Growth,tissue metabolites and digestive functionality in Sparus aurata juveniles fed different levels of macroalgae,Gracilaria cornea and Ulva rigida

The aim of this study was to evaluate the inclusion of two macroalgae as dietary ingredients on growth, body composition, intermediate metabolism, intestinal enzymes and gut histology of Sparus aurata juveniles. Six isocaloric and isonitrogenous diets incorporating 5%, 15% or 25% of Gracilaria cornea or Ulva rigida meal were tested against a control diet. Duplicate groups of fifteen fish (14 g) were fed the experimental diets for 70 days. Growth performance showed an inverse linear relationship with the inclusion level of Gracilaria. Feed conversion ratio increased significantly in fish fed 25% Gracilaria. Ulva presented a quadratic relationship with the lowest SGR at 15% of inclusion. Fish fed 25% Ulva exhibited significantly lower lipid content than those fed control diet. Overall, metabolites did not show differences with specimens fed control diet, except for plasmatic glucose and muscle lactate. Intestinal proteolytic activity was reduced with the macroalgae inclusion level, but all fish showed the same enzyme pattern. The histological study revealed no negative effect on liver and intestine structure by the macroalgae inclusion. The present work demonstrated that these ingredients can be used in practical diets for sea bream, but a more comprehensive research is needed to determine the optimum dietary level in a longer feeding trial.     

摄食不同饵料的大口黑鲈肠道菌群分析

为了探究投喂冰鲜杂鱼和人工配合饲料对大口黑鲈肠道菌群的影响,本研究采用变性梯度凝胶电泳技术(PCR-DGGE)对摄食不同饵料的2组鲈鱼肠道内容物的细菌群落结构进行了比较分析.结果显示,冰鲜组与饲料组的大口黑鲈肠道菌群组成相似性较低(相似性24％),且冰鲜组肠道菌群多样性高于饲料组.对2组大口黑鲈中共有条带和特异性条带进一步测序分析显示,2组大口黑鲈肠道共有菌有红螺菌、梭菌、保科爱德华菌、嗜麦芽窄食单胞菌和蓝细菌.冰鲜组大口黑鲈肠道中的特异性菌归属为拟杆菌、肉食杆菌、魏斯氏菌和邻单胞菌;饲料组大口黑鲈肠道中的特异性菌归属为黄杆菌、鲸杆菌、柠檬明串珠菌、缺陷短波单胞菌.研究表明,与投喂冰鲜杂鱼的大口黑鲈相比,人工配合饲料降低了大口黑鲈肠道菌群多样性,也抑制了拟杆菌等有益菌在大口黑鲈肠道内的分布.推测饲料组大口黑鲈生长较慢可能与其肠道菌群中拟杆菌与厚壁菌的比值下降有关.研究结果提示,在今后大口黑鲈养殖或者人工配合饲料研制过程中,添加拟杆菌等益生菌也许可促进大口黑鲈的生长和营养吸收.     

Influence of dietary soybean meal levels on growth, feed utilization and gut histology of Egyptian sole (Solea aegyptiaca) juveniles

The present study was undertaken in order to determine the effect of a dietary incorporation of soybean meal (SBM) on growth performance, nutrient utilization and gut histology of Egyptian sole Solea aegyptiaca. This species, which is genetically and morphologically close to Solea solea, is currently present in the Mediterranean Sea and reared in some Italian farms, using the same techniques for S. solea production and it could represent a promising new species for Mediterranean aquaculture.Three isoproteic and isolipidic extruded diets (crude protein, 47%, crude fat, 20%) were formulated containing 0 (diet 1), 18 (diet 2) and 30% SBM (diet 3). 540 juveniles (initial body weight of 6.9 ± 0.2 g) were equally distributed into nine 150 l squared tanks (bottom surface: 520 cm²) connected with a recirculation system (temperature 20 ± 1 °C and dissolved oxygen above 6 ppm). Animals have been hand-fed twice a day (at 9.00 a.m. and 5.00 p.m.) at a daily ratio of 1.5% body weight^− 1. Diets were tested in triplicate for 87 days. The fish were weighed at days 28, 57 and 87, and they were sampled at days 0, 57 and 87 for nutrient retention determination. One-way ANOVA with Newman-Keuls' post-test were used to analyze data (P ≤ 0.05).No differences in terms of palatability were observed and any diet has been consumed completely all throughout the experiment. At day 87, fish fed diet 1, 2 and 3 reached weights of 25.8 ± 0.8 g, 27.5 ± 0.9 g and 26.1 ± 1.5 g, respectively and no differences in performances or nutrient retention were found.Histopathological gut examinations have revealed no noticeable differences in the appearances of the intestines between any of diet groups. Intestinal mucosal cells presented well vacuolated upper, with well defined microvillar brush borders. On the basis of the results of this trial, SBM seems to be a good protein source for Egyptian sole and can be added in the diet up to 30% without any reduction in growth rate and no adverse effect on gut histology.     

