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Feeding Acipenser persicus larvae by microenriched D. magna

In another study, the D. magna were used to feed Acipenser persicus larvae. D. magna was used as a vector to carry yeast of Saccharomyces cerevisiae to digestive tract of Acipenser persicus larvae and were bioencapsulated in different levels of yeast suspensions (8, 8.30, 8.48 and 8.60 Log cells/liter) for 10 h. Sturgeon larvae were fed based on 30% of their body weight with enriched D. magna six times a day. Experimental period was 30 days. The results indicated that the microenriched D. magna by S. cerevisiae significantly increased final body weight (FBW), final body length (FBL), and specific growth rate (SGR) in the experimental fish in comparison with the control (Jafaryan, et al., 2007). Results of this study also showed that different concentrations of

probiotic yeast in bioencapsulation broth of D. magna could have different effects on growth performance and feeding efficiency of Acipenser persicus larvae.

Using enriched D. magna by probiotic bacillus in Persian sturgeon larviculture

Reared D. magna were employed for early feeding of Persian sturgeon larvae. In this study the D. magna were enriched in 3 levels (2x 107, 4x

7 7

107, 6x 107 CFU/ Liter) of Bacillus circulans and Bacillus licheniformis, and used as a vector to carry these probiotic Bacillus to digestive tract of Persian sturgeon larvae. The probiotic bacilli were successfully incorporated into the D. magna. The highest level of was 5.6x104 CFU/ D. magna after 24 hours inoculation in the bacterial broth. The Persian sturgeon larvae were fed on with bioencapsulated D. magna with three 3 levels of Bacillus for 4 weeks. In termination of experiment, results showed that the carcass proximate compositions of the larvae were significantly increased. Enriched D. magna had significant positive effects on relative food intake, nitrogen retained efficiency, protein gain and energy retained (Faramarzii et al., 2011). The results of the challenge tests with stress of alkalinity (pH=12), acidity (pH=2) and thermal stress (33°C), indicated that the best healthy rate and survival time were obtained in probiotic treatments in comparison with control.

Using bioencapsulated D. magna with the baker's yeast (A-MAX) extract as probiotic for Acipenser persicus larvae

D. magna was used as a vector for the delivery of the baker's yeast (A-MAX) extract into the digestive tract of Acipenser persicus larvae. The fish treated with the Saccharomyces cerevisiae extract and this probiotic showed positive response in the values of their growth parameters such as weight gain (WG), food conversion efficiency (FCR), conversion efficiency ratio (CER %), total net gain (TNG) and specific growth (SG). A significant positive correlation observed between the concentration of

probiotics in bacterial broth suspension of the D. magna and the growth parameters. (Lashkarblooki, et al., 2009a). This resulted in a higher cultural performance, with the best results in the trial in which the fish larvae were fed on bioencapsulated Artemia nauplii with 3x108 yeast and Bacillus pre liter. The superb performance of A. urmiana nauplii to carry the baker's yeast and probiotic Bacillus proved its high potential as a vector.

Feeding Rainbow trout (Oncorhynchus mykiss) larvae with bioencapsulated

D. magna In another research it was been elucidated that D. magna had plentiful capability in grazing the profitable bacteria and had a high rate of inoculation with them (Jafaryan et al., 2008). The Rainbow trout larvae indicated high capability in using bioencapsulated D. magna in larviculture. Significant positive results for the growth parameters, consisting of conversion efficiency ratio (CER), condition factor (CF) and daily growth (DG) were obtained in the fish fed with probiotics via bioencapsulation of D. magna.

Discussion and Conclusion

There is not enough information regarding its performance in the enrichment of D. magna with bacteria and yeasts. Using enriched D. magna by useful microorganisms helps in the optimization and development of gut microbial flora of the host animal and let to increased growth parameters and survival rate and decreased mortality, and finally, decreases the cost of aquaculture production. This can be a major step in the development of the industry. D. magna had a notable grazing rate, carrying efficiency and very high potential for carrying the beneficial microorganisms into the digestive tracts of cultivable fish larvae. They also had a remarkable tolerance against the manipulation of bacterial flora.

Faramarzi, M., Jafaryan,H., Farahi, A., Lashkar Boloki, M. and Iranshahi,F. 2011. The effects on growth and survival of probiotic Bacillus spp. fed to Persian sturgeon (Acipencer persicus) larvae. Aquaculture, Aquarium, Conservation & Legislation. 4:10-14.

