnosis on site.
After the farm location has been selected and the farm is built, it is only the last three topics that are controlled by farm management and comprise what the specialist veterinarian may influence in the short term.
An overview of the effects to fish health and wellbeing of the topics above will be given here, as these relate to the different farming systems. It is not the scope of this lecture/presentation to deal in depth with any particular element within each specific topic. For example, we shall refer to fish vaccination as an important health management measure, but not deal with the fish immune response according to species, age, vaccine type and administration method, hence the selection process of the suitable vaccination scheme, which comprises a specific component of the overall health management strategy.
1. Net pens (sea cages) for on-growing
Fish on-growing in sea cages comprises the common farming system in the Eastern Mediterranean for table fish production. Fish fingerlings at around 1.5-2g are transported to the farm sites from the hatcheries/nurseries and off-loaded into the cages.
Farming fish in net pens/cages is an open system, vulnerable to seawater, sea life, weather and effluents from surrounding agricultural or industrial activities.
Farm management may do nothing to change the environment other than selecting a proper farm-site to establish the farm in the first place. Nevertheless, as explained below, there are plenty of tasks to fulfil in order to keep up with good husbandry practice and hygiene.
1.1. The basic structures of a cage farm at sea
Cages: Selection of cage type depends on the degree of exposure of the farm site to high waves. Other variables, such as the strength and direction of sea currents, the depth, the nature of the bottom are decisive for arranging the anchorage and the layout of the cages. The tendency is to move outwards from protected bays to more exposed waters where the stronger water exchange benefits fish welfare, health and hence growth performance.
Therefore, the small rectangular (5m x 5m) wooden cages as well as the larger (10m x 10m) steel cages have given way to the modern, very robust, circular plastic cages, suitable for more off-shore, exposed locations. [photoarchive]
Several combined technological advances, available at an affordable price, coincided to make the use of large reinforced plastic cages a possibility: a) hydraulic cranes, fitted on service boats or rafts, able to manoeuvre and carry large/heavy nets, b) service boats and barges used for automatic or operator controlled feeding [photoarchive] as well as for fish harvesting, c) antifouling agents delaying the development of algal and molluscal growth on the netting, mainly in the summer, d) suitably large revolving drum net washers.
Fish in large cages anchored further off-shore in deep waters enjoy a considerable health advantage. They have more space for exercise; stronger currents remove their metabolic products and flush them with fresh water rich in oxygen.
Nevertheless, a certain number of smaller, usually rectangular cages (7m x 7m), also plastic, are still maintained in order to receive the fry. These cages are useful for most handling operations necessary on the young fish, such as vaccination and grading. During the early stages of growth, often up to 50g of average weight, fish are more vulnerable to disease; hence their behaviour, feeding response and potential ill-symptoms or mortalities have to be closely watched. Only in relatively smaller cages such close monitoring and handling may be facilitated.
Nets: Netting should be knotless with the proper mesh size according to fish size and properly sewn and stretched in order to avoid folding under the force of currents and waves. Folding may trap fish which agonise to escape, may suffocate and suffer self-injury.
The large, deep nets fitted onto the circular cages should remain free from fouling for at least 12 to 18 months. This is facilitated by treating them with antifouling chemicals (dipped/impregnated and let to dry).
Eventually the nets must be changed and washed, that is, put in a revolving large steel drum washer with continuous supply of sea water. Remnants of antifouling as well as any algae or bivalve molluscs and other attached organisms are removed during the process.
Treating/sanitising and discharging the net washer effluent away from the site remains a difficult task, still unresolved on most farms.
Night lights: Strong floodlights are usually based on shore at elevated points overlooking the cages in order to beam on to the cages all night long when the weather is stormy. Lightning and thunder frightens sea bass which dart on to the nets and onto each other. Injuries around the head and severe damage of the cornea result in blind fish and a heavy death toll [photoarchive].
Equipment and measuring devices: Apart from boats, feeding machines, cranes and other heavy equipment, measuring devices of water parameters are important for maintaining records, such as of temperature and dissolved oxygen. These are important indicators for the diagnostician (see further). Such devices are delicate and in need of proper calibration, maintenance and storage.
1.2. Cage farm environment sensitivities
As an open system, any cage farm is vulnerable to the fluctuations of the environmental parameters as well as the physical characteristics of its particular location.
Sea depth, currents, nature of bottom: A suitable site is usually selected to deploy cages when it combines the following properties: 1) deep waters close to shore (next to steep slopes), 2) reasonable strength of sea currents for dispersing the organic remnants and flush the farm with fresh well oxygenated water, 3) convenient bottom for setting the cage anchorage as well as rich in demersal fauna able to naturally consume/re-cycle the organic load escaping from the growing fish (avoid muddy bottoms poor in marine plants), 4) reasonable natural protection in case of storms.
Seasonal fluctuations of water parameters: The smoother the seasonal changes of the water parameters are at any site the better for fish health. Caged fish are trapped and unable to seek suitable conditions should a sharp change of water quality occur. For example, sea bass is particularly vulnerable to vibriosis during spring and autumn when sea temperature is unstable. Sea bream as well as bass are prone to suffer pasteurellosis when water temperature rises well above 22°C. A site with waters rich in oxygen, even during the warm seasons, will show better growing and healthier fish.
Inflow of contaminants from agricultural land: When the surrounding land is suitable for intensive agricultural use, it is likely that streams flowing into the nearby area carry agricultural fertilizers and pesticides which leach from the land into the sea as well as effluents from rural factories (e.g. fruit canneries, olive presses) that may intoxicate the fish. It is possible that the development of epithelial tumors, such as papillomas, may be encouraged by the presence of toxicants [photoarchive].
Natural biota (plankton, parasites, wild fish shoals): In some areas toxic phytoplankton blooms are known to occur. Also a sudden influx of toxic jelly fish in the spring may be detrimental, especially to the relatively smaller of the fish (block gills, irritate gill and skin epithelia).
Areas naturally rich in pelagic wild sea life may have their disadvantages. Wild fish attracted around the cages, may be the vectors of ecto-parasites as well as bacterial or viral pathogens. When these pathogens are contracted to the farmed fish, the concentration of the latter in the cages acts as an amplifier for the pathogens, even when disease and mortality is not evident.
It is well-known, for example, that bogues (Boops boops) carry the isopod Ceratothoa oestroides which has become a menace for caged bass and bream. Other fish species such as the mullets, which feed on the algae and other organisms that flourish on the nets, are known to act as vectors of bacterial diseases among farms.
Presence of predators (seals, tunas, sea-birds, etc.): Large fish predators (e.g. tunas), fish eating mammals (dolphins, seals) and sea birds may not only cause direct losses and tear the nets, but also provoke great stress, loss of appetite, panic and self-injury to the growing stocks. Stress to the fish is not avoided even when double netting is used to protect against seals, or when bird nets are hung overhead. Hi-tech seal scarers emitting repelling sounds have not proved effective in most cases.
1.3. Epizootiology basics
Transmission of pathogens: Pathogens (bacteria, viruses, parasites) spread very easily in water. Some, such as parasites, develop stages which swim actively in search for a new host. Hence horizontal transmission of disease is common among the fish of the same farm but also to neighbouring farms. Infected caged fish populations act as amplifiers of disease spreading vast amounts of the pathogens. Wild fish may get infected from the caged fish as well as pass on infections to them. Wild fish shoals are important vectors of all pathogens from farm to farm in an area.
