Egg Enumeration, lncubation, Hatching and Development of the "Miracle Fish", Oreochromis niloticus, in the Sudan

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1 Egg Enumeration, lncubation, Hatching and Development of the "Miracle Fish", Oreochromis niloticus, in the Sudan Afra A-A.Ahmed, Musab S. Abdalla and Prof. Thomas T. George School of Fisheries Sciences, Faculty of Agricultural Technology and Fisheries Sciences, Al Neelain University, Khartoum, Sudan, ( s: afr c om; pr aho o. c a) Abstract The "Miracle Fish", Oreochromis niloticus, is a mouth brooder that exhibits a high degree of parental care. Brooding is strictly maternal and breeding females undergo physiological change before spawning begins - a ventral bulging of the hyoid region to increase the mouth capacity in order to accommodate the eggs after being fertilizedby the male in a circular bowl-like nest built at the shallow part of the river bank or the pond. There are no records on the number of eggs accommodated in the female's mouth nor the egg development after hatching until the fry stage. In this paper, therefore, egg emrmeration, artificial incubation, hatching and development are recorded for the first time in the Sudan to promote sound hatchery operation and provision of quality fish seed. Introduction The "Miracle Fish", Oreochromis niloticus, is one of 77 tilapia species described by Thys in 1968 and belongs to the Family Cichlidae of the Tribe Tilapiine.(1,2) It is indigenous to Sudan among other tilapia species namely, Tilapia zillii and Sarotherodon galilaeus. (2,3) As a matter of fact, O. niloticus is believed to have originated exclusively from the African continent and evolved in the River Nile.(4,5) Also, it is assumed to have evolved from a marine ancestor and its penetration to freshwater is secondary.(5,6) However, now it is one of the world's most important fresh/marine, warm water cultured food fish, farmed from extensive to super-intensive water recirculating (> 100 kg/m3) and integrated hydroponic systems.(2,7) This is due to its hardiness, tolerance to varying degrees of physical and chemical environmental factors and other characteristics for aquaculture: better food conversion efficiency, faster growth rates on low protein diets, higher resistance to diseases and better palatability with superb flavor and firm, moist nutritious meat. Every 100 grams (3.5 oz) of raw meat contains 19.2 g protein, 2 g fat, 100 calories calcium and 400 g omega-3 fatty acids.(4,5) In 1982, Ethelwyn Trewavas (British Museum) had placed O. niloticus in the genus Oreochromis mainly because it is a microphagous mouth brooder tilapia species that exhibits a high degree of parental care and brooding is strictly maternal. Before spawning begins, breeding females undergo a physiological change - a ventral bulging of the hyoid region to increase the mouth capacity for the accommodation of the eggs after being fertilized by the male in a circular bowl-like nest built at the shallow part of the river bank or the pond.(2,8,9) The number of fertilized eggs brooded in the female's mouth increases with the fish body weight.(lo) While there are a number of scientific publications available on the different culture aspects of O. niloticus, there is no one publication in Sudan which could serve as a reference to the embryonic development of the fertilized eggs up to the post-yolk sac fry stage during natural brooding or in artificial incubators. An investigation, 60 AAC Spec. Publ. No. 12 (2007)

2 therefore, was undertaken for the first time in Sudan at Jebel Aulia Fish Farm on the White Nile for enumeration of fertilized eggs, artificial incubation, hatching and development of O. niloticus in order to promote sound hatchery operation and provision of quality fish seed. Objectives The objectives of the investigation are: o collect and enumerate the fertilized eggs incubated in the mouth of O. niloticus females of different sizes and weights; r hatch the collected fertilized eggs artificially in different types of incubators and find which type is more efficient under recorded environmental factors; r find if the clutch-removal method of fertilized eggs from the mouth of incubating females is better than the natural broodins or not and why? o follow up and record the embryonic development of the fertilized eggs up to the post-yolk sac fry stage;. record any revelant information for sound hatchery operation and provision of quality fish seed. Materials and Methods The investigation was conducted at Jebel Aulia Fish Farm (Golden Arrow Co.) which is 73 ha in area and located on the White Nile, west of Jebel Aulia Dam, about 40 km south of Khartoum, the capital of Sudan. Broodstock of 75 females and 25 males, ranging in total lengths and weights from cm and g respectively, were collected from the White Nile and growing ponds and used in the investigation. They were put before the breeding time in hapas, each measuring 8 x 1.5 x 2 m. These hapas were erected in half a hectare earthen pond above the water level and above the pond bottom by a quarter of a meter respectively. Applying the clutch-removal method, the fertilized eggs were collected directly from the mouth of incubating females. The number of fertilized eggs and length/weight of each female were recorded. Number of eggs produced per gram body weight was estimated by dividing the total number of eggs collected from each female spawn by the weight of the fish. The eggs were then transferred indoor for hatching in three types of incubators: porcelain Zotg jars, round plastic containers and perspex Zotg jars. Each incubator was stocked with up to 2011 eggs per liter and shielded from sunlight to avoid any damage to the eggs by ultra-violet radiation. A steady current of plankton-free water was allowed to pass through each incubator after being mixed with oxygen as soon as it was out of the bio-filter. Temperature, dissolved oxygen and ph were monitored daily at 10 am, 1 pm and 5 pm. Ammonia, total hardness, Chlorine, salinity, iron and copper were also monitored biweekly at 8 am. Results The investigation achieved its goals. The clutchremoval method of fertilized eggs from the brooding females, hatching and rearing them artificially in incubators, provided the necessary quantity at the required time. Tables I and 2 show the average data record for the enumerated fertilized eggs with respect to the females' sizes/weights (cmlkg) and the water analysis with respect to the physical and chemical parameters respectively. The results and observations are summarized hereinafter: r the eggs are pear-shaped, very yolky, yellowish brown with average weight of g; o the data in Table 1 indicate that the number of fertilized eggs incubated in the mouth of each female increased as the total length and weight of the fish increased; it was also observed that the number in several females was less within the same parameters of length and weight; o the number of eggs per gram body weight increased in the smaller females, i.e. 5 eggs per gram in the female that weighed 50 grams as compared to 2 and 1 in those that weighed 150 and 250 grams respectively; AAC Spec. Publ. No. 12 (2007) 61