Effects of the partial substitution of dietary fish meal by differently processed soybean meals on growth performance, nutrient digestibility and activity of intestinal brush border enzymes in the European sea bass (Dicentrarchus labrax)

This study evaluated the effects of replacing fish meal (FM) in the diet of European sea bass (Dicentrarchus labrax) with differently processed soybean meals on growth performance; apparent digestibility coefficients (ADCs); the activity of alkaline phosphatase and the brush border digestive enzymes: leucine amino peptidase, maltase and γ-glutamyl transpeptidase; and the fish morphometric and flesh quality. European sea bass with an initial average body weight of 187.8 ± 1.4 g, were fed either a FM based (control) diet or a soy replacement isoproteic, isolipidic and isocaloric diet. The treatments consisted replacing FM protein by: 25% toasted, dehulled and solvent-extracted soybean meal (SE25); 50% dehulled and toasted soy seeds subjected to dry extrusion and mechanical oil extraction (ME50); 50% enzyme-treated soybean meal (ET50) and an inclusion of 60% composed of 30% toasted, dehulled and solvent-extracted soybean meal and 30% enzyme-treated soybean meal (SE + ET/60). ADCs values of dry matter, crude protein, crude lipid and gross energy did not significantly differ between ME50 and SE + ET/60 but both of them were inferior to the other treatments (P < 0.05). Compared with the control FM-based diet, only the group that consumed the ME50 diet had lower specific growth rate, feed conversion ratio and gross protein retention efficiency values (P < 0.05). The protein efficiency ratio did not differ between the FM-based control group and all other soy derivate groups (P > 0.05). In order to gain more insight on the effects of the diet on fish growth, activity of the brush border enzymes was measured. No significant differences were found in leucine amino peptidase and maltase activities, between the control group and the other soy derivate groups. The activity of γ-glutamyl transpeptidase was significantly higher in the upper section in fish that consumed the control diet compared with fish that consumed the soy derivate diets. Yet, the main activity of this enzyme was found in the lower intestinal section. In all the soy derivate diets except for the SE25 diet, the activity of alkaline phosphatase was significantly lower than its activity in the control diet. Feeding diets that include various types and levels of soybean derivatives did not affect the whole body composition and slaughter yield but significantly reduced liver weight. The results show that in methionine supplemented diets, FM can be replaced in terms of protein by 25% SE, 50% ET or 60% soy composed of 30% SE and 30% ET without hampering fish performance. The inclusion of 50% protein from ME resulted in significantly lower growth performance and feed utilization.     

Alternative pathways in the life history of Gracilaria gracilis (Gracilariales, Rhodophyta) from north-eastern Sicily (Italy)

The reproductive phenology of Gracilaria gracilis has been followed closely in a brackish water lake in Sicily. The life history of this red alga deviated from the typical Polysiphonia-type cycle and was characterized by the predominance of the tetrasporophytic phase over gametophytes, in situ development of germinated gametophytes and the presence of sterile pleustophytic thalli. Release of spherical agglomerates of cells produced by young tetrasporophytes was observed as well. Their ability to germinate forming new thalli appears to represent an alternative reproductive strategy of this organism. The population structure appears to reflect a predominance of the diploid phase except during the warmer summer season when only vegetative juveniles were found in the lake. Release and early stages of development of carpospores and tetraspores were critically observed in culture. A Dumontia type pattern of germination was observed for both types of spores, showing an erect thallus arising from a multicellular basal disk. Compared to tetraspores, carpospores are less readily released resulting in a subsequent delay in the development of new gametophytes.     

The effects of fish hydrolysate (CPSP) level on Octopus maya (Voss and Solis) diet: Digestive enzyme activity, blood metabolites, and energy balance

