The South African mariculture industry is developing the rearing technology of indigenous fish species. Dusky kob (Argyrosomus japonicus) has high-quality flesh and a good market demand. Research is needed to determine the environmental requirements under which growth of dusky kob is optimised. This study assessed the effect of temperature, light intensity, feeding frequency and stocking density, respectively, on growth, feed conversion ratio (FCR) and survival of juvenile dusky kob within a weight range of 10-60 g fish⁻¹) in a series of four growth trials. The effect of temperature on growth and FCR was assessed at 14 temperatures from 17 to 28°C. The temperature for best growth was estimated to be 25.3 °C, while 21.4 °C was the temperature at which the best FCR was achieved. A growth trial testing the effect of light intensity on growth showed that light intensity did not affect growth in the range of 23 – 315 lx. Fish fed a restricted ration of 3.6% body weight per day (chapter 2) or 3.41% body weight per day (chapter 4) had a better FCR than fish fed to satiation. A trial to assess the effect of feeding frequency on growth and FCR showed that fish fed both two or three times daily grew better than those fed once daily. FCR was best in fish that were fed once or twice daily. Preliminary analysis of the results from a stocking density trial showed that stocking density in the range of 10 – 50 kg m⁻³ did not affect growth of juvenile dusky kob. It is recommended to culture dusky kob at a temperature of 25.3 - 21.4 °C at stocking densities up 50 kg m⁻³ to maximise growth. However, a study is needed to determine the environmental conditions needed to maximise profit under commercial conditions. The results allow the South African industry the opportunity to assess the culture potential of this species. In addition, the results will help develop protocols that can be used in other South African candidate aquaculture species such as silver kob, yellowtail, white-stumpnose and sole.

Traditionally, fish oil (FO) has been used extensively in aquafeeds. The stagnation in global fish oil production coupled with an increased demand for its use in aquaculture feeds, especially salmonid feeds, has greatly inflated fish oil prices. Therefore, in order to sustain the rapid growth of the tilapia industry, the dependence on these commodities in feeds should be reduced through use of cheaper and more sustainable sources of dietary lipids, such as palm oil. This study therefore investigated several, previously poorly understood, effects of palm oil on reproductive performance of the commercial tilapia species, Oreochromis niloticus; which currently ranks as second most popular species in world aquaculture. In the present study broodstock were fed on experimental diets at full and half ration regimes throughout their entire life cycle from exogenous feeding. Studies were conducted in standardised and controlled hatchery conditions, thereby reducing the potential influence of environmental variations. First feeding O. niloticus fry were fed on four diets, cod liver oil (D 1), palm oil (D 2), mixed palm and cod liver oil (D 3) (9:1 ration) and a commercial trout diet as control (D 4) (Skretting, U.K.) on a reducing ration based on fish size. The present study investigated the effect of dietary lipid sources on (1) growth performance, (2) biochemical composition of eggs (total lipid and fatty acid composition), (3) morphological parameters of eggs (total and relative fecundity, egg size, egg weight and EW:BW), (4) larval quality (larval length and weight) and (5) oocyte recruitment and its associated sex steroid hormones. Experimental diets and feeding ration significantly influenced (p<0.05) the growth performance over a period of 120 days. Total lipid and fatty acid composition of eggs originating from broodstock fed on palm oil, mixed palm and cod liver oil (9:1) or a control diet were not significantly different (P>0.05) when fed at either full (3% BWday-1) or half ration (1.5% BWday-1). The present study, however, confirmed that fatty acid composition of fish eggs reflected the fatty acid composition of the diet, although specific fatty acids were selectively utilized or retained in the eggs. The mean inter-spawning interval (ISI) increased with increasing fish size and averaged 14, 19 and 24 days for fish fed on palm oil, mixed palm and cod liver oil or control diets, respectively. The shortest ISI observed was 7 days for fish fed a palm oil diet. Total fecundity ranged from 660 - 820 eggs/clutch. Mean total fecundity was 750, 820 and 660 eggs/clutch for fish fed a palm, mixed palm and cod liver oil or a control diet, respectively, but these differences were not significant (P>0.05). However, relative fecundity and egg weight to body weight rates as a percentage (EW: BW) were found significantly differ (p<0.05) between fish fed the control diet and experimental diets. Mean egg diameter (2.2 mm) was not significantly influenced (p>0.05) by experimental diets. The