FINAL REPORT TRAINING COURSE ON TROPICAL FISH HEALTH |
at the
FACULTY OF FISHERIES AND MARINE
SCIENCE, UNIVERSITY PERTANIAN
MALAYSIA
(UNDER NACA/FAO PROGRAMME)
4 July 1988 to 3 March 1989
BALBIR SINGH
CENTRAL INLAND CAPTURE FISHERIES RESEARCH INSTITUTE
(Indian Council of Agricultural Research)
BARRACKPORE-743101, WEST BENGAL, INDIA
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EXECUTIVE REPORT IN RESPECT OF Dr. BALBIR SINGH, SCIENTIST (SG) ON TROPICAL FISH HEALTH PROGRAMME AT MALAYSIA FROM 4 JULY to 3 MARCH 1989
Various protozoans diseases, helminth infestations and crustacean ectoparasites were dealt within sufficient depth in order to have a broad knowledge of parasitic disease affecting cultured fishes and shellfishes. The major bacterial diseases of tropical fishes and shellfishes i.e. haemorrhagic septicaemias, vibriosis, myxobacteriosis were studied in detail. Other bacterial diseases of less importance were also briefly reviewed.
Comprehensive studies of the fungal diseases of fish and shellfish, with emphasis on integumentary and systemic mycoses, including fish mycotoxicoses were done. The toxicoses of fishes were described with an emphasis on pathology and diagnosis. The course included an extensive study of the non-infectious diseases of fishes including nutritional diseases, malformations and deformations, toxic algal blooms, genetic diseases, malformations and deformations, toxic algal blooms, genetic disease, traumatic injuries, diseases caused by physical and chemical factors in the water, neoplasias and diseases of unknown aetiology, Diagnostic techniques and pathology applicable to invertebrate diseases, pharmacology, chemotherapy, anaesthesia, and the management aspects of disease prevention were studied in detail.
A review of current fish quarantine regulations and systems throughout the world was undertaken. The “ideal” quarantine system, diagnostic tests, recommended procedures and laboratory systems needed to impliment such a system were discussed.
The knowledge gained by the training will help in greater understanding of the relationship between the environment and ulcerative disease syndrome in fish and development of much needed facilities. Training in research methodology and techniques will enable me to conduct research into new methods and techniques of rapid diagnosis and improved methods in treatment and control of fish disease. The gained knowledge will also help in developing fish quarantine regulations and systems in India.
( Balbir Singh )
1. Introduction
One of the constraints identified in the rapidly expanding aquaculture industry of the Southeast Asian region is the lack of skilled resource personnel in fish health. The implimentation and management of fish quarantine and health extention services is dependant on the availability of specialists in fish health. The programme was offered as an attempt to meet present and future requirement for specialist fish health workers in government and private sectors. Throughout the programme emphasis was laid on the aetiology, epizootiology, pathogenesis, diagnosis, treatment and prevention of health problems affecting commonly cultured tropical food fishes.
The following aspects were studied during the course
2. Aquatic Biology
Reviewed the principles and concepts in ecology, water chemistry and their applied implication in aquaculture and fish disease management. Studied the ecosystem concept with special reference to energy flow, food chain/web, water quality criteria, air-water-soil chemistry, fertilizers, and accumulated waters in modified ecosystems and physical, chemical & biological phenomena. The general taxonomy and biology of cultured fish and shellfish of the region were discussed at length.
A detailed examination of the integument, skeletal, muscular, circulatory, respiratory, heamatopoietic, excretory, digestive, reproductive and central nervous systems and organs of special sense were studied. Basic histology and histology of the different organ systems of the fish, functions and adaptation of fish particular to the aquatic environment osmoregulation, respiration, circulation, excretion and endocrinology were also studied.
3. Aquaculture Science
The role of water and soil in aquaculture and its implication were focussed on aquaculture systems and practices in the Southeast Asian region in general and Malaysia in particular. Applied aspects of fish nutrition and genetics were also studied.
4. Biology of pathogens
Studied the procaryotic cell, its morphology, growth characteristics and requirements. Bacterial taxonomy and the classification of fish bacteria, microbiological methods and their application in the laboratory to identify bacteria isolated from disease fishes were also studied.
Various protozone diseases, helminth infestations and crustacean ectoparasites were dealt with in sufficient depth in order to have a brood knowledge of parasitic diseases affecting cultured fishes and shellfishes. The major bacterial diseases of tropical fishes and shellfishes, i.e. haemorrhagic septicaemias, vibriosis, myxobacteriosis and mycobacteriosis were studied in detail. Other bacterial diseases of less importance were also briefly reviewed.
Studies were also undertaken on principles of parasitology with emphasis on fishes, structure, mode of life and classification of viruses, tissue culture techniques and laboratory methods in fish virology.
5. Fish pathology
Comprehensive studies of the fungal diseases of fish and shellfish, with emphasis on integumentary and systemic mycoses, including fish mycotoxicoses were done. Toxicology and toxiconts important to aquatic organisms were also studied in detail. The toxicoses of fishes were described with an emphasis on pathology and diagnosis. The course included on extensive study of the non-infectious diseases of fishes including nutritional diseases, malformations and deformations, toxic algal blooms, genetic diseases, traumatic injuries, diseases caused by physical and chemical factors in the water, neoplasias and diseases of unknown aetiology. Diagnostic techniques and pathology applicable to invertebrate diseases, pharmacology, chemotherapy, anaesthesia, and the management aspects of disease prevention were also given due emphasis in the course.
6. Fish diseases
The importance, characteristics, cause, transmission control and therapy of some of the important tropical diseases were studied in detail in theory as well as in practical classes.
6.1 The importance of fish diseases
The infectious diseases are most important. These are caused by the:
viruses
Of these the non-treatable forms are of greatest concern - example:
drug resistant bacteria
Economic loss due to disease
the whole future of fin mariculture is subject to control of vibriosis.
Why does disease occur
6.2. Microbial Etiology of Infectious Diseases
Koch's Postulates
What it takes for infection to occur.
Establish new infection
How to best express disease
This states
Disease or health = Virulence of pathogen × no. present
Resistance of host
7. BACTERIAL DISEASES OF FISH
Bacteria | Disease |
Vibrio anguillarum | Vibriosis |
Vibrio ordalii | Vibriosis |
Aeromonas salmonlcida | Furunculosis |
Aeromonas hydrophila | Motile Aeromonas septicemia |
Pseudomonas | Bacterial haemorrhagic or Pseudomonas septicemia |
Pasteurella | pasteurellosis |
Edwardsiella garda | Edwardsiellosis |
Yersinia ruckeri | Enteric red mouth |
Flexibacter columnaris | Columnaris |
Cytophaga psychrophila | Low temperature or coldwater disease |
Cytophagal or Flavobacterial organisms | Bacterial gill disease |
Cytophagal or other bacteria | Fin rot |
Sporocytophaga | Salt water columnaris |
Renibacterium salmoninarum | Bacterial kidney disease |
Lactobacillus | Pseudo kidney disease |
Mycobacteria | Mycobacteriosis |
Nocardia | Nocardiosis |
Stereptomyces | Streptomyosis |
Streptococcus | Streptococcal septicemia |
Eubacterium tarentellus | Eubacterial meningitis |
7.1 Vibrio anguillarum and Vibrio ordalii
Introduction
1. Vibriosis - the disease caused by V. anguillarum and V. ordalli primarily affect fish in salt water
Pen - rearing of salmon was not successful until vaccines against this disease were developed.
Because of its economic importance, V. anguillarum is probably one of the most extensively studied bacterial pathogens of fish.
2. Characteristics of V. anguillarum
Morphology | Short curved rods. Motile by single polar flagellum Endopores absent. Nonencapsulated. |
Staining | Gram negative |
Culture | Many standard media (TSA, BHI) colonies are cream-colored, circular and convex. Optimum growth 18–30°C. Generation time of 45 minutes. |
Biochemical | Facultative anaerobe Catalase positive Oxidase positive Acid, but no gas from certain sugars. |
G+C contact of DNA | 43–45% |
Serology | Six serotypes are known, 3 are major pathogens of cultured fish. |
Pathogenicity | Highly virulent, can survive outside the host. |
Disease | Usually causes an acute Gram negative septicemia, can occur subacutely, rarely a chronic disease. |
Distribution | World wide in salt and brackish water can be found in some fresh water areas. |
Species infected | Probably most species of fin fish. Some crustaceans and shellfish. |
3. Transmission
4. Control
Prevention
presently there are several commercially available bacterin preparations.
