Photo: Stuart Rowland

Introduction

For aquaculture stock, optimum farm design, appropriate husbandry, water quality management, and regular health monitoring all help to minimise disease outbreaks on farms. However at times, the farmer will experience disease problems, often resulting in stock losses. When the disease cannot be diagnosed on-farm, samples of the infected fish need to be submitted to a diagnostic laboratory, and it is a condition of every land-based aquaculture permit that:

The permit-holder shall notify the department within 24 hours of the discovery of any declared disease …, unusual disease or any significant event associated with the welfare of the fish on the premises  (eg unexplained or significant fish mortalities, >5% of fish stock loss in a week).

In the event of a declared disease, unusual or otherwise significant mortality:

Within 24 hours phone either:

Samples should be prepared for submission to a diagnostic laboratory as described below, and as directed by NSW DPI in accordance with guidance received at the time of reporting the event.
The importance of packaging appropriate samples for diagnostic purposes cannot be over emphasised. The selection and packaging of the samples together with the accompanying information often determines the success or failure of the diagnosis.  

On-Farm Diagnosis

Disease diagnosis begins on the farm. With the use of suitable equipment including water quality meters, a microscope and a dissecting kit a number of diseases, such as some external parasitic diseases, can be diagnosed on site. As a result, farmers can quickly act to reduce impacts of diseases and parasites.

Specimen Selection

Many pathogens (disease producing organisms) die or leave soon after the death of the fish, or their presence is masked by decomposition. Consequently, fish found dead in the water are of no or little value for diagnostic purposes. If the disease is sufficiently severe to cause some deaths, the remaining living fish will be infected with some of the pathogens. Ideally a range of clinically affected individuals should be sampled including those in early stages of disease, those displaying unusual behaviour and moribund (nearly dead) individuals. If fish of all sizes are affected, small specimens are easier to transport.

Live fish are the most desirable for reliable diagnosis. If this is not possible the specimens should be kept on wet ice and sent unfrozen. A final option is to send specimens preserved in formalin or other fixative (contact NSW DPI for further advice). In all cases, the sample should be representative of the population and include affected specimens.

Shipment of Specimens

Live specimens- Using one or two strong plastic bags, fill 1/3 full of water (preferably using the water the fish came from). Place the sampled fish (approximately 5-10 fish, depending on the size of the fish, and the size of the bag) in the water, squeeze out the air, then fill the bag with compressed oxygen making sure the air does not escape during the removal of the air delivery line. Twist and double over the neck of the bags and securely tie using tape, cord, or rubber bands. The bags should then be packed in a watertight container, sealed and clearly labelled "Scientific Specimens - Perishable" followed by the delivery address. During hot weather the specimens may be kept cooler by using crushed ice next to the fish. Overnight transport is essential.

Iced specimens- After collecting the fish, wrap in a damp cloth or paper towel. Place the sample in a plastic bag and cover with crushed ice. Shipment should be in a well-insulated container with a generous quantity of coarse crushed ice. Place in a watertight container and clearly label. Do not ship iced fish if delays are expected, as decomposition will render the sample unsuitable for diagnosis. Specimens should be received for examination within 24 hours.

Preserved specimens- Place the specimens in a plastic bottle using at least 10 times the volume of fixative as tissue. If the fish is more than 5mm thick or greater than 5cm in length, an incision should be made anterior to the anus to allow the penetration of the preservative into the body cavity. In the case of larger specimens, the organs or lesions under investigation should be dissected out and transferred into fixative. Skin lesions sampled this way should also include an adjacent region of normal tissue. Fill the container with 10% formalin.  Seal the container, place it within another watertight container and label correctly.

Figure 1. General location of internal fish organs

Frozen specimens- Frozen samples should only be considered as a last resort for disease diagnosis as some viruses, bacteria and parasites are inactivated or lost during freezing making identification impossible. However freezing of specimens is recommended for toxicological analysis providing the samples are frozen as soon as possible following the suspected poisoning.

Information to Accompany Specimen

It is important to provide as much information as possible with the specimens. Where possible this should include:

• Name; address; phone number and date
• Species of fish; age and size
• Water quality including pH; temperature; dissolved oxygen; ammonia; salinity
• Size and type of production facility eg. pond; tank; raceway
• Stocking density
• Date mortality began and mortality rate
• Clinical signs eg. poor feeding; abnormal swimming; "flashing"
• Water source
• Diet; size and manufacturer; ration; storage
• Recent husbandry eg. harvesting; grading; water exchange; fish delivery
• System changes eg. new tanks; changes in diet; chemical treatments
• Environmental changes; weather conditions.

This information needs to be submitted along with the;

• Veterinary laboratory specimen advice form  107.9 kb
  

Further information relating to sample preparation and submission may also be found in the NSW DPI Vet Lab Manual, or by contacting:

What steps can be taken to control losses while waiting for results?

Individual laboratories vary in the amount of time required to process samples and communicate the results to the producers.
Microbiology (bacterial isolation, identification and sensitivity; virology; identification of virus), and histopathology (microscopic examination of specially prepared tissues) may take between 2 to 14 days for completion. Results of post mortem examination should be available sooner.

• While waiting on results, the best approach is to remove mortalities from the pond and improve water quality through additional aeration and water exchange.
• Affected ponds or tanks should be isolated and equipment quarantined from other ponds or tanks.
• Minimise stock movements both within the facility, and stocking to other water bodies, in case you may be spreading an infectious agent (eg virus, new parasite).
• Effluent water should not be recycled but disposed of in a biosecure manner, such as after appropriate disinfection and ensuring it does not enter waterways.

For market size fish, food safety issues should be considered. For further information on food safety issues contact the NSW Food Authority (www.foodauthority.nsw.gov.au).