As has been demonstrated in previous studies, Octopus maya can be fed on artificial diets. In the present study six different diets were assayed. Five diets were designed to test the effect of percentage of inclusion of fish protein concentrate (CPSP: 0, 5, 10, 15, and 20%) and were offered to octopuses as a specifically designed artificial diet. The sixth diet consisted of frozen crab (Callinectes spp) and was used as control diet. Blood metabolites and energy budget of octopuses were evaluated to determine how CPSP levels modulate the digestive capacity and allow retaining energy for growth. Wild animals (316.4 ± 9.8 g) were used in the study. Results showed that CPSP produced a positive specific growth rate (SGR, % day^− 1) with high value in octopuses fed 15% CPSP level. A maximum growth rate of 0.86% day^− 1 was recorded in these animals, a value that is extremely low when compared with the SGR obtained when animals were fed fresh crab (3.7% day^− 1). In general, blood metabolites were affected by diet composition, indicating that some metabolites could reflect the nutritional and/or physiological status of octopus. Preliminary reference values for O. maya fed crab were found for glucose (0.09 ±0.02 mg/ml), lactate (0.004 ± 0.002 mg/ml), cholesterol (0.16 ± 0.02 mg/ml), acylglycerol (0.14 ± 0.01 mg/ml), protein (0.37 ± 0.04 mg/ml), hemocyanin (1.85 ± 0.04 mmol/l), and digestive gland glycogen (1.86 ± 0.3 mg/g). Total energy content can be used as an indicator of tissue metabolic reserves. In the present study, higher energy content in the digestive gland and muscle was observed in octopuses fed crab, followed by animals fed 15% CPSP. Results from the digestive gland indicated that the retained energy derived from glycogen, suggesting that lipids and protein were the main sources of variation linked with energy content. In general, digestive gland proteases activity and trypsin were induced in octopuses fed 15% CPSP. The capacity of O. maya juveniles to adjust their digestive enzymes to different types of food was evidenced. Essential amino acid content (EAA) of the diet was not a limiting factor. When dietary EAA profiles were compared with O. maya EAA profiles, all dietary EAA resulted in a higher concentration than whole body octopus composition. In the present study, all experimental groups ingested between 3300 and 4106 kJ wk^− 1 kg^− 1 without statistical differences among treatments, indicating that experimental diets were as attractive as crab. Differences were recorded in the proportion of absorbed energy (Ab, %) between CPSP-based and crab meat diets, suggesting digestion limitations associated with artificial diets. The present results indicate that the 15% CPSP diet had characteristics that stimulate digestive enzymes and reduce energetic costs associated with its digestion (HiE or SDA), channeling more biomass production than the other experimental diets.     

饲料中不同维生素A含量对花鲈生长和血清生化指标的影响

以酪蛋白和明胶为蛋白源,鱼油和豆油为脂肪源,糊精为糖源并添加氨基酸混合物配制精制基础饲料,通过向基础饲料中添加维生素A醋酸酯使饲料中的维生素A含量达到0、1 000、2 000、4 000、8 000和16 000 IU/kg(实测含量为245、1 005、1 835、3 781、7 583和15 273 IU/kg),配制出6种等氮等能的实验饲料(饲料1～6).以花鲈为实验对象,在室内流水系统中进行为期9周的生长实验,探讨精制饲料中不同维生素A含量对花鲈生长和血清学生化指标的影响.结果表明:摄食不添加维生素A组(饲料1)饲料的花鲈在养殖后期表现出较高的死亡率,并出现眼球浮肿、膨胀、鳍基充血以及鳃盖扭曲等维生素A缺乏症,而在其他维生素A添加组(饲料2～6)并未出现类似症状.体成分分析表明:饲料中不同维生素A含量会显著增加花鲈体脂肪含量,但对蛋白质和水分的含量没有显著影响.对花鲈血清指标分析表明:饲料添加维生素A含量会显著增加花鲈血清碱性磷酸酶(AKP)、甘油三酯(TG)和胆固醇(TC)含量,但显著降低丙氨酸氨基转移酶(ALT)和天冬氨酸氨基转移酶(AST)活力.采用折线模型,当以增重率为评价指标时,花鲈对饲料中维生素A的需求量为1 934.8 IU/kg;当以肝脏维生素A含量为评价指标时,花鲈对饲料中维生素A的需求量为3 546.6 IU/kg.     

Effect of dietary taurine and cystine on growth performance of juvenile red sea bream Pagrus major

Two experiments were conducted to investigate the effect of dietary taurine and cystine on growth and body composition of juvenile red sea bream Pagrus major. In Experiment I, a casein-based semi-purified diet included a small amount of fish meal were supplemented with taurine at the levels of 0 (control) and 1.0%. The experimental diets in Experiment II were without fishmeal and supplemented with taurine at 0 (control), 0.5, 1.0 and 2.0% or cystine at 1.0 and 2.0%. These diets were fed three times a day for 6 weeks to fish (average body weight: 2.3 g in Experiment I and 2.5 g in Experiment II). In Experiment I, fish fed the taurine-supplemented diet showed significantly (P < 0.05) improved growth, feed efficiency and feed consumption relative to fish fed the unsupplemental diet. The whole body taurine content increased, whereas the non-essential amino acid contents decreased, in fish fed the taurine-supplemental diet compared to fish fed the unsupplemented diet. In Experiment II, the growth, feed efficiency and feed consumption of fish fed the taurine-supplemented diets, irrespective of the dietary taurine levels, were significantly higher than those of fish fed the control diet and the cystine-supplemented diets. Taurine content in the whole body increased with the dietary taurine level, while the taurine contents did not increase by the supplemental cystine. Other free amino acid contents in the taurine-supplemented diet groups followed similar trends to those in Experiment I. These results indicate that supplemental taurine to a casein-based semi-purified diet at more than 0.5% improved the growth and feed performance of juvenile red sea bream. It is also suggested that juvenile red sea bream cannot metabolize cystine into taurine.     

