Effect of dietary saturated and monounsaturated fatty acids in juvenile barramundi Lates calcarifer

Barramundi (Lates calcarifer), a catadromous teleost of commercial interest, perform well when fed a wide range of dietary oils. However, the range of alternative oils now being explored is typically rich in saturated and monounsaturated fatty acids (SFA and MUFA). In this study, the response of juvenile barramundi (47.0 g per fish initial weight) fed isolipidic and isoenergetic diets with 82 g kg^?1 added oil was tested. The experimental test diets had a 2 : 1 or 1 : 2 ratio of SFA to MUFA (SFA‐D and MUFA‐D, respectively) compared to a control diet (CTRL‐D) fed for 8 weeks. The diets containing mostly olive oil (dietary MUFA‐D) and mostly refined palm oil (dietary SFA‐D) did not impact the growth performance or feed utilization parameters of the barramundi. The in vivo beta‐oxidation activity was consistent with the dietary fatty acid composition, with the most dominant FA being heavily beta‐oxidized. Together, the in vivo whole‐body mass balance of fatty acids showed that n‐3 long‐chain polyunsaturated fatty acids (LC‐PUFA) were most efficiently utilized in the SFA‐D‐ and MUFA‐D‐fed fish. This study provides evidence that additional dietary MUFA and SFA are suitable lipid classes for juvenile barramundi and they are both equally efficient at sparing LC‐PUFA from an oxidative fate.     

Comparison of faecal collection methods and diet acclimation times for the measurement of digestibility coefficients in barramundi (Lates calcarifer)

This study aimed to investigate the effects of two faecal collection methods (stripping and settlement) on the apparent digestibility coefficients (ADC) of dry matter, protein and energy of three different diets fed to barramundi. In a second experiment, the effect of acclimation time (i.e. number of days fed the diet) on the calculation of ADCs was also investigated. Each tank of fish was fed one of three diets for 12 days. Faeces were collected by both stripping and settlement, although only settlement was used prior to day seven of the acclimation period. Faeces were collected using the settlement method at regular intervals from day 1 to day 12. Comparisons between faecal collection methods were only made based on faecal material collected over a similar acclimation period. The collection of faeces by stripping produced more conservative ADCs, which were also more consistent than those obtained using the settlement technique. The calculated ADCs typically fluctuated for the first 3 days of collection before the variability diminished. Barramundi should be acclimated to diets for a minimum of 4 days before collection of faecal material, and collection by stripping is recommended to obtain the most reliable digestibility data.     

Effects of two commercial microdiets on growth and survival of barramundi (Lates calcarifer Bloch) larvae within various early weaning protocols

Because of high costs and labour requirements along with the highly variable nutritional value of live feeds, we investigated the possibility of early weaning for barramundi (Lates calcarifer Bloch) larvae aimed at reducing the use of Artemia. Two commercial microdiets, Gemma Micro (Skretting, Australia) and Proton (INVE, Belgium) were compared for growth and survival of larvae using three weaning protocols, until 33 days posthatch (dph). Enriched rotifers were fed to larvae in all protocols through mouth opening until 21, 18 and 30 dph (protocols 1, 2 and 3, respectively). At 13 dph, enriched Artemia metanauplii were introduced to weaning protocols 1 and 2, and continued until 29 and 24 dph, respectively, whereas protocol 3 did not receive Artemia. Microdiet was initiated at 20, 16 and 13 dph in protocols 1, 2 and 3, respectively. Barramundi larvae grew successfully to 33 dph when co‐fed rotifers and microdiet, and significantly larger larvae resulted from feeding Gemma Micro rather than Proton, when Artemia were not used. However, larvae weaned onto Proton using a longer period of Artemia provision were significantly larger than larvae reared according to all other protocols. Survival was significantly higher in all Gemma Micro protocols when compared with Proton protocols. This was in part due to higher cannibalism when using Proton compared with Gemma Micro (22.8 ± 0.9% and 9.2 ± 0.6%, respectively). Cannibalism became more noticeable in all protocols when the larvae reached 7–8 mm standard length and further increased after the cessation of live feed. Tank biomass production was the highest when larvae were weaned onto Gemma Micro including a short period of Artemia provision as a result of a combination of high growth and survival. However, similar biomass production resulted when larvae were weaned directly from rotifers onto Gemma Micro and/or from a prolonged Artemia period onto proton. The success of weaning barramundi larvae directly to microdiet from rotifers, thus eliminating the need for Artemia, was influenced by the microdiet. Relatively higher levels of free amino acids and lipids were believed to contribute to increasing larval growth and survival. Larvae that were fed Gemma Micro showed higher growth when Artemia were utilized for a shorter period, while Proton‐fed larvae benefited from an extended Artemia feeding period.     

