Haematological Responses of Heteroclarias Fed Dietary Levels of Alchornia Cordifolia Leaf Meal.

Transcription

1 International Journal of Research in Pharmacy and Biosciences Volume 2, Issue 6, July 2015, PP ISSN (Print) & ISSN (Online) Haematological Responses of Heteroclarias Fed Dietary Levels of Alchornia Cordifolia Leaf Meal. Anyanwu a, D.C., Udedibie b, A.B.I., Osuigwe b, D.I., Offor a J.I., Onogu B. a a Department of Agric. Science, Alvan Ikoku University of Education, Owerri. b School of Agriculture and Technology, Federal University of Technology, Owerri. ABSTRACT The haematological responses of feeding 35% isonitrogenous dietary levels of 0%, 5%, 10%, 15% and 20% Alchornia cordifolia leaf meal on Heteroclarias (Heterobranchus bidorsalis x Clarias gariepinus) post fingerlings were assessed. These were fed to the fingerlings, randomly assigned to 5 treatments: 0%-control(TCN), 5%(TA1), 10%(TA2), 15%(TA3) and 20% (TA4) ACLM in 3 replicates of 15 post fingerlings each using 15 plastic aquaria of 250 x 150cm dimension. The fish were fed at 5% body weight twice daily within the experimental period of 56 days. Samples of blood were coollected from the treatments bi-weekly and analyzed to evaluate some of the haematological profile, and data subjected to a one way analysis of variance (ANOVA). The study revealed significant differences among the treatments. The haemoglobin level of the fish on the control diet (5.9g/100ml) and TA2 (5.9g/100ml) were similar, (p>0.05) to TA1 (5.8g) and TA3 (5.58g/100ml). There was no significant difference (p>0.05) between TA1, TA3 and TA4.The red blood cells for TCN (3.0 x 106mm3), TA1 (2.44 x 106/mm3) and TA2 (2.48 x 106/mm3) were significantly different and higher (p<0.05) than TA3 (2.23 x 106/mm3) and TA4 (2.07 x 106/mm3), while TA3 and TA4 were not different (p>0.05).the white blood cells of the fish TCN (4.75 x 104/mm3) were significantly (p<0.05) higher than TA1 (4.5 x 104/mm3), TA2 (4.5 x 104/mm3) and TA4 (4.58 x 104/mm3), while TA3 (4.1 x 104/mm3) was the least. Packed cell volume of the fish on TCN (23.58%) was not different (p>0.05) from TA1 (23.0 %) and TA2 (23.25%). This was significantly (p<0.05) higher than TA3 (22.25%) and TA4 (22.25%), whereas there was no significant difference between TA1 and TA2 (p>0.05).the MCV value of mm3 for TA4 was significantly greater (p<0.05) than other TA treatments. This was followed in ranking sequence by MCV values of 99.77mm3, 94.26mm3, 93.75mm3 and 79.2mm3, for treatments TA3, TA1, TA2 and TCN, respectively. TA3 was significantly (p<0.05) higher than TA1 and TA2, while these on the other hand were higher than TCN. The MCH value of 26.71pg for TA4 was significantly higher (p<0.05) than the other TA treatments, while 20.82pg for TCN was the least. TA3 (24.02pg) was significantly higher (p<0.05) than TA1 (23.89pg) whereas TA2 (24.85pg), TA1 and TA3 were similar (p>0.05).the MCHC values for A. cordifolia leaf meal ranged from 24.85% %, but the treatments, were not significantly different (p>0). INTRODUCTION Nutrient utilization and biological values of feeds and feedstuffs evaluation may seem inconclusive without adequate consideration on their implications on the physiological and health status of the animal for example fish. Haematological and biochemical indices of farm fish as with other farm animals namely - haemoglobin, red blood cells, white blood cells, packed cell volume, plasma protein, blood glucose, specific gravity of blood plasma and whole blood, coagulation time etc. however have been analysed and variously reported as useful tools in assessing the performance, viability and health status of farm fish and animals (Blakhall and Daisley, 1973; Bhaskar and Rao, 1990; Musa and Omoregie, 2001; Harikrishnan et al, 2003; Anyanwu et al, 2003; Hemre et al, 2007; Anyanwu et al, 2011 ). Unconventional nutrient sources using tropical browse plant leaf meals like Alchornia cordifolia and other terrestrial or fresh water aquatic plant leaf meals due to their easy availability, potentials for nutrients provision and low procurement cost, would seem to provide acceptable feeding regimes for species of fish like catfishes, hybrids, and other farm animals (Udedibie and Opara, 1998; Reyes and Fermin, 2003; Ochang et al, 2007 Anyanwu et al, 2011). It is against this backdrop that this study was designed to determine the effect of ACLM on the haematology of Heteroclarias (H. bidorsalis Clarias gariepinus) post fingerlings. *Address for correspondence ahamefula_dan@yahoo.com International Journal of Research in Pharmacy and Biosciences V2 I6 July