Successful culture of larvae of Litopenaeus vannamei fed a microbound formulated diet exclusively from either stage PZ2 or M1 to PL1

In three separate experiments, harpaticoid copepods Tisbe monozota (alive and dead) and a microparticulate microbound diet were evaluated as alternatives to live Artemia nauplii as food, beginning at either stage PZ2 or M1, in the larval culture of Litopenaeus vannamei. Larvae were cultured in 2 L round bottom flasks at a density of 150 L^− 1 (Experiment 1) and 100 L^− 1 ( 3.2 and 3.3) at 28 °C, 35‰ salinity and 12:12 LD photoperiod, and fed 4×/day^- 1. Larvae were initially fed a mixture of phytoplankton to stages PZ2 or M1 and then fed either live Artemia, live or dead copepods, or a microparticulate microbound diet. The experiments were terminated and all larvae were harvested when more than 80% of larvae had molted to postlarvae 1 (PL1) within any flask representing any of the treatments. The comparative value of the different diets and feeding regimes was determined by mean survival, mean dry weight and total length of individual larva, and percentage of surviving larvae that were PL1. Trypsin activity of samples of larvae from each treatment was also determined. The microparticulate microbound diet effectively served as a complete substitute for Artemia nauplii when fed beginning at stage M1. When fed at the beginning of the PZ2 stage, survival was comparable to that of larvae fed Artemia, but mean dry weight, mean total length, and percent of surviving larvae that were PL1 generally were significantly less. Responses to the feeding of copepods, whether fed dead or live, as a substitute were generally significantly less than those of larvae fed either the Artemia nauplii or the microparticulate diet. Values of trypsin activity (10^− 5 IU/μg^- 1 dry weight) corresponded to the relative proportions of the different larval stages within a treatment, with higher activity being characteristic of early stages. Previously demonstrated successful results with another species of crustacean suggest that the microparticulate microbound diet has characteristics that should be effective in the culture of the carnivorous stages of other crustacean and fish larvae that are currently fed live Artemia nauplii.     

吉富罗非鱼对饲料中维生素B1的需要量

采用维生素B1(VB1)含量为0.08(对照组)、0.57、1.13、2.09、4.11和8.09 mg/kg的6种纯化饲料,分别饲养初始体质量为(64.4±1.5)g的吉富罗非鱼12周,研究VB1对其生长性能、部分血清生化指标、肝脏VB1蓄积量及转酮醇酶基因表达量的影响,以确定其对饲料VB1的需要量.结果显示,随着饲料中VB1含量增加,吉富罗非鱼增重率先呈线性增加后趋于稳定,当饲料中VB1含量为1.13、2.09、4.11、8.09 mg/kg时增重率达最大.吉富罗非鱼肝脏VB1含量随着饲料VB1含量增加不断增大,当增加到2.09 mg/kg后趋于稳定.饲料中缺乏VB1显著提高血清丙酮酸含量(P＜0.05),但对全鱼水分、粗脂肪、粗蛋白、灰分无显著性影响(P＞0.05).饲料中添加VB1显著提高血清高密度脂蛋白胆固醇和肝脏转酮醇酶基因表达量(P＜0.05).饲料中VB1含量大于1.13 mg/kg各组的肝脏转酮醇酶活性显著高于VB1含量小于0.57 mg/kg组(P＜0.05).折线回归分析表明,吉富罗非鱼(64 ～325 g)获得最佳生长时对饲料VB1需要量为1.16 mg/kg;肝脏VB1蓄积量达到最大时,对VB1的需要量为2.06mg/kg.     

