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(Marine Science)/Vol.1/No.2/December 2010/10/1-10 Effects of Artificial Diets on Biological
Performances of Acanthopagrus latus
Broodstock in the Persian Gulf
Zakeri, Mohammad1*; Savari, Ahmad2; Kochanian, Preeta1; Haghi, Mahsa3 1- Fisheries Department, Faculty of Marine Natural Resources, Marine Science and Technology University, Khoramshahr 2- Marine Biology Department, Faculty of Marine Science, Marine Science and Technology University, Khoramshahr 3- South Iranian Aquaculture Research Center, Ahwaz 2010 Journal of the Persian Gulf All rights reserved. Abstract
Effects of artificial diets on growth, spawning performance, egg and larval biochemical compositions and
quality of Acanthopagrus latus in the Persian Gulf were studied. Nine diets representing a combination of
three protein levels (40, 50 and 60%) and three energy levels (22.5, 23.5 and 24.5 MJ GE/Kg) were tested
in triplicates. Each tank was stocked with 10 fish with a sex ratio of 1:1 and with an average weight of
415g and 236g for females and males, respectively. Fish were fed to satiation, twice daily. The growth of
Acanthopagrus latus broodstock was not significantly affected (P>0.05) by dietary protein and energy
levels. Biometry, relative fecundity and survival rate and crude protein and lipid of eggs, hatchling and 3
day post-hatch (3DPH) larvae were significantly affected (P<0.05), but spawning performance and body
composition of broodstock, with the exception of body crude protein and lipid were not significantly
affected by dietary protein and energy levels. This study revealed the best spawning performances of A.
latus
broodstock were achieved at 40% dietary protein and 23.5 MJ GE/Kg dietary energy.
Keywords: Artificial diets, Growth, Spawning, Egg quality, Acanthopagrus latus, Persian Gulf. 1. Introduction
al., 2003; Platell et al., 2007). Furthermore, diets of Sparid species often differ markedly with respect to The Sparide is a predominantly marine family location, reflecting the opportunistic nature of the frequented in the Indian, Pacific and Atlantic oceans feeding behavior of the members of this family (Nelson, 1994; Platell et al., 2007) and contains (Sarre et al., 2000; Mariani et al., 2002; Tancioni et many species of commercial and/or recreational importance and some that are used for aquaculture Exogenous nutrition of broodfish provides the (Ingram et al., 2002; Platell et al., 2007). Sparids essential nutrients required for gonadal development typically consume a wide range of benthic prey and of females and the performance of produced seed occasionally substantial amount of plant material (Gunasekera et al., 1997; El-Sayed et al., 2008). Thus, (Sarre et al., 2000; Mariani et al., 2002; Tancioni et egg yolk is considered the major source of nutrition for embryonic development in fish. Therefore, Zakeri, M. et. al. / Effects of Artificial Diets on Biological Performances of Acanthopagrus latus… inadequate food supply for fish broodstock will lead The fishes were maintained in concrete tanks (75m3) to poor spawning performance and seed production prior to the commencement of trial. Fourteen days (Gunasekera et al., 1997; El-Sayed et al., 2008). before commencing the trial, 270 fish were weighed Proteins and energy, the main components of egg yolk, are considered to play pivotal role in reproduction (Afzal Khan et al., 2005). Accordingly, The experiment consisted of nine treatments with the relationship between dietary protein and energy three replicates each. Each tank was stocked with 10 levels in fish feeds should be considered for optimum fish with a sex ratio of 1:1 and an average weight of fish performance. At inadequate energy levels, dietary 415±39.4g and 236±24g for females and males, protein may be used as an energy source (Garling & respectively. Fish were fed to satiation, twice daily, Wilson, 1976; Cho & Kaushik, 1985; El-Sayed et al., at 09:00 and 17:00 h for 129 days prior to first 2008). At adequate energy levels, dietary protein can