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The establishment of the early-life gut microbiota plays an important role in fish development and influences the host's health status and growth performance. Different rearing conditions can impact the initial colonization of the gut microbiota, while the addition of probiotics may also affect such colonization. However, how this may affect fish larvae survival and growth remains largely unexplored. In this study, 3-day old Nile tilapia embryos were hatched until 9 days post fertilization (dpf) in three systems, including one flow-through system (FTS) and two identical recirculating aquaculture systems (RAS). When feeding started at 10 dpf, tilapia larvae in the FTS and one of the RAS were fed with a control diet, while larvae in the second RAS were fed with the control diet coated with B. subtilis spores (RASB). The feeding trial lasted 26 days, from larvae to fry stage, during which the survival, growth performance and gut microbiota were analyzed. The larvae reared in FTS showed significantly lower survival than those in RAS and RASB, while no differences were observed in fish growth and apparent feed conversion ratio between treatments. Different rearing systems resulted in different gut microbiota compositions, which strongly correlated with the survival rate and standard body length at harvest. Cetobacterium was enriched in RAS and RASB, while was barely detected in the gut of FTS-reared tilapia fry. Probiotic supplementation increased the relative abundance of beneficial Bacillus in fish gut. Our findings indicate that rearing fish larvae in RAS supports better survival compared to FTS, while dietary probiotic supplementation further modulates the gut bacterial composition and stimulates presence of beneficial bacteria during early life.

To assess the potential for improving sustainability and efficiency in an important Vietnamese finfish culture, we quantified sustainability indicators for 2 downstream and 2 upstream 3 to 4 m deep ponds for striped catfish Pangasianodon hypophthalmus (Sauvage, 1878) production along the Mekong River, Vietnam. The resources used per kg fish produced were (downstream vs. upstream): 2.8 vs. 7.1 m3 water, 0.04 vs. 0.14 kWh energy, 0.06 vs. 0.11 h human labour and 0.06 vs. 0.15 g antibiotics (p < 0.05). The feed conversion ratio (kg feed dry matter per kg fish wet weight) was 1.50 vs. 1.65 and feed dry matter utilization efficiency was similar (28 vs. 30%) for both types of ponds (p > 0.05). Nitrogen and phosphorus utilization efficiencies were 44 vs. 40% for nitrogen (p < 0.05) and 17.7 vs. 17.6% for phosphorus (p > 0.05). An advantage of production in deep ponds is the breakdown of organic matter through denitrification and fermentation. Based on the difference between feed input and monitored outputs and sinks, an estimated 29–37% of dry matter volatilized as CO2 and 30–34% of nitrogen was lost as N2. A major disadvantage of the deep pond production system is its dependency on water exchange with the Mekong River, resulting in low biosecurity, which is aggravated by the lack of availability of disease-free fingerlings. Dealing with disease caused farmers to apply suboptimal feeding rations and to lengthen the culture period. Developing (semi-)closed systems that adopt elements from recirculation technology could improve both culture performance and sustainability.

This paper compares the quantity and quality of solid waste obtained from traditional ponds and recirculating aquaculture systems (RAS) for striped catfish and evaluates methane and compost production from these wastes. Striped catfish sludge was collected from four commercial ponds along the Mekong river and from three indoor RAS. The amount of sludge dry matter produced per kilogram of fish in ponds was 6 times higher than that in RAS. However, the concentration of nutrients in solid waste from RAS was much higher, with better compost quality and higher methane yield than that of sludge from ponds. Out of the collected 381 L biogas, the methane yield of striped catfish’s solid waste in RAS systems was 201 L per kg chemical oxygen demand (COD). In ponds, the collected 267 L biogas yielded 125 L CH4 per kg COD. The higher methane production from RAS sludge concurred with higher digestibility of COD: 58% for RAS versus 38% for ponds. The quality and quantity of methane from striped catfish sludge were lower than that for other animal manures, and the resulting electricity yield was low. Considering the higher nutrient concentration in RAS-sludge, we recommend the combination RAS and composting in reusing sludge, which is presently the best option for a more sustainable and cleaner striped catfish production system.