Influence of the dietary inclusion of <Emphasis Type="Italic">Gracilaria cornea</Emphasis> and <Emphasis Type="Italic">Ulva rigida</Emphasis> on the biodiversity of the intestinal microbiota of <Emphasis Type="Italic">Sparus aurata</Emphasis> juveniles

Partial substitutions of fish meal by 5, 15, or 25 % of Gracilaria cornea or Ulva rigida in experimental diets were evaluated to study their effects on biodiversity of intestinal microbiota composition in gilthead seabream (Sparus aurata) juveniles. The diets were offered to duplicate groups of 15 juvenile fish (14.0 ± 0.5 g) for 70 days, and at the end of the experiment the intestinal microbiota from four specimens of each treatment was analysed by denaturing gel gradient electrophoresis. Results showed that the substitution of fish meal by algae meal induced important modifications in the intestinal microbiota community, as a big reduction of the biodiversity when the highest percentage (25 %) of U. rigida was included. On the contrary, an increase on the number of species was detected when a 15 % of algae was included. Various Lactobacillus delbrueckii subspecies were selectively stimulated when G. cornea was included in the feed, and other bacterial species, such as those included in the Vibrio genus, were reduced.     

Effects of diets containing a bacterial protein meal on growth and feed utilisation in rainbow trout (Oncorhynchus mykiss)

Triplicate groups of rainbow trout with initial weight 361 g were fed either a fish meal based control diet or diets containing 9, 18 or 27% bacterial protein meal (BPM) or 9% of an autolysate (AU) of the BPM. No significant treatment effects were found on specific growth rates (SGR), feed intake, feed efficiency ratio (FER), or retention of nitrogen, amino acids or energy. The apparent digestibility coefficients (ADC) of nitrogen, energy and most indispensable amino acids decreased when BPM was included in the diet. The ADC of lipid, sum of amino acids, arginine, lysine, threonine and most of the dispensable amino acids were reduced at 27% BPM inclusion compared to the control, 9% and 18% BPM diets. None of the ADC estimated was different in the 9% AU diet compared to the 9% BPM diet. The loss of nitrogen and energy in faeces per kg gain increased as the dietary BPM or AU levels increased, and the energy used for activity and maintenance was higher in fish fed the 27% BPM diet than in fish fed the other diets (P < 0.05). There were no significant differences in the urea concentrations in plasma, liver and muscle, whereas the uric acid level in plasma was elevated in trout fed the 27% BPM diet. Histological evaluation of tissue from the stomach, pyloric caeca, mid-intestine and distal intestine did not reveal any diet-related morphological changes.In conclusion, no significant differences in growth and feed efficiency were found in the rainbow trout fed diets containing up to 27% BPM, and the AU did not increase fish performance compared to the BPM. Based on the data from this study, at levels up to 27% dietary inclusion, BPM is a good replacement for fish meal in diets for rainbow trout.     

The effects of algal diets on population growth and egg hatching success of the tropical calanoid copepod, Acartia sinjiensis

As natural diets of fish larvae, a number of calanoid copepod species are being investigated for use as live prey in aquaculture hatcheries. One of these, the tropical calanoid copepod, Acartia sinjiensis, has good potential as a live feed for tropical reef fish larvae. However, the rearing techniques for A. sinjiensis require further development to improve productivity. This study was carried out to investigate the population growth and egg hatching success of A. sinjiensis when fed a range of mono-and binary algal diets, including algae in the form of frozen paste.For the population growth experiment, the final A. sinjiensis population, including eggs, nauplii, copepodites and adults, was determined after feeding eight algal diets (two frozen algae, four live monoalgal and two live binary algal diets) for 8 days at temperature 28 ± 1 °C; salinity 34 ± 1 psu and photoperiod 12 L:12 D. Five replicates, with an initial 12 adult A. sinjiensis per replicate, were set up for each treatment. In a separate experiment, effects of diets on egg hatching success were examined after 48 h incubation of eggs produced by A. sinjiensis fed the same eight diets.The results showed that diet significantly affected both population growth and hatching success of A. sinjiensis. Of the diets tested, the binary algal diets were more successful than monoalgal diets, while the frozen algae had little dietary value. The highest population growth was recorded on A. sinjiensis fed a binary diet of Tetraselmis chuii and the Tahitian strain of Isochrysis sp. (T-ISO) (final population: 1091 ± 80), which was significantly higher (P < 0.001) than all other diets tested except for the other binary diet of Nannochloropsis sp. and T-ISO (final population: 897 ± 123). Diet also had a significant effect (P < 0.001) on egg hatching rate, though the highest hatch rate was recorded with eggs produced by A. sinjiensis fed binary diet Nannochloropsis sp. and T-ISO (88.1 ± 2.1%), this was not significantly different from that of eggs produced by A. sinjiensis fed either T-ISO alone (88.0 ± 1.7%) or the binary algal diet of T. chuii and T-ISO (76.4 ± 7.1%). The results of this study suggest that among the diets tested, a combination of live T. chuii and T-ISO was the best for the culture of A. sinjiensis.     

