Weaning Chinese perch Siniperca chuatsi (Basilewsky) onto artificial diets based upon its specific sensory modality in feeding

Chinese perch are one of the most valuable food fish in China, but the sole source of feed for intensive culture is live prey fish. Our previous studies on systematic sensory physiology revealed that this species have a mechanism for this peculiar feeding habit. In the present study, a specific training procedure was designed, and both experimental (initial body weight 171.0 g; 120 days) and commercial (initial body weight 52.4 g; 240 days) net-cage cultures were conducted to investigate the training success, growth performance and survival of the trained yearlings fed with nonlive or Oregon-type moist diet. The training successes of minced prey fish and the Oregon moist diet were 100 and 89.9%, respectively, in experimental culture, and 92.2 and 83.5% in commercial culture. In an experimental trial, the fish fed minced prey fish or the Oregon moist diet attained final body weights of 472.7 g or 344.7 g, although the specific growth rates of these groups were significantly lower than that of the fish fed live prey fish (final body weight 560.0 g). Mortality was not significantly related to dietary treatment. In commercial culture, the final body weights were as follows: 750 g on live prey fish, 705 g on minced prey fish and 651 g on the Oregon moist diet. Feed costs to produce 1 kg fish were estimated to be US$6.59 for live prey fish, US$1.76 for minced prey fish and US$2.07 for the Oregon moist diet. The results of the present study confirmed that sensory modality and associative learning appear to be critical factors in determining food discrimination of Chinese perch, indicating that both minced trash fish and Oregon-type moist diet can be substituted for live prey fish in intensive commercial production.     

洞庭湖水系鳜遗传结构

测定了采自洞庭湖水系6个地理位置(湘江常宁、沅江洪江、沅江桃源、澧水石门、洞庭湖万子湖和南岳坡)共100尾鳜(Siniperca chuatsi)的线粒体Cytb基因DNA序列.结果显示:1 140 bp的Cyt b基因共检测到55个变异位点,100个样本得到9个单倍型.单倍型多样性指数(Hd)和核苷酸多样性指数(Pi)分别为0.777±0.033和0.019 70 ±0.000 74.洞庭湖的万子湖和南岳坡2个群体遗传多样性指数最高,湘江常宁群体遗传多样性指数最低.分子变异分析(AMOVA)结果显示:FST=0.09803(P＞0.05),9.8％的变异来自群体间,91.2％的变异来自群体内.除常宁群体分化显著外,其它群体没有出现明显遗传分化(两两之间FST在-0.065 ～0.024之间).     

Impact of environmental and innate factors on the food habit of Chinese perch Siniperca chuatsi (Basilewsky) (Percichthyidae)

Laboratory and field investigations were conducted to study the food habit of Chinese perch Siniperca chuatsi (Basilewsky) from first feeding through adult stage. Only fish larvae were consumed by Chinese perch larvae (2–21 days from hatching), and the presence of zooplankton did not have any significant effect on their survival rate. The ability of Chinese perch to feed on zooplankton is clearly limited by some innate factor. Instead of gill rakers, Chinese perch larvae have well‐developed sharp teeth at the first feeding stage, and are well adapted to the piscivorous feeding habit unique to the larvae of Chinese perch, e.g. they bite and ingest the tails of other fish larvae. At the first feeding stage (2 days from hatching), daily rations were both very low, either in light or complete darkness. Although early‐staged Chinese perch larvae (7–17 days from hatching) could feed in complete darkness, their daily rations were always significantly higher in light than in complete darkness. Late‐staged Chinese perch larvae (21 days from hatching) were able to feed in complete darkness as well as in light, similar to the case of Chinese perch yearlings. Chinese perch yearlings (total length, 14–16 cm) consumed prey fish only and refused shrimp when visual cues were available (in light), but they consumed both prey when visual cues were not available (in complete darkness), suggesting that prey consumption by Chinese perch yearlings is affected by their sensory modality in predation. Both prey were found in the stomachs of similar‐sized Chinese perch (total length, 14–32 cm) from their natural habitat, suggesting that shrimp are consumed by Chinese perch at night. Prey selection of Chinese perch with a length >38 cm, which consumed only fish in the field, appears to be based upon prey size instead of prey type. These results suggest that although environmental factors (e.g. light intensity) affect prey detection by Chinese perch, this fish is anatomically and behaviourally predisposed to prey on live fish from first feeding. This makes it a difficult fish to cultivate using conventional feeds.     