Oxygen limitation and dietary imbalances are key aspects influencing feed intake (FI) and growth performance in cultured fish. This study investigated the combined effects of hypoxia and dietary electrolyte balance on the growth performance, body composition and nutrient utilization in a rainbow trout (Oncorhynchus mykiss) isogenic line. Fish were fed ad libitum two experimental diets: electrolyte-balanced or -imbalanced diets (DEB 200 or 700 mEq kg−1, respectively) and exposed to normoxia or hypoxia (7.9 or 4.5 mg O2 l−1, respectively) for 42 days. DEB did not affect FI, growth performance or body composition. Nevertheless, hypoxia had a negative impact, reducing FI (6%), growth rate (8%), oxygen consumption (19%), energy (5%) and lipid (42%) contents. Digestible energy intake and heat production were higher in normoxic fish (40% and 23%, respectively), retaining 64% more energy in lipid or protein. Hypoxia reduced the apparent digestibility of dry matter, ash, protein, lipid, carbohydrates and energy. Trout fed DEB 700 diet were energetically less efficient, reflected in higher heat production and energy requirements for maintenance. FI was inhibited by low dissolved oxygen levels, but not by electrolyte-imbalanced diet, in spite of the higher energy requirements for maintenance. This study highlights the importance that dietary-electrolyte content and DO levels have on energy balance and growth performance when fish are fed to satiation.

We evaluated the effect of low pH and low and high total ammonia nitrogen (TAN) concentrations on the physiology, stress status and the growth performance of turbot in RAS. Two experiments were conducted. In Experiment 1, turbot (466 g) were grown at control (pH 7.5; TAN ~0.5 mg/L) or low pH and high TAN (pH 5.7; TAN ~50 mg/L) for 55 days. In Experiment 2, turbot (376 g) were grown at control (pH 7.5; TAN ~0.5 mg/L), low pH and low TAN (pH 5.7; TAN ~5 mg/L) or low pH and high TAN (pH 5.7; TAN ~50 mg/L) for 59 days. In Experiment 1, final body weight, feed intake and growth were significantly lower and FCR significantly higher in turbot exposed to low pH and high TAN. In Experiment 2, only growth was significantly lower in turbot exposed to treatment low pH and high TAN as compared to fish in the control treatment and low pH and low TAN. Osmoregulation and stress indicators measured were within normal levels. In conclusion, turbot grew equally well in a water pH of 7.5 or 5.7 provided a low TAN. In contrast, low pH combined with a high TAN impaired turbot performance. The effect of low pH on physiology, stress status and growth performance of turbot (Psetta maxima L.) cultured in recirculating aquaculture systems

The accumulation of steroids released by fish in recirculating aquaculture systems (RAS) may potentially influence their physiology and behavior. The present study examined the release rate of cortisol and testosterone by Nile tilapia, Oreochromis niloticus, and their accumulation in six identical lab scale RAS operated at different water exchange rates (150 L/kg feed/day, (LowRAS) and 1500 L/kg feed/day, (HighRAS)) and how steroid accumulation is affected by grading and weighing induced stress. Water cortisol and testosterone concentrations during the experimental period ranged between 1.0 and 5.1 ng/L and between 1.4 and 9.4 ng/L, respectively. Water cortisol concentration was 34% and 43% significantly higher in LowRAS when compared with HighRAS at week 3 and week 4, whereas for water testosterone concentration the two treatments did not differ. Overall steroid release rates were 0.02 +/- 0.02 and 0.04 +/- 0.02 ng/g body weight/h for cortisol and testosterone, respectively. Two hours after grading and weighing water cortisol concentration increased 30% in the LowRAS whereas water testosterone concentration remained unchanged. Calculated cortisol and testosterone discharge from the system were, respectively, 87% and 89% lower in LowRAS than in HighRAS. These findings show that reduced water usage and acute stressors can induce significant accumulation of cortisol in the rearing water at levels close to olfactory detection. Accumulation of metabolites, in particular steroids, should be taken in consideration when designing and managing RAS to prevent exceeding allowable concentrations. Statement of relevance: RAS containing Nile tilapia, cortisol and testosterone are released to the culture water. Water cortisol concentration depends on the water exchange rate that is used and higher concentrations are expected in lower water exchange rates. Fish grading and weighing has a short-term effect on the water cortisol concentrations, after which concentrations return to the basal levels. These findings show that reduction in water usage and fish handling may lead to a significant increase of hormones in the rearing water, which emphasizes the importance of accounting for steroids in the design and management of RAS. (C) 2016 Elsevier B.V. All rights reserved. Portuguese institution FCT - Fundacao para a Ciencia e Tecnologia (QREN-POPH - Advanced Training - European Social Fund) [SFRH/BD/65673/2009] Portuguese institution FCT - Fundacao para a Ciencia e Tecnologia (QREN-POPH - Advanced Training - national funds MEC) [SFRH/BD/65673/2009]

