NSW Department of Primary Industries Fisheries website

DPI veterinary pathologists can be contacted for advice on the investigation of fish health problems.

Standard containers and equipment

Water sample containers

The types of containers recommended for collection of water samples include:

• polyethylene or glass for ammonia, nitrate/nitrite, pH, cyanide and algae;
• glass stoppered glass for oxygen, filled to exclude air;
• nitric acid washed polyethylene for metals;
• new glass bottle for pesticides;
• sterile 200 mL glass bottles for bacteriology, chilled at 4°C, and submitted within 24 hours of collection.

Water test kits are available for routine testing for nutrients.

Fixatives

(Neutral) buffered formalin

Buffered formalin solution is the recommended fixative because of its stability, longer life of preserved tissues, and a negligible fixation deposit. It is prepared as follows:
 

Buffer salts are available commercially in prepared packs.

*Commercial formalin is Formalin BP 34-38% formaldehyde. It should be used neat in this formulation.

Bouin's fixative

Davidson's solution

Fixation time: 24 hours (minimum)

History and diagnosis

History taking

1. A comprehensive history assists the investigation. Estimates of morbidity and mortality, based on stocking density on farms or the abundance of the species in natural waterways should be provided. Include measurements of length and weight, with an estimate of the age or stage of growth. Larger fish are more likely to suffer oxygen deprivation, whereas smaller fish tend to be more susceptible to toxic insults. If other inhabitants of the aquatic environment are affected, poor water quality or exposure to toxins should be suspected.
2. A description of the locality of the fishkill should include the type of habitat, whether natural or man made, river, stream, lake, pond, dam, tank, raceway, cage or aquarium. On fish farms the design and construction of facilities and the source of water should be related to the number and distribution of ponds affected. Freshwater, brackish, estuarine or ocean environments are important because of their differences in salinity. A general description of the landuse in the local drainage basin or watershed may be appropriate.
3. Describe the condition of the water in terms of colour, turbidity, odour and flow. Is the water level high, low or normal? Are algae, organic debris, froth or oil floating at the surface? Does the ecosystem appear "healthy"?
4. Record the temperature of the water at several sites and depths. Submit the results of any onsite water quality tests performed. These measurements, especially O₂ and pH, could be taken at various sites (inlet, middle and outlet) and depths (surface, mid level and bottom) in the body of water.
5. Details of composition of diet, history of dietary changes, source or manufacturer of feed, rate of feeding, storage conditions and duration of storage could allow nutritional deficiencies or toxicities to be identified.
6. List any treatments, including chemicals and antibiotics, which have been administered to the fish. On many occasions malachite green or formalin will have been used; although these treatments may have eliminated the causative agent, rendering subsequent examinations negative, they may have caused further direct or indirect injury to the fish and these potential complications should be considered. What dosage regime (time of treatment, duration of treatment and concentration or dose rate of chemical) has been applied?

Diagnosis

History, clinical signs, field assessment of water quality*, field evaluation of environmental and ectoparasitic causes of disease**, histopathology, parasitology, bacteriology, virology, toxicology.

*Water quality assessment is best performed in the field using commercial test kits for ammonia, nitrite/nitrate, acidity, hardness and copper.

**Microscopic examination of wet preparations of gills, skin and fins should allow common parasites to be identified.

Diagnostic hints

1. Mortality patterns can be fitted to the time of year, day or season. For example, deaths due to oxygen deprivation are most likely to be clustered in the early hours of the morning when water oxygen levels are depressed. Some diseases, such as infestations with the gill protozoan Chilodonella in many species or epizootic haematopoietic necrosis (EHN) virus in redfin perch, are recurrent seasonal problems.
2. The signs exhibited by diseased fish are readily observed in glass sided aquaria, but may be difficult to detect in open water, unless fish are surfacing. Diseased fish may change the depth which they normally occupy in the water column or their spatial relation to inlet or outlet points to a pond. Feeding patterns may be altered. Behavioural abnormalities such as gasping at the surface of the water, flashing or turning upside down may be displayed. Skin colour may change.
   • Superficial lesions affecting the skin, fins, gills, eyes, mouth or anus may be visualised in the live fish. The results of any clinical examination should be detailed.
3. Any lesions noted at autopsy or during handling should be recorded. If several fish are available for examination, an onsite autopsy should be performed on at least one. Collect fresh and preserved specimens from any autopsies and submit these along with any live affected or freshly dead fish which are available.

