Factors Affecting Fish Blood Profile: A- Effect of Nutritional Treatments

Press and Evensen (1999) mentioned that species variation in the morphology of the immune system is to be expected, given the large number and diversity of species within the teleost fishes. Several external and internal factors can influence the activity of innate immune parameters. Temperature changes, handling and crowding stress can have suppressive effects on innate parameters, whereas several food additives and immunostimulants can enhance different innate factors. There is limited data available about the ontogenic development of the innate immunological system in fish (Magnadóttir, 2006). Moreover, Sahan and Duman (2008) found that haematocrit, leucocytes, monocytes, and neutrophils were increased in common carp fed with beta glucan. Also, Aly et al. (2008) showed significant increase in haematocrit values in group of Nile tilapia fed the mixture of B. subtilis and L. acidophilus comparing with the control. However, the present study was designed to evaluate the effect of some nutritional treatments on some blood parameters.

Random fish samples were taken for blood collected from the caudal peduncle by special syringe, adequate amount of whole blood was withdrawn in small plastic vials containing EDTA (ethylene diamine tetra acetic acid) as anticoagulant and used to obtain the blood plasma by centrifuge at 3500 rpm for 15 min. Blood plasma samples were used for determination of creatinine (Tietz, 1986), triglycerides (McGowan et al., 1983), total proteins (Tietz, 1990) and albumin (Wotton and Freeman, 1982) concentrations as well as the activity of aspartate amino transferase (AST) and alanine amino transferase (ALT) using commercial test kits in a private lab. in Kafr El-Sheikh governorate, Egypt. Globulin level was calculated by subtracting albumin from total protein. The other samples of blood were used to determine the blood hematology as concentration of hemoglobin (Hb), total count of erythrocytes (RBCs), and total leukocytes (WBCS) (Natt and Herrick, 1952) and hematocrit (Hct) using Auto Counter (Decie and Lewis, 2006) in the same lab. The other hematological parameters were mathematically calculated. Five fishes from each treatment were chosen to withdraw blood samples for analysis to study the effects on blood profile of:

All obtained data were analyzed according to statistical analysis system software (SAS, 2006) for windows. Multiple range tests (Duncan's, 1955) were used to compare between the parameters of the different nutritional groups. The differences were significant at 0.05 levels.

A study was conducted over a 96 days in order to comprise between floating and sinking diets with and/or without a probiotic and their effects on growth performance chemical composition and food utilization with Nile tilapia (Oreochromis niloticus). The two studied factors were the diet type (floating and sinking diets) and probiotic (PRO-LYNE, at 0%, 1% and 2% of the diet). Food type (floating or sinking diets) significantly affected blood values of hematocrit (Hct), mean corpuscular haemoglobin concentration (MCHC), platelets and white blood cells (WBCs); whereas the probiotic level affected all tested haematological parameters (Table 1), except red blood cells (RBCs), mean corpuscular volume (MCV), and mean corpuscular haemoglobin (MCH). Meanwhile, the only significant interaction (food type x probiotic level) was calculated for Hct and MCH. Concerning the biochemical parameters (Table 2), food type affected significantly urea, triglycerides (TG), and low density lipoprotein (LDL) only. The probiotic level did not affect the biochemical parameters measured. Yet, the interaction was significant for aspartate aminotransferase (AST), alanine aminotransferase (ALT), and high density lipoprotein (HDL).

A feeding experiment for 57 days was conducted on Nile tilapia to evaluate the effects of dietary inclusion of graded levels (0, 25, 50, 75, and 100%) of sieving wastes meal of Egyptian clover seeds instead of soybean meal based on crude protein content. The replacement level significantly affected all tested haematological parameters, except MCH (Table 3). It affects also albumin (AL), urea, total cholesterol (TCH), and HDL (Table 4).