5. Therapy
Many bacterial pathogens are controlled by oral administration of antibiotics. It is possible to control vibriosis by feeding drugs.
7.2. Aeromonas salmonicida
1. Introduction
The disease caused by Aeromonas salmonicida is known as furunculosis.
This is a misnomer which has persisted even through the lessions are not true “furuncules”.
The lesions involve no outstanding leukocytic infilteration.
Aeromonas salmonicida is probably the most commonly encountered, bacterial fish pathogen in the culture of salmonid species.
In the past, nearly all hatcheries had some level of management.
More recently, this problem has decreased because of better management.
Because of its importance in freshwater culture of salmonids, Aeromonas salmonicida and its disease furunculosis have been extensively studied.
2. Chracteristics of A. salmonicida
Morphology | Short nonmotile rods Endospores absent Nonencapsulated |
Staining | Gram negative |
Culture | Many standard media (TSA, BHI), Furunculosis |
Agar used to enhance pigment production | |
Colonies are white, circular and convex Optimum growth 18–23°C. | |
Biochemical | Facultative anaerobe Oxidase positive |
Acid, but little if any gas from certain sugars. | |
G+C content of DNA | 58% |
Serology | Most strains very similar antigenically |
Pathogenicity | Highly virulent forms exist, is an obligate pathogen. |
Distribution | Worldwide in freshwater where salmonids are reared, except Tasmania and New Zealand. |
Species infected | All freshwater and marine fish are considered susceptible. |
3. Transmission
The primary mode of transmission seems to be by horizontal methods.
Can occur through contaminated water.
Contact with diseased or carrier fish.
Contaminated eggs.
Contact with contaminated equipment or clothing.
Surface of aquatic birds, not through faces because of the thermal sensitivity of A. salmonicida.
Vertical transmission may occur, but has not been shown definitively.
4. Control
1. Prevention
The most effective method of prevention is AVOIDANCE of the pathogen.
Because A. salmonicida is an obligate fish pathogen, one can avoid infection by obtaining fish eggs or live fish from sources certified free of furunculosis.
Such sources have been established in USA and Canada.
Also must have no carrier fish in water supply.
Another useful method is avoidance by disinfection of eyed eggs. This is most commonly done with iodophors.
pathogen free fish are not useful if water supply is contaminated with fish pathogens.
Vaccines have been tested for the control of A. salmonicida.
There have been efforts in selective breeding to produce furunculosis resistant strains of fish.
These have been somewhat successful in producing more resistant animals.
However, the fish lost desirable traits or became more susceptible to other pathogens (i.e. Renibacterium salmoninarum).
5. Therapy
Furunculosis was the first disease of fishes to be treated with modern drugs - sulfonimides and nitrofurans. Some of these were effective.
Oxytetracycline (terramycin, TM 50) is also effective.
It is a USDA requirement that treatment with drugs or chemicals be terminated 3 weeks prior to marketing or release of the fish.
This allows for clearance of the drug.
Clearance usually taken place very rapidly (within a few days levels are down).
Bacteria can become resistant to drugs.
There are numerous reports of drug resistant A. salmonicida.
Usually this resistance trait is carried on a plasmid of the bacterium and is called on “R factor”.
Many drugs have been used to control A. salmonicida. Only two are now FDA approved in the USA. These are terramycin and sulfamerzaine.
A potentiated sulfonamide (R05–0037) is being tested and shows great promise. The criteria for FDA approved have been met.
It is well to remember that drugs are effective only in the treatment of outbreaks.
Recurrence of furunculosis are likely as long as A. salmonicida is present and environmental conditions are suitable for its growth.
7.3 PSEUDOMONAS
Introduction
The etiology of Pseudomonas septicemia is not precisely defined. Generally caused by P. fluorescens, but other species are probably also involved.
Characteristics of Pseudomonas fluorescens
Morphology | Motile rods Endospores absent May be encapsulated, not usually |
Staining | Gram negative |
Culture | Many standard media May produce fluorescent green pigment Grows well at temperatures from 18–37°C |
Biochemical | Strict aerobe Oxidative in O/C medium Oxidase positive |
G+C content of DNA | 59–61% |
Serology | Not defined |
Pathogenicity | Opportunistic pathogen - disease is stressmediated |
Distribution | Worldwide in freshwater - have been some saltwater isolations. |
Species infected | Probably most species of freshwater fish |
Control
Because the disease is stress-mediated, the best method of control is to provide good environmental conditions.
Drug therapy is similar to that for other Gram negative fish pathogens. Streptomycin has been reported to be superior in some cases.
There has been no interest in producing bacterins against Pseudomonas for use in aquaculture.
7.4 Aeromonas hydrophila
Introduction
There are numerous biotypes and serotypes of A. hydrophila which are some times referred to as Aeromonas hydrophila complex.
This group of organisms causes disease which is called motile Aeromonas septicemia (MAS)
Aeromonas hydrophila is primarily a pathogen of warm water species of fish. In all species it is usually considered an opportunistic pathogen.
Characteristics of Aeromonas hydrophila
Morphology | Motile rods Endospores absent Nonencapsulated |
Staining | Gram negative |
Culture | Many standard media, Rimler-Shotts can be used Colonies are white circular and convex Usually is nonpigmented Grows well at 37°C |
Biochemical | Facultative anaerobe Oxidase positive Acid and gas from carbohydrates Fermentative in O/F medium |
G+C content: of DNA | 57–63% |
Serology | Many serotypes which are not well defined |
Pathogenicity | Usually considered a opportunistic pathogen |
Distribution | Widely distributed in freshwater |
Species infected | Many species of freshwater fish are susceptible. |
Transmission
The disease agent is transmitted horizontally
From intestinal tract and external lesions
Through water
External parasites (trematodes, copepods and leeches) may be vectors.
Vertical transmission has not been demonstrated.
Control
1. Prevention
Avoidance of the pathogen is always the best method of control. Avoid the transfer of fish from areas where infected animals are found.
A. hydrophila is an opportunistic pathogen and it has been stated that prevention of its disease is a matter of good house keeping.
In European carp culture, prophylactic injections of Chloromphenicol are given to fish in the spring after overwintering.
No effective vaccines are available.
Therapy
Drugs
Tarramycin is available for use in the USA.
Chloramphenicol has been used in Europe.
Furance has been used in Japan.
7.6 Bacterial gill disease
1. Introduction
Characterised by presence of large numbers of cytophagal bacteria on swollen gill lamellae.
Common disease- Causative agent unknown. Could be more than one bacterium.
Occasional disease of warmwater pondfish.
B. Etiology: Etiology not fully understood, induction of disease with bacterium isolated has never been achieved. Koch's postulates never completed.
Long filamentous, gram negative rods
Filamentous forms grow on cytophaga agar. Will also grow well in broth cultures.
Mechanisms of pathogenicity.
Is generally accepted as cytophagal disease.
Cytophagal bacteria (causing gill disease) not adequately described.
Serotypes
Bacteria isolated from salmonids with BGD
Differ serologically from those causing colummaris disease.
Those causing low temperature disease.
While a particular bacterial type may predominate in an outbreak of bacterial gill disease, serological studies have shown that different types may be involved
C. Epizootiology
Host: infects wids range of hosts - all cultured salmonids.
Common disease among these fish.
Disease occurs among warm water fish but may not be same bacterium.
Source and reservoir of infection BGD
Not established.
Likely fish with chronic infections.
Mud and silt in water.
Mode of transmission: assumed to be transmitted in water.
Incubation period: about 3 to 7 days.
Period of communicability
Unknown.
Likely as long as infected fish in water supply.
Susceptibility and resistance
Common and acute among hatchery-raised salmonids.
Yearing and older fish less susceptible then fry.
Most hatchery trout have a low antibody titar against bacteria isolated from gill disease.
Range
Apparently worldwide in salmonids.
D. Pathology- (gross external)
Fish with BGD show
Lack of appetite
Loss of orientation with current
Gill lamellae show proliferation of epithelium, which in advanced cases results in clubbing and fusing of
Lamellae
Filaments
Examination of affected lamellae shows
Long, thin bacteria
Covering gill epithelium
Some bacteria attached by just one end.