Growth, immune function, and disease and stress resistance of juvenile Nile tilapia (Oreochromis niloticus) fed graded levels of bovine lactoferrin

Juvenile Nile tilapia (Oreochromis niloticus) were fed nutritionally complete, practical basal diets supplemented with bovine lactoferrin (Lf) at 0, 200, 400, 800, or 1600 mg/kg diet to apparent satiation twice daily for 8 weeks. After the feeding trial, the effect of dietary Lf on growth performance, immune function, and resistance to Streptococcus iniae challenge and low-water stress was determined. Dietary Lf did not affect growth performance (weight gain, feed intake, feed efficiency ratio, or survival) or haematological parameters (haemoglobin, white and red blood cell counts, or haematocrit) (P > 0.05). Crowding stress produced significant increases in plasma cortisol, glucose, lactate, and osmolality from baseline values (P ≤ 0.001), but dietary Lf did not affect plasma glucose, osmolality, or cortisol concentrations (P > 0.05). The level of Lf in diet had a significant impact on survival following S. iniae challenge with fish fed the 800 mg/kg Lf diet having significantly higher survival than control fish (P ≤ 0.05). There was not a corresponding increase in activity of non-specific or specific immune parameters (plasma lysozyme and spontaneous haemolytic complement activities or agglutination antibody titer against S. iniae) with addition of Lf to diets (P > 0.05), but plasma iron decreased and total iron binding capacity (TIBC) increased significantly with increasing concentration of Lf in diet (P ≤ 0.05). The ability of Lf to sequester iron, an essential nutrient required for the growth of bacteria, is regarded as one of its key antibacterial properties. The increased survival with increasing dietary concentration of Lf seemed to correspond with a decrease in plasma iron concentration and not enhancement of non-specific or specific immune functions.     

Distribution of a fish pathogen Listonella anguillarum in the Japanese flounder Paralichthys olivaceus hatchery

The present study was undertaken to investigate the distribution of Listonella anguillarum in a Japanese flounder (Paralichthys olivaceus) hatchery. A total of 2704 isolates were obtained from the developing fish, live diets and artificial feeds of Japanese flounder and their rearing water, 439 of which were identified as L. anguillarum by the combining incubation on thiosulfate-citrate-bile salt-sucrose (TCBS) agar at 35 °C overnight with polymerase chain reaction (PCR) detection for the VAH1 hemolysin gene. L. anguillarum was detected in all seven rotifer samples, with densities of 2.5 × 10³ to 4.6 × 10⁶ colony forming units (CFU) g^− 1. Both the analyzed samples of Nannochloropsis oculata contained this bacterium at densities of 1.6 × 10⁴ to 1.4 × 10⁵ CFU g^− 1. L. anguillarum was detected in only one of four samples of Artemia nauplii with a density of 4.8 × 10⁵ CFU g^− 1 (35%) and it was not detected in the two analyzed artificial feed samples. L. anguillarum was detected in 11 of 18 specimens of larval and juvenile Japanese flounder at densities of 5.0 × 10¹ to 7.4 × 10⁵ CFU g^− 1, while it was not detected in the two analyzed egg specimens of Japanese flounder. These results indicate that L. anguillarum associated with the developing Japanese flounder is likely derived from rearing water and live diets such as rotifers. Further, it is strongly suggested that L. anguillarum is a transient bacterium of the intestinal microflora for the Japanese flounder but is a permanently indigenous one for the Japanese flounder hatcheries.     

The effects of supplemental dietary cholesterol on growth, development and survival of mud crab, Scylla serrata, megalopa fed semi-purified diets

The effects of varying levels of dietary cholesterol on growth, development time and survival of mud crab, Scylla serrata megalopa were investigated using semi-purified microbound diets (MBD). Five iso-energetic diets containing different level of cholesterol ranging from 0.14% to 1% of dry weight of the diet were tested. Fifteen megalopa were reared individually for each dietary treatment, and development time and survival were recorded on a daily basis. More than 25% of megalopa from all treatments were able to metamorphose into the first crab stage, suggesting that the endogenous level of cholesterol in the basal diet (0.14%) was sufficient to support development of the megalopa stage of this species. Widest mean carapace width (3.53 ± 0.08 mm) and highest mean dry weight (2.11 ± 0.22 mg) were recorded for juveniles that molted from megalopa fed live Artemia, whereas no megalopa in the unfed control treatment metamorphosed into crabs. The average development time from megalopa to the juvenile crab stage varied between the treatments, where megalopa fed live Artemia or MBD containing 0.2%, 0.4% or 0.8% total cholesterol showed the most synchronized molting (between 8.0 and 9.9 days). Longest development time was recorded for the megalopa fed diets containing 0.14% or 1% total cholesterol (both 11 days). Highest survival (74.3%) was recorded for the megalopa fed a diet containing 0.8% cholesterol. The results of this study are valuable in research to develop formulated diets for mud crab larvae as a replacement for live food in hatchery culture.     