2 MATERIALS AND METHODS The experiment was carried out in a farm s fisheries house of 8 x 6m 2 situated in Owerri, Imo State. A total of 15 plastic aquaria (250cm x 150cm), covered with mosquito mesh nylon screen to prevent fish from jumping out and possible predation were used. The Alchornia cordifolia leaves were harvested from bushes at the outskirt of the Owerri capital territory, along Owerri/Onitsha Road, Imo State. These were spread under the sun and dried for three days until they became crispy while still retaining the green colouration. The dry leaves were milled, using a hammer mill to produce to leaf meal. The leaf meal was used to make 4 35%CP Isonitrogenous diets at inclusion levels of 5%, 10%, 15%, and 20% for TA 1, TA 2, TA 3, and TA 4 respectively. Maize was used as the major source of energy in the diets, while soyabean meal and fish meal as major sources of protein (Table 1), besides, the use of lysine and methionine at 0.2% levels of inclusion. 1% bone meal was used, with Vitamin/mineral premix and common salt at 0.5% levels of inclusion as main sources of vitamins and minerals. Cassava starch was used at 2% level of inclusion as a binding material. Table1. Experimental Diets using Alchornia cordifolia leaf meal (ACLM) Dietary levels of ACLM Ingredients 0% 5% 10% 15% 20% Maize Fish meal Soyabean meal ACLM Cassava starch Palm oil Bone meal Lysine Methionine Vit./min premix * Common salt The feedstuffs were finely ground and mixed up into a dough form in a plastic bowl using hot water. The mixture was then pelleted by passing through a mincer of 2mm die to produce 2mm diameter size of the pellets. The pellets were then sundried to about 10% moisture content, packed in polythene bags and kept safely dry for use. Two hundred and twenty-five post fingerlings of Heteroclarias collected from the African Regional Aquaculture Centre (ARAC) fish farm, Port Harcourt were stocked in an experimental tank for acclimatization. The fish were acclimatized for 7 days during which they were fed with the control diet containing 35% crude protein and of zero Alchornia cordifolia leaf meal twice daily, h and h. At the end of the acclimatization period, the 225 post fingerlings were completely randomized in 3 replicates of 15 post fingerlings per replicate for the 5 treatments TCN (Control), TA 1, TA 2, TA 3 and TA 4. The initial weight of fish in each aquarium was taken and recorded. Feeding commenced an hour after weighing exercise and the fish fed at 5% of their body weight twice daily, morning ( h) and evening ( h). The water in the aquaria was regularly monitored for the physico-chemical properties, and was renewed completely every other day within the experimental period that lasted 56 days of culture. Temperature was determined using mercury in glass thermometer calibrated from o c; immersed 5cm deep on the water surface. The ph and dissolved oxygen readings were taken using ph and oxygen meters respectively. Biweekly blood collection and sampling of the fish were carried out in line with Nlewadim and Alum (1999). The fish was anaesthesized in benzocain solution, using 0.4g dissolved in 1ml of 98% alcohol, and then added unto 1 litre of water. The fish was placed on its back in a trough, and blood collected from the posterior end of the abdomen, towards the tail, using a 2cm3 sterile plastic syringes and no 21 needle. The blood was emptied into EDTA (Ethylene Diamine Tetra Acetic Acid) treated bottle from Chemisciences Nig. Ltd. Owerri. Red blood cell and white blood cell counts were determined in line with Conroy and Herman (1970). Haemoglobin concentration and haematocrit (packed cell volume) estimates were determined with the procedure described by Wedemeyer and Yasutake (1977) and Blakhall and Daisley (1973) respectively. Mean corpuscular volume, mean corpuscular 12 International Journal of Research in Pharmacy and Biosciences V2 I6 July 2015