spawning (from middle of October to late of be spared for anabolic functions (El-Sayed et al., February). Broodstock were weighed in each tank at 1987, 2008; Satpathy et al., 2003). Therefore, the the end of the spawning period to determine weight proper balance between dietary protein and energy is essential for optimum use of fish feeds so that the Water temperature, salinity, dissolved oxygen and dietary protein can be spared by dietary energy effects pH were recorded daily. Salinity ranged between (Garling & Wilson, 1976; Satpathy et al., 2003; El- 39‰ and 44‰ (41 ± 0.1) and temperature fluctuated between 14 to 22oC (15.7 ± 0.2) during the The most study on Sparids remain confined to experimental period. Average values for dissolved fry, fingerling and young fish (Vergara et al., 1996; oxygen and pH were 7.5 mgl-1 and 7.9, respectively. Santhinha et al., 1999; Gomez-Requeni et al., 2003; Experimental diets- Nine semi-purified diets Venou et al., 2003; Teshima et al., 2004; Sá et al., representing a combination of three protein levels 2006; Benedito-palos et al., 2007) and the effects of (40, 50 and 60%) and three energy levels (22.5, 23.5 the interaction between dietary protein and energy and 24.5 MJ/Kg) were tested. Biochemical contents on spawning performance of yellowfin sea bream of diets were determined according to AOAC (1990). has not been determined. This study was undertaken Formulation and biochemical composition of the test to investigate the effects of dietary protein and energy levels on growth, spawning performance, Spawning, egg and larval collection- Each tank egg and larval biochemical composition and quality was checked daily for eggs. Spawning started at late of February (water temperature>20oC). Spawning took place at night (between 02:00 to 05:00) and the 2. Material and Methods
eggs were collected the next morning from the tanks and transferred to beaker (1L). Total number of eggs Experimental trial - Nearly 7 months prior to was estimated volumetrically by 54 samples of 1 ml natural spawning season (late February- early March) (6 samples of 3 levels of beaker in triplicates) to wild broodstock yellowfin sea bream, A. latus, were assess relative fecundity. Twenty seven samples of 1 caught with hook from the northwest of the Persian ml (3 samples of 3 levels of beaker in triplicates) Gulf during August to September (2008) and were investigated microscopically in order to sort out transferred immediately to the Mariculture Research fertilized and unfertilized eggs and to calculate Station of South Iranian Aquaculture Research fertilization ratio. All eggs were left to remain in Center, Mahshahr, Iran in a aerated fiberglass tank. beaker for 20 minutes to completely separate sunken Journal of the Persian Gulf (Marine Science)/Vol.1/No.2/December 2010/10/1-10 and floating eggs and to record respective broodfish (two females and two males) from each proportions. After eggs hatching, 50 five ml samples tank were individually weighed, gutted and frozen were collected randomly to observe the hatched ones. for biochemical analyses. Biochemical composition To determine survival rate of larvae, 50 five ml of feed ingredients, experimental diets, broodfish samples of were collected randomly at 3DPH, as whole body, eggs and larvae were analyzed using well. Fifty eggs, hatchlings and 3DPH larvae were standard methods (AOAC, 1990). Each analysis was selected randomly from each tank and days of spawning period and were preserved in 10% Statistical analysis- Analysis of variance formalin solution for biometry using a compound (ANOVA) (One-way and two-way, 3¯3 factorial) microscope equipped with ocular micrometer. The and descriptive statistics were used to determine remaining eggs were labeled and frozen immediately differences among protein, energy levels and treatments, using SPSS 11.5 statistical software. Biochemical analyses- Initial body composition of and when differences were found a Tukey’s post fish was analyzed from 20 samples of fish (10 hoc test was conducted to