Effects of dietary vitamin C on survival, growth, and immunity of large yellow croaker, Pseudosciaena crocea

An 8 weeks growth experiment was conducted to determine the effects of dietary vitamin C on the survival, growth, tissue ascorbic acid contents and immunity of large yellow croaker (Pseudosciaena crocea) with initial weight of 17.82 ± 0.68 g. Seven practical diets were formulated to contain 0.1, 12.2, 23.8, 47.6, 89.7, 188.5 and 489.0 mg ascorbic acid equivalent kg^− 1 diet, supplied as l-ascorbyl-2-polyphosphate (LAPP). Each diet was fed to triplicate groups of fish in seawater floating cages (1.5 × 1.5 × 2.0 m), and each cage was stocked with 120 fish. Fish were fed twice daily (05:00 and 17:00) to apparent satiation for 8 weeks. The water temperature fluctuated from 19.5 to 25.5 °C, the salinity from 25 to 28‰ and dissolved oxygen content was approximately 7 mg l^− 1 during the experimental period. Results showed that the specific growth rate (SGR) (from 1.80 to 1.96% d^− 1) had an increasing trend with the increase of dietary vitamin C, but no significant difference was observed among dietary treatments. No gross deficiency signs were observed in any of the experimental fish. Survival rate, however, significantly increased with increasing dietary vitamin C (P < 0.05). The vitamin C contents in liver and muscle correlated positively with the vitamin C in diets. The vitamin C requirement was estimated to be 28.2 mg kg^− 1 based on survival, and 87.0 mg kg^− 1 on liver content of vitamin C. The activities of serum lysozyme and alternative complement pathway (ACP), phagocytosis percentage (PP) and respiratory burst activity of head kidney significantly increased with increasing dietary vitamin C. The challenge experiment with Vibrio harveyi showed that fish fed the diets with supplementation of vitamin C had significantly lower cumulative mortality compared to the control group (66.7%), and the cumulative mortality (16.7%) in fish with 489.0 mg kg^− 1 ascorbic acid was significantly lower than that (41.7%) in fish with 23.8 mg kg^− 1 ascorbic acid. These results suggested that vitamin C significantly influenced the immune response and disease resistance of large yellow croaker.     

Experimental integrated aquaculture of fish and red seaweeds in Northern Portugal

Three potentially valuable red seaweeds, Chondrus crispus Stackhouse, Gracilaria bursa pastoris (S.G. Gmelin) P.C. Silva and Palmaria palmata (L.) O. Kuntze, collected in northern Portugal, were cultivated using the nutrient-rich effluents from a local turbot (Scophthalmus maximus Linnaeus) and sea bass (Dicentrarchus labrax Linnaeus) farm. The algae were cultivated in a two level cascade system. Several arrangements of the cascade system, stocking densities (3, 5, 7 and 8 kg m^− 2) and water fluxes (140 and 325 l h^− 1) were tested to optimize biomass yield and nitrogen uptake rate and efficiency. The yield and the total ammonium nitrogen (TAN) uptake of the three species were highly seasonal. Palmaria could not survive culture conditions during the summer when water temperature was above 21 °C. In the spring, Palmaria had an average yield of 40.2 (± 12.80) g DW m^− 2 day^− 1 and a nitrogen uptake efficiency (NUE) of 41.0% (± 17.26%). NUE expresses, in percentage, the average reduction in TAN concentration between the inflows and the outflows of the tanks. Chondrus performed better in summer with an average yield of 37.0 (± 11.10) g DW m^− 2 day^− 1 and removing 41.3% (± 17.32%) of nitrogen. Gracilaria grew year round, but also performed better during spring/summer, producing an average of 29.1 (± 2.90) g DW m^− 2 day^− 1, and only 7.3 (± 5.08) g DW m^− 2 day^− 1 during autumn. Yield of C. crispus did not differ significantly when grown at two different stocking densities (5 kg m^− 2 and 8 kg m^− 2). On the other hand, Gracilaria had significantly higher yields at 5 than at 7 kg m^− 2. Better NUE, on average 76.7% (± 22.13%), was also obtained with 5 kg m^− 2 stocking density and only 63.8% (± 24.62%) with 7 kg m^− 2. The yield of Gracilaria increased significantly with the increase of water flux from 140 to 325 l h^− 1 and more nitrogen was removed from the water. However, NUE decreased from 48.4% to 33.4% at 140 and 325 l h^− 1, respectively. Biofiltration was highly improved by a cascade system with a NUE as high as 83.5%.     