Stained smeare show filamentous, Gram-negative bacteria about 0.5 um × 5–10 um.
Differentation
Combination of large numbers of filamentous bacteria about-0.5 and gill epithelial proliferation differentiates BGD from
E. Detection and laboratory diagnosis
Large numbers of long filamentous bacteria.
Swollen gills (clubbing and fusion of gills).
Growth on cytophaga agar.
F. Method of control
Water supply free from adult fish, silt, and mud.
Crowding, resulting in accumulation of fish metabolic products.
Most important factor contributing to disease.
Avoid crowding at all costs.
If water reused should be
Filtered
Amonia oxidized by nitrifying bacteria
Keep ponds clean (remove unused food and feces).
Chemoprophylaxis with quaternary ammonium and organic mercurial disinfectants were used.
3. Treatment: Oxytetracycline.
7.7 Fin rot
1. Introduction
Fin rot occurs among population of fishes often where intensive culture is practiced.
Bacteria are found in the fin lesion only.
B. Etiological agent
Causative agent unknown but often gliding bacteria are observed.
May be cytophagal, myxobacteria or flavobacteria.
In addition, Aeromonas salmonicida, A. hydrophila and V. anguillarum have been known to cause fin rot condition.
2. Morphology
May observe gliding bacteria or more well-known fish pathogens such as A. salmonicida
3. Cultivation
Gliding bacteria grown on cytophaga medium.
4. Mechanisms of Pathogenicity
Not known but associated with ponds which have not been kept clean and in which fish are crowded.
5. Serotypes- unknown.
C. Epizootiology
Attacks a variety of hosts, particularly salmonids; geographic range worldwide.
Reservoirs of infection.
Perhaps fish
Often the organism isolated from the fin rot lesion is the predominant organism in the incoming water supply.
Not known if it is a primary or secondary involvement.
Transmission
Carrier fish
Waterborne infection
Environmental factors.
Poor sanitary conditions associated with occurrence of disease.
Also crowding and nipping.
Nutrition may play a role in predisposing fish to disease.
D. Pathology
Bacteria found in fin lesions only.
Advanced cases of fin rot of the caudal fin resemble peduncle disease.
Signs of fin rot may occur during outbreaks of other diseases- Example- furunculosis.
Fins become opaque at the margine.
Opacity usually progresses towards the base of the fin.
Fins become thickned (proliferation of epithelium).
In advanced cases- are frayed with rays protruding.
In advanced cases of the tail fin, the whole caudal fin may be lost.
This may be followed by erosion of peduncle.
E. 1. Prevention - Control
Clean ponds
Avoid crowding
Therapy
Treat with oxytetracycline added to water 10 to 30 parts per million.
8. Viral diseases of fish
Virus | Disease |
Infectious Hematopoietic | Infectious Hemetopoietic |
Necrosis virus | Necrosis (IHN) |
Viral Hemorrhagic Septicemia Virus | Viral Hoemorrhagic Septicemia (VHS) |
Rhabdovirus carpio | Spring Viremia of Carp/Swim Bladder Inflammation |
Pike Fry Rhahdovirus | Red Disease of Pike |
Infectious Pancreatic Necrosis | Infectious Pancreatic Necrosis |
Virus | (IPN) |
Eel Virus European | Virus Disease of European Eels (EVE) |
Herpesvirus salmonis | Herpesvirus Disease of Trout |
Channel Catfish Virus | Channel Catfish Virus Disease (CCV) |
Oncorhynchus Masou virus | Oncorhynchus Masou Virus Disease (OMV) |
Erythrocytic Necrosis Virus | Viral Erythrocytic Necrosis (VEN) |
Lymphocystis Virus | Lymphocystis |
Chum Salmon Virus | Focal Necrotizing Hepatitis |
8.1 Tissue culture
Introduction
In the 1050s a major advance in virology was made with the development of methods for establishing and maintaining cells in vitro. The ability to culture cells provided a system for the isolation and propagation of many viral agents. In the late 1950s these techniques found application in fish virology. To date, over 60 cell lines of fish have been reported and many more have been established. The use of these cell lines has led to the isolation of over 20 viral agents from fish. Many fish viruses are important disease agents and the use of cell cultures has allowed the completion of Rivers' postulates in establishing the viral etiology of these diseases.
Cell Culture
Growth media
Formulations of various complexity depending on cells.
Defined medium.
Buffer.
Undefined medium
Exact components not known.
Sterility is essential - antibiotics may be employed.
Establishing cell cultures
Tissue of choice.
Rapidly proliferating tissues.
Neoplastic tissue.
Enzymatic treatment.
Plating of single cells in growth medium.
Explant technique
Growth of cells from explants to form monolayers.
Mophology and cultured cells.
Epithelial.
Handling cultured cells
Culture vessels- flasks, microplates, etc.
Virus Isolation and Propagation
Viruses as intracellular parasites.
pH and temperature for replication.
Choice of material for isolation.
Sexual fluids.
Preparation of samples.
Toxicity.
Inoculation of cells.
Depend on type of virus- somewhat characteristic.
Virus Identification
Serum neutralization.
Prevent CPE.
Fluorescent antibody.
Fluorescence in UV microscope.
8.2 Rhabdovirus carpio
1. Introduction
The most serious cause of loss among cultured cyprinid fishes in Europe is attributed to a disease complex variously known as infectious dropsy, hoemorrhagic septicemia, rubells and infectious ascites. The etiology of this complex has been controversial. A rhabdovirus (R. carpio) has been isolated and shown to reproduce symptoms of the acute stage of the disease now termed spring viremia of carp. A bacterial etiology (Aeromonas sp.) is generally accepted for the chronic or subacute form of the complex with the proposed name carp erythrodermatitis.
Additionally, a contageous and lethal disease of carp termed swim bladder inflammation has been described. The pathology of this condition is quite different and the disease has an acute and a chronic form. A virus has been isolated from fish with the acute form and shown to reproduce the disease. This virus is morphologically and serologically identical to R. carpio.
2. Characteristics of Rhabdovirus carpio.
Classification | Rhabdovirus |
Size | 190 × 70 mm |
Shape/symmetry | bullet/helical |
Envelop | Yes |
Genomc | SS RNA |
Proteins | 5 |
Sentitivity to | |
Other | + |
acid | + |
heat | + |
Cell lines for isolation | BB, EPC, RTG-2, FHM |
Temperature for incubation | 20–22°C |
Cytopathic effect | cell rounding and lysis |
Serotypes | 1 |
Natural host | Carp |
Geographic range | Europe |
Disease | spring viremia of carp and swim bladder inflammation |
3. Transmission
Natural route not determined. Waterborne infection has been experimentally demonstrated and is the most likely source.
Vertical transmission in or on eggs is unlikely.
Experimentally, the virus has been transferred by injection and bath but not feeding.
4. Control
Prevention.
Avoidance by use of virus-free broodstock and water source.
Destruction of infected stocks and hatchery disinfection.
Carp surviving infection are resistant to R. carpio.
Attenuated vaccines have been developed which show promise.
IP inoculation of killed virus also stimulated immunity.
Some carp strains are more resistant to challenge and have been selected for culture.
Chloramphenicol has been reported to be bebeficial in treating SBI. This has confused the etiology and suggests secondary bacterial infection may be important.
Protozoan disease of fish
Protozoans | Disease |
Ciliophorans | |
Ichthyophthirius multifiliis | White spot disease, ichtyophthiriasis. |
Trichodina sp. | Trichodinosis |
Epistylis sp. | |
Chilodonella sp. | Chilodonellosis |
Tricophrya sp. | |
Flagellates | |
Trypenosoma sp. | Trypanosomiasis |
Cryptobia sp. | Cryptobiasis |
Hexanuta sp. | Hexamitasis |
Ichthyobodo (Costia) sp. | Costiasis |
Sarocodina | |
Amoebae (ameba) | |
Myxosporida | |
Myxosoma cerebralis | Whirling disease |
Ceratomyxa shasta | Ceratomyxosis |
Myxobolus insidiasus | Muscele myxosporidosis |
Henneguya salmonicola | Tapioca disease |
Myxoscma squamalis | Scale pocket myxosporidosis |
Myxidium minteri | |
Parvicapsula sp. | Kidney myxosporidosis |
Microsporida | |
Pleistophora salmonis | Gill microsporidosis |
Pleisophara ovariae | |
Glugea stephani | Intestinal microsporidosis |
CILIOPHORANS
9.1 Ichthyophthirius multifiliis
Introduction
This ciliate causes ichthyophthiriasis (“ich”) or white spot disease in freshwater fishes throughout the world with the possible exception of Antarctica. Can parasitize the gills, eyes, skin and fins.
