Fish Diseases and Management

Gastrointestinal examination

Followed by external examination and  gill observation, the gut examination has to be carried out. Following procedure is adopted.

• For examination of hepatopancreas, head is separated from abdomen to expose midgut and hepatopancreas and it is examined for atrophy
• squash hepatopancreas between fingers and note colour and texture:

1. if white and fluid filled indicates severe atrophy;
2. presence of black streaks could indicate necrotising hepatopancreatitis (NHP), septic hepatopancreatitis (SHPNS, probable Vibrio sp. Infection), or aflatoxicosis;
3. orange to reddish usually will be normal.
4. For microscopic examination, remove HP and place it on a clean slide and bisect on mid line.
   • Make impression smears and fix with methanol for 6 min and air-dry prior to staining. Giemsa staining is carried out as follows:
   • Preparation of Giemsa stock solution - Mix 66.0 ml of glycerin and 1.0 g Giemsa powder. Place in 60 °C oven for 2 hours. Add 66.0 ml methyl alcohol and mix.
   • Giemsa working solution – Mix 1.25 ml Giemsa solution, 1.50 ml methyl alcohol, 2 drops of sodium carbonate and add 50 ml distilled water.
   • Preparation of Rosin alcohol stock solution – dissolve 10.0 g Rosin, white in 100.0 ml Absolute alcohol
   • Rosin alcohol working solution – Mix Rosin stock solution 5.0 ml and 40.0 ml of 95% alcohol
     
   1. Procedure for Giemsa staining
      • Stain the slide in working solution of Giemsa at room temperature for overnight or 30 min at 60 °C (use fresh solution, discard used solution)
      • Differentiate in working rosin alcohol solution until smears assume a purplish pink colour.
      • Dehydrate in absolute alcohol and clear in xylene and mount in DPX .
      • Examine for colour, melanized foci or streaks, atrophy, and other signs that may be abnormal
5. Remove with forceps several medial HP tubules and place in a drop of clean seawater on slide.
6. If BP, MBV, or BMN are suspected, add small drop of 0.01% malachite green, squash with coverslip, and examine several items over next 5 min for baculovirus occlusion bodies and / or hypertrophied HP cell nuclei (as described earlier).
7. If baculovirus is not suspected, coverslip the tissues and examine for lipid droplets (quantify grade of lipid droplets present. A few to none are sign of poor health and disease), melanized HP tubules, abnormally high numbers of motile rod shaped bacteria and gregarine trophozoites.

For gut examination, check the gut contents in midgut (MG) and stomach. For microscopic examination, dissect and remove MG from its junction with HP in head and hindgut in the sixth abdominal segment. Place MG on a dry clean glass slide with forceps and a coverslip on edge, strip MG to exude its contents onto slide. Add a drop of clean seawater to the contents, spread as needed, place coverslip and examine for

• Gregarine trophozoites and/or gametocytes
• Baculovirus occlusion bodies (MBV or BP)
• Melanized masses of haemocytes that could indicate haemocytic enteritis or vibriosis