Introduction

Synthetic androgens are used in fish culture as sex controlling agents and as growth promoters if energy is shut away from developing ovaries towards growth of somatic tissues (Rizkalla et al., 2004). However, the recent consumer demand for farmed fish has increasingly stressed quality and safety, and the absence of pollutants, antibiotics and carcinogens. Thus, along with growth performance, the fish rearing strategy needs to focus on food hygiene. Since the European Union ratified a ban in 2006 for the use of all sub-therapeutic antibiotics (Regulation 1831/2003/EC), scientists have intensified efforts to identify and develop safe dietary supplements and additives that enhance the life activity, health and immune system of farmed fish (Ji et al. 2007; Shim et al. 2009). Plants are natural sources of safer and cheaper chemicals. Plant products have been reported to promote various activities like antistress, growth promotion, appetite stimulation and immuno­stimulation in aquaculture practices (Citarasu et al., 2001, 2002; Sivaram et al., 2004).

Garlic (Allium sativum) has been proven effective as a hypolipidemic (Sumiyoshi 1997), antimicrobial (Kumar and Berwal, 1998), antihypertensive (Suetsuna, 1998), hepatoprotective, and insecticidal (Wang et al., 1998) agent in various human and animal therapies. Garlic was studied in different forms of extracts: aqueous, ethanol and dried powder (Shin and Kim, 2004). In aquaculture operations, garlic promotes growth, enhances immunity, stimulates appetite, and strengthens the control of bacterial and fungal pathogens. These effects of garlic are due to the presence of various organosulphur compounds, including allicin (Augusti and Mathew 1974). Many reports (Fo et al., 1990; Hu 1999; Aly et al., 2008; Aly and Mohamed 2010) have documented the effect of allicin as a growth promoter.

Garlic (A. sativum) contains at least 33 sulfur compounds, several enzymes and the minerals germanium, calcium, copper, iron, potassium, magnesium, selenium and zinc; vitamins A, B1 and C, fiber and water. It also contains 17 amino acids: lysine, histidine, arginine, aspartic acid, threonine, swine, glutamine, proline, glycine, alanine, cysteine, valine, methionine, isoleucine, leucine, tryptophan and phenylalanine (Josling 2005). One of the most biologically active compounds in garlic is allicin (diallyl thiosulfinate or diallyl disulfide). The most abundant sulfur compound in garlic is alliin (S-allylcysteine sulfoxide), which is present at 10 and 30 mg g-1 in fresh and dry garlic, respectively (Lawson 1998). Although allicin is considered the major antioxidant and scavenging compound, recent studies show that other compounds may play stronger roles; such as polar compounds of phenolic and steroidal origin, which offer various pharmacological properties without odor and are also heat stable (Lanzotti 2006; Maniat et al., 2014).

Garlic inclusion in fish feeds has also been reported to increase growth performance in fish (Metwally, 2009). According to Sheela and Augusti (1992) and Diab et al. (2002) garlic has the ability of enhancing catalase activity in serum and lowering the levels of plasma glucose in fish. Dietary garlic as a growth promoter in Nile tilapia (Oreochromis niloticus) improved body weight gain, feed intake and feed efficiency (Diab et al., 2002; Shalaby et al., 2006). However, dietary garlic as a growth promoter coupled with the optimum graded level needed for the growth, survival and nutrient utilization of red belly tilapia Tilapia zillii is yet to be well tested. On the other hand, early sexual maturity of this species is a well-recognized problem. The problem of precocious sexual maturity and unwanted reproduction has long been accepted as a major constraint to further development and expansion of Tilapia culture (Ajiboye et al., 2015).There is a paucity of information on the optimum dosage and duration regime of garlic as growth promoters in red belly tilapia T. zillii. Thus, the necessity for a laboratory experiment to optimize the optimum graded level for the growth, nutrient utilization and survival of T. zillii reared in a glass aquaria tanks for 28 days.

Methodology

This study was conducted under laboratory conditions at Aquaculture Department, Nigerian Institute for Oceanography and Marine Research to optimize the optimum graded level for the growth, nutrient utilization and survival of T. zillii reared in a glass aquaria tanks for 28 days. The experimental fish were acclimated under wet laboratory conditions for two weeks and fed the basal diet. One hundred and eighty Tilapia zillii fry with average body weight of 0.03 g were randomly stocked in each aquarium with average water holding capacity of 0.01m³ and at a stocking density of 15 fish / glass aquarium, where three aquaria were referred to every dietary treatment. Aquaria were daily cleaned, and the water exchange rate per day, including fish feces and remaining food, was approximately 25% of the total volume. Then, each aquarium was refilled to a fixed volume also using stored and well-aerated freshwater.