determine specific females and 10 males) frozen prior to the differences in protein and energy levels and commencement of trial. After spawning, four Table 1- Formulation and biochemical composition of the test diets (Dry-weight basis) 8.39 7.56 7.11 8.14 7.36 6.52 7.73 7.29 6.42 11.43 9.8 8.62 8.82 6.76 5.58 10.48 3.61 2.43 23.39 20.71 18.02 17.21 12.87 12.61 4.59 6.5 5.28 19.62 21.23 22.59 20.62 22.70 23.91 21.62 24.10 25.34 a Per kilogram: vit. A, 600,000 IU; vit. D3, 400,000 IU; vit. E, 40,000 mg; vit. K3, 1,000 mg; vit. B1 (Thiamin mononitrate), 3,000 mg; vit. B2 (Ribofelavin), 5,000 mg; vit. B6 (Pyridoxine hydrochloride), 3,000 mg; vit. B12 (Cyano cobalamine), 8,000 mg; vit. C, 52,000 mg; Nicotinic acid, 30,000 mg; D-calcium pantothenate, 9,000 mg; Folic acid, 1,600 mg; D-biotin, 160 mg; Inositol, 24,000 mg; Antioxidant, 5,000 mg. b Per kilogram: Manganese, 2,600 mg; Copper, 6,000 mg; Ferrous, 4,000 mg; Zinc, 6,000 mg; Selenium, 500 mg; Iodine, 2,000 mg; Cobalt, 500 mg; Choline chloride, 120,000 mg. c Value are means of three replicate samples per diet. d Nitrogen-free extract (calculated by difference). e Gross energy, calculated based on 0.17, 0.398 and 0.237 MJ/g for carbohydrate, lipid and protein, respectively. 3. Results
table 2. Growth of A. latus broodstock was not affected but, biometry of eggs, hatchlings and 3DPH Growth- Results on growth and biometry of eggs larvae were affected significantly (P<0.05) with and hatchlings larvae of A. latus fed with different dietary protein and energy levels are summarized in Spawning performance - Results indicated that Zakeri, M. et. al. / Effects of Artificial Diets on Biological Performances of Acanthopagrus latus… spawning performance criteria of A. latus broodstock, body crude protein and lipid, body composition of except fecundity and survival rate in 3DPH larvae broodstock was not significantly affected (P<0.05) (%), were not significantly affected (P<0.05) by by dietary protein and energy levels (Table 4). dietary protein and energy levels (Table 3). Biochemical composition of eggs, hatchlings and Biochemical composition- With the exception of 3DPH larvae are summarized n in Table 5.
Table 2- Growth and biometry of eggs, hatchlings and 3DPH 1 larvae of Acanthopagrus latus fed different dietary protein and energy levels 181.9±2.7a 175.6±1.6ab 171.1±1.7bc 178±1.3a Mean ± SE values with different superscript in each row are significantly (P<0.05) different. 1 Days post-hatching Table 3- Spawning performance of Acanthopagrus latus broodstock fed different dietary protein and energy levels P40E22.5 P40E23.5 P40E24.5 P50E22.5 P50E23.5 P50E24.5 P60E22.5 P60E23.5 P60E24.5 94.9±4.0a 93.9±4.4a 60.0±6.3bc 21.1±6.4d 91.6±5.2a 50.1±6.4c 58.1±6.3c Mean ± SE values with different superscript in each row are significantly (P<0.05) different. 1 Days post-hatching Table 4- Biochemical composition of whole body of Acanthopagrus latus broodstock fed different dietary protein and energy levels P40E22.5 P40E23.5 P40E24.5 P50E22.5 P50E23.5 P50E24.5 P60E22.5 P60E23.5 P60E24.5 Initial Moisture 65.3±1.9 67.0±0.6 63.6±2.1 66.0±1.0 66.6±0.4 64.9±1.1 64.1±2.7 63.9±1.8 65.7±0.6 63.7±0.8 Crude 59.9±0.6ab 59.5±0.8ab 58.1±0.9b 62.7±0.8a 58.6±1.0ab 57.1±1.4b 60.1±1.2ab 58.9±1.0ab 57.4±1.2b 55.8±0.8b 19.6±0.9bc 20.6±0.9bc 27.0±1.3a 18.2±0.7c 21.0±1.0bc 21.2±1.1bc 18.5±1.0c 23.9±0.9ab 22.9±0.7ab 21.4±1.1bc 1.9±0.2 1.0±0.1 1.0±0.1 1.6±0.1 2.0±0.1 2.0±0.2 2.0±0.2 1.3±0.1 1.8±0.2 1.9±0.2 13.2±1.1 15.1±1.0 13.4±2.1 13.0±0.7 13.3±1.1 14.0±1.2 13.3±1.0 12.3±0.8 12.5±0.6 10.5±0.8 1 Dry-weight basis; mean ± SE values with different superscript in each row are significantly (P<0.05) different. Journal of the Persian Gulf (Marine Science)/Vol.1/No.2/December 2010/10/1-10 Table 5- Biochemical composition of eggs, hatchlings and 3DPH1 larvae of Acanthopagrus latus fed different dietary protein and energy levels (Dry-weight basis) P40E22.5 P40E23.5 P40E24.5 P50E22.5 P50E23.5 P50E24.5 P60E22.5 P60E23.5 P60E24.5 Hatchling 54.0±0.6a 45.5±0.4bc 51.1±0.1ab Hatchling 7.5±0.1f 14.8±0.3a 12.9±0.2b Mean ± SE values with different superscript in each row are significantly (P<0.05) different. 4. Discussion