Vertical transmission of pathogens (from brood-fish to larvae and fry via the fertilised ova) is important in hatcheries and shall be referred to in that section.
Factors predisposing to infections (temperature, stress): Pathogen activity and hence the rate and intensity of infections are mostly temperature dependent, thus there is a clear seasonal pattern for particular disease incidents.
Striking examples comprise pasteurellosis, which breaks out in late spring and summer, nodavirosis, which is seen mainly towards the end of summer and early autumn, myxosporidioses, trematodiases and isopod infestations, which develop and intensify from early spring throughout the summer.
Apart form the natural conditions which may elevate the infectivity and multiplication of many pathogens, the resistance of the fish themselves is vital. Well nourished not crowded fish in seas with stable temperature and rich in dissolved oxygen are considerably less prone to suffer disease. In addition, immunisation/vaccination against specific pathogens blocks disease.
The inherent defences of the fish (anosopoetic system, humoral responses) break down under stress and hardship [photoarchive]. Environmental upsets, such as weather extremes, excessively high temperature combined with poor dissolved oxygen and especially, temperature fluctuations in excess of 1.5°C predispose to disease by compromising the fish own defenses.
Species sensitivity and age related resistance: Fish species and age are important epizootiology determinants and influence any policies as regards the prophylactic measures to be taken. Some species are resistant to diseases (albeit may become latent carriers of the pathogens), which devastate other species. Prime examples comprise bream which resists vibriosis and nodavirosis, both being deadly for sea bass.
Age/size related sensitivity to pathogens is also well documented in practice. For example, sea bream suffers pasteurellosis with grave consequences until the size of about 8g and sea bass until about 60g. Then on, for reasons not yet researched, both species, although may get infected, resist well with minimal losses.
The host specificity of the parasites is also well known. Monogenetic trematode worms show a close relation to particular fish species. Sea bream is "preferred" mainly by Microcotyle chrysophrii and Furnestinia echeneis, sea bass is found infested by Diplectanum aequans, sharp snout sea bream by Lamellodiscus spp [photoarchive].
Nevertheless, some parasitic protozoa do not seem to show host specificity [photoarchive]. Among the fish parasitic metazoa, prime example among these being the Myxosporeans, some have exploited the opportunity of farming, providing such a concentration of hosts, to shift across species, like Enteromyxum leei, which causes problems to bream, bass and sharp snout bream alike.
Others however, insist as yet in their host specificity, such as Sphaerospora dicentrarchi infesting the gut epithelium of sea bass and Polysporoplasma sparis infesting the kidney of sea bream. Nonetheless their host shifting potential is unknown [photoarchive].
1.4. Health and production management considerations
Fry transportation and offloading: Fry transportation is critical to the wellbeing of the fry since it comprises a very strenuous procedure. It involves 3 stages: 1) capture, weighing, counting and loading onto the transportation tanks at the nursery, 2) transportation, usually by road haulage, often involving several days and one or two water exchanges en-route, 3) off-loading at the cage site subsequent to water/temperature adjustment for few hours.
All handling should be delicate; constant monitoring of water quality throughout transport and gradual water exchanges are a must during transport. Crowding of approximately 2g bream or bass fry in the transportation tanks should not exceed 40kg/m3.
Despite all attention, stress and a great opportunity for bacterial pathogens to establish on gills and external epithelia are unavoidable. Handling trauma is often evident in the form of dermal lesions and fin erosion [photoarchive]. These usually heal within few days under proper care (vitamin C supplements, occasional prophylactic antibiotic treatment). Normally, the fish are expected to resume their normal behaviour and actively accept feed after 24 to 48 hours of acclimatization to their new environment.
Initial stocking of fry per cage: The initial stocking of the cage is crucial. What matters most is the absolute number of the fry stocked rather than their number and biomass per unit volume of the net, since early life in the cage is characterised by close association and fierce antagonism. The use of relatively smaller cages is helpful at this stage (e.g. 7m x 7m).
Stocking density by fish size and net space: As the fish grow, the biomass per volume ratio gains in importance and hence the fish need to be transferred into larger more spacious cages, most often subsequent to grading by size and vaccination. For healthy fish with firm flesh texture the ultimate density at harvest should not exceed 8kg/m3. (That is, in a 12m diameter circular cage with a 12m deep net there should be 27,000 fish at 400g average weight, or 20 fish/m3).
Feed quality, storage and feeding pattern: Feed manufacturers provide fish feed of adequate quality, hence diet related health problems are non-existent, other than those due to inappropriate feed storage. Fish feed should be fresh, stored in a well ventilated, dry place, away from direct sunlight, with proper control against rodents or other pests. Thus, fats will not get rancid or moulds develop. Feeding depends on season/water temperature, the adequacy of dissolved oxygen and the size of the fish. The smaller the fish the more frequent feeding they need. Thus, a common rule of thumb is 3-4 meals a day for young fry, reduced to 2 at around 10g of average weight, down to 1 meal a day beyond 50g average weight, presuming feeding to satiation (ad lib.).
Observing fish behaviour during and in-between meals is the best indicator for adjusting the feeding pattern. Avoid full reliance on computerised feeding systems. Given the water conditions, "the fish themselves tell what they need to eat".
Feed quantity (usually expressed as a percentage of biomass) relates to feed composition in terms of energy content at a given season. Three variables need to be accounted for in order to evaluate feeding performance: 1) the growth rate, 2) the feed conversion ratio and 3) the accumulation of fat in the peritoneal cavity of the fish.
Vaccination and grading: Intensive farming of a large number of fish may not succeed without active immunisation of the population against major bacterial epizootics. It is unfortunate that so far, vaccines exist for only a couple of bacterial diseases, namely vibriosis and pasteurellosis, the later with limited efficacy. Fish vaccination requires handling of the fish and hence stress and injuries occur. Oral fish vaccination has not proved consistently efficacious. Usually, vaccination is combined with grading in order to avoid duplication of effort.
Vaccination requires fish entrapment in tarpaulins, sedation and/or deep anaesthesia, immersion in a dilution of vaccine, or individual injection, grading and release. The process is prone to human error and fish hardship. The following basic precautions apply here: 1) fast the fish until no feed remains in their gut (empty gastrointestinal tract eases handling stress); 2) ensure that the fish are healthy -no latent disease; 3) never hurry when manoeuvring nets and tarpaulins or netting the fish out; 4) add anaesthetics gradually watching fish behaviour closely (some anaesthetics induce an initial short but dangerous phase of excitement prior to sedation).
Disinfection: It is common sense that all implements (nets, injection guns, other tools) as well as boats, rafts, graders, pumps, feeding systems, etc., should be maintained clean and properly disinfected. (Spilt fish feed left to decay on boats and rafts attracting insects comprises a common sight.) The same applies for land facilities, such as feed storage areas. Staff outfits/boots and personal hygiene (hand wash/antisepsis) should not be overlooked either.
Unsuitable equipment, hurried maneuvers of nets, insufficient sedation when grading or vaccinating may cause scale loss or corneal damage and apart from stress, opportunist bacteria may establish on injured epithelia [photoarchive].
Overfeeding is a common error. Misled by the fish appetite, farmers may feed excessive amounts of energy rich extruded diets. Fatty degeneration of the liver and a large amount of fat deposits in muscle and the peritoneal cavity occur as a result. Fish with compromised liver function are perfect candidates to go down with disease [photoarchive].
A less common problem is copper toxicity by the antifouling chemical compounds used to impregnate the nets. Treated nets should remain in the sea for no less than 5 days prior to receiving fish. Otherwise, copper-based compounds that leach in the water may intoxicate the fish [photoarchive].