To evaluate the effects on Nile tilapia of replacing 25 and 50% of the diet's corn by meals of fruit skin and leaves of Teen Barshomy (Teen Shoky). Hundred all-males mono-sex Nile tilapia fish (17 g initial body weight) were adapted for one week. Thereafter, feeding trial duration was 75 days. Five experimental diets were formulated to replace 25 and 50% of dietary corn by fig skins and fig leaves besides the control one. The replacement type (pee/waste) significantly affected WBCs only; whereas, the replacement level affected significantly all tested haematological parameters, except MCV, MCH, and MCHC. The interaction (replacement type x replacement level) was significant only for platelets (Table 7). The replacement type significantly affected TCH, HDL, and LDL. Moreover, the replacement level affected significantly albumin (AL), globulin (GL), total protein (TP), and LDH (Table 8). The interaction (type x level of replacement) was significant only for TCH. 

Gray and Hall (1930) gave normal blood sugar value for some fish species [makrele (Scomber scombrus) and Opsamusta] as 63.5 and 15.4 mg%, respectively. Yet, El-Houssiny et al. (1999) reported no significant (P≥0.05) differences in non-specific immunity factor (serum proteins) nor humoral immune response in Nile tilapia fed different protein levels (20, 25, and 30 %), feeding levels (3 and 5 %), and feeding frequency (2 and 3 times) for 170 days. They gave the following ranges for total serum protein 2.51-2.91, albumin 0.99-1.17, globulin 1.48-1.83 g/dl, and albumin/globulin 0.57-0.86. Hussein et al. (2001) found that canola meal increased significantly Hb, serum total protein, globulin, triglycerides, AST, ALT, and thyroid hormones levels. Dietary yeast strains increased Hb, serum total protein, globulin, triglycerides, glucose, AST, ALT, and thyroid hormones levels. They gave the following ranges: Hb (6.4-9.3 g/dl), PCV (22.0-25.3 %), serum total protein (4.83-10.5 g/dl), albumin (1.74-2.64 g/dl), globulin (2.57-8.68 g/dl), triglycerides (248-375 mg/dl), glucose (93.5=128 mg/dl), total cholesterol (146-165 mg/dl), AST (38.8-66.4 u/l), and ALT (22.5-37.9 u/l). El-Ebiary and Zaki (2003) registered increases in the Hb (6.36-8.39 g/dl), PCV (23.1-25.0 %), serum total protein (4.85-9.46 g/dl), albumin (2.26-264 g/dl), globulin (2.59-6.85 g/dl), glucose (92.5-148 mg/dl) and triglycerides' (255-356 mg/dl) concentrations but decrease of the total cholesterol level (156-143 mg/dl) by increasing the level of active yeast in the Nile tilapia diets. Red tilapia fed sesame hulls by-product reflected lower plasma glucose (78.6-67.2 mg/dl) and total protein concentrations (5.10 vs. 2.69 g/dl) and ALT activity (39.5-10.9 u/dl)  comparing with the control (Abd Elmonem et al., 2004). Ayyat et al. (2004) fed Nile tilapia different levels of fish meal and zinc.

They found that serum total protein (5.00/5.69 g/dl), albumin (2.90/3.65 g/dl), and AST (30.4/36.6 u/l) values significantly increased with increasing fish meal level in the fish diet, while urea-N (3.83/3.65 mg/dl) and ALT (16.2/12.8 u/l) values significantly decreased. Also, serum total protein (4.60/5.84 g/dl), albumin (2.68/3.39 g/dl), and creatinine (0.90/1.02 mg/dl) concentrations and AST activity (29.6/37.2 u/l) significantly increased with increasing Zn level in the fish diet, while ALT activity significantly decreased. Moreover, El-Dakar et al. (2004) found that dietary inclusion of graded levels of fennel seeds meal as a feed additive affected significantly some serum biochemical parameters, particularly total lipids (3.33-5.51 g/dl) and AST (72-116 u/dl) but did not affect glucose (34.3-59.6 mg/dl),  total protein (5.03-5.43), and ALT (34-45 u/dl). Abdel-Tawwab et al. (2005) represented that Nile tilapia reflected slightly lower Hb and PCV values under crowding-stress condition but there was no significance in them due to dietary CP level. They added that RBCs count and glucose concentration in chronically stressed fish were elevated but plasma cortisol value was reduced particularly with increasing the dietary protein level. However, both transaminases (AST & ALT) were not affected. That means that Nile tilapia may quickly adapted to high rearing density by enhancing feed quality especially protein level in the diet to prevent the deleterious effect in fish farm. The recorded the following ranges: RBCs 1.26-1.61 x 106µl-1, Hb 6.06-8.40 g dl-1, PCV 13.8-16.5 %, glucose 64.9-128 mg dl-1, protein 2.07-3.45 g dl-1, ALT 6.08-8.75 ul-1 and AST 17.4-65.9 ul-1. 