3 haemoglobin and mean corpuscular haemoglobin concentration, expressed in fento litres, Picogram and grams per 100ml respectively, were also calculated as reported by Anyanwu (2008). The proximate analysis of the test feedstuff and diets were carried out to determine the moisture content, ash, lipid, crude protein, crude fibre and nitrogen free extract, using the A.O.A.C (2000) methods. Experimental results were subjected to analysis of variance (ANOVA) as described by Steel and Torrie (1980). Test of significance was by Duncan multiple Range Test (DMRT) at 95% confidence level, using statistical package for social sciences (SPSS) for windows (version 15). RESULT The proximate composition for the Alchornia cordifolia leaf meal were 19.02%, 4.50%, 6.30%, 14.30%, 47.13% and 8.75% for crude protein, ash, lipids, crude fibre, nitrogen free extract and dry matter respectively. Water quality condition in the experimental aquaria of mean values ± C, 6.30 ± 0.08 and 4.72 ± 0.17mg/l for temperature, ph and dissolved oxygen respectively, showed little variations throughout the experimental duration (Table 2). Table2. Water parameters of the experimental aquaria Variable Parameters TCN(0%) TA 1 (5%) TA 2 (10%) TA 3 (15%) TA 4 (20%) Mean Temperature ( o C) ±0.01 Ph ±0.08 Dissolved Oxygen (mg/l) ±0.17 The dietary feeds chemical compositions are shown in table 3. The energy level of the diets decreased with increased levels of leaf meal. Table3. Percentage Proximate Composition of Experimental Diets Dietary levels of ACLM Nutrients 0% 5% 10% 15% 20% Crude protein (%) Crude fibre (%) Ether extract (%) Ash (%) ME (Kcal/kg) The haematological responses of Heteroclarias (Heterobranchus bidorsalis x Clarias gariepinus) fed varied levels of Alchornia cordifolia leaf meal are shown in Table 4. Table4. Haematological responses of Heteroclarias (Heterobranchus bidorsalis x Clarias gariepinus) fingerlings fed varied levels of A. cordifolia leaf meal. Variable parameters Alchornia cordofolia leaf meal SEM TCN(0%) TA 1 (5%) TA 2 (10%) TA 3 (15%) TA 4 (20%) haemoglobin (g/100ml) 5.9o a 5.80 ab 5.90 a 5.58 ab 5.53 b 0.16 Red blood cells (10 6 /mm 3 ) 3.00 a 2.44 ab 2.48 ab 2.23 bc 2.07 c 0.08 Packed cell volume(%) a ab ab b b 0.51 White blood cells(10 4 /mm 3 ) 4.75 a 4.58 b 4.50 b 4.10 c 4.58 b 0.07 MCV(mm 3 ) d c c b a 1.40 (MCH) (pg) d c bc b a 0.50 (MCHC) (%) NS NS NS NS NS 0.35 a, b, c, d Means within a row with different superscripts are significantly different (p< 0.05). RBC-red blood cell, WBC-white blood cell, HB-haemoglobin, PCV-packed cell volume, MCH-mean corpuscular volume, MCH-mean corpuscular haemoglobin MCHC- mean corpuscular haemoglobin concentration. The studies revealed significant differences among the treatments. The haemoglobin level of the fish on the control diet (5.9g/100ml) and TA 2 (5.9g/100ml) were similar, (p>0.05) to TA 1 (5.8g) and TA 3 (5.58g/100ml). There was however no significant difference (p>0.05) between TA 1, TA 3 and TA 4.The red blood cells for TCN (3.0 x 10 6 mm 3 ), TA 1 (2.44 x 10 6 /mm 3 ) and TA 2 (2.48 x 10 6 /mm 3 ) on the other hand were significantly different and higher (p<0.05) than TA 3 (2.23 x 10 6 /mm 3 ) and TA 4 (2.07 x International Journal of Research in Pharmacy and Biosciences V2 I6 July

