Hematological and serum biochemical reference intervals of rainbow trout, Oncorhynchus mykiss cultured in Himalayan aquaculture: Morphology, morphometrics and quantification of peripheral blood cells

Abstract

De novo reference intervals (RIs) for a total of thirty two hematological and serum biochemical attributes were established for rainbow trout (Oncorhynchus mykiss) cultured in Himalayan aquaculture system. For this purpose, long term assessment of hemato-biochemical parameters was carried over a period of one year from March 2019 to February 2020 and a total of 444 blood samples were analysed. Blood examination results were recorded systematically and reference intervals were established, notably for erythrocyte parameters: hematocrit (Hct) 29-40%, hemoglobin 8.32-12.28 g/dL, red blood cell (RBC) count 1.01-2.04 (×10⁶/mm3); leukocyte parameters (x 10³/mm3): total leukocytes 31.32-90.60, neutrophils 4.21-18.85, total lymphocytes 20.55-63.63, small lymphocytes 14.86-46.50, large lymphocytes 6.35-22.34 and monocytes 1.22-7.56; thrombocyte count 23.00-68.00 (×10⁶/mm3). RIs were also established for red blood cell indices, vital serum constituents involved in carbohydrate, protein, lipid and nitrogen metabolism including the less known, diagnostically important, serum enzymes and electrolyte concentrations. Principal component analysis revealed that certain serum components were more efficient at distinguishing between the life stages (juvenile, adult) of fish by explaining about 92.7% of variation in the whole dataset compared to the principal hematological components which explained only about 80% of the variation. Significant (P < 0.05) differences were noted for RBC count, total leukocyte count (TLC), total protein, total cholesterol and uric acid with respect to the sex of fish. Moreover, clearly differentiable morphometric and morphological attributes were also noticed among erythrocytes, leukocytes (lymphocytes, neutrophils and monocytes) and thrombocytes. To our knowledge, the present study is the first of its kind that elucidates blood chemistry of cultured rainbow trout, O. mykiss in accordance to the guidelines framed by the American society of veterinary clinical pathologists (ASVCP). RIs reported here can help monitor the fish health status by improving the use of non-lethal diagnostics in piscine medicine.

Keywords: Aquaculture; Blood biochemistry; Cell morphology; Rainbow trout; Reference values.

Fig. 1

Map showing the location and layout of the study site at Dachigam, Kashmir, India.

Fig. 2

Peripheral blood cells of rainbow trout, O. mykiss stained with Wright Giemsa. (A) Erythrocytes (EØ) depicting prominent eccentric nucleus surrounded by a clear cytoplasmic region. (B) A large lymphocyte (LL) and a small lymphocyte (SL) with nucleus surrounded by deep blue cytoplasm demarcated by a fine whitish rim at the cell periphery. (C) A neutrophil (NØ) with ribbon shaped, light blue coloured nucleus surrounded by pale lavender coloured cytoplasm. (D) A neutrophil (NØ) with heavy multi-lobed blue coloured nucleus surrounded with pale cytoplasm. (E) Monocyte (MØ) with indented nucleus and vacuolated cytoplasm; small lymphocytes (SL). (F) Thrombocytes (TØ), rod shaped with spindle-shaped or fusiform nucleus that conforms to the shape of the cell ending with transparent cytoplasmic protrusions. (G) Thrombocytes (TØ) with horse shoe shaped nucleus. (H) Thrombocytes (TØ), short statured with oval shaped nucleus and a protruding cytoplasmic end. Scale = 10 µm.

Fig. 3A

Line plot depicting percentage variation in erythrocyte indices between life stage and sex.

Fig. 3B

Dot plot showing distribution pattern of erythrocyte indices between life stages and sex.

Fig.4A

Score plot of hematological parameters for juvenile and adult fish.

Fig. 4B

Loading plot of hematological parameters for juvenile and adult fish.

Fig. 5A

Variation in percentage composition of leukocytes between life stages and sex.

Fig.5B

Box plots depicting reference intervals for TLC and DLC between life stages and sex.

Fig. 6A

Score plot of serum biochemical parameters for juvenile and adult fish.

Fig. 6B

Loading plot of serum biochemical parameters for juvenile and adult fish.