Characteristics of Ichthyophthirius multifiliis, the causative agent of ichthiophthiriasis or white spot disease.
Morphology | Uniformly ciliated Temites oval 30–45 um Trophozoites oval to round 50 μm -1mm |
Culture | Can be maintained alive for a period of time but cannot complete life cycle in fish mucus or pulverized skin and gills. |
Pathogenicity | Obligate pathogen occurring in the epidermis. |
Disease | Highly temperature dependent and characterized by white spots caused by hyperplasia of cells around parasite. |
Distribution | Worldwide in freshwaters. |
Species infected | Most freshwater fishes. |
Transmission
Fish to fish. Trophozoites drop off fish, encapsulate and divide. Release tomites which penetrate and infest fish.
Control
1. Prevention
Avoidance
Disinfect equipment.
Alterations of environment
Increase water temperature if fish species allows.
Vaccines
Vaccine using Tetrahymena pyriformis cilia has given promising results. Natural immunity is short lived unless fish in constant challenge and community probably due to antibody being secreted in the fish's mucus thus stopping the infective stage before penetration.
Prophylactic agents
Chlorination-dechlorination of water supply.
Therapy
Drugs
Quinine sulfate 1 ppm daily
Chemical agents
9.2 Other ciliophorans
Ciliophorans bear cilia during some stage of their development and all possess two types of nuclei, a macronucleus and a micronucleus. Reproduction is both a sexual and sexual which consists of conjugation with micronucleus exchange. Ciliates bear cilia throughout their life while sucturians possess them only during early developmental stages. Most ciliophorans are freeliving but several symbiotic forms, including many parasites, exist. In fishes the majority of the parasitic ciliophorans are ectocommensal, but a few endocommensal species have been described.
The majority of these parasites do not feed on fish tissues but merely use the fishes' surface as enchors or feeding grounds. The primary food sources for these organisms are bacteria and other protozoans. Problems arise when large numbers attach to the respiratory or skin surfaces of fish or grase over these surfaces creating respiratory blockage and irritation.
Trichodina spp. are ectoparasite and endoparasitic in many fish species. These ciliates move about freely over the surface of the skin, gills or within the digestive, urinary, or urogenital tracts.
Epistylis spp. are bell-shaped, stalked, sessile, facultative ectoparasites of fish usually occurring in colonial arrangements. This ciliate has been implicated as a factor in red sore disease of warm water fishes.
Chilodonella (Chilodon) spp. are ectoparasitic on the skin of several fishes, including salmonids. These ciliates are ovoid and move about freely over the surface of the fish.
Tricophrya spp. are attached suctorians which possess cilia only during their swarmer stage. As the parasite matures the cilia are lost and tentacles develop which are used for obtaining food. Primarily attached to gills of fish.
Treatment for ciliophorans consists of one of the following:
Malachite green (0.1 ppm)
Formalin (167–250 ppm for one hour)
Copper sulfate (concentration variable depending on water hardness
Malachite green-formalin (0.1 ppm-25 ppm).
9.3 ADDITIONAL PARASITIC DISEASES OF FISHES
The following parasites may be very destructive to fish. Infected fish should not be transferred to fisheries facilities where such parasites do not exist. Highlights of host range, diagnosis, and other pertinent facts for these diseases are given below.
Monogenea (gill and body flukes)
Gyrodactylus
Many species, usually species-specific for fish host. Possesses two large posterior anchors, no eye spots, and an embryo with anchors and hooks in utero.
Dactylogyrus (gill flukes)
Many species, usually on cyprinids. Goldfish Dactylogyrus is probably the most important. Possesses two large posterior hooks, four anterior eye spots, and egg(s) in utero.
Cleidodiscus (gill flukes)
Many species, on many fishes. Catfish Cleidodiscus can be dangerous to fry. Possesses four large posterior anchors, four anterior eye spots, and egg(s) in utero.
Digenetic trematodes
Adults or metacercariae in fish.
Diplostomulum spathaceum (eye fluke)
Found in many fishes. The metacercaria is not encysted, possesses forebody and hindbody, oral sucker, ventral sucker, ventral holdfast organ, and two anterolateral pseudosuckers. May cause blindness.
Ornithodiplostomum ptychocheilus (brain fluke of fathead minnow)
Found in the viscera of many cyprinids, but in the cranial cavity and brain of Pimephales promelas, Notropis heterolepis, and Notropis cornutus. Can be serious in cultured fatheads. Is encysted, has forebody and hindbody, anterior oral sucker, ventral sucker, and ventral holdfast.
Clinostomum complanatum (Clinostomum marginatum, yellow grub)
Unsightly, usually yellowish grub. Body with large ventral sucker and anterior “shoulders”. Encysted; large, 1–2 mm long.
Sanguinicola sp.
Adult Worms
Sanguinicola davisi, fully developed individuals are flattened and spindleshaped flukes, having a length of 8.5 mm and a width of 0.21 mm. There is no oral sucker, acetabulum or pharynx. The mouth is subterminal and leads into a long slender esophagus that ends in an X-shaped cecum with four short, rounded lobes. The single testis is large, irregular in shaped and approximately equoatorial in position, the sperm duct leads to a crook like thick-walled cirrus sac. The bilobed ovary located at the posterior margin of the testis opens by means of a short oviduct into a gourd-shaped ootype. Genital pores open separately near the posterior end of the body with the female pore being anterior to the male pore. Yolk glands fill the entire body. Eggs are oval, nonoperculate, and measure 63 by 35 μm.
Major gill arteries, dorsal aorta, and heart chambers from at least 60 fish should be examined for blood flukes fitting the above description. Squeeze contents from blood vessels, splitting large ones if necessary.
Cestodes (tapeworms)
Adults in intestine, plerocercoids in viscera and musculature.
Proteocephalus ambloplltis (Bass tapeworm)
Adult in intestine of Micropterus spp., plerocercolds in viscera of bass and small fish. Globular scolex has four suckers and an anterior vestigial apical sucker. The eggs are dumbell-shaped.
Nematodes (roundworms)
Goezia, a spined nematode, produces modules in the intestine of Micropterus salmoides in the Southeastern United States. Eustrongylides sp. cause swollen abdomen in tropical fish and small balt fish in extreme South-western and Southeatern United States.
Acanthocephala (thorny headed worms)
Acanthocephalus jacksoni
Acanthocephalus jacksoni of trout in Northeastern United States may cause concern among fishermen finding them in lower intestine of fish.
Copepods
Lernaea elegans (L. cyprinacea, anchor parasite) No host specificity. Adult female with large Y-shaped dorsal arms.
Ergasilus spp. (hay fork claspers)
On gills of many fish. Ergasilus labracis of striped bass (Morone saxatilis) can be lethal.
Branchiura
Argulus (fish louse). Occasionally causes problems. Try to avoid Asian Argulus japonicus and European Argulus follaceus and Argulus glordanii.
10. MYCOTIC DISEASES OF FISH
Saprolegnia sp | External fungus |
Achlya sp. | External fungus |
Phoma herbarum | Internal fungus |
Branchyomyces | Gill rot |
10.1 OOMYCETE FUNGI
Saprolegina and Achlya
Introduction
Oomycetes are the most common and widely distributed fungi infecting fishes.
The ability to produce motile spores separates the Oomycetes from the other fungi.
Produce a cottony mycelium on the surface of the affected animal.
Characteristics of Oomycete fungi producing infections in fish.
Morphology | Mycelia composed of hyphae with few septae. Zoosporangia which produce biflagellated zoospores and gemmae. Antheridial cells and oogonia which produce oospores upon fertilization. |
Staining | Polyvinyl Lactophenol blue to stain hyphae and reproductive stages. |
Culture | Sabouraud dextrose agar, nutrient agar, hemp seeds. Development of white mycelial mats. |
Cell wall | Some cellulose present. |
Pathogenicity | Some species pathogenic, others saprotrophs. |
Diesease | Epidermal and dermal tissue destruction with expanding edges from site of infection. Destruction of gill epidermal layer. |
Distribution | Worldwide. |
Species Infected | Many species of fish. |
Transmission
Fish or substratum to fish via zoospores, gemmules, or oospores.
