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Effect of dietary supplementation of biofloc meal on growth and survival of GIFT tilapia

A 60 days indoor growth trial was conducted to study the effect of dietary supplementation of biofloc meal on growth and survival of juvenile GIFT tilapia. Four isonitrogenous and isoenergetic experimental diets (32% crude protein) were prepared using biofloc meal at different inclusion levels viz., 0 (T0), 20 (T1), 30 (T2) and 40% (T3). A commercial diet (T4) was used for comparison. The feeding trial was conducted in 15 nos. of 40 l plastic troughs in triplicate, utilising GIFT tilapias weighing an average of 2 g. During the experimental period, water quality parameters were measured and recorded daily. The mean value of water temperature, pH, dissolved oxygen, salinity, ammonia-N, nitrite-N, nitrate-N, hardness and alkalinity recorded in the experimental systems were 28.5 °C , 8.1, 5.5 ppm, 4.5 ppt, 0.03 ppm, 0.07 ppm, 11 ppm, 630 ppm and 162.5 ppm respectively. Among the biofloc meal enriched diets, diet T1 with 20% biofloc yielded the best results in terms of average body weight gain, feed conversion ratio (FCR), specific growth rate (SGR), protein efficiency ratio (PER) and feed efficiency ratio (FER). The mean body weight gain recorded in T1 was 25.28±0.81 g. The results demonstrated that biofloc meal is a potential ingredient that can be incorporated in GIFT tilapia diet at 20% level for better growth performance.


Introduction
Aquaculture is the fastest growing sector of global fish production (FAO, 2010). Approximately 40% of fish from aquaculture originates from tilapia production (Scorvo-Filho et al., 2010). Today, tilapia has become the shining star of aquaculture, which is also popular as 'aquatic chicken' and the rate of its consumption has increased across the globe (Fitzsimmons, 2005). Annual global production of cultured tilapia has increased consistently in recent years (Ahsan et al., 2013). The GIFT strain was developed by International Centre for Living Aquatic Resources Management (ICLARM; presently World Fish Centre, WFC) through several generations of selection from the base population involving 8 different strains of Nile tilapia Oreochromis niloticus (Eknath and Hulata, 2009). Feed costs in aquaculture can be reduced by the adoption of biofloc technology, since the expenses with commercial diets account for more than 50% of the production cost. Biofloc contains up to 30% crude protein and about 2% lipids (Azim and Little, 2008;Ballester et al., 2010;Xu and Pan, 2012;Luo et al., 2014). Biofloc contains adequate levels of protein, lipid, carbohydrate and ash and is suitable for use as an ingredient in aquaculture feed (Crab et al., 2010). In this context, the present study was planned to evaluate inclusion of biofloc meal at different levels in the diet of GIFT tilapia.

Experimental fish
GIFT tilapia seeds were procured from the State Fisheries Department, Krishnagiri, Tamil Nadu, India. They were acclimatised in FRP tanks, by nursing for 15 days feeding on a commercial diet. They were graded according to weight prior to the experiment. One hundred and fifty numbers of GIFT tilapia having average weight of 2 g were selected for the trial.

Experimental design
The trial was conducted in the indoor biofloc laboratory of Fisheries College and Research Institute, Thoothukudi, Tamil Nadu, India. The 60 days indoor growth experiment carried out in plastic troughs of 40 l, consisted of five dietary treatments with three replicates each. Water was filled in the troughs upto 3/4 th of the volume. All the troughs were provided with proper aeration facility.

Biofloc collection and analysis
Biofloc was collected using a plankton net of 100 µm mesh size from Hi-Tide sea farm, Mahendrapally, Nagapattinam District, Tamil Nadu, India. Totally 23 kg of wet biofloc was collected and dried under sunlight for 8 h. The total quantity of dried biofloc meal obtained was 2.1 kg. It was powdered into fine particles and stored in an air tight container under refrigeration. The proximate composition of the biofloc meal was analysed following standard protocols (AOAC, 1995).

Experimental diets
Four isonitrogenous and isoenergetic experimental diets were formulated viz., T0, T1, T2 and T3. A control diet (T0), without biofloc was compared against the three prepared diets formulated with different levels of biofloc at 20 (T1), 30 (T2) and 40% (T3) by manipulating fish meal and soybean meal level. A commercial diet (T4) was used as an external control. The ingredient composition of the experimental diets is presented in Table 1. method (APHA, 2005) was adopted to estimate dissolved oxygen. Total alkalinity was determined as per the method described in APHA (2005). Total ammonia-N, nitrite-N, nitrate-N, water hardness and turbidity were assessed twice a week, following standard methods (APHA, 2005).

Sampling
Fish sampling was done every fortnight, collecting all the stocked fish from each tank to measure total length and body weight.

Growth performance
Growth performance was assessed in terms of feed conversion ratio (FCR), feed efficiency ratio (FER), protein efficiency ratio (PER), specific growth rate (SGR), The proximate composition of all the experimental diets (T0 -T4) was estimated following standard protocols (AOAC, 1995).

Stocking and feeding
Fish of 2 g mean weight were stocked at 10 nos. per plastic trough. After stocking, the experimental troughs were covered with plastic net on the top in order to prevent fish jumping out of the troughs.
Feeding was done thrice a day (at 9: 00; 12: 00 and 16: 00 hrs) ad libitum. Each of the experimental diets was fed by hand slowly to avoid wastage. Feed was given until apparent satiation.