Management should always adjust its actions in line with fish biology and environment (at least in accordance with existing knowledge). A metabolic disease of sea bream, known as "winter syndrome", is the result of the insistence in feeding fatty diets to bream when water temperature drops bellow 14°C for several days during winter. Bream is unable to metabolise fats under such low temperatures and is best left without feed during these -relatively short- periods [photoarchive].
1.5. Data collection on site
Fish history of stocking, feeding and growth performance: Like the medical records of human patients, which are of paramount importance for doctors to reach diagnosis and suggest action/treatment, historical and recent fish records are equally important for both day-to-day management decision-making as well as for a sound diagnosis by the veterinarian, should a health problem be evident.
Problems encountered at transport, initial fry stocking, the duration of the adjustment period to the new cage environment, appetite and behaviour soon after fry delivery should be noted. Likewise the growth performance of the young fish and the establishment or not of an unacceptably wide size distribution, comprise early important determinants of the overall growth potential/quality of the fry batch.
Fish movements (traceability); history of handling; accidents at handling: All handling, such as grading, net changes, vaccination, movements to other cages and mixing with fish of similar size but from batches of different origin must be logged onto diaries or computerised records. Ability to trace back fish of a particular origin (traceability) comprises a contemporary requirement for product quality assurance. It is imperative that accidents at handling with the associated consequences should be detailed in the records. This latter is occasionally difficult as staff try to disassociate from errors.
Adverse weather (storms, strong winds, lightning): Hardship caused by adverse weather conditions should be noted, since ill-symptoms are usually widespread (e.g. self injuries due to lightning or fish entrapment in a folding net pushed by strong waves) and may be confused with an epizootic. Since the fish are contained in situ, prolonged bad weather itself may decapacitate the fish defences against disease.
Water parameters (temperature, dissolved O2, plankton blooms): Records of at least water temperature and dissolved oxygen, taken twice daily, should be maintained preferably on a computer, together with the feeding and growth records, with which they associate closely. Measurements should best be taken from at least two distant points (one closest to shore, the other further off-shore and from outside and inside a cage after a meal). Other occasional incidents, such as plankton blooms (phyto-planktonic flagellates, irritating jelly fish, etc.) as well as cases of environmental contamination (oil spills, toxic run-off from agro-industries) should be kept at least on diaries.
Recent pathologies and treatments: Anything that has been diagnosed to affect fish health and all associated treatments should be kept alongside the veterinarian's written reports and prescriptions. Farmers should not rely (as they often do) on their vets' own archives for their farm.
Epizootiological archives: Not only recent disease and growth performance are essential, but also the perennial epizootiology and climatic archives. Only through these the consultant may assess the risk of disease re-occurrence, associate particular disease outbursts with specific conditions and hence, suggest prophylactic treatments, lay out the proper strategy for vaccination, arrange the optimal timing for fry deliveries and even propose the most appropriate insurance policy.
In all, the more precise and consistent the farm records are the more likely it is for the pathologist to reach a sound diagnosis, especially when he/she is called upon obscure, rapidly worsening cases when differential diagnosis is imperative and the time to perform additional laboratory tests is scarce.
Finally, if data collection is to be computerised, the following principles must apply when selecting an off-the shelf software programme, or when deciding to build bespoke software: 1) easy data input; 2) simple, comprehensible data processing and straightforward report presentation; 3) easy/cheap software upgrading, preferably modular; 4) transferability across computer operating systems; 5) software code un-locked from proprietary rights of programmers.
1.6. What does a Vet do when called on farm?
Shoaling behaviour, reactions to stimuli and feeding: Watches for some time, as he/she considers adequate, the behaviour of the fish, their reactions to external stimuli and in particular their response to feeding (care should be taken not to misinterpret a weak response in cold weather, or soon after staff have fed the fish). Attention should be given to potential cannibalistic behaviour. The overall body condition of the fish is important (existence of weak, emaciated fish).
Water conditions (temperature, dissolved oxygen, clarity, plankton): Measures, or asks to see the recent measurements of water parameters. For example, poor availability of dissolved oxygen in the summer is evident by loss of group swimming behaviour and aggregation of fish close to the net; the presence of small toxic jelly fish provokes irritation and swimming bursts.
Presence of dead and moribund fish: Checks for the presence of moribund fish dying near the surface, or of dead fish on the surface or aggregates of mortalities on the net bottom (white net bottom). The location of the dead or moribund fish is diagnostically important for some diseases. Diseases that cause swim bladder distension result in many sick fish on the surface (vibriosis, nodavirosis) [photoarchive]. Acute diseases without swim bladder distension result in sudden "white net bottoms" with few lethargic fish near the surface (pasteurellosis) [photoarchive].
External lesions (ulcers, exophthalmia, reddening, fin erosion, abdominal distension): Inspects the ill-behaving fish to see characteristic disease symptoms and lesions, such as particular swimming behavioural patterns, the existence or location of external lesions, like skin ulceration, reddening, exophthalmia, lip or fin erosion, dangling of mucous pseudo-faeces from the anus, etc.
Recent history (onset of disease, empirical treatments, handling, bad weather): Asks the farmer/fish supervisor to supply the detailed recent history as regards the fish batch in trouble, that is, any problems at delivery, establishment of normal behaviour and appetite post delivery, growth rate and day-to-day behaviour and feeding response, any recent handling (grading, net change, vaccination), onset of disease, daily mortality pattern, spread of disease to other fish populations on farm, any other observations (e.g. presence of predators) plus the environmental records. It is important to tactfully elucidate whether there has been any empirical antibiotic administration and if so, which medication, at what dose rate and for how long as well as whether there have been accidents at handling or equipment failures (e.g. flood lights failing to dim properly).
Past epizootiological archives (related to season and species): Examines the past epizootiological archives of the farm, especially the seasonal periodic occurrence of health problems and the diagnosed cause.
On site necropsy (skin, fins, gills, internal organs): Picks random moribund fish for on-site necropsy. Usually, there is no suitable room on farm to perform this task properly, so frequently, the veterinarian has to endure heat/cold, wind, insects and above all the curiosity and constant questioning of staff! In such cases, a suitable sample has to be taken and packed on ice to be carried to the laboratory for proper examination including microscopy (see below).
Basic necropsy on-site comprises external appearance and lesions (muscle and skeletal development, skin, fins, eyes, mouth and lips, bucal and branchial cavity, gills) existence of visible parasites, internal organs (liver, gall bladder, spleen, gut, swim bladder, kidney), accumulation of peritoneal fat or perhaps, ascitic fluid and/or peritonitis. A clear note is taken on all observations.
If a secluded room with a microscope is available on farm, gill tissue and scrapings from skin and fin lesions may be examined on site to judge the bacterial load as well as the existence of parasites (trematodes, crustacea, protozoa). Fresh preparations of gut contents, or homogenised gut segments as well as kidney parenchyma squashes should also be examined for parasites (myxosporea, coccidia). Giemsa staining of blood smears fixed with methanol and air dried, may also be performed and checked on the spot.
Under suitable conditions, bacteriology testing is performed by plating spleen, liver, kidney, and/or brain on non-selective solid medium (usually T.S.A.) or, according to judgement, on selective solid media (e.g. TCBS agar, McConkey agar, BHI agar). The plates are sealed and placed in an insulated box.