Effects of dietary fish oil replacement on growth and carcass proximate composition of juvenile barramundi (Lates calcarifer)

An experiment was carried out to determine the effects of dietary fish oil replacement at an optimal level on the growth and carcass proximate composition of juvenile barramundi or sea bass (Lates calcarifer). Ten fish feeds were formulated to contain iso‐ingredients with theoretically identical dietary lipid levels but with different sources of lipids (fish oil, soybean oil, canola oil and linseed oil) replacing dietary fish oil. Three hundred fish were equally divided into 30 70‐L plastic tanks (three replicate tanks for each treatment). Fish were fed with experimental diets for 40 days within closed recirculating freshwater systems (70 L h^?1, 28°C and 12L:12D). Growth, specific growth rate, food conversion ratio (FCR) and carcass proximate composition were determined. Dietary lipids significantly affected (P<0.05) the growth of the fish. Good growth and low FCR were observed in all treatments. Therefore, fish oils can be partially replaced by vegetable oils to reduce the feed cost. There were no significant differences (P>0.05) in lipid, moisture and protein content in fish carcasses among the group. However, juvenile barramundi showed increasing lipid and moisture content in muscle, whereas decreasing protein content when compared to the initial fish.     

A determination of the quantitative requirements for docosahexaenoic acid for juvenile barramundi (Lates calcarifer)

A series of diets with varying docosahexaenoic acid (DHA; 22:6n‐3) inclusion levels (1 g kg^?1 3 g kg^?1, 6 g kg^?1, 10 g kg^?1, 15 g kg^?1 and 18 g kg^?1) were fed to juvenile barramundi (Lates calcarifer) for 6 weeks. Two additional diets examined the addition of arachidonic acid (ARA; 20:4n‐6) or eicosapentaenoic acid (EPA; 20:5n‐3) to the diets at 10 g kg^?1 when DHA was also included at 10 g kg^?1. Fish were fed the diets on a pair‐fed feeding regime to eliminate feed intake variability. Fish were weighed, and blood and tissue samples were collected after 6 weeks. Behavioural parameters were also assessed. Improvement in growth was seen with increasing inclusion of DHA up to a maximum at 10 g kg^?1 inclusion, albeit the response was minor. However, the addition of ARA to the diet reduced the growth response, while the addition of EPA improved the growth response. An improvement in feeding behaviour was also seen with increasing DHA up to a peak at 10 g kg^?1, while those animals fed diets low in DHA showed increasingly cryptic behaviour. With the increasing inclusion of DHA, a range of pathologies were observed, but the addition of an EPA component to the diet limited these pathologies, while the addition of ARA made little improvement and in some cases exacerbated the pathologies.     

Potential of meat meal to replace fish meal in extruded dry diets for barramundi, Lates calcarifer (Bloch). II. Organoleptic characteristics and fatty acid composition

The organoleptic quality of barramundi fed for 66 days on pelleted diets containing varying amounts of fish meal and meat meal was determined in two experiments (E1 and E2). Each compared four diets: a 430 g kg^?1 crude protein (CP) control diet (containing 35% Chilean anchovy fish meal); two diets containing high inclusions (40% or more) of meat meal; and a proprietary barramundi diet. In E1, the two meat meal diets contained 10% Chilean fish meal whereas the two meat meal diets in E2 had no marine protein ingredients. Panellists identified and rated the colour of flesh, and scored odour, flavour and texture characteristics and overall liking on structured graphic line scales (0–100). Fish fed the high‐meat meal diets were sweeter and firmer than those fed the high‐fish meal control diet in E1 (P < 0.05). Scores for fishy flavour were also highest for the meat meal diets and lowest for the proprietary diet. In both E1 and E2, scores were high (> 60) for overall liking and low (< 10) for undesirable odours and tastes. Exclusion of all sources of marine protein from the diet in E2 did not detract from the sensory value of the fish. The influence of diet on the fatty acid profile of the fish was examined in E2. Compared with fish fed the control diet, the neutral lipid fraction of those fed the meat meal diets had higher proportions of saturated and short‐chain monounsaturated fatty acids at the expense of longer chain fatty acids, especially 22:6n‐3. Polar lipids showed only subtle dietary effects, which were confined to the long‐chain unsaturated fatty acids.     