Magnadóttir (2006) overviewed the innate immunity of fish. He cited that the innate immune system is divided into physical barriers, cellular and humoral components. Humoral parameters include growth inhibitors, various lytic enzymes and components of the complement pathways, agglutinins and precipitins (opsonins, primarily lectins) natural antibodies, cytokines, chemokines and antibacterial peptides. Several external and internal factors can influence the activity of innate immune parameters. Temperature changes, handling and crowding stress can have suppressive effects on innate parameters, whereas several food additives and immunostimulants can enhance different innate factors. There is limited data available about the ontogenic development of the innate immunological system in fish. Osman et al. (2010) registered that feeding with probiotics improved the growth performance parameters, feed efficiency, RBCs count (1.159 x 106 vs. 0.999 x 106/µl), Hb concentration (5.42 vs. 4.77 g/dl), phagocyte activity, A/G ratio, and serum total protein level of Nile tilapia. Nile tilapia fed with spirulina (Arthrospira fusiformis) supplementation significantly decreased plasma AST (108 vs. 115 u/dl) and ALT (41 vs. 46 u/dl) activity, but significantly increased plasma glucose (63.87 vs. 58.11 mg/dl), total protein (7.58 vs. 5.10 g/dl), and total lipid (5.28 vs. 4.23 g/dl) (Belal et al., 2012).

Shortage of food supply and animal feedstuff gape forced the nutritionists to find out and evaluate novel or unconventional food resources such as corn gluten meal (Metwalli, 2013) and insect (black soldier fly) larvae to replace the expensive fish meal in aqua feed (Makker, 2015). Al-Ashrm (2017) mentioned among the properties of an ideal narcotic agent, it must be not toxic to fish and human being, do not affect fish behavior or its physiological functions, and to be cheap. He added that clove oil (extracted from the plant Eugeaia caryaphyllata) is an local product, safe, effective, and cheap narcotic agent well recommended for fish narcosis. Moazenzadeh et al. (2017) investigated the effect of dietary Zn levels (0, 5, 10, 20, and 40 mg Zn /kg) on hematological parameters of juvenile Siberian sturgeon (Acipenser baerii, Brandt 1869). Erythrocyte number, hemoglobin concentration, and hematocrit were increased with raising dietary Zn level (P≤0.05), whereas a regular trend was not observed in blood indices, including MCV, MCH, and MCHC.

A significant increase was also found in the leucocyte number followed by their differential counts, except for eosinophil percentage. In spite of well correlation with dietary Zn level, no break point was observed to estimate the juvenile Siberian sturgeon requirement based on the erythrocyte numbers. Hassaan et al. (2018) fed Nile tilapia, Oreochromis niloticus (average initial weight, 5.91 ± 0.04 g) isonitrogenous and isolipidic diets for 84 days. The diets contained four levels of yeast extract rich in nucleotides and β-glucan: 0 (control), 5, 10 and 15 g/kg diet. No significant (P>0.05) differences were found in haematocrit, haemoglobin or total protein contents among the treatment groups. Blood sample profiles showed an increase in white and red blood cells in fish fed 15 g/kg yeast extract in comparison with the other treatment groups. The fish fed the diets with 10 and 15 g/kg yeast extract had significantly higher albumin and globulin levels than the control group, whereas decreased levels of cholesterol and triglycerides, aspartate aminotransferase and alanine aminotransferase were noted in fish fed the diet with 15 g/kg yeast extract.

Conclusively: The interpretation of blood data of fish must be done on light of the specific individual experiment condition because of absence of referenced ranges for fish haematological and biochemical measurements.