Rapid diagnostic testing (rapid test-kits): When particular diseases are highly suspect, performs rapid ELISA tests on target tissues of sick fish in order to verify the diagnosis on-site.
Sampling for laboratory examination (whole fish, organs): Samples moribund fish, showing ill-symptoms, to be carried to the laboratory. When fish are relatively small (up to 80g) and the symptomatology is variable, at least 20 fish are needed in order to extract safe conclusions. Whole fish are packed in plastic bags and placed in an insulated box with ice flakes (often polystyrene boxes are used).
If histology is deemed necessary, tissues or whole organs may be extracted immediately after lifting the fish out of the water and preserved/fixed in 10% buffered, neutralised formalin or 70% strong ethyl alcohol (depending on test). The w/v or v/v ratio of tissue to preservative in the jar should be 1:20.
Land based farming comprises intensive fish cultivation in semi-controlled environments. The farms are built on the coast and water is either pumped ashore from the sea or pumped from deep wells (boreholes).
Systems, which produce Mediterranean marine species for the table (mainly sea bream and sea bass), are found in northern European countries, such as the northern coast of France, in Denmark, but also in southern countries, especially Italy for bass and bream, but also Spain for flatfish. They utilise arrays of long concrete raceways with automated feeding and water aeration.
Land-based marine fish production for the table is a highly energy demanding enterprise as regards continuous water pumping and treatment (filtration, sanitation) as well as water temperature and dissolved oxygen regulation. Therefore, in the north of Europe they are located close to nuclear power stations where heated sea-water effluent is available. This effluent is used through heat exchangers to elevate the temperature of the cold sea water pumped from the sea, thus saving energy.
In the major producing countries, such as Greece and Turkey, land-based systems are relatively uneconomic for table fish production, therefore, fish on-growing has been developed in sea cages. Land installations are utilised only as hatcheries to produce fry for stocking the cages. Water is sourced mainly through boreholes and rarely by pumping it ashore from the sea. Borehole water pumped close to shore from an average depth of about 60m has the same qualities as sea water but offers great advantages for fish cultivation. It is naturally filtered passing under pressure through deep ground layers; it is close to sterile and has stable salinity and temperature, not to mention locations with natural geo-thermally heated underground water at a constant 18-24°C. A disadvantage may be the excessive content/high pressure of dissolved gases, mainly nitrogen and secondarily carbon dioxide. De-gassing is needed.
Whatever the scope of production, a land-based system benefits from a greater control over the environmental conditions it offers to the growing species. With a good water quality source to start with, technology provides the means to sanitise, re-use as well as adjust, monitor and safeguard water parameters, light and ambient temperature in order to optimise fish comfort. Obviously, investment is considerable and the need for staff with focused expertise inevitable. Human errors cost dearly.
2.1. Design, structures and equipment
Since a land-based system is also "technology-based", its structural design as well as the application of up-to-date mechanisation and computer controlled automation must be planned ahead of construction.
Water source (pump-ashore, borehole, recirculation): The "watering point" is crucial for the location of a hatchery. A good borehole source of warm, sterile sea-water is ideal. Availability of a good water source overwhelms landscaping costs or remoteness. On the other hand, a strategic location and the availability of electricity and suitable land may diverge planning in favour of pumping seawater ashore and/or partially re-circulating this water. There are large differences in technology investment for water treatment depending on the incoming water quality.
Site characteristics (climate, landscape): The climate/micro-climate and landscape impose different demands as regards the lay-out of the facility and the control of ambient conditions indoors. For example, a natural ground inclination usually exploits gravity for water flow as well as for easy flow-transport of fish juveniles through the different hatchery compartments. Strong sunshine or excessive summer heat or winter cold requires special outdoor tank covers, insulation and air-conditioning indoors.
Water treatment (filtration, degassing, UV-ozone treatment, bio-filtration): Prior to supplying the various compartments in a hatchery, pumped ashore water from the sea necessitates sedimentation in large tanks or basins, filtration, usually through sand filters first and then through disk filters, temperature adjustment through heat exchangers or coolers, sanitation (UV radiation or ozonization) and finally oxygenation by injecting liquid oxygen, deposited in large outdoor tanks. Then, if the water outflow is to be re-used to conserve energy, prior to its return in the system it has to pass through drum and/or disk filters for the removal of suspended solids and through bio-filters for the removal/consumption of any organic matter left, sterilised and oxygenated. Disposition of sediments and filtered out solids is necessary. In case of borehole water coming from deep underground layers, de-gassers are necessary to adjust the relative pressures of the dissolved gases. Obviously, the size/scale of such structures/equipment relates to the scale of production and the volume of water flow through the system. Besides, all such functions must be monitored through automated control mechanisms with alarms. Back-ups in case of mechanical failure are required.
Ergonomy (interdepartmental flow of people and inputs): Apart from the flow of water and fish, the planners should consider the "flow of people" and the "flow of inputs" within the aquaculture facility. Staff movements should be minimised and there should be no "cross-roads" of people working in different departments, nor complicated pathways of inputs from storage to supply points (e.g. inert and live feeds). All implements, medications and chemicals (nets, buckets, siphons, detergents and disinfectants) as well as staff footwear, gloves and gowns should be assigned proper storage places avoiding unnecessary mobility. Specifically assigned, detached or semi-detached departments should be used for the maintenance of machinery, piping or any other mechanical work and associated tool and spares storage.
Easy maintenance and disinfection (modular disassembling): Such a complicated and technology intensive system is in need of almost constant testing of function, maintenance and frequent repair. Any structure should be obvious, clearly labelled or colour marked, easily disassembled and preferably made of light non-corrosive materials suitable for effective disinfection (plastic, stainless steel). Departments should be able to be shut-off from others if need be (e.g. contamination hazard alarm), hence a modular design is most appropriate. A good example is the network of water pipes. The design of the piping system should account for partial shut-off and dismantling. All piping should be laid out over-head with colour marks and arrows indicating the pipes carrying heated water, cold water, fresh water, etc. and the direction of flow. Concise plans/maps of all networks (electrical, water, sewerage, etc) and engineering designs of all equipment should be available to the expert mechanics if need be.
All above have a strong bearing on fish wellbeing in enclosed systems, since the optimised environment should be maintained and any failure reinstated fast. Any upset to the system (e.g. a de-gasser or UV filter failure, a failure to monitor dissolved oxygen, lack of proper disinfection) may cause domino-like detrimental effects to the growing stocks. Unnecessary staff movement and mixing of implements among departments spreads pathogens quickly.
2.2. Environmental sensitivities
In land-based aquaculture the ability to control most environmental parameters comes at a considerable cost. Therefore, the closer the natural conditions to the required optima the smaller the investment in complex adjustment and monitoring systems and the less risk of production upsets.
Water quality and bacterial load (naturally filtered): If the water pumped is devoid from suspended matter, bacterial load or other pathogens (e.g. borehole water naturally filtered through underground geological strata) it may be used with minimal processing. Pumped-ashore sea water on the other hand, requires passage through all those treatment procedures described.
Water dissolved gas pressure (gas bubble, nephrocalcinosis): There are four principal gases dissolved in hatchery water; oxygen (O2), carbon dioxide (CO2), nitrogen (N2) and argon (Ar). Each has a different solubility in water and when air is in contact with water they dissolve until the pressure of each gas in the water equals its partial pressure in the air (saturation). Pressure (sum of atmospheric and hydrostatic pressure), temperature and salinity govern naturally the concentration of each gas in the water. Super-saturation occurs when the concentration of a gas in water exceeds what the water should hold under a given set of temperature, pressure and salinity.