Potential of meat meal to replace fish meal in extruded dry diets for barramundi, Lates calcarifer (Bloch). I. Growth performance

Juvenile barramundi (～220–280 g start weight) were fed extruded dry‐pelleted diets containing varying amounts of fish meal and meat meal in three experiments (E). E1 and E2 were each 66‐day farm studies utilizing 16 floating cages (400 fish per cage) in an aerated freshwater pond. E3 examined the same diets as fed in E2 but under controlled water temperature (28 ± 0.7 °C) and photoperiod (12:12) laboratory conditions in a 42‐day study involving 24 aquaria (eight fish per aquarium). In all studies, the same 430 g kg^?1 crude protein (CP), 15 kJ g^?1 digestible energy (DE) control (Ctl) diet (containing 35% Chilean anchovy fish meal) was compared with two high‐inclusion meat meal diets and a proprietary diet. The meat meal diets evaluated in E1 were a high‐ash (260 g kg^?1) meat meal that contained 520 g kg^?1 CP and a low‐ash (140 g kg^?1) meat meal that contained 600 g kg^?1 CP when included at either 450 or 400 g kg^?1, respectively, in combination with 100 g kg^?1 Chilean fish meal in diets that were isonitrogenous and isoenergetic with the Ctl diet. Growth rates and feed conversions were similar (P > 0.05) for all diets. In E2 and E3, the 520 g kg^?1 CP meat meal was included at 500 g kg^?1 without any marine protein source in diets formulated to provide either 15 or 16.2 kJ g^?1 DE and the same CP/DE ratio (29 mg kJ^?1) as the Ctl diet. Fish performance ranking of diets was similar in both experiments, with the 16.2 kJ g^?1 DE diet supporting better (P < 0.05) growth rates than the Ctl diet and feed conversion ratios equivalent to the Ctl diet but better (P < 0.05) than all other diets.     

Effects of sodium diformate on growth performance,gut microflora,digestive enzymes and innate immunological parameters of Asian sea bass (Lates calcarifer) juveniles

This study was conducted to evaluate the effects of dietary sodium diformate (NaDF) on growth performance, gut microflora, digestive enzyme activities and immune response parameters of Asian sea bass (Lates calcarifer) juveniles. Fish with initial weight of 12.5 ± 0.4 g were fed with five experimental diets contained 0.0 (control), 5, 10.0, 15.0 and 20 g NaDF kg^?1 in triplicate for 6 weeks. Fish fed diet containing 5 g NaDF kg^?1 had significantly the highest final body weight and feed intake among different treatments. The gut total viable bacterial counts gradually decreased with increasing dietary NaDF level. Specific activity of chymotrypsin improved in fish fed diets administered with NaDF compared to the control group. Fish fed 5 g NaDF kg^?1 diet showed the highest serum lysozyme level among different treatments. The serum classical pathway activity of complement showed higher level in fish fed diets contained 5 and 10 g NaDF kg^?1 than other groups. According to break‐point regression method analysis, the optimum inclusion of dietary NaDF in L. calcarifer juveniles was estimated between 4.6 and 5.1 g/kg, when specific growth rate and feed conversion ratio were plotted against dietary NaDF levels.     