Control
1. Prevention
Avoidance
Remove dead fish and debris from ponds, remove dead eggs.
Alterations of environment
Use ground water.
Vaccines, mechanism of immunity
Little or no inflammatory response against fungi has been observed.
Prophylactic agents
Ultraviolet irradiation of water supply.
Therapy
Drugs
None
Physical, chemical and biological agents
Biological control by crustaceans.
10.2 BRANCHIOMYCES SPP.
Introduction
This fungus is the causative agent of gill rot in fish. Two species described B. sanguinis and B. demigrams.
Characteristics of Branchiomyces sp., the causative agent of branchiomycosis disease of fish
Morphology | Hyphae 8–30 μm in diameter with spores ranging from 5–17 μm in diameter. |
Staining | Polyvinyl lactophenol blue |
Culture | Sabouraud maltose agar, other mycological media. Brownish pellicle-like colonies with no aerial or submerged hyphae. |
Pathogenicity | Appears to be an obligate pathogen, can occur intravascular or on the surface of cell lamellae. |
Disease | Acute or chronic, characterized by obstruction of blood circulation and necrosis of gill tissue. |
Distribution | Eastern and Western Europe, Mediterranean countries, Japan and United state. |
Species infected | Several species of fish, mostly cyprinids. |
Transmission
Fish to fish via spores or gemmae.
Control
1. Prevention
Avoidance
Do not introduce infected fish.
Alterations of environment.
Reduce organic load in the water.
Vaccines, mechanism of immunity
Some species appear to be immune in certain situations.
Prophylactic agents.
Quicklime.
Therapy.
Drugs.
None
Physical and chemical agents.
11. VITAMIN DEFICIENCY SYNDROMES
Vitamin | Symptoms in salmon, trout, carp, catfish |
Thiamine | Poor appetite, muscle atrophy, convulsions, instability and loss of equilibrium, oedema, poor growth. |
Riboflavin | Corneal vascularization, cloudy lens, haemorrhagic eyes - photophobia, dim vision, incoordination, abnormal pigments ion of iris, striated constrictions of abdominal wall, dark colouration, poor appetite, anaemia, poor growth. |
Pyridoxine | Nervous disorders, epileptiforn fits, hyperirritability, ataxia, anaemia, loss of appetite, oedema of peritoneal cavity, colourless serous fluid, rapid postmortem rigor mortis, rapid and gasping breathing, flexing of opercles. |
Pantothenic acid | Clubbed gills, prostration, loss of appetite, necrosis and scarring, cellular atrophy, gill exudate, sluggishness, poor growth. |
Inositol | Poor growth, distended stomach, increased gastric emptying time, skin lesions. |
Biotin | Loss of appetite, lesions in colon, colouration, muscle atrophy, spastic convulsions, fragmentation of erythrocytes, skin lesions, poor growth. |
Folic acid | Poor growth, lethargy, fragility of caudal fin, dark colouration, macrocytic anaemia. |
Choline | Poor growth, poor food conversion, haemorrhagic kidney and intestine. |
Nicotinic acid | Loss of appetite, lesions in colon, jerk or difficult motion, weakness, oedema of stomach and colon, muscle spasms while resting, poor growth. |
Vitamin B12 | Poor appetite, low haemoglobin, fragmention of erythrocytes, macrocytic anaemia. |
Ascorbic acid | Scoliosis, lordosis, impaired collagen formation, altered cartilage, eye lesions, haemorrhagic skin, liver, kidney, intestine, and muscle. |
p-Aminobenzoic Acid | No abnormal indication in growth appetite, mortality. |
12. PRAWN DISEASES
1. Viral disease
Monodon Baculovirus (MBV)
Infectious Hypodermal and Haematopoietic Necrosis Virus (IHHNV)
Hepatopancreatic parvo-like Virus (HPV)
Reo-like Virus.
Monodon Baculovirus (MBV)
Found in PL5 to adult.
Bright green spherical bodies in hyperthrophied nuclei.
Necrosis and cytolysis of hepatopancreas and gut epithelium.
PL small, dark, weak, lose appetite reduced preening and mortality.
Avoidance
Proper sanitation, proper nutrition, eradication.
Treatment
No treatment but to eradicate.
Infectious Hypodermal and Haematopoietic Necrosis Virus (IHHNV)
Found in PL and juvenile-possible latent infection in larvae.
Transferred from P. stylirostis.
Cowdry type interamuscular inclusion bodies.
Necrosis of hypodermis, hemocytes, haematopoietic organs, connecting tissues.
80–90% commulative mortality.
Avoidance
Absolute quarantine and eradication.
Hepatopancreatic Parvo-like Virus (HPV)
Found in juveniles.
Necrosis/atrophy of hepatopancreas
Poor growth rate, anorexia.
Reduced preening and mortalities.
Reo-like Virus
Sign of disease in P. japonicus
Poor growth
Anorexia, reduced preening.
Gill/surface fouling.
Opacity of muscles.
Effect on host
Necrosis and atrophy of hepatopancreas.
Prevention
Avoidance by quarantine.
Destruction of contaminated stocks, depopulation and disinfection of contaminated facilities.
Treatment
Not known
Rickettsial infection
Cause - Rickettsia or Rickettsia like organism.
Stages affected - Juvenile
Sign of disease (P. merguiensis)
Heavily infected shrimp lethargic, off feed with atrophy, pale colouration of hepatopancreas.
Sign of disease (P. monodon)
Heavy infected shrimp lethargic, off feed, swim along side of pond, brown gills, deffused opacity of abdominal muscle and mushy texture of hepatopancreas.
Effect on host
Impaired digestion and absorption of nutrients.
Reduced growth rate and high mortality.
Prevention
Avoidance by quarantine.
Destruction of contaminated stocks and disinfection of contaminated facilities.
Treatment
None responded.
2. Bacterial diseases
Shell disease.
Necrosis of appendages
Necrosis/erosion of appendages
Chemoprophylaxis/Chemotheraphy
Erythromycin phosphate
Streptomycin - Bipencillin
Tetracyclin - chlorohydrate
Sulphamethazone, Furanace and chloromphenicol.
Shell disease
Cause : Chitinolytic (Chitin degrading) species of Vibrio and Aeromonas.
Stages affected - Post larvae to adult
Sign of disease - Brownish/blackish area on shell
Effect on host - Erosion of shell, underlying tissues may be affected.
Prevention
Avoid over crowding and minimise handling to prevent injuries to prawn.
Luminous bacterial infection
Cause - Vibrio harveyi and V. splendidus
Stages affected - Larvae and PL.
Sign of disease - Prawn weak & luminace in dark
Effect on host - 100% mortality.
Prevention
Disinfection of rearing water, rigid water management and sanitation.
Treatment
Furazolidine at 10 ppm for 12 hrs. for 4–5 consecutive days.
Filamentous bacteria
Cause - Leucothrix mucor and other filamentous spp.
Stages affected - PL to adult
Sign of disease - Fine filamentous growth on gills, setae, shell and appendages.
Effect on host - Hypoxia, impaired moulting and mortalities.
Treatment
- Cutrine plus at 0.1 mg cu/l for 24 h. or 0.25–0.5 mg Cu/l 4–8 h.
3. Fungal disease
Larval mycosis
Cause - Lagenidium callinectes, Lagenidium sp. Haliphthors phillipinsis and Sirolpidium sp.
Stages affected - Eggs, larvae and PL
Sign of disease - Prawn whitish and weak, mortalities.
Effect on host - Fungal mycelia replace internal tissues mortality 100% in 2 days.
Prevention
Disinfection of rearing facilities with Tide detergent at 100 ppm for 24 h.
Chemical prophylaxis of spawners with Treflin at 5 ppm for 1 h. or eggs with tide detergent at 20 ppm for 2 h.
Treatment
Treflin or trifluratin at 0.2 ppm for 24 h.
Fussarium infection (Black gill disease)
Cause - Fussarium solani
Stages affected - Adult
Sign of disease - Scanty white cottony growth on prawn, black gills, reduced feeding and preening.
Effect on host - Destruction of gill tissue and mortality.
Prevention
Disinfection by Chlorination
Treatment
Harvest prawn if size is adequate
4. Protozoan disease
Microsporidiosis.