De-gassing mechanisms should be installed irrespective of water source, but more so in case of pumping water from deep boreholes. Super-saturation may also occur when cold, saturated water is heated, thus reducing gas solubility, or in the water pipes if "venturi tubes" are accidentally created (e.g. at connection points) and improper sealing of the pipeline allows air to leak into the water stream. Gaseous nitrogen, which is neither consumed nor produced by metabolic processes, is the common problem in the incoming water or in such accidental situations. Gas bubble disease results when fish live in gas super-saturated conditions [photoarchive].
In aquaculture, dissolved oxygen (DO) should be maintained at close to saturated conditions despite its consumption by the fish (plus bacteria and bio-filters) and remain between a minimum of 80% and a maximum of 100% saturation. Hence, oxygen must be diffused into the water.
Carbon dioxide is very soluble in water and difficult to force out of solution (vacuum de-gassers are needed). It is produced by fish and bacteria and if not removed, or displaced by DO saturation, it may accumulate up to toxic levels.
Fish excrete 1.4g of CO2 for every 1g of O2 consumed through their gills. Elevated CO2 concentrations in water (approaching 30mg/lt.) impair effective CO2 gill excretion, hence blood acidity is increased and the ability of haemoglobin to bind with and transport O2 to the tissues is decreased. Under such conditions nephrocalcinosis develops, comprising multifocal deposition of white calcium salts in the kidney parenchyma, even in the form of renal stones up to 1mm in size and occasional granulomas in the stomach and gut walls [photoarchive].
The configuration of the hatchery's rearing space dictates the types of oxygenation and de-gassing units that are required. If a liquid oxygen transfer unit is used for oxygenation, then the de-gassing system must be included ahead of the oxygenation unit. Systems exist that serve the dual role of de-gassing and oxygenation.
Temperature control (ambient, water): Water temperature governs the rate of metabolic processes and should be maintained close to the optimal for the species and age of the fish. For example, 10 day old bream and bass larvae favour 18°C, whereas nursery stage fish enjoy 20-22°C. Temperature fluctuations should never exceed 0.5°C in 24 hours when the fish are young, hence even ambient air temperature should be stabilised within 1°C, especially at night.
Water temperature should be reduced in several occasions, such as for bass and bream brood-stock spawning induction, or when pumped in water is warmer than the optimum for the development stage of the fish. At some unfortunate occasions of temperature-related disease outbreaks, such as pasteurellosis in bream juveniles, reducing temperature below 17°C is in itself an adequate measure to arrest the epizootic.
Light intensity and photoperiod (controlled indoors): Adjustments of light intensity and photoperiod are critical for the proper development of fish larvae and differ according to species and age. For example, complete darkness is suggested during the first 5 days of sea bass larvae. Photoperiod also relates to water temperature, hence feeding intensity of larvae. Photoperiod control together with temperature adjustments are used for spawning induction of brood-fish, thus emulating seasonal patterns and "fooling" the fish to produce off-season gametes. It is very important to avoid overwhelming stress to young fish and panic among brood-stock during lights on/off operations. A dimmer switch must be in use, controlled by a timer, which provides for 10 minutes twilight effect.
Salinity control (availability of fresh water): Salinity control is a blessing when fresh water is available (rarely). Apart from its use for cleaning and disinfection as well as in the live feed production (e.g. the rotifer cultures), gradual establishment of hyposalinity for a few hours can kill off any parasitic protozoa that may have established in the tanks and adversely affect the fish (flagellates, ciliates, amoebas).
Disinfection measures (foot baths, implements, tanks, pipes): Cleaning and disinfection measures should be routinely applied. Any enclosed system is prone to pathogen establishment. Staff, visitors and inputs should be regarded as being contaminated. Continuous water flow creates suitable environment for algae, protozoa and bacteria to flourish on pipe and tank walls. Rich organic matter in the form of feed is added daily and the natural mortality of live feed (rotifers, Artemia nauplii) and fish larvae create ideal substrates for potential pathogen growth and spread [photoarchive]. Tank bottoms are daily siphoned for the removal of such organic debris, floors are cleaned and sprayed with disinfectants, tanks (fish tanks, live prey culture tanks) are meticulously washed and disinfected when emptied, temporarily unused pipes are dismantled and disinfected, nets and siphons are immersed in disinfectants, foot-baths and special clothing and footwear for staff and visitors are mandatory as well as frequent hand sanitation with 70% alcohol dilutions [photoarchive].
2.3. Health/production management
2.3.1. Larvae, juveniles and fry
Managing hatchery production is based on sound scientific evidence but also depends greatly on the experience and "feel" of expert staff. There may be manuals describing what, how and when to do things, but producing healthy marine fish fry from egg remains an "art" which adapts to the particular hatchery designs and conditions. Proof of this is the greatly variable survival rates achieved in different hatcheries, but also from egg batch to egg batch in the same hatchery.
In addition, knowledge as regards the adaptations of applied methodologies for producing species other than bass and bream is rather scarce and scattered, treated by hatcheries as confidential.
Managing production in hatcheries may not be distinguished from maintaining healthy young fish with adequate survival rates. Every single task, from incubating fertilised eggs to producing suitably enriched plentiful live prey, to dispersing the feed in a tank, to adjusting tank aeration, light and photoperiod has a direct effect on fish health.
Therefore, in order to diagnose correctly and tackle fish health problems, veterinarians need to have a good knowledge of the hatchery production processes on top of their "traditional" expertise. They must be able to distinguish a "healthy tank" from a "bad tank" by observing the larvae/juvenile behaviour and dispersion in the water. They must be in a position to realise technical errors (e.g. abnormal light, water flow, or aeration rate) and must be able to fluently discuss technical matters with hatchery staff assigned different responsibilities.
The minuscule size of the "patients" (e.g. larval fish) obviates the need to work with a stereoscope and a microscope and makes identification of lesions, dissection and isolation of organs a very strenuous exercise.
Hatchery staff has to focus on a particular work section if the tasks are to be performed with the necessary precision. Therefore, different responsibilities are assigned to experts working in different departments, such as:
•Algae production (phyto-plankton).
•Rotifer cultures and artemia nauplii hatching and enrichment (zoo-plankton/live prey).
•Egg incubation and pre-weaned larvae fed live-prey.
•Weaned larvae and juveniles fed inert feeds.
•Nursery stage fry.
Major concerns of management in order to maintain good health throughout the production chain comprise:
Age and stocking number (per tank or raceway): Stocking indoor larval or juvenile tanks or nursery raceways with the proper number of fish in order to avoid over crowding and excessive accumulation of organic matter. This is a difficult task with very young stages, requiring experience, since counting larvae is impossible.
Control of algae and live prey production: The control of continuous production of algae and zoo-plankton as well as the suitable enrichment of live prey with the necessary nutrients. Algae and rotifers are very sensitive to water quality, oxygen and temperature as well as the supply of proper nutrients in their culture tanks in order to sustain continuous multiplication of their numbers. Obviously, insufficient quantity/quality of live prey directly affects the survival of otherwise healthy fish larvae.
Feeding pattern of live and inert feeds; nutritional quality: Frequent, adequate feeding with quality live or inert feeds according to the age of fish. Dispersion of feed in the tanks, oxygenation and light are main variables to watch for when feeding very young fish. A clear idea of the fish population in a tank as well as the concentration of prey organisms left in the tank from the previous feeding determines the feed quantity supplied (number of prey organisms). Prior to supply live prey has to be filtered through fine mesh membranes and washed with sterilised water in order not only to reduce the bacterial load but also to remove any unwanted ciliates or Artemia cysts which usually exist in the culture tanks.