3 Aquacultur e CanadaoM the data in Table 2 indicate that the eggs when put in three types of incubators needed a constant flow of plankton-free water mixed with dissolved oxygen in a concentration of 6.3 ppm, an appropriate temperature of 25 C, a ph of 8.7 and incubation in a well shaded place to avoid ultra-violet rays especially during early cell cleavage ofthe eggs; hatching of fertilized eggs in the three artificial incubators was highest in the Perspex Zoug jars followed by the porcelain Zottg jars and the round plastic containers; after the eggs were transferred in the incubators, they started to develop through a sequence of events which can be briefly defined into three phases from the start of incubation till after the yolk sac was completely absorbed (Plate l): phase 1: from fertilization until the appearance of the eyes ('eyed eggs'); Table 2. Average Data Record for Water Analysis. Parameter Plate l. Sequence of the Egg Embryonic Development of O. niloticus During the First 15 Days (Afra). i.i Average data Temperature ( C) 25.1 Dissolved Oxvsen (onm) 6.3 ph 8.7 Total Hardness (ppm) Ammonia (ppm) 2.0 Chlorine (onm) 0.0 Salinitv (oom) 0.0 Copper (ppm) 0.0 Iron (nom) 0.0 phase 2: from the appearance of the eyes until hatching (yolk sac fry); phase 3:from hatching until the absorption of the yolk sac (post-yolk sac fry). Table 1. Average Data Record of Enumerated Fertilized Eggs of O. niloticus with Respect to the Female Parent Size (cm) and Weight (kg). Total length (cm) Weight (ke) Number of fertilized eggs Actual-Pergbody weisht - 4.6: 5 :2-2.0: 2 : t L s s : 1 lta* Day 1: yellow brown and yolky fertilized egg; Day 2: small dark spots collected on the egg surface; Day 3: eyes and hair-like tail appeared; Day 4: head appeared; Day 5: head and tail more developed, the yolk sac decreased and the embryo started slight movement; Day 8: yolk sac fry started to swim out of the incubators with the water flow; 62 AAC Spec. Publ. No. 12 (2007)

4 7 Aquacultur e CanadaoM 2006 I k ffi & s F h -*J Day 12: skeleton and fins more developed and yolk sac decreased in size; Day 13: fry transferred from water trough to small frberglass tanks with slow water flow and left until day l5; Day 15: post-yolk sac stage whereby yolk sac was fully absorbed and f.y started to feed on supplementary food at the rate of 10% while average body weight reached grams. Discussion Development of any aquaculture species requires research into several areas such as broodstock management, egg incubation, larval rearing, juvenile-on growing and marketing. The present investigation dealt only with egg enumeration and incubation. It supports an earlier finding that the number of eggs per clutch increases with the length and weight of the spawner (10, 1l) However, it was observed that within the same parameters of weight and length, the number of eggs could be less. This may be attributed to the fact that the broodstock used in the investigation was a mix of females from the White Nile and the growing ponds. It is a known fact that in ponds, tilapias mature much sooner (earlier age) than under nafural conditions and produce smaller and more numerous eggs for a given body weight because of the unfavorable pond conditions, a homeostatic device in response to the environment. ( I 0, I 1) The number of eggs per gram body weight increased as the weight of the brooding female decreased. Therefore, it is advisable to use females of smaller size (50 g) for breeding. O. niloticus spawns easily and frequently but one of the major constraints to large-scale commercial farming is the scarcity of quality seed in large numbers required at any time. That is why, the clutch-removal method was used in the present investigation. Also, because previous studies ascertained that it is superior to natural mouth brooding which produces poor seed due to cannibalism of eggs and fry by the adults and besides, it increases the spawning frequency of the females and consequently, the overall production of fry or seed.(l1) It is very important to shield the eggs from direct sunlight especially during early development to avoid damage by ultra-violet radiation. A previous study recommended that hatcheries should be equipped with subdued lighting. (tz) f'urttrerrnore, the concentration of dissolved oxygen in the plankton-free water for artificial egg incubation was found to be extremely important and more effective than the 'churning' action of the female during matemal incubation. The temperature maintained during the egg incubation period served the purpose. However, there is need for more studies on this aspect because it was ascertained in other species that small temperature changes can cause considerable variation in the development and physiology of the developing fish. The water flow into the incubators in liters per minute was not carried out in the investigation. This is another important factor because it is reported to remove harmful metabolites (CO2, NH3) excreted by the developing eggs and prevent the lipids in the heavy yolk of the eggs from settling and disrupting development.(10, 11) Finally, the embryonic (brooding) period begins with the fertilization of the egg and terminates with the transition from endogenous to exogenous feeding i.e. up to the post-yolk sac fry. In O. niloticus, egg development is a fast process whereby the main distinguishable stages are swelling, germ development and embryonic development stages. These stages are clearly illustrated in Plate 1 and demonstrate for the first time in Sudan, a detailed sequence of development in the fertilized egg of O. niloticus. However, it is interesting to know that only Morrison et.al.(l3,) of Dalhousie University, Faculty of Medicine, Halifax, N.S, Canada published a detailed histological study of the development of the embryo and early larva of Orechromis niloticus. AAC Spec. Publ. No. 12 (2007) 63