4 10 6 /mm 3 ), while TA 3 and TA 4 were not different (p>0.05). The white blood cells of the fish TCN (4.75 x 10 4 /mm 3 ) were significantly (p<0.05) higher than TA 1 (4.5 x 10 4 /mm 3 ), TA 2 (4.5 x 10 4 /mm 3 ) and TA 4 (4.58 x 10 4 /mm 3 ), while TA 3 (4.1 x 10 4 /mm 3 ) was the least. Packed cell volume of the fish on TCN (23.58%) was not different (p>0.05) from TA 1 (23.0 %) and TA 2 (23.25%). This was however significantly (p<0.05) higher than TA 3 (22.25%) and TA 4 (22.25%), whereas there was no significant difference between TA 1 and TA 2 (p>0.05).the MCV value of mm 3 for TA 4 was significantly greater (p<0.05) than other TA treatments. This was followed in ranking sequence by MCV values of 99.77mm 3, 94.26mm 3, 93.75mm 3 and 79.2mm 3, for treatments TA 3, TA 1, TA 2 and TCN, respectively. TA 3 however was significantly (p<0.05) higher than TA 1 and TA 2, while these on the other hand were higher than TCN. The MCH value of 26.71pg for TA 4 was significantly higher (p<0.05) than the other TA treatments, while 20.82pg for TCN was the least. TA 3 (24.02pg) was significantly higher (p<0.05) than TA 1 (23.89pg) whereas TA 2 (24.85pg), TA 1 and TA 3 were similar (p>0.05). The MCHC values for A. cordifolia leaf meal ranged from 24.85% %, but the treatments, were not significantly different (p>0.05). DISCUSSION The heamatological responses of the experimental fish as summarized in Table 4 revealed significant differences in the various haematological indices of the treatment groups, (TCN, TA 1, TA 2, TA 3 and TA 4 ) except for the mean corpuscular haemoglobin concentration (MCHC) that showed no significant differences (p>0.05). The haemoglobin level of the fish on TCN (5.9g/100ml) and TA 2 (5.9g/100ml) were the highest, but however not different (P>0.05) from TA 1 (5.8g/100ml) and TA 3 (5.8g/100ml).These were significantly (P<0.05) higher than TA 4 (5.53g/l00ml) which was the least. Similarly, the red blood cells for TCN (3x10 6 /mm 3 ), TA 1 (2.44x10 6 /mm 3 ) and TA 2 (2.48x10 6 /mm 3 ) were significantly (p<0.05) higher than TA 3 (2.23x10 6 /mm 3 ), and TA 4 (2.07x10 6 /mm 3 ) which was the least. ln the same vein, the packed cell volume for TCN (23.58%), TA1 (23%) and TA 2 (23.25%) were similar, but TCN was significantly (P<0.05) higher than the value of 22.25% for TA 3 and TA 4.The white blood cells almost followed similar trend with TCN (4.75x10 4 /mm 3 ) being significantly higher (p<0.05) than the rest of the treatments, while TA 3 (4.1x10 4 mm 3 ) was the least. The trend for these observations were suggestive of retarding responses in the haemoglobin, red blood cells and packed cell volume profile of the experimental fish with increase in A. corilifolia leaf meal dietary inclusion levels. Higher inclusion levels of the leaf meal up to 15% - 20% seemed to have a deleterious effect on these haematological profiles of the experimental fish. ln their studies in relation to stocking and feeding conditions, Bhasker and Roa (1990) reported values of between g/100m1, x10 6 /mm 3 and % for haemoglobin, red blood cells and pack cell volume, respectively. Similarly, the values of x10 4 /mm 3 were also reported for the white blood cells. Blakhal and Daisley (1973) reported similar ranges, and these were in consonance with the ranges of g/100ml, x10 6 /mm 3, % and x10 4 mm 3 for haemoglobin, red blood cells, packed cell volume and white blood cells volumes respectively of the experimental fish fed varied dietary inclusion levels of Alchornia cordifolia leaf meal in this study. The mean corpuscular volume of fl for treatment TA 4 was significantly (P<0.05) the highest while TCN (79.2fl), the least. The same was applicable for the mean corpuscular haemoglobin with the value of 26.71pg for TA 4 and TCN (20.82pg) the least. The mean corpuscular haemoglobin concentration ranged from 24.85% % and were not significantly different (P<0.05). The trend of these erythrocyte indices was indication of the deleterious effect of higher levels of the leaf meal on the haematological profile and performance of the experimental fish. Nlewadin and Alum (1999) reported ranges of fl, pg and % for MVC, MCH and MCHC respectively in their studies on culturable fish species. On the other hand, Bhasker and Rao (1990) observed MCV, MCH, and MCHC ranges of fl, pg and %, respectively in their haematological studies in relation to stocking and feeding condition. These values seemed to be in consonance with the MCV, MCH and MCHC observed in this study. However, there seemed to be retarding responses in the haemoglobin, red blood cells and packed cell volume profile of the experimental fish with increase in Alchornia. corilifolia leaf meal dietary inclusion levels. This suggests that higher inclusion levels of the leaf meal up to 15% - 20% would have a deleterious effect on the haematological profiles of the experimental fish. 14 International Journal of Research in Pharmacy and Biosciences V2 I6 July 2015