Dietary composition manipulation to enhance the performance of juvenile barramundi (Lates calcarifer Bloch) reared in cool water

Barramundi Lates calcarifer reared in cool water (20–22°C) grow slowly and feed is used poorly compared with fish in warm water (28–32°C). Two comparative slaughter growth assays were carried out with juvenile barramundi to see if increasing the digestible energy (DE) and/or the n‐3 highly unsaturated fatty acid (n‐3 HUFA) content of the feed would improve growth of fish raised in cool water. Increasing the DE content of the feed from 15 to 17 or 19 MJ kg⁻¹ while maintaining a constant protein to energy ratio in Experiment 1 brought about significant improvements in feed conversion ratio (FCR) (from 2.01 to 1.19) and daily growth coefficient (DGC; from 0.69 to 1.08%/day) for fish at 20°C. For fish at 29°C, improvements, while significant, were of a lesser magnitude: from 1.32 to 0.97 for FCR and from 3.24 to 3.65%/day for DGC. Increasing the absolute amount of dietary n‐3 HUFA, expressed as the sum of eicosapentaenoic and docosahexaenoic fatty acids, from 0.5% to 2.0% in Experiment 2 improved DGC linearly and FCR curvilinearly for fish at 29°C whereas at 20°C, DGC was not affected while FCR improved slightly (from 1.83 to 1.68). Feed conversion ratio was optimized with a dietary n‐3 HUFA of about 1.5%. Providing barramundi with a feed that is high in DE (viz 19 MJ kg⁻¹) and a digestible protein to DE ratio of 22.5 g MJ⁻¹ is a practical strategy for improving the productivity of barramundi cultured in cool water whereas increasing dietary n‐3 HUFA conferred very little additional benefit.     

Estimating reproductive success of brooders and heritability of growth traits in Asian sea bass (Lates calcarifer) using microsatellites

Asian sea bass (Lates calcarifer) is one of the most important marine food fish species in Asia and Australia. To estimate the reproductive success of broodstock and heritabilities of growth‐related traits, two independent full‐factorial crosses (PI and PII) were created by crossing 10 males and 10 females. At 90 days post hatch (dph), the body weight (BW) and total length (TL) of 804 individuals from PI and 900 individuals from PII were measured and tissues samples of each fish were collected. Parents and offspring were genotyped with nine polymorphic microsatellites. Out of 1704 offspring from the two crosses, 98.7% were assigned to their parents. In PI, 19 of 20 parents produced offspring, while in PII, only five parents contributed to offspring. Low contribution of parents to offspring could lead to reduced genetic variation in the next generation. Heritabilities for growth‐related traits were estimated using the pedigrees reconstructed using microsatellite genotypes. The estimates of heritability were 0.22±0.16 and 0.25±0.18 for BW, 0.31±0.14 and 0.24±0.21 for TL and 0.22±0.22 and 0.15±0.09 for Fulton's condition factor in the two crosses respectively. Body weights at 90 dph and at harvest (289 dph) were significantly correlated (r=0.601, P<0.01). Therefore, growth‐related traits could be improved by exploiting additive genetic effects through selective breeding, and broodstock candidates could be selected early in the production cycle.     

Partial utilization efficiencies of protein and methionine by barramundi (Lates calcarifer) in response to dietary methionine source and form

An experiment was conducted with barramundi (Lates calcarifer) juveniles (initial weight 10.3 g ± 0.3 g) to examine the partial efficiency of utilization of methionine (Met) from intact protein (fishmeal or lupin protein concentrate) and a crystalline DL‐Met source. Fish were fed at one of three ration levels: Low (0.4 g/fish/day), Moderate (0.8 g/fish/day) and High (1.6 g/fish/day). Those fed the fishmeal‐based diet (Diet FML) at the highest ration level grew to an average weight of 37.3 ± 0.46 g, whereas those fed the Lupin Protein Concentrate (LPC)‐based diet fortified with all EAA (Diet LPCM) at the highest ration level grew to 25.4 ± 2.27 g. The weight of the fish fed the LPC diet with no additional Met (Diet LPC) even at the highest ration level declined over the course of the experiment resulting in a final weight of 9.2 ± 0.88 g, clearly demonstrating the impact of dietary Met deficiency. The partial efficiency of protein utilization was also significantly reduced when Met was limiting (a coefficient of 0.06, compared to 0.39 in the Met‐supplemented LPC diet). The results suggested that the partial efficiencies (coefficients) of both Met and protein utilization in diets where crystalline Met is the primary source of Met (Diet LPCM: 0.26 and 0.39 respectively) were significantly poorer than from an intact protein source (Diet FML: 0.89 and 0.67 respectively) when Met is provided in excess.     