Gatesoupe, F. J. 1991. Bacillus sp. Spores: A new tool against early bacterial infection in turbot larvae, Scophthalmus maximus ln: larvens, p., Jaspers, E., Roelands, I. (Eds), Larvi -fish and crustacean larviculture symposium. European Aquaculture Society, Gent, pp. 409-411, Special publication no. 24.

Keskin, M., and H. Rosenthal. 1994. Pathways of bacterial contamination during egg incubation and larval rearing of turbot, Scophthalmus maximus. Appl. Ichthyol.

10: 1-9.

Jafaryan, H., Makhtomy, N., Ahmadi, M. and Mahdavi, M. 2007. Evaluation of the effects of yeast (Saccharomyces cerevisiae) as a probiotic on the growth, feeding parameters and survival rate of Acipenser persicus larvae which was fed on by bioencapsulated Daphnia magna. Aquaculture Europe. 24-27 october,2007.

Istabul, turkey. P: 260-261.

Jafaryan, H., Morovat, R. and Shirzad, H. 2008. The use of bioencapsulated Daphnia magna by probiotic bacillus and their effect on the growth of Rainbow trout (Oncorhynchus mykiss) larvae. Iranian Journal of Biology. 21: 24-35.

Lashkar boloki, M., Jafaryan, H., Faramarzi, M. and Adineh, H. 2011. The effects of Amax yeast fed to Persian sturgeon (Acipencer persicus) larvae via bioenrichment of Daphnia magna. Aquaculture, Aquarium, Conservation & Legislation. 4:361-367.

The replacement of Daphnia magna flour in feeding of Rainbow trout (Oncorhynchus mykiss) larvae by using

blend of probiotic

Jafaryan, Hojatollah1*, Jafaryan, Samira 2 and Shahi, Godratollah1

1 University of Gonbad kavoos, Golestan, Iran

2 Gorgan University of Agricultural Sciences and Natural Resources, Faculty of fishery

3 Center of domestic animal research of Gorgan

*Author for correspondence e-mail: Hojat.Jafaryan@gmail.com


The application of probiotics in aquaculture was increasinig rapidly (Gatesoupe, 1999) and some papers were associated with the effect of probiotics in fish (Sharma and Bhukhar, 2000). The use of probiotic bacillus in many research indicated that the growth performance and feeding efficiency were promoted. Many probiotic bacteria have been proposed improve growth performance and helth in rainbow trout. Daphnia magna is one of the most important animals which attention by fish nutritionistis around the wourld. The use of Daphnia flour as a possible feed substitute to reduce the cost of fish feed is employed in this research. Appropriate of probiotic applications were shown to improve intestinal microbial balance, thus leading to improved food absorption (Fuller, 1989). This study was desingned to evaluate the replacement of Daphnia magna flour in feeding of rainbow trout (Oncorhynchus mykiss) larvae using blend of probiotic bacillus.

Materials and methods

Five diets were formulated to be isoenergitic (4500 cal/g), isolipid (19.99% Crude lipid) and isoprotein (48.84% Crude protein), but contain different levels of concentration of probiotics (B. licheniformisi, B. subtilis, B. polymyxa, B. laterosporus and B. circulans). Four concentrations of bacillus (6, 6.30, 6.48 and 6.60 Log cells per g in these diets were used respectively. These experimental diets were prepared using the 500 g of

starting feed of Biomar and 500 g of Daphnia magna flour respectively. The homogenous mixtures of different diets were spread over aluminium foil and dried at 40°C in a hot air oven. The dried mixture was powdered and sieved to obtain micropellets. This experiment was conducted in a completely randomized design with five treatments (trial 1-3 and control) with three replicates for treatment and control. The trout fry were fed on the base of the 10 percent of their body weight for 6 times per day for a period of 30 days. The control treatment was fed on unbacterial supplemented diet. On the first time of experiment, 10 fish from each tank randomly selected and anaesthetized and the total length, body weight of fish were mrasured. The feeding trial was conducted by 15 circular fiberglass tanks. In the termination of experiment (4 weeks), all fish were sampled and measured of their total length, body weight. The growth performance was calculated by using of mathematical model


The maximum body weight was obtained in treatment of diet F2 (6.30 Log cells per g). The values of specific growth rate (SGR) in trial 2 (diet F2) treated with the probiotic bacillus, were significantly higher than the control treatment (p<0.05). The highest protein efficiency ratio (PER) and Lipid efficiency ratio (LER) were obtained in experimental treatment of F2. These values were 0.90 and 3.22 respectively. The positive significant correlation was obtained between the concentration of bacillus in experimental diets and body weight gain (r=0.515, p<0.05).