each level. Similarly, results of effects of dietary protein and energy on growth gain of broodstock Growth and anabolism of fish are supported by have been reported in Dicentrarchus labrax (Ceredà energy obtained from the catabolism of dietary et al., 1994), Oreochromis niloticus (El-Sayed et al., protein and energy sources. Similarly, dietary lipids 2003), Labeo rohita (Afzal Khan et al., 2005) and also influence influence the dietary protein Ictalurus punctatus (Sink and Lochmann, 2008). requirement of fish profoundly. Many studies have These differences could be attributed to the species- been conducted to study the potential sparing of specific growth rates. A. latus has been reported to dietary protein by increasing the concentration of have a low growth rate (Hesp et al., 2004) and this non-protein energy. These studies have indicated that probably might be another reason for the low growth at inadequate energy level, dietary protein may be used as an energy sources, whereas at high protein Egg diameter is considered an important criterion level, a proportion of this protein will be deaminated for the assessment of spawning performance in fish. and carbon skeleton used as an energy source (El- The effect of dietary protein and energy levels on Sayed et al., 2008). At adequate energy level, dietary egg diameter of A. latus was significant. Similar protein can be spared for anabolic functions (Morais finding have also been reported in Cyprinus carpio et al., 2001; El-Sayed et al., 2008). Therefore, the (Manissery et al., 2001) and O. niloticus (El-Sayed et proper balance between dietary protein and energy al., 2003). Afzal Khan et al., (2005) found that must be considered if cost-effective diets are dietary protein level did not affect the diameter of spawned eggs of L. rohita and Sink and Lochmann Growth in A. latus broodfish is not affected by (2008) reported that egg diameters were not different dietary protein and energy levels because most of for eggs from I. punctatus broodfish fed different feeding energy is used for reproductive activities instead of somatic growth in broodfish (Bone et al., The present results indicated that dietary protein 1995). There is inconsistency in the reported results significantly affected length of hatchlings and 3DPH about the effect of dietary protein and energy on larvae of A. latus. Hatchling length was increased growth gain of broodfish. El-Sayed et al. (2008) with decreasing dietary protein from 60 to 40% found that the growth of Oreochromis niloticus regardless of energy levels. Also, the length of 3DPH broodstock was significantly affected (P<0.05) by larvae was significantly affected by energy levels, dietary protein and energy levels, and weight but effects of dietary protein and energy showed increased with increasing dietary protein from 30 to irregular trends. Similar results have been reported 40% and energy level from 16.7 and 18.8 MJ/Kg at by El-Sayed et al., (2003, 2008) where growth of Zakeri, M. et. al. / Effects of Artificial Diets on Biological Performances of Acanthopagrus latus… Nile tilapia fry was found to increase with increasing Although, Afzal Khan et al. (2005) expressed that dietary protein content of broodstock diets from 30 low dietary protein levels (250 and 300 g Kg-1) to 40%. Also Duray et al., (1994) showed that produced higher relative fecundity in L. rohita. increasing lipid levels from 12% to 18% in The percentage of buoyant eggs on the water broodstock rabbit fish produced large newly hatched surface has been broadly used to evaluate egg quality larvae. However, Sink and Lochmann (2008) in marine fish (Izquierdo et al., 2001). However, in reported that there was no difference in weight or A. latus buoyant eggs (%), despite being at their length of I. punctatus fry at 14 day from broodstock maximum number in fish fed with high (60%) fed with different lipid diets. Results of OGD