CRISPR-based platform for rapid,sensitive and field-deployable detection of scale drop disease virus in Asian sea bass (Lates calcarifer)

Scale drop disease virus (SDDV) is a major pathogen of Asian sea bass that has emerged in many countries across the Asia Pacific since 1992 and carries the potential to cause drastic economic losses to the aquaculture sector. The lack of an approved vaccine for SDDV necessitates timely prevention as the first line of defence against the disease, but current diagnostic platforms still face challenges that render them incompatible with field applications, particularly in resource-limited settings. Here, we developed a novel detection platform for SDDV based on a CRISPR-Cas12a-based nucleic acid detection technology combined with recombinase polymerase amplification (RPA-Cas12a). Using the viral adenosine triphosphatase (SDDV-ATPase) gene as a target, we achieved the detection limit of 40 copies per reaction and high specificity for SDDV. The coupling with fluorescence and lateral flow readouts enables naked-eye visualization and straightforward data interpretation requiring minimal scientific background. Compared with semi-nested PCR in field sample evaluation, our RPA-Cas12a assay is more sensitive and capable of detecting SDDV in asymptomatic fish. Importantly, the entire workflow can be carried out at a constant temperature of 37°C within an hour from start to finish, thus removing the need for an expensive thermal cycling apparatus and long turnaround times associated with PCR-based methods. Therefore, owing to its high accuracy, rapidity and user-friendliness, the developed RPA-Cas12a platform shows the potential for diagnosis of SDDV at point of need and could be a valuable tool to help protect fish farming communities from large-scale epidemics.     

The effect of culture on morphology, lipid and fatty acid composition, and polyunsaturated fatty acid metabolism of rainbow trout (Oncorhynchus mykiss) skin cells

Rainbow trout (Oncorhynchus mykiss) skin cell cultures were obtained by trypsinization of the tissue and grown in Leibovitz L-15 medium. Lipid class compositions, and fatty acid profiles of total lipids and individual phospholipid classes were determined at different times of culture. The metabolism of polyunsaturated fatty acids (PUFA) was investigated by incubating primary cultures after 7 and 14 days with [1-¹⁴C]18:2n-6 and [1-¹⁴C-]18:3n-3. The change in morphology between epithelial-like primary cultures and fibroblastic-like secondary subcultures was accompanied by alterations in the lipid composition. Polar lipids became predominant by 14 days in culture. The relative proportions of phosphatidylcholine (PC), the most abundant phospholipid, phosphatidylinositol and cholesterol increased significantly, while sphingomyelin decreased. Saturated fatty acids, 18:1n-9, n-6 and n-9PUFA were more abundant in total lipid in cultures at 14 days and 4 months than in cells initially isolated which contained higher percentages of longer chain monoenes and n-3PUFA. The changes in fatty acid composition with time in culture were observed in all the major phospholipid classes. Rainbow trout skin cells in culture desaturated and elongated both 18:2n-6 and 18:3n-3, with 20:4n-6 and 20:5n-3 being the most abundant products, respectively. PC presented the highest incorporation of radioactivity, especially following incubation with 18:3n-3. Lipid metabolism in general increased with the age of primary cultures, with both the amount of C18 PUFA incorporated and metabolized by desaturation/elongation significantly increased in 14 day cultures compared to 7 day cultures. Product/precursor ratios calculated for both n-6 and n-3 fatty acids showed that, while 6 desaturase activity was increased significantly with cell age, 5 desaturase activity was more affected by the fatty acid series, with 18:3n-3 being more readily transformed to 20:5n-3 than 18:2n-6 to 20:4n-6. Further desaturation of 20:5n-3 to hexaenes was low. Overall, the data suggested that the trout skin cell cultures were more similar to mammalian skin fibroblasts than mammalian epidermal/keratinocyte cultures.     