5 Recommendations The following recommendations are suggested: 1) Initiate finfish reproduction and broodstock development program in the Faculty of Agricultural Technology and Fisheries Sciences, Al Neelain University, Sudan. 2) Develop a genetic-based broodstock management approach for O. niloticus to optimize egg and fry quality through artificial diets, selective breeding, photoperiod and temperature manipulation. 3) Hatchery operators are advised to breed smaller spawners, follow the clutch-removal method of eggs from the incubating O. niloticzs females and hatcurear them artificially in efficient incubators to increase the spawning frequency of the females and avoid the problem of cannibalism of eggs and fry. a) Study the selection effects of O. niloticus females in ponds for spawning frequency, number and diameter size of eggs per clutch as compared to natural brooding females. 5) Study the influence of egg size on the progeny with respect to natural and pond broodstock. 6) Study the effects of water flow (liters/min), dissolved oxygen concentration, temperature, ph and photoperiod on the incubation /hatching time and embryonic development of the fertilized eggs of O. niloticus. Acknowledgments We are very much indebted to the conference organizers for accepting this paper to be presented at Aquaculture Canada OM 2006 and to Prof. Hassan A. Alsaouri, Vice-Chancellor, Al Neelain University, Khartoum, Sudan for approving financial support to participate in the conference. Sincere thanks are also extended to the manager of Jebel Aulia Farm for providing pond and hatchery facilities and also to the technicians and fishermen for their cooperation and assistance. References 1. Thys van den Audenaerde DFE An annotated bibliography of Tilapia (Pisces: Cichlidae) Mus. A. Afr. Cent. Doc. Zool. No. 14,406 p. 2. George TT The most recent nomenclature of Tilapia species in Canada and the Sudan. Bull. Aquacult. Assoc. Canada, Sp. Pab. No l0: Sandon H An illustrated guide to the freshwater fishes of the Sudan. Sudan Notes and Records 31: George TT Tilapia - a potential culture species in Canada. Bull. Aquacult. Assoc. Canada 96-3: Pullin RSV, Lowe-McConnell RIl.l982. The Biology and Culture of Tilapias. ICLARM, Manila, Philippines, 432 p. 6" George TT.I997. Saltwater culture of tilapias and possible commercial application in Canada. Bull. Aquacult. Assoc. Canada 97-2: Fitzsimmons K The most important aquaculture species of the 21" Century, pp.3-8, In, Tilapia in the 21" Century, (K. Fitzsimmons, ed, Proc. Fifth Int. Symp. on Tilapia Aquaculture, Yol. 1, Rio de Janeiro, BrazII. 8. Trewavas E Tilapias: Taxonomy and Speciation pp. 3-13, In, The Biology and Culture of Tilapias (RSV Pullin, RH Lowe- McConnel, eds), ICLARM Conf. Proc. 7, 432 p., Manila, Philippines. 9" Trewavas E Tilapiine Fishes of the Genera Sarotherodon, Oreochromis and Danakalia. British Museum (National History) London. 563 p. 10. Fryer G, Iles TD The Cichlid fishes of the Great Lakes of Africa - Their Biologt and Evolution. Oliver and Boyd. Edinburgh, 64I p. t 1" Balarin JD, Hatton n Tilapia. A Guide to their Geology and Culture in Africa. Unit of Pathobiology. University of Sterling. Scotland. r74 p. 12. Shepeard CJ, Bomage NR Intensive Fish Farming. Blackwell Scientific Publication, 436p. 13. Morrison CM, Miyake T, Wright Jr. JR Histological study of the development of the embryo and early larva of Orechromis niloticus (Pisces: Cichlidae). Journal of Morphology 247: AAC Spec. Publ. No. 12 (2001)