Ciliate infestation
Cause - Zoothamnium, Epistylis, Vorticella Acineta, Ephelota.
Stages affected - PL to adult
Sign of disease - Fuzzymat on shell, dark brown gills.
Effect on host - Respiratory/locomotory difficulties, loss of appetite, mortality
Prevention
Avoid build up of organic matter, silt and sediment.
Maintain optimum dissolved oxygen.
Treatment
Chloroquin diaphosphate at 1.1 ppm for 2 days.
For Zoothamnium - 50–100 ppm. formalin for 30 min.
For Epistylis - 30 ppm formalin for juveniles only.
Gregarine disease
Cause - Gregarine.
Stages affected - Larvae and adult.
Sign of disease - Under microscope gliding through the digestive tract or attached to the intestinal wall.
Effect on host - Large number of protozoans may interfere with particle filteration through digestive tract.
Prevention
Eliminate molluscans which act as intermediate host.
Treatment
None
Microsporidiosis
Cause - Microsporidia
Stage affected - Adult
Sign of disease - Whitening of affected areas, e.g. ovaries & abdominal muscles.
Effect on host - Microsporidian spores and other stages of parasite replace affected tissue, sterility in female spawners with white ovaries.
Prevention
Disinfect with chloride or iodine containing compounds.
Dinoflagellate infestation
Cause - Peridinium sp.
Stage affected - PL
Sign of disease - Shrimp moribund and depigmented microorganism fill ventral sinus and hemolymph spaces.
Effect on host - Mortality
5. Ectozoic algal bloom
Cause - Schizothrix calcicola
Stages infected - Mysis, PL, juvenile, adult.
Sign of disease - Sluggishness, inactivity, surface fouling.
Effect on host - Locomotory and respiratory difficulties, gill disease, mortality.
6. Nematodes infestation
Cause - unidentified nematodes.
Stages affected - Mysis, PL, adult.
Sign of disease - Presence of nematodes.
Effect on host - mortalities.
Prevention
Maintain good water quality and reduce organic load.
7. Copepod infestation
Cause - Caligus epidemicus
Stages affected - Adult
Sign of disease - Presence of ectoparasite, on pleopods and tail.
Effect on host - Morphological deformatics.
WATER QUALITY DISEASES
1. Temperature
This is probably the single most important factor in both the farming situation and in fancy fish keeping. Fish are ectotherms (poikilotherms) and with few exceptions (muscles of fast swimming species) mirror the temperature of the surrounding water. Nevertheless, each species has its own preferred range. Sustained periods at the extremes of this range, or probably more important, rapid changes (even very small ones sometimes) within this range, represent stressful conditions. While metabolic rates alter with varying water temperature (roughly doubling for each 10°C) this does not happen immediately. Advantage is taken of this enhanced metabolic rate in aquaculture where waste heat is available from eg. power stations. Thus the fish will grow faster in warmer water.
In general terms, fish will tolerate a temperature drop better than a rise. Some species are more susceptible to temperature stress than others and this is thought to be due to a poorer ability to osmoregulate at the new temperature. Higher temperatures cause an increased metabolic rate and hence an increased O2 demand necessitating increased “irrigation” rates. The increased water flow past the gills causes an increased water influx and thus possible compromise of the osmoregulatory mechanisms.
Fish also appear much more susceptible to bacterial diseases in conditions of rising water temperatures. Even a small change from as low as 5°C can be sufficient to promote overt furunculosis in Atlantic salmon fry. The reason for this susceptibility is unknown in precise terms but it is thought to be the result of an enhanced rate of replication or enzymic production by the pathogen which is not matched by that of the fish's immune mechanisms.
Increasing water temperature in general reduces the survival time of fish in pollutants. With heavy metals, their toxicity is enhanced, but the fish also has an increased respiratory rate and therefore increased exposure - a synergistic effect.
2. Oxygen
The efficiency of extraction by the gills is high, and it has to be when one considers that at 15°C one litre of freshwater contains 7 cc O2 and seawater even less as compared to almost 21 volumes per cent in air. In warm or polluted waters, where O2 levels are low, the cost of extraction may be too high i.e. where the energy required for ventilation exceeds that released by the O2 so obtained. Under these circumstances, the fish enter Respiratory Distress Syndrome. Different species of course have different requirements for O2: salmonids for example have a high requirement, catfish a lower one. Some species have an accessory breathing apparatus, such as a modified swimbladder, and in some cases rely heavily upon them although they are able to survive in even totally anoxic conditions eg. snakeheads. Small fish have higher requirements than older larger ones: this translates in an aquaculture situation into placing small fish in the best quality water available.
Fish will normally demonstrate an O2 lack by gathering at inlets or by gasping at the surface and by decreased activity. Although O2 levels may be higher in surface waters, the gills collapse on contact with the air, so reducing surface area.
Increasing water temperature cause an increased demand for O2 for the increased metabolic rate so produced, but a decrease in the holding capacity of water for O2. Appetite also increases, but this extra intake of food may place a burden on the ability of the fish to extract enough O2 from the reduced levels available to metabolize the food properly. At 28°C (a high temperature, but is encountered in some trout farms in Europe for short periods of time) O2 consumption just about matches that which is obtainable and there is thus no O2 left over to properly metabolize food. In general terms therefore, with high water temperatures, a reduced food intake is advisable, and a complete cessation of feeding is indicated if furunculosis or other bacterial diseases appear. In some situations it may be necessary to match feeding frequencies with O2 levels: these are normally highest at mid-day due to photosynthesis, but surprising variations do occur - best therefore to directly measure.
Fish can acclimate to lower O2 levels to a certain extent by increased hematocrit. Diel variations, however, depress appetite and growth.
The LC50 of some toxicants is lower at high O2 levels, largely due to the reduced irrigation rate over the gills. Similarly, uptake of many toxicants correlates positively with O2 uptake: thus methyl mercury or endrin uptake increase with increased swimming speed, due to the increased oxygen uptake. The rate of uptake depends on the physiochemical properties of the toxicant: Lipid solubility, small molecular size and un-ionized molecules favour uptake.
High fecal levels or other organic detritus utilize available O2.
A minimum of 5 mg/L is considered a satisfactory level for most stages under most conditions, although higher is certainly preferable for salmonids.
3. Supersaturation
Rapid increases in water temperature or reduced pressure may lead to a situation of supersaturation. Both of these may be seen when water from a tap is used to fill a domestic aquarium. Supersaturation may also be seen in the wild associated with very rapid photosynthesis by plants and algae, or more commonly in an intensive culture operation, associated with leaky valves or pumps, in those farms where pumping is a feature (air is forced under pressure into the water supply - so called Venturi principle). Water which is supersaturated with gas, either O2 or N2, may cause the condition gas-bubble disease. This is the fish equivalent of divers “bends”. Fish normally equilibrate quickly with supersaturated water and it should therefore cause little problem. The reason for the gas in the blood coming out of solution is not therefore completely understood but is thought to be associated with the great drop in pressure experienced by the blood when crossing the gills. Whatever the reason, bubbles of gas cause emboli in the vessels of the gills pseudobranch, choroid gland and elsewhere. These are often difficult to see, and a “candling” procedure may be found advantageous in diagnosing the condition. An apparent contraindication, vigorous aeration or agitation of the incoming water, or replacement of the leaky valves, are considerations in curing the condition.
4. Suspended solids
5. Ammonia
fish excrete this via the gills, and where there is plenty of water to remove it there is no problem. Toxicity arises however, in situations of overcrowding, or where for example, chicken slurry is added to the water. Young fish are relatively quite susceptible.
6. Nitrite
7. Nitrate
8. Carbon dioxide
9. Chlorine
10. Alkalinity, hardness, salinity and pH
11. Light intensity
12. Heavy Metals
13. Pesticides and Herbicides
species of organism is also important. In general, fish are more resistant than invertebrates, although there are exceptions (the herbicide trifluralin).
Comparison of 96 hr LC50's to OP's given in ug/L | ||
Pesticide | Inverts | Fish |
Malathion | 49 | 162 |
Ethyl parathion | 24 | 1391 |
Methyl parathion | 11 | 5411 |
Diazinon | 7 | 640 |
Chlorpyrifos | 4 | 81 |
14. Algal blooms (include dinoflagellates, phytoflagellates and blue-green algae)
15. Electrocution
Occasionally encountered with badly insulated pumps, or aerators, or due to improper use of electro-shocking equipment.