egg volume, egg dry and wet weight, fertilisation and hatching rate were also not significantly different between fish fed the experimental diets. Oocyte development was classified into distinct stages based upon oocyte size, biochemical properties and structure. The recrudescence to these stages was not significantly influenced by broodstock fed experimental diets either at full or half ration. Steroid hormones and histological analyses provided valuable data concerning the oocyte development and recruitment in this species. Levels of 17ß-oestradiol (E2) and testosterone (T) peaked within 6 days of spawning, suggesting that vitellogenesis began as early as day 2 or 3 post-spawning. By day 6, ovaries were dominated by large late-vitellogenic/maturing oocytes (stages 6 & 7) occupying about 70% of the ovary. Gonadosomatic index (GSI) reached maximal levels by day 6. It is suggested that pre-vitellogenic oocytes are recruited into vitellogenic growth immediately after spawning and complete vitellogenesis on day 6 post-spawning. Finally, the present study investigated the effect of food restriction at two rations (full and half) on broodstock reproductive performance. Oreochromis niloticus were rationed from first feeding and throughout their life-cycle. The dietary regime, full ration (3%) and half ration (1.5%), influenced fish size but despite this variation no significant differences (p>0.05) were detected in total lipid and fatty acid composition in the eggs, total fecundity, egg diameter, total egg volume and larval size. These results suggested that despite large differences in food availability throughout their life cycle, investment in reproduction had remained remarkably consistent. It appeared that during food restriction, O. niloticus sacrificed body weight and growth so as to maintain reproductive investment. In summary, this study provides valuable information using a novel experimental design on the effects of dietary lipid sources on reproductive performance of female O. niloticus. Substituting palm oil for fish oil as the dietary lipid source and reducing ration by half (1.5% BWday-1) had no significant effect on reproductive performance. Therefore it is suggested that under controlled conditions, lipids of non-marine origin, such as palm oil, can be successfully substituted for broodstock diets. Halving feed requirement should also increase profitability of seed production. KEYWORDS: Tilapia; O. niloticus; palm oil; diet; fecundity; spawning periodicity; oocyte recruitment; reproductive performance.

Intensive and semi-intensive aquaculture systems are dependent on nutrient input either in the form of supplemental or complete feeds. Most complete diets still include high fish meal (FM) levels (≥10%). However, as the industry attempts to reduce its reliance on FM, feeds must now be formulated with much lower levels especially for omnivorous species such as tilapia. By 2015, mean FM inclusion in tilapia diets was projected to fall below 3% and be further reduced to 1% by 2020. In the global context of competition for crops, finding suitable plant-based replacers for FM and meeting the increasing demand for seafood, lower-cost and under-utilised plant feedstuffs are now receiving greater attention. The study was divided into three distinct components – field survey, growth experiments, and life cycle assessment. Field surveys were used to contextualise the growth experiments and assess commercialisation opportunities for multi-enzyme inclusion in tilapia feeds. Two sets of digestibility and growth experiments were designed to evaluate the feasibility of using high inclusions of plant-based ingredients sourced from locally available feedstuffs in Thailand to substitute FM at low inclusion levels (0 – 5%). The research evaluated the hypothesis regarding the potential of exogenous enzymes (protease, xylanase and phytase) to minimize anti-nutritional effects on nutrient digestibility of proteins, polysaccharides and phosphorus in tilapia. The research also assessed the secondary effects of enzyme supplementation on economic efficiency and life cycle environmental impacts. Tilapia is the second most cultured finfish globally and Thailand is the sixth largest producer. Based on the findings of the field survey, feeding practices of Thai tilapia farmers were confirmed to be diverse. Feed inputs included, but were not limited to, agro-industrial by-products (e.g. rice bran, corn bran etc.) and commercial diets. Commercial diets contained 15 – 30% crude protein and lower protein livestock diets (i.e. pig ration) were often used for supplemental feeding or “fattening”. The experimental low FM diets were therefore formulated as grow-out or “fattening” diets for semi-intensive green-water systems, a prominent feature (>60%) of Thai tilapia farming. In Phase 1, the digestibility experiment assessed the digestibility and growth in tilapia fed 0%, 3% and 5% FM diets with and without xylanase (0.385 g kg-1) and phytase (0.075 g kg-1). Performance decreased