Specimens required, storage and despatch

1. Live moribund or affected fish (preferred specimens)
   Fish to be submitted are placed in a strong plastic bag inside another plastic bag, together with a half volume of water from the source environment. If likely to be in transit for any length of time, the plastic bag should be inflated with oxygen before twisting, double folding and sealing with strong rubber bands. The plastic bag is then placed inside an esky and sent to the laboratory.

   Fish should never be fed before transport; this increases metabolic rate which increases oxygen consumption and may lead to fouling of the water. Sick fish should also be protected from changes in temperature.

   Despatch of several survivors and/or several healthy control fish is often beneficial in reaching a diagnosis, as this allows the pathologist to compare affected and unaffected/recovered animals in greater detail.
    

2. Dead fish (rarely of much diagnostic value, unless they are examined on site)
   Dead fish are virtually useless after transport, even if chilled, because decomposition in fish occurs rapidly and many ectoparasites will detach from the host soon after death. If only dead fish are available for submission to the laboratory, submit:
   • Fixed tissues for histopathology (must be fixed on site)
   • Frozen tissues for toxicology/virology. If only dead fish are submitted, bacteriology is precluded (culture must be undertaken within 1 hour of death to be of value). Frozen fish are unsuitable for histopathology.
     
3. Tissues in fixative (submitted in addition to live fish)
   This ensures that serviceable specimens will be available should there be transportation delays that result in death or deterioration of other specimens. Small fish may be immersed in fixative whole, with a section of abdomen removed to allow penetration of fixative to internal organs. A range of tissues could be taken from larger fish at autopsy, concentrating especially on those tissues which exhibit gross pathology.

   In addition to internal organs such as heart, liver, kidney, spleen and intestine, consider submitting brain, skin, muscle and gonad. As autolysis is rapid in the gills, sections of the gill arches should be placed in fixative at the start of the examination.

   Ten percent neutral buffered formalin or 5% formol saline are suitable fixatives for fish tissues. Bouin's fixative can be used if the specimens are small, for rapid penetration and optimal preservation. The tendency of some ectoparasites to detach from gills and skin, even in the presence of formalin, may be overcome by the use of Davidson's solution
    

4. Fresh tissues (for bacteriological examinations, virology and toxicology)
   Tissues from large fish (which have been freshly killed) may be removed and transported chilled in sterile containers. A pool of fresh chilled brain, liver, kidney and spleen is often suitable for isolation of viruses. Swabs are less suitable.
   
5. Smears of blood and body fluids (for haematology and bacteriology)
   Many examinations for ectoparasites are best carried out in the field using wet preparations. Rapid blood stains can also be used for smears from fish.

In special cases and after consultation specimens can be forwarded to the Australian Fish Health Reference Laboratory, Geelong, Victoria.

Charges for laboratory tests commonly applied to aquatic animals

Further information on laboratory fees

Some diseases of fish, crustaceans and shellfish encountered in NSW

Fish Kills

Attributed to:

• Anoxia
• Environmental toxins e.g. superphosphate
• Infectious diseases e.g. Epizootic haematopoietic necrosis (EHN) in redfin perch.

Skin lesions in fish

Often multifactorial; may involve nutritional, parasitic, bacterial and viral pathogens. Histopathology of lesions has been of value. Causative agents include:

• Bacterial infection with Aeromonas hydrophila in hatcheries, seen in stressed adult fish.
• Yellow spot
• Red spot (Epizootic ulcerative disease, EUS) in estuarine fish on the North Coast
• Goldfish ulcer disease due to Aeromonas salmonicida or Vibrio infections. The former is an important pathogen of Salmonids

Shellfish diseases

• Discolouration in abalone

Crustacean diseases

• Protozoal myositis with ataxia in yabbies