5 REFERENCES [1] Anyanwu, D. C., Umesiobi, D. O. and Orji, B. O. (2003). Diurnal Climate pressures on haematology and blood Biochemistry of West African Dwarf sheep. The Nigerian Agric. Journal, 34: [2] Anyanwu, D. C. (2008). Effects of some tropical plants leaf meals on the performance of Heteroclarias (H. bidorsalis x C. gariepinus). PhD Thesis, Federal University of Technology, Owerri, Imo State, Nigeria, 203pp + xvii. [3] Anyanwu, D.C., Udedibie, A.B.I. and Osuigwe, D.I., Ogwo, V.O. (2011). Haematological responses of hybrid of Heterobranchus bidorsalis and Clarias gariepinus fed dietary levels of Carica papaya leaf meal. World Observations, 3(1): [4] A.O.A.C. (2000). Association of Official Analytical Chemists International (AOAC) Official methods of analysis, 17th edition. A.O.A.C International, Gaithersburg, Maryland. [5] Bhaskar, B. R. and Rao, S. K. (1990). Use of haematological parameters as diagnostic tools in determining the health of milkfish (Chanos chanos (forskall), in Brackish water culture. Aquaculture and fisheries mgt., 21: [6] Blakhall, P. C. and Daisley, K. W. (1973). Routime haematological methods for use with fish blood. Jorn. Fish boil, 5: [7] Conroy, D. A. and Herman, R. C. (1970). Textbook of fish diseases. T. E. H. Publication Inc., Jersey City, 302pp [8] Harikrishnan, R., Nisha, R. M. and Balasundaram, C. (2003). Hematological and Biochemical parameters in common carp, Cyprinus carpio, following herbal treatment for Aeromonus hydrophilia infection. Aquaculture, 221; [9] Hamre, G. I., Sagstd, A., Bakke Mekellep, A., Acierno, M., Maffia, M., Osiash, M., Ystad, A. and Krogelahi, A. (2007) Nutritional, Physiological and histological responses in Allantic Salmon, Salmo salar I. fed diets with genetically modified maize. Aquaculture Nutrition, 13: [10] Musa, S.O. and Omoregie, E. (1999). Haematological changes in mudfish, Clarias gariepinus (Burchel) exposed to malachite green. Journal of Aquatic Sciences, 14: [11] Nlewadim, A. A. and Alum, O. U. (1999). Haematological studies of some culturable fish species in Umuahia area of Abia State. Proceeding of the 4th Annual Conference of Animal Science Association of Nigeria, September 14 16, 1999, Ibadan, pp [12] Ochang, S. N., Fagbenro, A. O. and Adebayo, T.O. (2007). Growth performance, body composition, haematology and product quality of the African catfish (Clarias gariepinus) fed diets with palm oil. Pakistan Journal of Nutrition 6(5) : [13] Ochang, S. N., Fagbaro, A. O. and Adebayo, T. O. (2007) Influence of dietery palm oil on growth response, carcass composition, haematology and organoleptic properties of Juvenile nile tilapia, Oreochromis niloticus. Pakistan Journal of Nutrition 615: [14] Reyes, O.S. and Fermin, A.C. (2003). Terrestrial leaf meals or fresh water aquatic fern as potential feed ingredients for farmed abalone Haliotis asinine (Linnaeus 1758). Aquaculture Research, 34(8): [15] Steel, G. D. and Torrie, J. H. (1980). Principles and procedures of statistics, McGraw Hill Book Co. Inc. New York, 633pp. [16] Wedemeyer, G. A. and Yasutake, W. T. (1977).Clinical Methods for the assessment of environmental trees on fish health. Technical paper, 89 Us Department of interior, fish and wildlife services, Washington D. C.pp [17] Udedibie, AB.I. and Opara, C.C. (1998). Responses of growing broilers and laying hens to the dietary inclusion of leaf meal from Alchornia cordifolia. Animal Feed Science and Techn., 71: International Journal of Research in Pharmacy and Biosciences V2 I6 July