The bulbs of garlic were cleaned, peeled and cut into small pieces and dried in a Gallenkamp oven at 50°C for 14 hours at the Main Agriculture Laboratory of the Nigerian Stored Products Research Institute, Sapele out-station, Delta State, Nigeria. The dried garlic (GP) was grinded in electric mill into fine particle size (< 200 μm), then stored in a dry, air-tight transparent plastic container until composing of the experimental diets. GP was added in experimental diet at levels 0, 1, 2 and 3% / kg diet. Fish were fed with test diet at a ratio of 3% of their body weight for a period of 28 d, six days a week and twice daily at 0800 hour and 1600 hour. The present experiment was designed as 4 treatments for 4 weeks factorial replicated thrice to assess the effect of dried garlic as growth promoter in Tilapia zillii diet at levels 0, 1, 2 and 3% / kg diet. Estimation of experimental data on growth such as weight gain (g), specific growth rate (SGR) was conducted. Survivability (%) test of fishes in each aquarium was also conducted. Water quality parameters were regularly monitored throughout the experimental period for both control and treatment aquarium using LaMotte test kit. Data generated from this study were analyzed by using SPSS version 11.5 (Chicago, USA). Significant differences were determined among treatments at the 5% level (P < 0.05).

Results

Growth responses of T. zillii after 28 d of feeding are summarized in Table 1. The results of this present study revealed that fish group fed on garlic powder (GP) at 3% graded level had higher final weight (FW) of 7.59g followed by fish group fed on GP at 2%/kg diet (4.34g) while the least final weight of 3.18g was recorded in the control diet (0%). Growth responses of T. zillii in terms of FW and MWG were significantly affected by the GP (P < 0.05) as shown in Table 1 and Figure 1. Fish in all treatments gradually grew with time, and the highest individual weight was obtained in the 4^th week (28 d) as shown in Figure 1, and there was also a steady increase in the average mean weight gain from week zero to the 4^th week.

The highest amounts of dry feed intake (5.51 g/fish/day) were seen in fish groups fed on GP at 2% graded level while the least (5.21g/fish/day) was recorded in the fish group fed the basal diet (control group at 0% graded level) as shown in Figure 2.In this study, it was observed that the feed intake increased with increasing Allium sativum with the exception of the group fed at 3% graded level.

Fish fed GP at 3% graded level had the best feed conversion ratio (FCR) of 0.69 and were able to convert the feed better to flesh while the worst FCR of 1.65 was recorded in fish fed basal diet without garlic (0% inclusion rate).  Similarly, fish fed GP at 3% graded level had the best feed efficiency ratio (FER). However, there was a significant difference (P<0.05) between FCR of fish group fed on diet containing 3% graded level of GP and that of the other fish groups. It was also observed that the FCR recorded in this experiment decreased with increasing Allium sativum levels.

Survival rate exceeded 80% in all treatments (Figure 3) but there were significant differences (P < 0.05) among the treatments. The highest survival rate of 93.30% was recorded in the treatment fed GP at 3% / kg diet followed by treatment fed GP at 2% / kg diet which recorded 86.70% survival rate while the least survival rate of 66.7% was recorded in the basal diet (control group) without garlic. In general, the high survival rate recorded in this present study is an indication that the garlic-supplemented diets improved the growth and survival rates of T. zilli.

Physical and chemical water quality parameters are presented in Table 2. In general, there was no significant difference (P > 0.05) in the water quality parameters recorded in all the treatments. Results showed that all tested physical and chemical parameters were within the permissible levels required for tilapia growth.

Discussion

The significant (P < 0.05) FW and MWG recorded in the fish group fed on the GP at 3% graded level in this study is contrary to the opinions of Irkin et al., (2014) who reported that the efficacy of different doses, particularly at high doses, appeared to be worse based on investigations of toxicology. Similarly, Yilmaz and Ergun (2012) reported that high concentrations of phyto-additives may result in negative effects on fish health which is also contrary to the data generated in this study in which high survival rates was recorded at the end of this experiment. These may explain the fact that allicin, the major bioactive compounds in garlic, which enhances growth response, stimulate appetite, improve anti-oxidant status and act as immune-stimulants and anti-stress in fish may cause different synergetic effects on different finfish species. Results in Table 1 are in agreement with that reported by Soltan and El-Laithy (2008) who reported that the incorporation of garlic in diets for growing Nile tilapia significantly improved MWG and SGR.