Observing fish behaviour and feeding adequacy: Judging the adequacy of feeding. This is a continuous process and involves observation of the fish themselves in terms of gut content and hunting behaviour as well as sampling of tank water to determine the concentration of prey organisms per ml left over from the previous feeding.
Water clarity (surface skimmers, siphoning): Cleaning tank water from floating organic matter and removing organic debris accumulating on the bottom. Accumulation of lipids which form a thin surface film and protein nutrients frothing on the surface can be detrimental. They block the exchange of gases between air and water and prevent fish larvae from coming to the surface and gulping air in order to fill and develop their swim bladders (around the 5-8th day of age). For this purpose, skimmers are installed to isolate floating matter and frequent siphoning of tank bottoms ensures the removal of sinking debris, such as dead organisms and fish excreta or uneaten inert feed.
Bacterial load monitor in water, algae, live prey cultures: Careful watch of the bacterial load in fish tank water, algae and live prey cultures. No mater the incoming water treatment and disinfection measures, bacteria flourish in the rich in nutrients warm water of plankton cultures. These are then transferred through feeding in the fish tanks. Inert feeds are also far from being sterile. Hence, routine monitoring of the bacteria developing in the water is crucial. Bacterial flora has to be quantified (cfu/ml should not exceed 104) and frequently identified in order to monitor the presence of potential pathogenic strains, mainly belonging to the genera Vibrio spp., Photobacterium spp. and Aeromonas spp.
Fish sampling and microscopic watch of development: Routine sampling and inspection of the anatomic development of the fish under magnification. Despite efforts to optimise environment, conditions in captivity and nutrition may not emulate nature accurately. Embryonic development, while eggs are being collected and left to hatch, is prone to stress from any change in water conditions plus vibrations/movement shocks. Even the slightest of error going undetected (e.g. stronger than normal water flow) may affect the anatomic development of the sensitive fish embryos and larvae.
Vaccination and grading (size variation, anatomic disorders): Grading juveniles and fry for quality as well as active immunisation. Weaned juveniles and fry at the nursery stage of development undergo regular quality checks. They are graded by size and screened for anatomic disorders (fin and opercula development, skull and spinal deformities). Floating is a process for ensuring swim bladder development. Fish are anaesthetised and left to float in hyper-saline sea water (epiplefsis). Fish that sink are rejected. Finally, vaccination by immersion or long bath should be performed at least 200 degree-days prior to releasing the fry for sale. Stress is considerable during these procedures; hence the experience of the operators is important.
Establishment of emergency procedures: Planning for emergencies. All sections/departments in a hatchery depend on one another and all demand accurate performance of delicate tasks which are in constant danger of unforeseen upsets with serious repercussions. Therefore, despite adherence to the principles already described (ergonomic design, water treatment, disinfection, etc.) emergency remedial procedures have to be in place and contingency plans ready to put to action should a department fail temporarily. Some examples among the many may be: delay in spawning and lack of enough good quality fertilised eggs for stocking, break down in algae or rotifer production chains, sudden disease and mortality of larvae and/or juveniles, abnormally higher rate of anatomic disorders.
2.3. Health management
2.3.2. Brood fish and fertilised ova
Only with a well organised brood-stock department may a hatchery secure adequate and reliable supply of good quality fertilised fish eggs.
The broodstock unit/department should ensure no shortage of eggs in order to avoid imports and hence the risk to introduce disease (see epizootiology as regards vertical transmission of pathogens). The relatively high running costs are justified through control over egg quality and disease free status as well as the potential to carry out research with species other than bass and bream and perform phenotypic genetic improvements.
Major management tasks with a direct bearing not only on the production of healthy quality fertilised ova, but also on the health status of the brood fish themselves comprise the following:
Age, number, sex ratio per unit (tank, basin): "Stock dimensioning", that is proper population age, sex ratio and number of fish in each tank or basin and the number of tanks in order to obtain safely the quantity of fertilised ova, hence the surviving larvae required. As a rule of thumb 150,000 viable larvae may be obtained per kg body weight per year from a female sea bass brood-fish and 350,000 from a female sea bream respectively. Usually, tagging is used to distinguish the parent fish and refer to their history. Tagging may be with visual tags (colour polymers injected under transparent epithelia, such as the fins) or with programmed microchips injected in the dorsal muscle next to the dorsal fin. Information is read from these tags by passing the fish close to a reading device.
Feed quality and feeding pattern: Feed and feeding is performed according to the ovulation/spermiation stage of the fish and is adjusted in line with the induction of spawning. A "maintenance diet" is provided to fish breeders until the onset of gametogenesis. Then the "boosted diet" is supplied to spawning fish. Feeding is performed by hand once daily in order to observe behaviour. Special dry feeds are available, which amply provide the necessary energy, nutrients and micro-nutrients that are used by the fish for gamete production. Nevertheless, fresh feed, such as molluscs (squid, mussels) are indispensable for brood-fish. Their source must be disease free areas and they undergo treatment, such as deep freezing and thawing, in order to reduce contamination risks. Feeding fresh feed necessitates frequent tank cleaning.
Behaviour: Brood-fish appearance and behaviour is indicative of their health and spawning status and has to be regularly monitored by experienced staff, responsible for the brood-stock department.
Vaccination: Annual vaccination of brood-fish secures their own health but it may pass on some degree of resistance to the offspring. Passage of antibodies to the gametes is open for research. Vaccination by intraperitoneal injection must precede the spawning period and requires sedation of the fish in their tanks and careful handling. Ambient light must be dimmed and the fish head and eyes covered with a wet soft cloth in order to reduce stress and avoid violent reactions.
Antiparasitic treatments: Like vaccination, antiparasitic baths comprise part of the annual prophylactic treatments for conditioning brood-fish in advance of spawning. Formalin is often used (suggested concentration 250 ppm for one hour) under continuous aeration in order to rid gills and skin from monogenetic trematodes and protozoa, such as ciliates, some of which, like Cryptocaryon irritans, are particularly dangerous for brood-fish [photoarchive].
Spawning management (photoperiod control, hormonal induction): Spawning management aims at providing hatcheries with quality gametes at the required timing, often off natural spawning season. Sea bass and sea bream are seasonal breeders spawning in winter and early spring. Induction of spawning may be done either via manipulation of natural parameters, such as water temperature and photoperiod and/or via injection of hormones (e.g. LH-RH luteinising hormone-releasing hormone, or better its synthetic analogue LHRH-A).
Fertilised ova collection system: Ova are released and fertilised in the fish tank where they float (salinity >35ppt for egg buoyancy). They are collected via continuous surface water over flow which is gently directed into a fine mesh basket. The collection system has to ensure minimum possible disturbance to the eggs, because the first 10 hours post fertilisation, which commences in the spawning tank, are critical for embryonic development. Exposure to physical shocks, such as thermal and salinity change, direct sunlight, or vigorous water splashing in the egg collecting basket should be avoided.
Egg inspection, disinfection, incubation, hatching rate: Collected ova are sampled and microscopically inspected for quality and development. They are surface disinfected prior to being left to hatch. Iodophor dilutions are used for egg disinfection (50ppm active iodine per litre). However, surface egg disinfection is no guarantee against vertically transmissible diseases where the pathogen is located inside the egg. The eggs are subsequently placed either in 100-300 lt. incubators, or directly stocked in the larval rearing tanks where they hatch after 48 hours at 18°C water. Hatching rate is a crucial determinant of egg quality and is recorded for every egg batch. A good rate approaches 80% of eggs.