The effect of taurine supplementation to a plant‐based diet for barramundi (Lates calcarifer) with varying methionine content

The effect of variable taurine inclusion (Tau) (1 g/kg DM to 15 g/kg DM) in the diet of juvenile barramundi (Lates calcarifer) on growth and nutrient utilization was investigated at three levels of dietary methionine (Met) supplementation. Diets were fed to juvenile barramundi (starting weight: 26.8 g) twice daily under a restricted pair‐fed regime for a period of 42 days. No significant effect of dietary Tau supply on growth or nutrient utilization was observed at any level of Met inclusion. Numerical variations suggested a positive effect of Tau provision at the mid‐level of supplementation (6–8 g/kg DM). The best‐fit response model (5‐SKM), fitted to the percentage body weight gain data of fish fed diets with an adequate level of Met, suggested a relatively weak pattern of response (R² = 0.183) and predicted a Tau requirement of 5.47 g/kg DM (9.64 g/kg CP) similar to that reported for several other species. It was concluded that taurine appears to be conditionally essential to barramundi, whereby provision in the diet when sulphur amino acid supply is insufficient to meet biosynthetic demands may be beneficial, but that the predicted requirement is likely reasonably accurate for use as a minimum level of inclusion.     

The influence of a dietary lipid source on growth, muscle fatty acid composition and erythrocyte osmotic fragility of hybrid tilapia

An 8-week feeding trial was conducted to determine the effects of various dietary lipids on the growth, tissue proximate composition, muscle fatty acid composition and erythrocyte osmotic fragility of red hybrid tilapia, Oreochromis sp. Five isonitrogenous and isoenergetic semipurified diets were supplemented with 10% of either cod liver oil (CLO), sunflower oil (SFO), crude palm oil (CPO), crude palm kernel oil (CPKO), or a combination of 5% CLO with 5% palm fatty acid distillates (PFAD), respectively. There were no significant effects (P > 0.05) of diet on growth but fish fed the CLO diet showed a significantly (P< 0.05) poorer feed efficiency ratio compared to fish fed the CPO diet. Lipid deposition in fish muscle was mostly similar among fish fed the various diets but bone ash was significantly higher in fish fed the CPO and CPKO diets. Muscle lipids of fish fed palm oil-based diets did not increase in saturated fatty acids content but showed significantly lower polyunsaturated fatty acid (PUFA) concentrations compared to fish fed the CLO diet. The concentrations of individual PUFA in muscle lipids were strongly influenced by dietary PUFA concentrations. Dietary lipids did not markedly affect the structural integrity of erythrocyte membranes but the erythrocytes of tilapia fed the CPO diet were slightly more resistant to osmotic lysis. It was concluded that palm oil products, especially CPO, could be successfully used in the diet of hybrid tilapia based on its availability, cheaper costs and its potential ability to enhance oxidative stability due to its low PUFA content and high natural concentrations of antioxidants.     

Influence of fish size and water temperature on the metabolic demand for oxygen by barramundi, Lates calcarifer (Bloch), in freshwater

Oxygen demand by all animals is driven primarily by their needs for sustaining metabolism. Typically, larger animals require more oxygen and cellular fuel to carry out respiration than smaller animals. This relationship in most cases is not linear and is usually described by a coefficient and exponent (e.g. ax^b): the exponent b showing the relationship between live‐weight and energy/oxygen demand and is often termed the metabolic body weight (MBW) exponent, while the coefficient (a) tends to be temperature specific and describes the relationship between MBW and maintenance metabolic energy and oxygen demand at that specific temperature. Across all temperatures (range 26.0–32.0°C), the relationship between barramundi (Lates calcarifer) live‐weight (x; g) and relative oxygen consumption as standard metabolic rate (y; mg O₂ kg^?1 h^?1) at 29.4±1.5°C (mean±SD) was described by the exponential curve: y=710.19 x^?0.3268, R²=0.6875 (n=222 assessments). Examination of the same data but on a gross oxygen consumption (mg O₂ h^?1) basis showed a relationship between live‐weight (x; g) and gross oxygen consumption (y; mg O₂ h^?1) that was described by the exponential curve: y=0.710 x^0.6732, R²=0.9033. Evaluation of the combined relationship between fish live‐weight (y; g) and water temperature (x; °C) on gross oxygen consumption rate (z; mg O₂ h^?1) was described by the equation: z=(?20.7818+1.4017x?0.0227x²) ×y^0.673.