of eggs dietary protein level and intermediate (23.5 MJ/Kg) and hatchlings were noticeable and the effect of energy level diet, were not affected by dietary dietary protein at 50 and 40% levels were highly protein and energy levels. Ceredà et al. (1994) reported that eggs of D. labrax fish fed with low Fecundity, another important parameter to assess (34%) protein diet had significantly lower buoyancy spawning performance of fish, is known to be than those fed with high (51%) protein diet. affected by nutritional deficiencies in broodstock diet Nutrition influences the fertilizability and (Izquierdo et al., 2001). Diet containing high (60%) hatchability (%) of fish eggs; Results showed these dietary protein level and intermediate (23.5 MJ/Kg) criteria, however, were not significantly affected by energy level produced higher relative fecundity in A. dietary protein and energy levels. Although, latus. Also, relative fecundity was significantly hatchability (%) was at its maximum in fish fed with affected by energy levels. However, the increase in P40E23.5 diet with noticeable values. Similar relative fecundity was comparable among increasing findings have also been reported by Afzal Khan et al. dietary protein levels, presumably dietary protein (2005) in L. rohita. Irregardless, El-Sayed et al. levels used were above the minimum requirement, (2008) reported that fertilized eggs produced from O. and any increase in dietary protein did not further niloticus broodstock fed with 30 and 35% protein affect the relative fecundity. These results appear to diet displayed lower hatchability than the eggs of be in agreement with the results of Sink and fish fed with a 40% protein diet. However, Lochmann (2008) who reported spawning success in increasing energy level at each dietary protein level punctatus increased as dietary energy did not significantly affect these parameters. concentration increased. Fernàndez-Palacios et al., Conversely, Sink and Lochmann (2008) described (1995) stated that in gilthead sea bream, the total that eggs from I. punctatus broodfish fed with the number of eggs produced / Kg of female was more 10% lipid diet showed greater hatching success than sensitive to the dietary essential fatty acid (EFA) eggs from broodfish fed with 4% lipid diet. Ceredà et levels than the other parameters. Similar observation al. (1994) reported that the eggs of D. labrax fish fed was made in Paralichthys olivaceus (Furuita et al., with 51% protein diet had significantly higher 2000). However, El-Sayed et al. (2008) described at hatchability than those fed with 34% protein diet low (30%) and intermediate (35%) dietary protein fish. In addition, increased egg quality, fecundity, levels, increasing dietary energy levels resulted in a fertilization success and egg hatching success in fish significant decrease in spawning performance of O. have been linked to diets enriched with highly niloticus, probably because the P/E ratio of these unsaturated fatty acids (Izquierdo et al., 2001). diets was not appropriate for optimum performance. In C. carpio maximum fecundity was at high dietary significantly affected by dietary protein and energy protein levels (350 g Kg-1) (Manissery et al., 2001). levels. Furuita et al. (2000) stated that survival at Journal of the Persian Gulf (Marine Science)/Vol.1/No.2/December 2010/10/1-10 3PDH was positively correlated with dietary lipids Afzal Khan et al. (2005) stated that the fat content in P. olivaceus, and Fernandez-Palacios et al. in eggs of L. rohita did not fluctuate with variations (1995) found a significant decrease in the survival in dietary protein. However, egg protein was of gilthead sea bream larvae at 3DPH when increased with increasing dietary protein levels. broodstock were fed with 21% lipid diet. Similar Ceredà et al. (1994) showed that the egg proximate trends have been reported by Duray et al., 1994 for composition of D. labrax was the same regardless rabbit fish and Sink and Lochmann ( 2008) for I. of dietary treatment. Similar observations