As a consequence of water re-use and high stocking densities, Recirculating Aquaculture Systems (RAS) may lead to an accumulation of substances released by the fish into the water, e.g. cortisol and alarm pheromones. This study investigated the effect of stressing fish on the feeding motivation of other fish not subjected to stress but sharing the same water of stressed fish. Two identical RAS were used (operated at 30 L kg feed-1 day-1) and contained grouped (stressed fish) and individually (receiving water from stressed fish) housed Nile tilapia. A stress test was applied in grouped housed fish on days 17 and 55. Feeding behaviour (intake and latency) was recorded in the individually housed fish 3 days before, during and 3 days after the stress test. The results showed no effect on feeding behaviour in fish receiving the water from stressed fish. These results could be a consequence of insufficient cortisol/alarm cues' release by the stressed fish into the water or inactivity of such substances, either due to a trapping effect of humic acids or due to degradation in the nitrification and denitrification processes. Future research is needed to clarify how these processes may affect the water concentration of cortisol and alarm pheromones and should be extended by measuring other behavioural and physiological traits

The accumulation of substances in Recirculating Aquaculture Systems (RAS) may impair the growth and welfare of fish. To test the severity of contaminants accumulated in RAS, early-life stages of fish were used. Ultrafiltered water from two Recirculating Aquaculture Systems (RAS), one RAS with a high accumulation of substances (water exchange rate 30 L/kg feed/day) and one RAS with a low accumulation of substances (water exchange rate 1500 L/kg feed/day), was used to incubate eggs and rear larvae of common carp Cyprinus carpio . A broad range of read-out parameters was used to determine the effect of accumulation level on the development of the early-life stages; from hatching dynamics to larvae length and dry weight. The water quality (temperature, pH, dissolved O 2 , conductivity, total bicarbonate, ortho-phosphate-P, TAN, NO 2 − –N, NO 3 − –N and minerals) was compared between the 2 treatments. Carp eggs developing in the high-accumulation water had higher mortality percentages (both for eggs and larvae), reduced hatching percentages, delayed hatching dynamics and reduced larvae length and body weight. However, these larvae exhibited fewer deformities than larvae incubated in the low-accumulation water. Furthermore, an accelerated development both of the embryo (appearance of heart beat, pectoral fin bud and tail movement) and yolk-sac larvae (depletion of the yolk sac) was observed in the high-accumulation water. The high-accumulation water had significantly lower pH and higher conductivity, NO 2 − –N, NO 3 − –N and ortho-phosphate-P. Most of the minerals (As, Cu, Mn, Ni, Zn, K, Mg, Na, P and S) including heavy metals, were present at a higher concentration in the high-accumulation water. The influence of these parameters on the embryonic and larval development of fish is discussed. It is suggested that in the high-accumulation water, the concentration of ortho-phosphate-P, nitrate and of the heavy metals arsenic and copper is likely to have impaired the embryonic and larval development and therefore deserves further research as potential growth inhibiting factors in RAS.