Tablel. Growth parameters in raibow trout fry in different treatments Parameter


FBW (g)






2.23 ±0.04b

3.41 ±0.49b

1.74 ±0.29a




2.32 ±0.08ab

3.53 ±0.1ab

1.68 ±0.56ab




2.44 ±0.17a

3.68 ±0.27a

1.59 ±0.11b




2.35 ±0.09ab

3.57 ±0.13ab

1.65 ±0.07ab




2.36 ±0.06ab

3.58 ±0.1ab




No significant difference in growth parameters between trials F1, F3 and F4 were observed (p>0.05). The probiotic bacillus decreased the food conversion ratio (FCR) of rainbow trout larvae in treatment of diet F2 in comparison with the control treatment (p<0.05).


The nutritive value of Daphnia magna flour as an alternate protein source has been demonstrated in the present study. The different levels (Log cells per g) of probiotics indicated different growth performance in raibow trout larvae.

In the probiotics supplementation diets results in growth performance and feed utilization better than of the control basal diets. The best performance of fish in terms of growth performance and feed utilization efficiency was recorded at the bacterial supplementation of 6.30 Log cells per g in diet. Similar results were observed by Ghosh et al. (2003) and Bairagii et al. (2004) in Indian carp. Suggesting that the addition of probiotics reduced the culture cost of fishes in cultivation systems (Yanbo and Zirong, 2006). The resuts indicated the suitable concentration of probiotic bacillus in this experiment, for bacterial supplementation of diet, was 6.30 Log cells per g. In similarity with these results, Ghosh et al. (2003) indicated that Bacillus circulans was employed in diets of rohu, had the best growth performance in use of 2x105 cells per 100 g of feed. The using of Bacillus subtilis and Bacillus circulans in formulated diets for rohu fingerling, increased the PER, LER and decreased the FCR in experimental treatments (Bairagi et al., 2004).

This study also showed that the Daphnia magna flour as an alternate protein, lipid and energy source can use in formulate diet and different levels of probiotic bacillus could cause the different effects on growth parameters in rainbow trout larvae.

Bairagi, A., Ghosh, K.S., Sen, SK. and A.K. Ray. 2004. Evaluation of the nutritive value of Leucaena leucocephala leaf meal , inoculated with fish intestinal bacteria Bacillus subtilis and Bacillus circulans in formulated diets for rohu, Labeo rohita (Hamilton) fingerlings. Aquaculture Research. 35: 436 - 446.

Fuller, R. 1989. Probiotics in man and animals .J. Appl. Bacteriol. 66 :365-378 .

Gatesoupe , F.J. 1999. Review: The use of probiotics in aquaculture. Aquaculture. 180:

147- 165.

Ghosh, k., Sen, S.K. and A. K. Ray. 2002. Characterization of Bacillus Isolated from the gut of Rohu, Labeo rohita, fingerlings and its significance in digestion . Appl. Aquaculture. 12:33-42.

Ghosh, k., Sen , S.k., and A. k. Ray. 2003. Supplementation of an isolated fish gut bacterium , bacillus circulans , in Formulated diets for Rohu, Labeo rohita, Fingerlings. Aquacu lture- Bamidgeh. 55(1): 13-21.

Sharma, O.P. and S.K.S. Bhukhar. 2000. Effect of Aquazyn-TM-1000, a probiotic on the water quality and growth of Cyprinus carpio var. communis (L.). Indian J.

Fish. 47:209-213.

Yanbo, W. and X.Zirong .2006. Effect of probiotic for commom carp (Cyprinus carpio) based on growth performance and digestive enzymes activities. Animal feed science and technology. 127:283-292.

