Nutrition and reproductive performance of African catfish fed bitter kola (Garcinia kolal)

Fish serves as a major source of protein for human, providing signifi cant portion of nutrient to a large proportion of people particularly in the developing world [1]. The global production of fi sh and other aquatic animals for human use occurs either by commercial fi shing or through aqua cultural and farming techniques. According to FAO [2], the world production of fi sh in 2005 consists of 93.2 million tons captured by commercial fi shing in wild plus 48.1million tonnes produced by fi sh farms. Despite the outrageous increase in numbers of people yearly with the production rate of fi sh, Nigeria, like most third world countries, is not able to meet her animal protein requirement which is traceable to our fi sh production which has fallen below expectation. Many fi sh hatcheries in Nigeria are functional at low capacity; producing only a total some of 30million fi ngerlings per year, although the total existing capacity could easily be 1 billion fi ngerlings per year. Based on 1992 United Nations Development Project (UNDP) assisted base line study in the total annual fi ngerlings requirement for Nigerian was 250,000 million while the domestic production stood at 7.2million [3]. In Nigeria, the most common fi sh reared are tilapia and catfi sh because they are mostly found in fresh water habitat. Therefore, in fi sh reproduction under controlled conditions, attempt are made to obtain sperm from the fi sh with high quality seeds although, several factors affect fi sh seed quality such as different strains, genetics, nutrition, content of feed and deposition of organic matter, chemical fertilizer into water used for cultured medium and for hatchery purposes [4]. According to [5], there are some common hatchery practices such as handling, cleaning, use of chemicals and water quality problems which do have negative effect on fertilization success in artifi cial reproduction thereby producing low quality fi sh seeds Therefore, the need to research into various ways of enhancing fi sh fertility to meet the growing demand. Abstract


Introduction
Fish serves as a major source of protein for human, providing signifi cant portion of nutrient to a large proportion of people particularly in the developing world [1]. The global production of fi sh and other aquatic animals for human use occurs either by commercial fi shing or through aqua cultural and farming techniques. According to FAO [2], the world production of fi sh in 2005 consists of 93.2 million tons captured by commercial fi shing in wild plus 48.1million tonnes produced by fi sh farms. Despite the outrageous increase in numbers of people yearly with the production rate of fi sh, Nigeria, like most third world countries, is not able to meet her animal protein requirement which is traceable to our fi sh production which has fallen below expectation. Many fi sh hatcheries in Nigeria are functional at low capacity; producing only a total some of 30million fi ngerlings per year, although the total existing capacity could easily be 1 billion fi ngerlings per year. Based on 1992 United Nations Development Project (UNDP) assisted The use of medicinal plants as fertility enhancer has now gained much ground in aquaculture. Plants such as bitterleaf, kigelia [6], Garcinia kola [7], have been used to enhance fertility and it is generally accepted that medicine derive from plant products are safer than their synthetic counterparts [8]. Bitter kola (Garcinia kolal) belonging to the family of Clusiaceae highly esteemed by the native of Africa as Negroes chewed and it is a widespread tree of evergreen forest valued in Nigeria for its medical nuts [9]. Negroes chewed it as a powerful aphrodisiac and as masticatory; they administer them in treating common colds, cough and throat infections. G. kola stem bark has been shown to contain a complex mixture of phenolic compounds such as tannin guttiferin, biofl avonoids, xanthenes, benzophenone, kolafl avanone and garciniafl avanone [10], all of which have antimicrobial activity. Besides, G. kola exhibits purgative, anti-parasitic, anti-infl ammatory, anti-bacterial and anti-viral properties [11]. Therefore, considering the importance of G. kola, this study is carried out to investigate the effect of Garcinia kola on the growth performance haematology and sperm quality of African catfi sh.

Experimental site
The experiment was carried out at the Fishery unit of Teaching and Research farm Ladoke Akintola University of Technology (LAUTECH), Ogbomoso, Oyo State, Nigeria.

Experimental fi sh and management
A total number of ninety (90) healthy African catfi sh were procured from a reputable farm at Ibadan, Oyo State, Nigeria. The fi sh were acclimatized for two (2) weeks in tanks containing aerated water and they were fed fl oating feeds to empty their gut so as to maintain a uniform stomach condition in preparation for the experiment. At the end of the acclimatization period, a total number of sixty (60) male fi sh were randomly distributed into twelve plastic tanks at stocking density of 5 fi sh per tank and replicated two times. The fi sh were fed twice daily, both in the morning and evening (9:00 hours and 17:00hours), weighed every two week and the daily feeding rate (5% of the total biomass) were adjusted.

Collection and processing of test ingredient
Bitter kola seeds were procured from a local market in Ogbomoso, Oyo State, Nigeria. The outer coats of the bitter kola were removed and the seeds were sundried, milled to a fi ne powder and stored in an air tight nylon prior use.

Experimental diets
The feed ingredients were obtained from a reputable feed mill in Ogbomoso, Oyo State, Nigeria. The ingredients include; Fish meal, maize, soya bean meal, groundnut cake, wheat offal, bone meal, oyster shell, vegetable oil, lysine, methionine and salt.
Six isonitrogenous diets containing 40% CP were formulated and Garcinia kola meal was included at varying levels (50, 100, 150, 200, 250g/kg) in the diets and they were represented as follows:, D2, D3, D4, D5, D6 respectively. The control basal diet (D1) contained 0% bitter kola Table 1.  at the end of the feeding trial from the caudal peduncle of both the test and control fi sh with a sharp surgical blade, the blood samples were dispensed into tubes containing ethylene diamine tetra acetate (EDTA) and empty bottles to determine the following blood parameters.