It has been suggested that fish cultured in recirculating aquaculture systems (RAS) grow less as compared with fish cultured in flow-through systems due to the accumulation of substances. In the Netherlands, the commercial culture of Nile tilapia Oreochromis niloticus in 300 and 600 MT's systems is done exclusively in RAS operated at water exchange rates as low as 30 L/kg feed/day due to nitrate control by single-sludge denitrification reactors. The use of such nearly closed RAS raises the question whether growth retardation (GR) is present in Nile tilapia. This study is the first to investigate the existence of growth retardation in Nile tilapia by comparing the growth, feeding behaviour and stress response of Nile tilapia cultured in RAS with different levels of substances accumulated. Three RAS, operated at 30 L/kg feed/day (HIGH accumulation), 70 L/kg feed/day (MIDDLE accumulation) and 1500 L/kg feed/day (LOW accumulation) were used. Each RAS contained 24 glass aquaria with individually housed fish. To determine whether GR is size-dependent, per RAS 3 fish size categories were tested in the 57 day experimental period: large (288.7 ± 34.2 g; N = 8), medium (162.4 ± 23.4 g; N = 8) and small (81.4 ± 21.0 g; N = 8). Experimental fish were fed ad libitum , twice per day. Feeding behaviour was determined once per week and was measured as the time taken by each fish to eat the first pellet (latency, LAT) and the total time spent feeding (total feeding time, TFT). Temperature, pH, conductivity, alkalinity, dissolved oxygen, dissolved CO 2 , nitrogen compounds (TAN, NO 2 -N and NO 3 -N), chemical oxygen demand and orthophosphate-P were measured over time. At day 57 fish were weighed, blood sampled and returned to their tanks for an extra experimental period of 15 days, and subjected at day 72 to an acute stress followed by blood sampling. Blood was analysed for glucose and cortisol. Results showed that the water quality parameters measured in the 3 RAS (with the exception of alkalinity) were still within the optimum range for growth of Nile tilapia. Large individuals showed a tendency to grow more in the LOW treatment (2.66 ± 1.35 g/kg 0.8 /day) as compared with the MIDDLE treatment (0.93 ± 1.63 g/kg 0.8 /day). On the contrary, small individuals grew significantly less in the LOW treatment (3.60 ± 1.74 g/kg 0.8 /day) as compared with the HIGH (7.22 ± 1.58 g/kg 0.8 /day) and MIDDLE (6.82 ± 4.54 g/kg 0.8 /day) treatments. Small fish were more motivated to eat (lower latency) in the MIDDLE (4.63 ± 5.24 min) as compared with the LOW treatment (8.94 ± 6.41 min). In the HIGH accumulation treatment higher glucose levels were observed in the small fish, before and after acute stress, as compared with the LOW accumulation treatment. In conclusion, this study showed that the extent to which the accumulation of substances in RAS affects growth depends on fish size: large individuals show a trend towards growth retardation in the highest accumulation RAS while small individuals, on the contrary, seem to grow better in such systems.

One of the challenges that Recirculating Aquaculture Systems (RAS) are still facing is the risk that in RAS fish grow less than in flow-through systems due to the accumulation of substances originating from feed, fish or bacteria associated with the water re-use. The present study investigated whether RAS with high and low accumulation levels of these substances affect feed intake and growth of Nile tilapia Oreochromis niloticus, African catfish Clarias gariepinus, and European eel Anguilla Anguilla. One-hundred and twenty individuals of each species were used (start body weights: Nile tilapia 264.8 ± 8.3 g; African catfish 253.2 ± 2.1 g and European eel 66.6 ± 1.3 g). For a period of 39 days, growth and feed intake were compared between high and low accumulation RAS. HIGH accumulation RAS was designed for maximal accumulation of substances in the water by operating the system at nearly-closed conditions (30 L/kg feed/d), using mature biofilters and high feed loads; and (2) LOW accumulation RAS was designed to be a proxy for flow-through systems by operating at high water exchange rates (1500 L/kg feed/d), new biofilters and low feed load. HIGH accumulation RAS induced a reduction in feed intake (42%) and growth (83%) of Nile tilapia, as compared to systems that are a proxy for flow-through conditions. This effect was not observed in European eel and African catfish. The cause of this reduced feed intake and growth rate of Nile tilapia is still unclear and should be addressed in further studies.