significantly with declining FM levels. No differences in feed intake, feed conversion ratio (FCR), specific growth rate (SGR) and weight gain were observed between the enzyme and control diets. Nevertheless, tilapia fed the enzyme supplemented 3% FM and control 5% FM performed similarly (P < 0.05). No enzyme-related effects were noticed for protein digestibility but phosphorus (P) digestibility improved by 9%, except at 0% FM level (P > 0.05). The enzymes had no apparent influence on nitrogen (N) retention contrary to previous studies, however, higher retention for P was observed. Villus length decreased with declining FM levels yet no improvements were seen in tilapia fed enzyme diets. In a simultaneous grow-out experiment, the six experimental diets were compared to an industry 10% FM standard. Conversion ratio was the lowest (1.66) in adult tilapia fed 10% FM diet however the enzyme supplemented 0% FM fed fish had a comparatively low FCR of 1.67. There were no significant enzyme-related effects on weight gain¸ SGR and protein efficiency. Proximal villi results were inconsistent. The cost of feed decreased with declining FM levels but increased with enzyme inclusion. Nevertheless, the economic returns per kg of whole fish produced were better using enzyme supplemented diets compared to the controls. Though the size of the effects on growth and nutrient utilisation were modest, the findings suggested that xylanase and phytase had some level of synergistic action on the targeted anti-nutrients. However, further research was required. In Phase 2, two control diets (2% FM, negative control (NC) and 10% FM, positive control (PC)) were compared with three enzyme supplemented 2% FM diets (NO-PRO, 0.385 g kg-1 xylanase and 0.075 g kg-1 phytase only; LO-PRO, xylanase + phytase + 0.2 g kg-1 protease and HI-PRO, xylanase + phytase + 0.4 g kg-1 PRO). Growth performances improved with enzyme supplementation compared to the NC (P < 0.05). Of the enzyme supplemented diets, the LO-PRO diet showed the highest improvements in weight gain (26%) and feed intake (19%), the latter comparing statistically to the 10% FM PC diet. The HI-PRO diet had the best FCR (1.88), again comparable to the PC (1.73). The NO-PRO diet had the highest protein, P, lipid and energy digestibility, suggesting no additive effect of protease on these coefficients. In terms of gut histomorphology, the LO-PRO and PC diets had the highest measurements and were statistically similar which may have explained similarities in feed intake. Compared to the NC, the HI-PRO diet produced the highest level of change in net profit due to gains in feeding efficiency however, the LO-PRO showed better improvements in terms of growth. Based on these findings, the ternary combination of protease with xylanase and phytase (LOPRO) has potential in limiting FM use for tilapia grow-out feeds, however, the economic efficiencies were still below that of a 10% FM diet. Future considerations for research should target the indigestible dietary components in order to optimise enzyme dosages and maximise the benefits of each enzymes. In conclusion, a comparative life cycle assessment (LCA) was used to evaluate the environmental impacts of low FM diets and commercial feeds associated with tilapia production in Thailand. The study showed that the low FM enzyme supplemented diets had lower impact potentials and were environmental superior to the average (10% FM) commercial standard. LCA modules are recommended for least-cost formulation programmes as an option going forward. Additionally, LCA can be used as a predictive tool to guide farmers, especially small-scale producers, on the potential impacts of feed input choices and feeding practices. This will ensure higher product quality but also demonstrate environmental responsibility on the part of aquafeed and fish producers to final seafood consumers.

The gilthead seabream (Sparus aurata) and European seabass (Dicentrarchus labrax) require n-3 long-chain polyunsaturated fatty acids (LC-PUFA), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), for optimal growth and health. Due to the rapid growth of global aquaculture the quantity of marine oils used in aquafeeds has been limited, yet the overall quantity of oil in an aquafeed has increased by the addition of vegetable oil (VO) to supply dietary energy. For aquaculture to continue to grow more fish must be produced with less marine ingredients, yet EPA and DHA must be maintained at levels above fish requirements. This project set out to re-evaluate the requirement for EPA and DHA in gilthead seabream and European seabass. Two dose-response studies were designed and executed where juvenile seabream and seabass were fed one of six levels of EPA+DHA (0.2 – 3.2 % as fed). Biometric data were collected and analysed to determine new requirement estimates for EPA+DHA for fish of two weight ranges (24 – 80 g and 80 – 200 g). The effects of the dietary LC-PUFA gradient on lipid composition and metabolism were also considered.