饲料蛋白质和脂肪水平对牛蛙生长性能的影响

为研究饲料中不同蛋白质和脂肪水平对牛蛙生长、体组成和肝脏健康的影响,以红鱼粉、豆粕和酪蛋白为蛋白源,鱼油和豆油为主要脂肪源,α-淀粉为主要糖源,微晶纤维素为填充剂,并补充无机盐和维生素等配制实验饲料。实验设3个蛋白水平(35%、40%和45%),每个蛋白水平下设4个脂肪水平(4%、7%、10%和13%),共12种饲料,饲料可利用能变化范围为12.4~17.8 kJ/g,蛋白能量比变化范围为21.9~32.0 mg/kJ。每种饲料设置3个重复,每个水族缸放养12只牛蛙[初始平均体质量为(91.5±1.1)g],每日饱食投喂2次,实验周期56 d。结果发现,饲料蛋白质和脂肪水平对牛蛙的存活率无显著影响,而13%脂肪组的牛蛙肝脏出现颜色不均一和肝体脆化无韧性的现象。饲料蛋白质和脂肪水平显著影响牛蛙的增重率、特定生长率、摄食率、饲料效率和蛋白质效率。其中,40%蛋白组牛蛙的增重率和特定生长率显著高于35%蛋白组,与45%蛋白组无显著差异。随着蛋白质水平的升高,饲料效率显著升高,而摄食率和蛋白质效率显著降低。随着脂肪水平的升高,增重率、特定生长率和摄食率显著升高,饲料效率、蛋白质效率和氮保留率无显著差异。饲料脂肪水平与牛蛙全体的脂肪含量和总能水平正相关,但不影响牛蛙全体的蛋白质含量。随着饲料脂肪水平的升高,牛蛙肝脏丙二醛含量显著升高,过氧化氢酶和脂蛋白酯酶活力先升高后降低。研究表明,牛蛙饲料适宜的蛋白质和脂肪水平分别为40%和7%,适宜蛋白能量比为27.7 mg/kJ。     

Dietary copper requirements of juvenile grouper, Epinephelus malabaricus

To quantify dietary copper (Cu) requirements in grouper, Epinephelus malabaricus, copper sulfate was added to the basal diet at 0, 1, 2, 4, 6, 8, 10 and 20 mg Cu/kg diet providing the actual dietary value of 0.11, 1.66, 2.41, 4.37, 6.56, 8.97, 11.03 and 20.05 mg Cu/kg diet, respectively. Each diet was fed to triplicate groups of grouper (initial body weight 13.35 ± 0.01 g) in a recirculated rearing system for 8 weeks. The Cu concentration in rearing water was monitored during the feeding period and ranged from 1.0-1.5 μg/l. Weight gain (WG) and feed efficiency (FE) were higher (P < 0.05) in fish fed diets with 4.37 and 6.56 mg Cu/kg diet than those in fish fed diets with ≥ 11.03 and ≤ 1.66 mg Cu/kg diet. Hepatic thiobarbituric acid reactive substances (TBARS) value was lowest in fish fed diets with 4.37 and 6.56 mg Cu/kg diet, intermediate in fish fed diets with 11.03 and ≤ 1.66 mg Cu/kg diet, and highest in fish fed the diet with 20.05 mg Cu/kg diet. Differences between each of the three groups were statistically significant (P < 0.05). Both hepatic copper-zinc superoxide dismutase (Cu-Zn SOD) and plasma ceruloplasmin activities in fish fell into four groups. Cu-Zn SOD value was highest in fish fed the diet with 6.56 mg Cu/kg diet, second highest in fish fed the diet with 8.97 mg Cu/kg diet, intermediate in fish fed the diet with 20.05 mg Cu/kg diet, and lowest in fish fed the diet with 0.11 mg Cu/kg diet; plasma ceruloplasmin was highest in fish fed the diet with 6.56 mg Cu/kg diet, second highest in fish fed diets with 2.41, 4.37 and 8.97 mg Cu/kg diet, intermediate in fish fed diets with ≥ 11.03 mg Cu/kg diet, and lowest in fish fed diets with ≤ 1.66 mg Cu/kg diet. All values in each of the four groups were significantly different from the values of three other groups (P < 0.05). Analysis by broken-line regression of WG, hepatic SOD activity and TBARS value and linear regression of whole-body Cu retention of the fish indicate that the adequate dietary Cu concentration in growing grouper is about 4-6 mg Cu/kg diet.