Effects of replacing dietary fish meal with different protein sources on growth performance of rainbow trout (Oncorhynchus mykiss) fry

Reza Jalilia*, Naser Aghb, Reza Asgarib, Forouzan Bagherzadeh Lakania

aDepartment of Fisheries, Faculty of Natural Resources, Urmia University, Iran bDepartment of Fisheries, Artemia and Aquatic Animals Research Institute, Urmia University, Iran

* Corresponding author. E-mail address: Re.Jalili @ gmail.com


An experiment was performed to examine the effect of replacing fish meal with different protein sources on growth performance in rainbow trout fry. Five experimental diets containing different protein sources were tested: 1), 100% fish meal (FM), 2), 50% fish meal + 50% plant protein (50FM/50PP), 3) 100% plant protein (PP), 4) 50% poultry by-product meal protein + 50% plant protein (50PP/50PM) and 5) 100% poultry by-product meal protein (PM). Rainbow trout with a mean initial weight of 0.1±0.005 g were fed experimental diets for 60 days. Results showed that 50% fish meal replacement with plant protein (50FM/50PP) and 100% with combination of plant proteins and Poultry by-product meal (50PM/50PP) did not negatively affect the growth indices. However fish fed 100% plant protein (PP) and 100% poultry by­product meal (PM) resulted in decreased final body weight and SGR. Hepatosomatic index (HIS) was significantly higher in fish fed diet with PP, 50PP/50PM and PM groups compared to control group (P < 0.05). No significant differences were observed in HIS among 50FM/50PP and control group (P > 0.05). Viscerosomatic index (VSI) was significantly higher in fish fed diet with plant protein sources (PP) compared to control group (P < 0.05). However no significant differences were observed in VSI among other feeding treatments (P > 0.05).

Key words: Protein source, Growth, Rainbow trout, Fry.

Feed cost is the major expense in fish culture. One of the challenges is to develop less wasteful and more economic diets. Fish meal has long been the major protein source in feeds for trout, salmon, and marine fish. In order to reduce feed costs and improve sustainability of culture of these fishes, fish meal is increasingly being replaced by more economical protein sources. The production of successful fish feed formula which rely less on fish meal, requires accurate information on the nutritive value of more economical protein sources. The increasing demand of ingredients for aquaculture feeds all over the world has driven an important research effort towards the nutritive evaluation of other protein sources.

Many plant protein sources can be used to partially or almost totally replace dietary fish meal (Kaushik et al. 1995; Robaina et al. 1995), provided that the essential amino acid requirements of the fish species are met, the palatability of the diet is improved and the levels of anti-nutritional factors (ANFs) are reduced (Francis et al. 2001). Another alternative ingredient to fish meal is poultry by-product meal (PBM). PBM is made of ground and clean parts of the carcass of slaughtered poultry.

Previous works have shown good potential of the combination of PBM, FM and blood meal (BM) (Fowler, 1991) and PBM and FM (Steffens, 1994) in diet of various fish species. Fowler (1991) reported PBM could replace about 50% of fish meal in the diets for chinook salmon and rainbow trout. Higss et al. (1979) found that defatted PBM and PBM mixed with hydrolysed feather meal could replace up to 33% and 75 % of fish meal, respectively, in coho salmon diets. About 50 % of fish meal was successfully replaced with PBM in chinook salmon and rainbow trout (Steffens, 1994). Moreover, PBM has been tested at varying success so far in sea bream (Nengas et al., 1999), European eel (Appelbaum et al., 1996), channel catfish (Sadiku and Jauncey, 1995), common carp (Hasan and Das, 1993) and sunshine bass (Webster et al., 2000). Since there are few researches regarding the effects of fish meal

replacement with PMB and combination of different protein sources on growth performance in rainbow trout larvae, the present study was performed to study the probable effects of replacement of fish meal with different protein sources on growth performance rainbow trout fry.

Materials and methods

Fish husbandry and diet preparation

The rainbow trout (Oncorhynchus mykiss) fry used in this experiment came from a batch of eyed (embryonic) eggs from a commercial fish farm of domestic origin. Fry were reared in our laboratory hatchery from eggs to the beginning of the experiment, according to Rollin et al. (2003). Hundred fish with average weight of 100±5 mg were stocked in 15 tank (8 L) supplied with freshwater at a flow rate of 1.5 L min-1. Light/dark cycle was 12 L:12 D. The pH (7.3-7.7), temperature (14-15°C) and dissolved oxygen level (7.5-7.8 mgL-1) of each tank were monitored daily

Five experimental diets with similar protein, lipid and energy content were formulated to contain different protein sources to replace fish meal (Table 1). The control diet contained only the Kilka meal (Clupeonella sp.) as the primary sources of protein (FM). The experimental diets contained: (1), 100% fish meal (FM) (2), 50% fish meal + 50% plant protein (50FM/50PP) (3) 100% plant protein (PP), (4) 50% poultry by­product meal protein + 50% plant protein (50PP/50PM), (5) 100% poultry by-product meal protein (PM). The experimental plant protein sources included wheat gluten, corn gluten and soybean meal.