Lightning may cause massive fish kills: the amount of current needed to kill fish depends on several factors, but water hardness is one of the most important. Soft waters are poorer conductors than hard, and thus the fish's body with its higher conductance will be a selective path. Thus, lethal voltages need to be higher in hard than soft waters.
Outer ponds can be affected, but indoor facilities are not immune, as lightning can enter via pipes or drains.
Fish so affected may be killed outright; but survivors may show hyperesthesia or spinal damage due to overcontraction of muscles. A common location is the fulcrum just beneath the dorsal fin in salmonids.
FISH HEATLH AND NECROPSY EXAMINATION
Examination Outline
HISTORY
SPECIMEN EXAMINATION
External Examination:
growths, deformities, abnormalities
skin - coloration, mucus production, scale examination, scraping and wet mount, vital staining, parasites
fins - color, condition, wet mount preparation
gills - color, condition, mucus production, foreign bodies or parasites, wet mount preparation
eye, opacities, exophthalmia.
Internal Examination
body cavities - color, condition, fluid accumulation (wet mount of body fluid)
visceral organs - size, shape, color, location consistency, cut surface, impression smears, wet mounts
LABORATORY EXAMINATION
Bacteria:
tissue or organ submitted
examination requested
gram stain
Blood
hematology
plasma chemistry
parasite examination
Virus
tissue or organ submitted, method of preservation (do not fix)
Parasita - tissue, organ, or preparation submitted, method of preservation gross description of organism, behavior of organism, location
Histology - tissues submitted, method of fixation, principal findings, incidental findings
DIAGNOSIS
Tentative Diagnosis
Differential Diagnosis:
principal findings
supplemental findings
proposed pathogenesis
relationship of history, clinical findings, necropsy and laboratory results
RECOMMENDATIONS
Therapy - type and amount of treatment and dosage schedule
Prevention & Control - recommendations for shipping quarantine water quality control, disinfecting husbandry, record keeping
FISH EXAMINATION - HISTORY
SPECIES: | ||
SOURCE: | ||
AGE: | SIZE: | SEX: |
Environmental Conditions:
1. | date of capture | ||||
2. | length of time in present facility: | ||||
3. | water: | Source - | |||
Flow and Temperature - | |||||
D.O. and hardness, or salinity - | |||||
NH3 | NO3 | NO2 | pH | ||
4. | Past episodes of diseases: | ||||
5. | Feed: | storage | |||
additives | |||||
type | |||||
date acquired and source | |||||
size and amount fed | |||||
feeding frequency | |||||
feeding method | |||||
feeding response | |||||
6. | Sanitation: | frequency of cleaning | |||
cleaning compounds and equipment | |||||
dead fish removal and disposal | |||||
7. | Events of 10 days preceding disease | ||||
transferring fishes | |||||
changes in environmental conditions | |||||
changes in husbandry practice | |||||
climatic conditions | |||||
8. | Clinical signs | ||||
flaring of opercula | |||||
gulping | |||||
surface breathing | |||||
scratching | |||||
darting | |||||
circling |
PREPARATION OF A SKIN SCRAPING
(Figure 1)
Using a clean scalpel, scrape the skin from head to tall to prevent dislodging many scales. In this way, mucus mixed with epidermis cells and any skin parasites is obtained. The best skin smears are obtained from the lateral sides of the body, the caudal fin and the base of the fins.
The material collected is then smeared on a microscope slide and covered with a coverslip. The smear should not be too thick or identification of parasites may be difficult
Introduce freshwater (freshwater fish) or 1 to 2% saline (salt water fish) under the coverslip with a Pasteur pipette until the area beneath the coverslip is completely saturated. A well made preparation will be free of air bubbles, and will have no excess fluid at the edges of the coverslip. Methylene blue for contrast or a vital stain may be substituted for water or saline.
Examine under a light microscope. A duplicate smear may be airdried, fixed in methyl alcohol (3–5 minutes) for preparation of a permanent mount and stained with Wrights stain or Wright's Giemsa.
Figure 1
Preparation of a skin scraping
PREPARATION OF A GILL SMEAR
Remove part of a gill arch with scissors and holding the tissue in forceps, smear the lamellae on a microscope slide. The material which adheres to the slide may contain mucus, blood cells, epithelial cells and any bacteria present on the gills.
Allow the smear to air dry and pass the slide through an open flame 2–3 times to fix the smear on the slide.
Stain using the Gram stain method and examine under a compound microscope. Gill smears should be made when ever the gills are examined.
PREPARATION OF A GILL MOUNT
(Figure 2)
Remove a gill arch from the fish using forceps and scissors. Avoid damaging the filaments to be examined as much as possible.
Using a scalpel cut the cartilage away from the filaments.
Place a section of the gill on a microscope slide and coverslip it.
Introduce freshwater (freshwater fishes) or 1 to 2% saline (salt water fishes) under the coverslip until the area beneath the coverslip is saturated. If more contrast is desired, Wrights stain or a vital stain can be used in place of the saline or freshwater.
Examine under a light microscope for parasites, swollen gill lamellae or white spots on the filaments.
Figure 2.
Preparation of a gill wet mount
PREPARATION OF A TISSUE IMPRINT AND TISSUE SMEAR
(Figure 3)
Tissue Impriat:
Cut a small block (1 cm3) of the tissue and press it to a microscope slide Leaving an imprint. Repeat 2–3 times with each piece of tissue. Allow the imprint to air dry and then fix it to the slide by passing it through a flame 2–3 times. Dip the slide into 70% ethyl alcohol for about 5 minutes and when dry, stain with Wright's or Geimsa stain.
Tissue Smear:
Scrape the tissue with a clean scalpel blade as shown in Figure 5.2. Smear the material on to a microscope slide and allow to air dry. Do not make the smear too thick or it will be difficult to see anything. When the smear is dry, pass it through a flame 2–3 times to adhere it to the slide and fix in 70% ethyl alcohol for 5 minutes. Stain with Wright's or Gram's stain when the smear is dry.
Figure 3.
Preparation of a tissue imprint (1) and a tissue smear (2)
PREPARATION OF A TISSUE SQUASH
(Figure 4)
Cut a small section of tissue and place on a microscope slide. Any cysts or abnormalities in the tissue should be examined.
Cover the tissue with a coverslip and press down gently allowing the tissue to spread out on the slide. If the tissue is hard or will not spread fairly easily, tease it apart carefully using two mounted needles.
Using a pasteur pipette, introduce saline (or fresh water) under the coverslip until the area under the coverslip is saturated. Examine under a dissecting or compound microscope. Bacterial cysts, sporozoa, fungal cysts and encapsulated larvae of various worms may be encountered.
Figure 4. Preparation of a tissue squash
Preparation of Tissue for Histopathology
Specimen Collection
Fixation is the process of killing and hardening tissue. Tissues from poikilotherms tend to decompose rapidly after death, and therefore should be placed in fixative immediately upon removal from the carcass. If fixative is not at hand then try to keep the tissue iced or refrigerated. Do not freeze, as this will destroy most of the cellular detail.
Small fish, up to 5 or 10 cm in length, may be fixed whole. Slit open the muscle along the ventral surface and remove the covering muscle on one side of the abdomen in the larger fish, to allow fixative to reach internal organs quickly. Keep fish flat - do not bend or twist while fixing.
If fish is larger than 10 cm, or in the case of large lesions, tissue sections should be cut thick enough so that the fixing fluid can penetrate within a reasonably short time. Tissues should be no more than 0.5 cm in thickness, and should be immersed in at least 20 times their volume in fixative. Smaller ratios may be required under field conditions, with some sacrifice in quality of the final product. If the fish is considered to be diseased, fix a portion of all tissues. Include some adjacent normal tissue with the lesion when fixing.
If possible photograph the condition and/or the lesion before processing the tissue.
The accompanying figure (page 61) lists the tissues which should be submitted routinely for histopathologic examination.
fin (s)
skin with underlying muscle
kidney (includes head kidney and posterior kidney)
spleen
intestine
pyloric caeca (includes pancreas)
stomach
liver
heart - (atrium and ventricle)
eye
gill
* Always submit one or two gill arches
Submit brain when feasible.