The survival rate was significantly greater in all garlic-supplemented groups (1% - 3% graded levels of GP) when compared with the control group (0% graded level) at the end of the experiment. However, the survival rate was significantly higher in the group fed GP at 3% graded level. Although the use of garlic resulted in good survival rates, feeding the higher doses of garlic for extended periods gave better results (Figure 3). The current results showed the stimulatory effect of garlic on the immune system that correlates with improved fry survival. In general, the significant (P < 0.05) growth rates recorded in the group fed A. sativum when compared to the control group fed the basal diet in this present study could be attributed to the fact that the various organosulphur compounds, including allicin in garlic enhance various activities such as growth, appetite, and also play pivotal role as immunostimulant and antistress.

There is a paucity of information on the optimum dosage and duration regime of garlic as growth promoters in different finfish species and with more emphasis on the red belly tilapia, T. zillii. For instance, Shalaby et al. (2006) reported a significant increase in MWG, FER, PER, and SGR of Nile tilapia fed diet containing 3% GP. Similarly, Diab et al. (2002) mentioned feeding diet with 2.5% garlic resulted in the highest growth performance in O. niloticus. In the same species, Abou-Zeid (2002) found a positive improvement in biomass and SGR with garlic supplementation. Metwally (2009) also mentioned that the best performance was obtained in Nile tilapia fed diet with 3.2% GP. On the other hand, Mehrim et al. (2014) conducted a study on the dietary addition of garlic lobes in O. niloticusat 0, 1, 2, and 3% inclusion levels for 60 day regime and they observed significant growth in O. niloticus at 1% / kg diet which is a growth pattern different from those obtained in T. zillii in this present study. The different growth pattern in the O. niloticus recorded by Mehrim et al. (2014) for 60 d regime and those recorded in T. zillii in this present study for 28 d regime could be due to differences in fish species to metabolize the active bioavailable dose in the garlic-containing diets. According to Pollack et al. (2003), differences in the growth response to a phytochemical may be due to differences in fish species’ ability to metabolize or utilize a particular substance to which it has been exposed in the diet. The significant (P < 0.05) growth responses and feed utilization recorded in T. zillii fed the highest dose of 3% graded level of A. sativum is contrary to reports of Mehrim et al. (2014) who recorded significant growth in O. niloticus fed A. sativum at 1% graded level and least significant growth at 3% graded level. This may explain the variable effects of phytochemicals on certain endpoints in different species and differential levels of growth performance by a particular fish species at various concentrations of the phytochemical (Chakraborty et al., 2013). Further research to analyze the functional mechanism behind the physiological activity of garlic-supplemented diets in different finfish species needs to be investigated.

The water quality parameters measured were within the acceptable limits for tilapia growth as stated by El-Sayed et al., 1996; Milstein and Svirsky, 1996 and Salem, 2006. The physico-chemical parameters monitored in the glass aquaria tanks under the laboratory conditions showed that water temperature, dissolved oxygen, and pH values varied among all the treatments and this is in accordance to Frank 2000 and Marjani et al., 2009 findings who stated that DO should be above 3mg/l, pH of 6.0-9.5 and temperature (28+10C). No significant difference in water temperature, pH and DO was observed among the treatments.

Conclusion

There is a paucity of information on the optimum graded level of garlic (A. sativum) as growth promoters for different finfish species. Thus, the necessity for this present laboratory experiment to optimize the optimum graded level for the growth, nutrient utilization and survival of T. zillii reared in a glass aquaria tanks for 28 days. The results of this study revealed that fish group fed on GP at 3% graded level had significant (P < 0.05) higher final weight (7.59g), mean weight gain (7.56), average daily gain (0.27mg/fish/day),survival rate (93.30%) and better feed conversion ratio (0.69) when compared to other treatments. The study therefore recommends that T. zillii should be fed on a garlic-containing diet at 3% / kg diet to achieve significant growth rates and to reduce the stunted growth which is the bottleneck in tilapia industries. Further research to analyze the functional mechanism behind the physiological activity of garlic-supplemented diets in different finfish species needs to be investigated.

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