Briefly, all dry ingredients were thoroughly mixed in a mixer. Oil was added and thoroughly mixed for 5 min and then moistened by adding cold distilled water until stiff dough yielded. The strands were dried at 50°C for 8 h using an oven, manually crumbled into appropriate size and sieved. Pellets were stored at 4°C during the experiment. Fish were fed three times per day at 7-9 % body weight for 8 weeks.

Specific growth rate (SGR, %d-1)=100x[(lnWf-lnWi)xT-1]; Condition factor (CF)=100x(WxTL-3); days reared. Where: Wf and Wi are the final and initial body weights (g), T—time of rearing (days), FB and IB are the final and initial absolute weights (g).

Collection of samples

At the start of the experiment and at the end of the growth period, 12 fish/tank were sampled for measurements (weight and total length). Nine fish per tank were removed to weigh the liver and digestive tract for measurement of hepatosomatic index (HSI) and viscerosomatic index (VSI).

Proximate composition of diets

Feeds was determined by drying in oven (Iran khodsaz Co, Iran) at 105°C for 25 h to a constant weight; ash was determined by incineration in a muffle furnace (Iran khodsaz Co, Iran) at 600 C for 6 h; crude protein was determined by the Kjeldahl method (N x 6.25) using an automatic Kjeldahl system (Behrotest WD 40, Germany); Crude lipid content determination was conducted by ether extraction. Gross energy content of the diets was calculated on the basis of 5.64, 9.43, and 4.11 (kcal/g diet) of protein, fat, and carbohydrate, respectively (NRC 1993).

Statistical analysis

The results were analysed using analysis of variance, one way ANOVA, for which the homogeneity of variances and the normal distribution were tested according to the Levene and Shapiro_Wilk tests, and comparison among the means was made using Duncan's multiple range test (DMRT) (Sokal & Rohlf 1969). All statistical analyses were conducted using SPSS (version16) and tested at P < 0.05.

Table 1 Ingredient and proximate composition of experimental diets

Ingredients (g kg-1 diet)

Dietary treatment 1






Kilka fish meal






Wheat gluten






Corn gluten






Soybean meal






Poultry by-product






Blood meal






Kilka fish oil






Wheat meal






Wheat starch


















Vitamin premix1






Mineral premix2


















Di-calcium phosphate






Calcium carbonate






Proximate composition (% dry matter)







Crude protein






Crude lipid






Crude starch






Gross energy (kcal/g)3






Digestible energy (kcal/g)4






1Vitamin mixture: (mg or IU/kg of diet) Vitamin A (as acetate) 1600000 IU; vitamin D3, 400000 IU; choline chloride.12000; niacin, 4000; riboflavin, 8000; pyridoxine, 4000; folic acid, 2000; vitamin B12, 8000; biotin, 1; inositol, 20000; vitamin C, 60000; vitamin H2, 2.4; vitamin B2, 8000; vitamin K3, 2000; vitamin E,40000.

2Mineral mixture (g/kg): zinc, 12.5 g; iron, 26 g; manganese, 15.8 g; copper, 4.2 g; cobalt, 0.48 g; selenium, 2 g; iodine, 1 g.

3Calculated on the basis of 5.64, 9.43, and 4.11 (kcal/g diet) for protein, fat, and carbohydrate, respectively (NRC 1993).

4Calculated using apparent coefficients of digestibility of 0.9, 0.85, and 0.8 for crude protein, crude fat, and carbohydrates (NFE), respectively.

Growth indices in different treatments are shown in table 2. No significant differences were detected in growth of fish fed control diet (FM) and those fed on diet containing 50% fish meal protein + 50% plant protein (50FM/50 PP) and 50% poultry meal protein + 50% plant protein

(50FM/50PP). However fish fed 100% plant protein (PP) and 100% poultry meal protein (PM) resulted in decreased Final body weight and SGR.

Table 2 Growth indices of rainbow trout fed experimental diets for 60 days (n=9


Dietary treatment 1

Performance parameters -

FM        50FM/50PP        PP        50PP/50PM PM

Initial body weight (g)






Final body weight (g)

































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