Include with the specimen your name and address, your specimen identification number and species of fish. Supply any pertinent information that is available, eg.:
location, date, and method of catch
gross description of the lesion-where found etc.
size (length/weight), sex, and age of fish
evidence of abnormal behaviour
whether wild or cultivated fish
the number of fish with similar lesions
NOTE:
Bacteriological or virological identification cannot be undertaken on formalin-fixed tissue. However, it is possible to observe cellular changes which indicate infection by microorganisms.
FORMAL IN PRESERVATION OF TISSUES FOR HISTOPATHOLOGY
13. Quarantine
Quarantine, a period of isolation of newly transplanted stocks until the possibility of the introduction any pathogens they may carry can be eliminated, is the most potent weapon at the disposal of fish health authorities, as well as of every fish producer. Fish imported from abroad or moved from one locality to another within a single country should be placed in quarantine on arrival and should remain there until all danger has passed. The quarantine period should exceed the length of the longest: latent period of the pathogens. Russian authors recommend a period of one year's quarantine for all imported fish. However, under the higher temperatures occurring under tropical conditions this period can be much shorter.
Quarantine ponds must be safely isolated and must be located downstream from all other ponds on the farm to minimise the danger of penetration of pathogens into them.
Fish markets can become centres for the disposal of pathogens. To avoid this danger, fish should be disinfected upon arrival at the market. Disinfection should aim at destruction of the most dangerous ectoparasites (e.g. Lernaea, Gyrodactylus, etc.) and, so as not to impede normal market activities, should be organized to take as little time as possible. Short duration baths should be experimentally developed and bath techniques should be taught to market personnel. Water temperature/ bath duration charts should be developed and made available to market managers.
14. Regular prophylactic survey
Periodic checks on the status of health in fish production facilities can detect dangerous conditions before they become too great to be handled without incurring serious economic losses. Ideally, their frequency will be determined by the manpower and resources available.
Utilization of knowledge in the country
The knowledge gained by the training will help in greater understanding of the relationship between the environment and ulcerative disease syndrome in fish and development of much needed facilities. Training in research methodology and techniques will enable me to conduct research into new methods and techniques of rapid diagnosis and improved methods in treatment and control of fish disease.
February 27th to March 3rd 1989 (Thailand)
27.2.1989 | - | Opening ceremoney |
- | Introduction of participants | |
- | Reception | |
- | Visit to N.I.F.I. Laboratory | |
28.2.1989 | - | Travel to Chachoengsao, Chonburi and Samutprakan District, Chachoengsao province. |
- | Visit to Chachoengsao Fisheries Station, Bangpakorg District. | |
- | Visit to Artemia Farm, Chonburi province (Overnight stay in pattaya) | |
01.3.1989 | - | Visit to Marine Aquarium, Bangsaen District, Chonburi province. |
- | Visit to Institute of Marine Science, Srinakharinwirot University at Bangsaen. | |
- | Return to Bangkok. | |
2.3.1989 | - | Visit to Catfish farm, Bangkruay District. |
- | Visit to Snake head farm, Bangkhon - tee District. | |
- | Visit to Crocodile farm, Samutprakan province. | |
- | Retun to Bangkok. | |
3.3.1989 | - | Overall discussion of the training course. |
- | Certificate distribution and closing ceremony. | |
- | Dinner party. | |
4.3.1989 | - | Departure for India. |
ACKNOWLEDGEMENT
I express my deep sense of gratitude to Dr. A.G. Jhingran, Director, Central Inland Capture Fisheries Research Institute, Barrackpore for deputing me to undergo such a valuable training. I acknowledge the financial aid rendered to me by FAO/NACA. I thank ICAR for permitting me to participate in the training programme. I am grateful to Dr. Mohamed Shariff, Coordinator, FPSS, University Pertanian, Malaysia for providing excellent facilities during the training period. Finally I remember with gratitude the whole hearted help extended by Dr. Chen Foo Yan, Coordinator, NACA and Dr. J. Richard Arthur, Programme Officer (Fisheries) IDRC.
ANNEXURE 1
Teaching Schedule for M.S. Tropical Fish Health Program 1988/89
Week | Course | Dates | Lecturers |
1 : | Aquatic Biology I | July 4 – 8 | Dr. Phillip Fatimah/Law |
2 : | Aquatic Biology I | July 11 –15 | Dr. Mohsin |
3 : | Aquatic Biology II | July 18 –22 | Ms. Siti Khalijah |
4 : | Aquatic Biology II | July 25 –29 | Dr. Chan Hooi Har |
5 : | Aquaculture Science | Aug 1 – 5 | Dr. Ang K.J/Salleh |
6 : | Aquaculture Science | Aug 8 –12 | Mr. Aizam/Salleh |
7 : | Aquaculture Science | Aug 15 –19 | Ms. Siti Shapor/Mr. Cheah |
8 : | Seminars+ Holidays | Aug 22 –26 | Coordinator |
9 : | Bofp III Pathology | Aug 29-Sept.2 | Dr. Supranne/Sharrif |
10 : | Bofp II parasitology | Sept 5 – 9 | Dr. Faizah Shahrom |
11 : | FPI-parasitic Diseases | Sept 12 –16 | Dr. Faizah Shahrom |
12 : | Bofp I Bacteriology | Sept 19 –23 | Dr. Gordon Bell |
13 : | Bofp I Bacteriology | Sept 26 –30 | Ms. Mariana |
14 : | FPI-Bacterial Diseases | Oct 3 – 7 | Ms. Mariana |
15 : | Bofp II parasitology | Oct 10 –14 | Dr. Jiri Lom/Faizah |
16 : | Bop II + FP II - Virology | Oct 17 –21 | Dr. Kinkelin/Hariyadi |
17 : | FP II Viral Diseases | Oct 24 –28 | Dr. Kinkelin/Hariyadi |
18 : | Examination Week | Oct 31-Nov 4 | Coordinator |
19 : | Farm visits | Nov 9 – 13 | Coordinator |
20 : | Farm visits | Nov 16 – 20 | Coordinator |
21 : | Seminars by guest lecturers | Nov 21 – 25 | Coordinator |
22 : | Seminars by guest lecturers | Nov 28-Dec 3 | Coordinator |
23 : | Bop III/FP II Mycology/Toxicology | Dec 5 – 9 | Ms. Gilda Lio Po |
24 : | FP II Non/inf diseases/Pharmoco | Dec 12 – 16 | Dr. Rohana |
25 : | Biostatistics | Dec 19 – 23 | Mr. Liew H.C. |
26 : | Biostatistics | Dec 27 – 31 | Mr. Liew H.C. |
27 : | FP III Public Health | Jan 2 – 6 | '89Dr. Abdullah/Gulam |
28 : | Bop III Haematology/Immunology | Jan 9 – 13 | Dr. Patrick Woo |
29 : | Bop III Immunology | Jan 16 –20 | Dr. Patrick Woo |
30 : | FP III Orn Fish/Invert disease | Jan 23 –27 | Dr. Gary Nash |
31 : | Workshop Histopathpathology | Jan 30 -Feb 4 | Dr.H.Fergusen/Arthur |
32 : | Fish Quarntine Systems | Feb 6 –10 | Dr. John Copland |
ANNEXURE - II
(To be filled in by the Scientist deputed abroad to attend Training course/Seminar/Symposium/Conference)
1. Name of the Scientist | : | Balbir Singh |
2. Grade | : | Rs.3,700–5,700 |
3. Designation of present post | : | Scientist (Sel. grade) |
4. Institute/Organisation | : | Central Inland Capture Fisheries Research Institute, Barrackpore, West Bengal, India. |
5. Discipline (i.e. subject of specialisation) | : | Fish and Fishery Science |
6. Details of deputations in the last 5 years (subject, period, name of the last organisation, whether sponsored by selection or nomination) | : | Not applicable |
7. Present deputation | : | To attend NACA/FAO training course |
a) Purpose | : | Tropical Fish Health programme |
b) Period | : | 4 July 1988 to 3 March 1989 |
c) Subject | : | Fish pathology |
d) Countries visited | : | Malaysia and Thailand |
8. Details about sponsors of visit. | : | |
a) Whether expenses were met by organisers/Outsiders/ or Institute/ICAR | : | By organisers |
b) Whether sponsored by selection in the ICAR or nominated by name by the host organisation. | : | Sponsored by ICAR |
( Balbir Singh )