Red Blood Cells (RBC):
The blood of the fi sh was diluted in an improved newbauer pipette with formal citrate fl uid at 1:200. The diluted blood was introduced into a newbauer counting chamber and the red blood cells counted under microscope [12].

White Blood Cells (WBC):
For the white blood cell count, blood was diluted at 1:20 with diluting fl uid. The resulting mixture was introduce into newbauer counting chamber and counted under the microscope [12].

Packed Cell Volume (PCV):
To determine the packed cell volume, a haemotocrit tube was three quarters fi lled with blood and the ends sealed with critaseal, the tube was then centrifuged in a microhaematocrit for 5minutes at 2000g. The PCV was read by a microhaematocrit reader and expressed as the volume of the erythrocytes per 100cm 3 [12].

Mean Corpuscular Volume (MCV):
The mean volume of each red blood cell was estimated using the following formular 10 Haemoglobin MCH Erythrocyte count   Haemoglobin (Hb): The cyamethaemoglobin method was used. 0.02cm 3 of blood was placed on 4cm 3 of drabkins reagent in a test tube and mixed. After 30minutes, the optical density was read calorimetrically at 540μm. values of haemoglobin were determined by comparing with cyamethaemoglobin standards [13].

Sperm quality determination:
Male fi sh were randomly selected from all the treatments for milt collection. The male fi sh were sacrifi ced and the testes were removed, then the sperm were examine and snapped and viewed under the computer aided sperm analysis microscopic system.
Milt count: Concentration of sperm was determined by counting the numbers of spermatozoa in sample dilute with distilled water (100×) in a Burker haemocytometer under 400×magnifi cation [14].
Percentage motility: Each sample was estimated using large microscope at 400×magnifi cation, immediately after addition of 20ul distilled water as an activating solution.

Results
The proximate composition of Bitter Kola Seed Meal (BKSM) is presented in the Table 2. Bitter kola seed meal had a crude protein of 2.45%, crude fi ber 6.50%, ether extract 2.50%, ash 6.10% and dry matter 93.46%.

Discussion
The use of bitter kola in the diet of African cat fi sh in this study has revealed its ability to enhance sperm quality as well as improving growth better than the control diet with no inclusion level of bitter kola seed meal. Differential growth among the control diet and other diets with various inclusion levels of bitter kola seed meal(BKSM) as observed in the study is defi nitely not due to protein since isonitrogenous diet was used for the study however, variation in this study is strongly linked to the presence of biofl avonoid in Garcinia kola which stimulates growth in fi sh as previously reported by Braide [16]. Kocour, et al., [17], revealed that biofl avonoid is plant    feed intake in ruminants while low consumption seem not to have any affect [18].
A retarded growth effect was reported by Akpantah, et al., [19], on rat treated with Garcinia kola seed extract for six weeks.
However, the results observed shows that there was signifi cant differences (p>0.05) in the mean weight of fi sh fed with  [20].
Haematological parameters are routinely used for the evaluation of physiological environment and husbandry stressors in fi shes [21]. In recent year good management practices have been advocated as effective's ways of reducing stress in fi sh culture [22]. Many researchers have proved that change in blood characteristics of African catfi sh is due to stress, exposure to the environmental pollutant or by pathogen [22][23][24]. Also, haematological component of blood are valuable in the monitoring of feed toxicity especially with feed constituents that affects the formation of blood in culture fi sheries [25]. Packed cell volume (PCV) range of 27.00 to 37.00% observed in this study is within range of 20 to 50% reported by Piestse, et al., [26]. And rarely do values above 50% being reported [27,28]. Reduction in the concentration of the PCV in the blood usually suggests the presence of toxic factor example of which is haemagglutin which has adverse effect on blood formation [25]. White blood cells (WBC) and lymphocytes result recorded in this study showed a decrease as the level of bitter kola seed meal increases in the diet. The highest value 17.10×10 3 /μ for WBC was recorded in the control diets as well as that of lymphocyte 65.25%. WBC and LYM are the defense cells of the body. Douglas and Jane [29], demonstrated that the amount has implication in immune response and the ability to fi ght infection. High WBC is usually associated with microbial infection or the circulating system [25]. The value recorded in this study (11.75-17.10) is also within the range recommended value of (16.13×10 3 to 16.39×10 3 mm) as reported by Sotolu and Faturoti [30]. and correlates strongly with fertilization process [33,34].
Moreover, reproductive capacity is the most conclusive way of testing sperm motility [5].
In this study, the sperm count, sperm motility, sperm morphology and live percentage were higher in treatment  [7], that rats fed Garcinia kola seed extract exhibited increased libido (sexual instinct) for the male rats justifying the use of Garcinia kola by native as an aphrodisiac, but did not improve pregnancy rates in female rats as a measure of the male fertility index.
The fi ndings of Adeparusi, et al., [6], also agrees that male Clarias gariepinus fed Kigelia africana had signifi cantly higher sperm count than the control.

Conclusion
In conclusion, this study showed that:  Garcinia kola at all inclusion levels was insignifi cant on the haematological parameters of Clarias gariepinus meaning that the fi sh can tolerate higher inclusion of Garcinia kola without any adverse effect.
 Inclusion of Bitter kola seed meal at the rate of 150g/ kg (T4) in the diet of African catfi sh enhances sperm

Recommendation
It is therefore suggested that: • Bitter kola (Garcinia kola) seed meal at 150g/kg inclusion level can be used by fi sh farmers to enhance sperm quality and thereby increasing the production rate of African catfi sh (Clarias gariepinus).
• The use of Bitter kola (Garcinia kola) seed meal for improving sperm fertility is not exorbitant in price (with respect to the quantity required by the farmers) therefore it is of high economic value because it reduces the cost of production compare to hormonal drugs that are very expensive.