Quarantine and conditioning of incoming brood-fish: Fecundity and egg quality increase after the first spawning and remain appropriate until the age of 5 years for female fish, whereas for the males the optimal age is about 2-4 years. Therefore, continuous selection and conditioning of fish as brood-fish candidates is required in a secluded "quarantine section" of the hatchery. Farmed as well as wild fish are used as parents. Selection is phenotypic. Wild fish are introduced in order to avoid "consanguinity" and are selected according to desired shape, size and pigmentation. Farmed fish are selected in addition for their good growth rate, feed conversion and domestication. The period under observation should be sufficiently long for all prophylactic treatments to take place (repeated antiparasitic and antibacterial baths or oral medications) in order to ensure a good health status, behaviour, feed acceptance and overall proper acclimatisation of the broodstock candidates. Quarantine facilities must not come into contact with other sections of the hatchery through effluents, shared equipment or staff.
2.3. Health management
The "dry period" coincides with the end of production season for a hatchery, whereby after a series of production batches from egg to fingerling the system is left to rest. The dry period is timed to coincide with the end of summer when the last introductions of fingerlings to the cage on-growing farms occur.
Why? The scope is to deplete the micro-organisms, some potentially pathogenic, which build up in the wet environment gradually, but inevitably day after day. Only by drying the facilities it is possible to radically sanitise the system.
How? All departments are emptied and dismantled for servicing and thorough cleaning and disinfection. Besides allowing for repairs and maintenance, the dry period also allows for staff holidays, time to install new investments and for a thorough assessment of productivity [photoarchive].
Exceptions are the brood-stock unit and the algae and rotifers starter cultures, which continue to operate albeit isolated from the other departments. Brood-fish undergo all necessary conditioning and prophylactic treatments (repeated antiparasitic baths, vaccination, grading and replacement).
The modular design of a land-based aquaculture system provides a compromise, whereby some sections/modules "rest" while others continue to operate in a rotational basis.
Is it necessary? Drying out of all facilities is a critical part for system hygiene. In addition, it comprises the most radical and effective way to break the cycle of any pathogen in the system, should persistent disease occur and the treatments fail. Frequently, the cause of disease and mortality is obscure in a complex system, such as a hatchery. Then "switching-off" production and starting over provides the only safe way for a solution.
Is it worth the forgone production? A land-based system by-passing the annual dry period soon breaks down with disease. On the other hand, it is false economy to insist in keeping a system in operation under persistent health problems in the hope that they may disappear automatically. Surely enough production drops below target while operating costs as well as fixed costs accumulate.
Land-based semi-closed aquaculture systems may help prevent pathogen entry through continuous water treatment and disinfection, nevertheless they are prone to pathogen establishment should a pathogen manage to enter and be carried around the system. Hence, knowledge about disease transmission and about those conditions which may counter pathogen multiplication is important.
Vertical transmission of pathogens: Vertical transmission of pathogens, especially viral, should not be considered accidental when the gametes are produced in-house through own brood-fish. Fertilised egg disinfection in iodophor dilutions does not ensure complete sanitation therefore the good health status of the breeders may safeguard off-spring health as regards the vertically transmitted pathogens. In the Mediterranean hatcheries of sea bream and bass, the Lymphocystis Irridoviruses as well as the Viral Nervous Necrosis (VNN) or Viral Encephalopathy and Retinopathy (VER) Nodavirus are of main concern. Non-lethal periodic sampling of brood-fish with PCR techniques and rejecting carrier fish may be an option (expensive and hence rarely applied). Stringent quarantine of incoming brood-fish candidates as well as vaccination are required procedures.
Pumped in pathogens: Pumping in contaminated water by-passing treatment or passing through inadequate sanitary treatment (filtration, UV, ozonisation) may allow entry and establishment of parasitic protozoa, bacteria and viruses.
Inadequate hygiene: Slack management may also allow pathogen entry. Examples may be: visitor movements, in particular cage farm workers or staff from neighbouring fish packing plants, the introduction of improperly conditioned broodstock suffering latent disease, introduction of fertilised ova without health certification, the use of contaminated feeds, such as badly treated fresh molluscs for broodstock.
Temperature controlled diseases (mainly bacterial and viral) and salinity dependent pathogens (usually parasites): Should a pathogen be established in a closed system and disease occur, management has few options in its disposal to take advantage of the pathogen's weaknesses and attempt to block its spread. Among these, water temperature regulation/chilling and salinity control/lowering are the most important. However, the technical means and energy to regulate water temperature and the availability of fresh or hyposaline water are required.
For example, pasteurellosis, which devastates production, may be controlled by lowering water temperature to just below 17°C. The causative bacterium (Photobacterium damselae subsp. piscicida) is not infectious at low temperatures.
The build up of parasitic protozoa in the fish tanks, such as trichodinas, Ichthyobodo, or amoebas may be stopped by reducing salinity by more than 5ppt (i.e., from the usual 39-40ppt to around or below 34ppt. The parasites may not tolerate the hypo-osmotic environment and their cells burst by the incoming water and the loss of ions through their cell membranes.
Stamping out, temporary shut down: Radical measures comprise stamping out of populations that show signs of disease and thorough disinfection of the tanks, pipes and implements associated with them. Last resort of failed curative attempts is the abrupt temporary shut down of the system. Even the bio-filters have to be cleaned and re-conditioned (costly and troublesome) since pathogenic bacteria may find sanctuary in them.
2.5. Data collection
Daily records form the important background for decision making by hatchery production managers and pathologists alike.
Egg hatching rate and larval survival: The starting point comprises the fertilised egg quality expressed by their hatching rate and the survival of the larvae. These results depend on brood-fish health and conditioning as well as the environmental conditions as regulated by management.
Water parameters (verification of regulated conditions): Water quality and tank hydrology plus other physical conditions, such as light and ambient temperature are in need of regular measurements in order to verify their stability around the optimal values. Alarms and monitoring systems must be in place to ensure that no diversions have occurred.
Handling and medical treatments (transfers, grading, vaccination): Young fish are stress sensitive, hence health problems usually relate to mishandling. Records of handling and any mishaps should be available to the visiting veterinarian as well as any medical treatments (e.g. formalin baths, vaccination).
Fish feeding and growth rate: Nutrition, feed quality, supply rate, feeding behaviour and satiation of larvae are important. The hatched larva is still an "out of the egg embryo" which develops gradually into a fish. It is in need of high nutritional value feed, initially in the form of live prey. It is a "feeding machine" and any scarcity of food or improper conditions obstructing hunting (e.g. poor light, increased water flow) are detrimental. Proper/expected anatomic development and growth is proof that the young fish obtain the required nutrition. Records of age-related feed supply and growth show the wellbeing of the populations. For example, if live prey remains largely uneaten in a larval tank, it shows either an under-populated tank or sudden anorexia and disease.
Daily records of phyto- and zoo- plankton production: Live prey comprises the natural food source for hatchlings. Under artificial rearing conditions, the natural diversity of prey is unavailable. Specific zoo-plankton organisms (rotifers, Artemia) are cultivated for feeding to the larvae. However, it is through the diversity of planktonic organisms that the larvae secure in nature the adequate nutrients for their survival and growth. This lack of natural food variety is balanced by enriching/feeding the rotifers and Artemia nauplii themselves with nutrients just prior to offering these as prey. Thus, they become carriers of the essential nutrients, like "feed capsules", to their hunters.