were made in O. niloticus (El-Sayed et al., 2003), Body crude lipid was significantly affected by Plectorhynchus cinctus (Li et al., 2005) and I. energy and protein levels. Body lipid increased with punctatus (Sink and Lochmann, 2008). increasing energy. Interestingly body crude protein In the present study, at 40 and 60% dietary protein was found to be significantly affected by energy levels, increasing energy levels resulted in levels. This might be because all diets contained significantly increased crude protein of hatchlings efficient dietary protein levels. Similarly, increasing and 3DPH larvae. Also, crude lipid of hatchlings and protein in high dietary protein levels (300, 350 and 3DPH larvae significantly increased by increasing 400 g Kg-1) in L. rohita did not significantly affect dietary protein levels at 23.5 MJ/Kg energy level. muscle protein of females (Afzal Khan et al., 2005). However, Li et al. (2005) showed the levels of total On the contrary, El-Sayed et al. (2008) stated that lipid in larvae of P. cinctus was independent of body protein of O. niloticus was significantly dietary treatment and El-Sayed et al. (2003) reported increased with increasing dietary protein levels. that the chemical composition in fry of O. niloticus There are however, relatively few studies on the was not significantly affected by dietary protein influence of dietary protein and lipid on body It is concluded the best reproductive performance The chemical composition of eggs is often of A. latus broodstock was achieved at 40% dietary examined to evaluate egg quality, as the egg must protein and 23.5MJ/Kg energy. This finding is of satisfy nutritional needs for embryonic and larval practical implication and in light of scarce development (Furuita et al., 2002). This study knowledge on Acanthopagrus nutrition, signifying showed that protein in eggs was significantly the necessity for more research under different increased with decreasing dietary protein levels. However, increasing energy levels at each dietary protein level did not significantly affect this 5. Acknowledgment
significantly affected by dietary protein and energy Authors are thankful to Director, South Iranian levels. Similarly, El-Sayed et al. (2008) reported Aquaculture Research Center, Ahwaz, Iran for that only egg crude protein was significantly financial grant No. 4-74-12-87042, and Director and affected by protein and energy content of O. Staff at Mariculture Research Station, Mahshahr, niloticus broodstock diets and indicated at all Iran for providing necessary facilities for the energy levels, eggs crude protein significantly experiment and the University of Marine Science and increased with increasing dietary protein levels. Technology, Khoramshahr, Iran, for its support These differences might be related to the difference during the tenure of this project. We would like to in species, diet composition, dietary protein and express our thanks to Mr. Udaya Sankar Sethi for energy levels, P/GE ratio and experimental design. Zakeri, M. et. al. / Effects of Artificial Diets on Biological Performances of Acanthopagrus latus… References
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2 Runsonrun M Robaldo 5 SANADOR snabbt till ledningen där han omedelbart blev 3 Her Birthday L Santos Resultat 21/10 uppvaktad. Blev rejält utmanad över upploppet och såg 4 Hurry Harry J Gustafsson dessutom slagen ut en bit men kämpade väl och vann med 5 My Pal Al M Rodriguez minsta möjliga marginal. 6 MARGAUX bra iväg och direkt molnigt, 14 5 Montgomery J J

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Page 1 of 2 Methyl Methacrylate & Butyl Methacrylate Kit Background: Blending methyl methacrylate (MMA) and butyl methacrylate (BMA) will yield a clear and firm cast. The plastic can be removed from the sections before staining with xylene or acetone treatment. Polymerization can be obtained with the addition of benzoyl peroxide at 60°C or with the addition of benzoin methyl ether,

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