Water quality parameters
During the experimental period, water quality parameters such as temperature, dissolved oxygen, pH, and total alkalinity were measured daily. Water temperature was measured using a thermometer with an accuracy of 0.1 o C. pH of water was measured using the laboratory model Elico pH meter. Modified Winkler's titration mean weight gain and survival estimated using the following formulae:

Statistical analysis
Data were analysed by one-way ANOVA using the statistical software SPSS 16.0 for windows (SPSS Inc., Chicago, IL, USA) and also by Duncan multiple range test (Duncan, 1995).

Water quality parameters
The range of water quality parameters such as temperature, dissolved oxygen, pH and total alkalinity recorded in the experimental systems are presented in Table 2.

Proximate composition of experimental diets
Proximate composition of the experimental diets is presented in Table. 3. Dried biofloc contained 17.92% crude protein, 0.15% crude fibre, 0.41% ether extract, 51.28% total ash, 10.06% moisture and 1864 kcal kg -1 gross energy. The percentage crude protein content of the prepared experimental diets ranged from 32.54±0.07 to 32.74±0.05.

Discussion
Nutritional composition of biofloc varies according to the environmental condition, carbon source applied, total suspended solid level, salinity, stocking density, light intensity as well as with changes in phytoplankton and bacterial communities. Protein content of biofloc in the present study was 17.92% which tallies with the earlier findings of Soares (2004), Hende et al. (2014) and Neto et al. (2015), whereas lesser values of biofloc crude protein was reported by Magondu (2012) and Megahed (2014). The lipid content of biofloc in this study was 0.41%,    (2015), Hargreaves (2013) and Himaja (2016). Biofloc is usually a poor source of total lipids with concentrations ranging from below 1 to 25 g kg -1 (Azim et al., 2008;Avnimelech 2009;Kuhn et al., 2009;Bauer et al., 2012). The ash content of biofloc in the present investigation was 51.28%, which agreed with earlier reports (Soares, 2004;Maica et al., 2012;Mangodu, 2012;Hende et al., 2014;Neto et al., 2015;Suita et al., 2015). High ash content of biofloc may be related to the abundance of acid-insoluble oxides and mixed silicates (Tacon et al., 2002). Accumulated minerals and metals may also contribute to the high ash content. Biofloc may thus be a good source of essential minerals and trace elements (Tacon et al., 2002;Ju et al., 2012) and it may contribute to animal growth (Avnimelech, 2006).
In the present study, the highest mean body weight of GIFT tilapia was recorded in the control, followed by T1, T2, T3 and T4. However, the mean body weight of fish in T1 group fed 20% biofloc meal did not differ significantly from that of the control. Increase in the dietary biofloc meal level resulted in significant decrease in growth rate and higher FCR when compared with the control. Minimum biofloc meal inclusion (20%) produced better weight gain and lower FCR than the maximum inclusion level (40%) which is in agreement with Ajiboye et al. (2012) and Kissling and Askbrandit (1993) who observed reduction in growth rate of fishes at higher levels of microbial supplementation, as microbial products at higher levels tend to reduce feed palatability and digestibility. In the present study, the survival of fish was 95%, with the exception of T4 (86.6±0.88%). FCR was lowest in the control, followed by T1, T2, T3 and T4 that showed higher values. These values fell in the range from 1.10 to 1.50 as reported by Moss et al. (2006), Samocha et al. (2007), Taw et al. (2009, Xu et al. (2012), Hussain et al. (2014) and Dantas et al. (2016). PER is used to assess protein utilisation and turnover, where it is related to dietary protein intake and its conversion into fish protein. Ahamed et al. (2004) observed that PER is significantly affected by dietary protein level and ranged from 1.58 to 2.35 for fry, 1.19 to 1.92 for fingerlings and 0.99 to 1.53 for adult Nile tilapia. PER values in the present study ranged from 4.90±0.74 to 7.41±0.10.
Earlier studies have reported increase in growth rate, general welfare and survival of fishes reared in microbial floc based systems (Tacon et al., 2002); however, studies that have used flocs as ingredients in aquaculture diets are limited. Kuhn et al. (2009) replaced 37% of fishmeal in shrimp feed, achieving similar performance as diets containing lower replacement levels. In another study, Kuhn et al. (2010) reported no deleterious effects on growth when L.vannamei were fed with diets containing biofloc inclusion from 10 to 30%, regardless of fishmeal substitution of 0 -67%. The results of the present study suggest that 20% biofloc meal in the diet is optimum for the growth of GIFT tilapia. This finding is supported by the results of Himaja (2016) who obtained higher growth of catla by replacing fish meal in the diet with 20% biofloc meal. Dantas et al. (2016) suggested that fish meal replacement by biofloc meal at 20% improves the growth of L. vannamei, whereas Bauer et al. (2012) stated that fish meal in L. vannamei post-larve diet could be replaced by biofloc meal up to 30%.
The positive effect of the experimental diets on the growth of GIFT tilapia clearly demonstrates that biofloc meal is a viable alternative ingredient and can be used in the diet up to 20% level. Biofloc meal thus represents an important, alternative ingredient for the development of cost effective feed for fishes, in particular by reducing, dependence on fishmeal as well as environmental impact.