Recording the variables affecting the production chain of plankton (population density, multiplication rate, vitality) and their enrichment process is necessary to judge the quality of live feed.
Brood-stock health, treatments, fecundity and egg quality: Brood-stock records as well as records of the incoming candidate breeders under quarantine show the status of the breeding department which provides the "raw materials" for production. Genetic selection and tagging, nutrition, conditioning, treatments, fecundity, egg batch quality, hatching rate and survival as well as the quality parameters of the finished fry batches (growth rate, anatomic deformities etc) should relate to the parental populations -broodfish tanks- and signify any necessary genetic improvements or need for replacements.
Fish movements (traceability): Any recording system should refer to particular stock batches starting from egg batches put to hatch and following the fish development up to finished fry ready for sale. At best, batches should not be mixed with grading in order to facilitate traceability back to broodfish tanks. Thus, genetically related disorders (undesired phenotype) may be uprooted.
If on-growers maintain also proper records, then the combination of both recording systems should allow traceability of table fish down to their parental stock in the hatchery of origin.
Routine disinfection checks: The importance of routine disinfection should be reflected in the recording system. Daily records should me maintained in order to prove that the daily schedule has been followed as planned (jobs check-marked) and that disinfectant baths/dilutions are fresh and potent.
In their crude form, data boards should be hung on each tank. They should show stocking day, fish age from hatch (the hatching day is considered as day 0), feeding rate and type, population estimate, mortality, water parameter measurements. More detailed data, kept on computer and processed in report form should depict the evolution of each fish batch. These records may be distinguished according to age and development phase of the populations, such as records for larvae, for weaned juveniles, for nursery fry, etc. allowing performance comparisons and hence evaluation of parental stocks.
2.6. On site clinical inspection, microscopy, testing
As mentioned already, fish pathologists should combine technical experience in hatchery processes, larval evolution, behaviour and nutrition with their "traditional" veterinary training. During either a routine or an emergency visit to a marine fish hatchery, the veterinary consultant performs wide ranging tasks in addition to those that focus on any particular problem. Microscopic examinations are required, but in the hatcheries laboratory facilities exist, because they are necessary for the daily checks on algae and live prey by staff. Sampling of fresh not fixed specimen of very young fish for transportation to outside laboratories is not a common option. Post mortem tissue degeneration processes are rapid despite chilling.
In the hatchery facility the veterinary consultant evaluates the following main aspects:
Presence of dead and/or moribund fish: Inspects the fish tanks for the presence of floating moribund or dead fish or for the accumulation of dead fish on the tank bottom. Verifies that the surface skimmers are conditioned and functional. Tank bottoms are also observed for proper cleaning (siphoning).
Fish behaviour and ill-symptoms; water parameters: Checks for the presence of visible external lesions and symptoms (ulcers, shining heads, exophthalmia) or common signs of disease (e.g. darkened skin, lethargy, fin erosion). This task may be relatively straightforward for brood-fish and for juveniles beyond 1g of body weight, but requires special techniques and microscopy for the larvae.
Assesses the fish dispersion in the water, the swimming and feeding behaviour, the response to external stimuli as well as the water and ambient conditions, according to fish age and stage of development. The macroscopic observation of larvae concentrates on their swimming and hunting pattern (e.g. sick larvae with abnormal movement often present shining eyes/heads under a beam of torchlight) and the presence or not of adequate gut content or distended swim bladder (also obvious under torchlight). Fast response to feed supply and characteristic hunting behaviour is important particularly for the larvae. Population reactions to external stimuli indicate good health for juveniles and fry (e.g. tapping on tank sides or moving an arm over the surface).
Ensures that de-gassers, filters, UV radiators, bio-filters are accompanied with adequate records proving proper maintenance and function. Measures at random water temperature and dissolved oxygen (saturation level as well as concentration) in order to verify the recorded data.
Evaluation of feeding status; necropsy; microscopy: Evaluates adequacy of feeding by random measurements of live prey concentration in the tank water as well as by sampling larvae and juveniles to examine the gastric and gut content. Inspection of these transparent young fish is performed under a stereoscope or a microscope. Under the stereoscope/microscope samples of larvae of various age/stock batches are examined for an evaluation of proper anatomic development (skeleton, fins, swim bladder) and of the expected status of parenchymatic organs, such as the liver and spleen, as well as the urinary system [photoarchive]. Gills, skin, fins and muscle are examined for bacteria or parasitic protozoa on fresh squash preparations (filamentous bacteria, myxosporea, ciliates and flagellates). Obviously these tasks are much less tiresome on the bigger specimen. For example, an ichthyoscope may be utilised to observe the anatomic development of older/larger juveniles and fry [photoarchive].
Rapid diagnostic tests; microbiology: Rapid diagnostic ELISA tests may be performed when available. Fertilised eggs, whole homogenised larvae, or target organs from larger fish may be used. Care should be taken that the number of samples is of statistical significance. Bacteriology is an option when fish are large enough to aseptically isolate organs under a dissection microscope. Crude but sound bacteriology testing on larvae may be performed by plating whole squashed larvae subsequent to a short dip in 70% strong ethyl alcohol dilution and air drying.
Water and live prey bacterial loads: The bacterial load of the water, both in the pipeline and in the tanks is crucial for larval survival. (The bacterial load of tank water should not exceed 106 cfu/ml.) Excess numbers of bacteria and the presence of potentially pathogenic strains (Vibrio spp., Aeromonas spp.) often lead to disease outbreaks due to the colonisation of surface epithelia as well as of the gut mucosa with abnormal bacterial flora. The contribution of the live prey cultures in contaminating the tank water with bacteria is significant (bacterial loads of 1012 cfu/ml are frequently found in the rotifer and Artemia nauplii cultures). Hence, water samples from several sampling points as well as samples from live prey cultures should be serially diluted in sterile saline 0.9% and their bacterial load assessed by plating onto non selective solid media (TSA is a good option) -plate count method. The most dominant colonies on these plates may be visually distinguished (preferably under a stereoscope), sub-cultured and the strains identified [photoarchive].
Live prey quality (parasitic loads, vitality and enrichment status): Contamination of the rotifer cultures with foreign ciliates may present a problem for the development of the cultures themselves, but also for the larval tanks they are added to. These ciliates compete in culture with the rotifers and colonise the larval tanks. They are either larger or smaller in size from the rotifers. Hence, the veterinarian should microscopically assess rotifer samples for purity and instruct filtration for the removal of unwanted ciliates. For example, a fine mesh size of about 50μ will hold the much larger rotifers, but allow flushing away of the much smaller ciliates, such as Cyclidium spp. at around 20μ and vice versa for ciliates larger than the rotifers, such as Euplotes spp. Sampling of live prey aims also at assessing quality by means of their vitality (density in culture, lively motility and egg bearing) as well as adequacy of their enrichment. Rotifers and Artemia nauplii should present themselves "packed" with the enrichment medium used.
Author: Dr. Panos Varvarigos
Freelance Veterinarian – Fish Pathologist, Athens, Greece.
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Every effort has been made to ensure that the information is accurate until the date of last editing. It is based upon the accumulated personal experience of applied veterinary work. The author cannot take responsibility for incorrect interpretation or any resulting consequences. The contents may be used as an educational guide and are definitely not meant to become a stand-alone diagnostic tool or operations manual.
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