This paper analyses nutrient conversions, which are taking place in integrated intensive aquaculture systems. In these systems fish is cultured next to other organisms, which are converting otherwise discharged nutrients into valuable products. These conversions are analyzed based on nitrogen and phosphorous balances using a mass balance approach. The analytical concept of this review comprises a hypothetical system design with five modules: (1) the conversion of feed nutrients into fish biomass, the “Fish-Biomass-Converter”; (2) the separation of solid and dissolved fish waste/nutrients; the “Fish-Waste-Processor”; (3) the conversion of dissolved fish waste/nutrients, the “Phototrophic-herbivore-Converter”; (4 and 5) the conversion of solid fish waste, the “Bacterial-Waste-Converter”, or the “Detrivorous-Converter”. In the reviewed examples, fish culture alone retains 20–50% feed nitrogen (N) and 15–65% feed phosphorous (P). The combination of fish culture with phototrophic conversion increases nutrient retention of feed N by 15–50% and feed P by up to 53%. If in addition herbivore consumption is included, nutrient retention decreases by 60–85% feed N and 50–90% feed P. This is according to the general observation of nutrient losses from one trophic level to the next. The conversion of nutrients into bacteria and detrivorous worm biomass contributes only in smaller margins (e.g. 7% feed N and 6% feed P and 0.06% feed N 0.03 × 10−3% feed P, respectively). All integrated modules have their specific limitations, which are related to uptake kinetics, nutrient preference, unwanted conversion processes and abiotic factors.

Little information is available on steroid concentrations in the rearing water of aquaculture systemsand whether they accumulate in recirculating aquaculture systems (RAS). Therefore this study aimedat determining (1) the concentrations and variation of cortisol and sex steroids in RAS, (2) the con-tribution of fish rearing conditions to steroid concentrations in seven commercial RAS. Each RAS wassampled twice at three different points: (1) make-up water; (2) influent and (3) effluent of the rear-ing unit. The results showed significant higher steroid concentrations in the influent and effluent whencompared with the make-up water. On average cortisol concentration was 15.7% higher in the efflu-ent when compared with the influent. Mean steroid concentrations in the rearing unit effluent variedbetween: 3.8–217.0 ng/L for cortisol, 3–12.5 ng/L for testosterone, 0.9–7.1 ng/L for 11-ketoteststerone and1.8–12.8 ng/L for 17,20 -dihydroxypregn-4-en-3-one. Stocking density, Total Ammonia-Nitrogen con-centration and orthophosphate-P concentration (a measure of make-up water usage) showed a positivecorrelation with sex steroids in the water. The steroid concentrations from the present study were ordersof magnitude lower than initial estimations indicating a water treatment efficiency of >99%. The resultssuggest that an intensification of fish production through decrease of make-up water use and increaseof stocking density will lead to a build-up of steroids in the water. Although intensification is critical forthe economical success of RAS, this ultimately could affect fish performance as steroids accumulates inthe water of RAS at levels that can potentially be detected by some fish species. Institution FCT – Fundação para a Ciência e Tecnologia

This review deals with the main mechanisms and parameters affecting design and performance of trickling filters in aquaculture. Relationships between nitrification rates and easily accessible process parameters, like bulk phase concentration of TAN, O2, organic matter (COD), nitrite, temperature, HCO3−, pH and the hydraulic loading of the trickling filter, are discussed in relation to the design and operation of such filters. Trickling filter design procedures are presented and one of them, a model describing the nitrification performance of trickling filters by plug-flow characteristics, is discussed in greater detail. Finally, practical aspects in relation to filter design and operation are presented.

The effluent from the drumfilter of a recirculation aquaculture system was used as substrate to produce heterotrophic bacteria in suspended growth reactors. The effects of organic carbon supplementation (0, 3, 6, 8 g/l sodium acetate) and of hydraulic retention times (11-1h) on bacteria biomass production and nutrient conversion were investigated. Bacteria production, expressed as volatile suspended solids (VSS), was enhanced by organic carbon supplementation, resulting in a production of 55-125 g VSS/kg fish feed (0.2-0.5 g VSS/g carbon). Maximum observed crude protein production was approximately 100 g protein/kg fish feed. The metabolic maintenance costs were 0.08 Cmol/Cmol h, and the maximum growth rate was 0.25-0.5 h(-1). Ninety percent of the inorganic nitrogenous and 80% of ortho-phosphate were converted. Producing bacteria on the drumfilter effluent results in additional protein retention and lowers overall nutrient discharge from recirculation aquaculture systems.