HOW TO ASSESS THE HEALTH OF YOUR OYSTERS

History

The history of each group of oysters being examined should be closely followed, particularly information related to water source, pond, cage, rack, bag or system number, species, age, stocking rate, growth performance, survival rates, grading history and parasite or pest burdens. Any abnormalities in any of these areas should be noted when they are observed so that these data are available for future reference if required.

Water quality

Basic water quality parameters such as temperature and dissolved oxygen should be regularly monitored. Other water variables such as salinity, pH, microbiology (enteric virus or faecal coliform counts) and presence or absence of toxic phytoplankton should also be recorded. Ideally these records should be available for the entire history of each group of oysters being examined.

Gross examination

The oysters should be examined for abnormal behaviours such as gaping or other visible signs of stress. The presence of mudworm, flatworm or boring sponge infestation of the shell should also be noted on data sheets .

Shucking

Measure the oysters for appropriate biological data (shell height, width, wet weight) prior to shucking - add this data to the data sheets. Then shuck the oyster, taking care not to contaminate the internal organs by ripping them with the knife. Look at its condition, note whether it is fat, medium or watery. Also look for other abnormalities on the shucked shell, such as evidence of mantle recession, boring sponge, pustules, as well as symbionts such as pea crabs, copepods etc. Dead oysters are of limited value, they should be noted, but not included in any subsequent diagnostic techniques.

Heart or digestive gland imprints in oysters

Stained imprints of the heart are useful for examining for the presence of Bonamia sp. in flat oysters (Angasi oyster Ostrea angasi, Bluff Oyster Ostrea chilensis, see Figure 1). Stained imprints of the digestive gland are useful for examining for Marteilia sp. (QX disease) in cupped oysters (Pacific oyster Crassostrea gigas, Sydney rock oyster Saccostrea glomerata).

Prior to starting, label an appropriate number of clean, frosted microscope slides with:

Site number / oyster number (e.g 03 / 09) = oyster number 9 at site 3

For detection of Bonamia sp. in flat oysters:

1. Remove a small part of the black region of the heart (dark organ adjacent to adductor muscle) with fine forceps. Do not remove the entire heart as only a small amount of tissue is required for imprints.

2. Place heart on filter paper for a moment (no more than 5 seconds) to remove excess water/fluid, This is just enough time to wipe the forceps on filter paper to remove excess water wicked up the forceps shafts.

3. Pick up heart with forceps and make 3 or 4 rows of 8 to 10 imprints per row on the appropriate slide. The imprints should be done gently, or else excessive tissue will be deposited on the slide and cell integrity will be disrupted.

4. Place finished slides in slide rack for at least 5 minutes to air dry.

5. Take slide racks to dry laboratory for staining with Hemacolor or DiffQuick (see Recipes page), following the manufacturers recommended method.

For detection of Marteilia sp. in cupped oysters:

1. Remove a small part of the digestive gland with fine forceps.

2. Place the piece of digestive gland on filter paper for a moment (no more than 5 seconds) to remove excess water/fluid.

3. Make 3 or 4 rows of 8 to 10 imprints per row on the appropriate slide. The imprints should be done gently, or else excessive tissue will be deposited on the slide and cell integrity will be disrupted.

4. Place finished slides in slide rack for at least 5 minutes to air dry.

5. Take slide racks to dry laboratory for staining with Hemacolor or DiffQuick (see Recipes page), following the manufacturers recommended method.

Histopathology

Histopathology is a routine method used to detect a variety of oyster pathogens, such as Bonamia sp. Figure 2, and all the other internationally notifiable disease agents of molluscs listed by the OIE (see Links page). Using a scalpel or sharp scissors, cut a standard cross section 3 to 5 mm thick located to include all major organs of interest (see figures 4-8 . Place the excised section into a labelled histology cassette and fix in 10% seawater formalin (see Recipes page). Ensure that the volume of fixative used is at least 5 times that of the tissues being fixed. If grossly visible abnormalities are observed in any organs, make sure the abnormal tissues are included in the samples excised for histopathology.

Bacteria testing

Bacteriology uses specialised techniques which should be conducted using methods recommended by the diagnostic laboratory which will undertake the sample screening. Contact DigsFish Services or the laboratory responsible for processing your samples for more information on their desired methods of sample collection.

Electron microscopy

Excise small sections (1-2 mm3) of digestive gland, gill, mantle, gonad or any other organ of interest and place them in a 1.5 ml eppendorf tube containing 2.5% gluteraldehyde (see Recipes page). Store them at 4C for 24-48 hours. Rinse once in seawater filtered to 0.22 microns and send to DigsFish Services for processing.

Molecular techniques

Excise small sections (1-2 mm3) of digestive gland, gill, mantle, gonad or any other organ of interest and place them in a 1.5 ml eppendorf tube containing 70% ethanol (see Recipes page). Store them at 4C for 24-48 hours, then send to DigsFish Services for processing Figure 3.