鳜UCP1,2基因结构、序列分析及组织表达

从分子水平上探讨了鳜解偶联蛋白1、2 (UCP1, 2)基因结构、组织表达水平及与产热、脂肪代谢等生理机能的关系。通过与脊椎动物UCP1、UCP2基因序列进行比对,设计简并引物与特异引物进行PCR和RACE扩增、测序、拼接序列,获得UCP1、UCP2的基因组序列和内含子/外显子结构。基因组步行法克隆鳜肝脏UCP1、UCP2 基因5′侧翼序列。应用半定量RT-PCR的方法,以β-肌动蛋白作为外参照,在其指数期增长的范围内得到鳜不同组织UCP1、UCP2的相对表达水平。结果表明,UCP1基因组全序列为3 146 bp, 5′侧翼调控区为1 333 bp,含有5个内含子和6个外显子,开放阅读框(ORF)长942 bp,编码一个大小为313个氨基酸的蛋白质。UCP2基因组全序列为2 890 bp, 5′侧翼调控区为1 800 bp,含有7个内含子和8个外显子,ORF长939 bp,编码一个大小为312个氨基酸的蛋白质。UCP1、UCP2间隔外显子的内含子皆符合“GT-AG”规则,内含子的数目与哺乳动物一致。系统进化分析表明,鳜UCP1、UCP2氨基酸序列分别与鱼类UCP1、UCP2氨基酸序列聚为一支,且与UCP3、UCP4、UCP5分支区分明显。鳜UCP1、UCP2基因不同组织表达水平的高低可能与鳜本身的生态习性及各器官在产热、脂质代谢中的作用相关,但明确的分子机制尚待进一步研究。     

鳜胃泌素与胆囊收缩素基因cDNA全长的克隆与表达特征

为探明鱼类消化液分泌的相关调控因子,采用RACE技术分别克隆了鳜(Siniperca chuatsi)两种肠道激素——胃泌素(gastrin,GAS)与胆囊收缩素(cholecystokinin,CCK)基因的cDNA序列。鳜GAS基因cDNA全长581 bp,5′非翻译区长100 bp,3′非翻译区长145 bp,开放阅读框长336 bp,编码111个氨基酸;鳜CCK具有2种类型:CCK1与CCK2,CCK1 cDNA全长为843 bp,5′非翻译区长60 bp,3′非翻译区长369 bp,开放阅读框414 bp,编码137个氨基酸;CCK2 cDNA全长846 bp,5′非翻译区长112 bp,3′非翻译区长332 bp,开放阅读框403 bp,编码134个氨基酸。鳜GAS与CCK成熟肽C末端具有相似的八肽结构(DYQGWVDF/DYLGWMDF),仅在C末端第3位和第6位氨基酸发生替换。荧光定量PCR分析表明,鳜GAS mRNA主要表达于肠和幽门垂,CCK1与CCK2 mRNA在脑中表达量最高,在肠和幽门垂也有较高表达,表明GAS与CCK同为消化调控因子,而CCK还是神经分泌因子。鳜GAS与CCK mRNA表达贯穿于整个幼体早期发育阶段(孵化后0～22 d),前期表达水平较高,后期表达水平较低,并趋于平稳,GAS与CCK mRNA发育表达水平变化可能与这一时期消化道生长发育旺盛有关。     

Characterization of β‐catenin 1 during the gonad development in the common carp (Cyprinus carpio)

β‐catenin gene is a pivotal gene for gonad development and maintenance of ovarian function in mammals. However, little is known about its expression and function in gonad development of fish. In this study, a complete cDNA (3342 bp) sequence of β‐catenin 1 was cloned from the common carp, Cyprinus carpio, by RACE PCR, which encodes a 780‐amino‐acid protein. Quantitative real‐time PCR demonstrated that β‐catenin 1 mRNA expressions were high in the testis and ovary tissue and the expression increased as the testes developed and the early stage ovaries developed. Western blot results revealed a single immunoreactive band with an estimated molecular weight of 90 kDa in testes. Immunohistochemistry analysis revealed that the β‐catenin 1 protein was concentrated mainly in the cytoplasm of early development stage of oocyte cells and in the cytomembrane of developing and mature sperm cells. 17β‐Ethinylestradiol injecting intraperitoneally into the fish decreased the relative β‐catenin 1 mRNA expression level except 1 μg/g 72 hr and 5 μg/g 48 hr of treatments in the ovary by real‐time PCR. These results suggest, for the first time, that β‐catenin 1 is an essential protein in gonad development and might be involved in ovarian early development of C. carpio.     

Immunomodulation of rainbow trout (Oncorhynchus mykiss) fry by bath exposure to a β‐glucan from Euglena gracilis

Early developmental stages of fish mostly depend on innate immune factors for their protection. Augmenting these factors by application of different immunostimulatory substances may be beneficial for rearing and survival of the early life stages of fish. Bath administration of stimulants leads to a uniform exposure of fish independent of feed intake and reduces the individual handling. The present study demonstrates the immunostimulatory effect of β‐glucan (bath exposure) in rainbow trout fry at different dosages and exposure time. Rainbow trout fry (avg. wt. 770 mg; 87 days post hatch) were exposed to three different concentrations of β‐glucan (10, 100 and 1000 μg mL^?1) by bath exposure for 1 and 24 h. Expression of immune related genes from pooled internal organ samples of individual fish were analysed using a real time qPCR assay. Expression of complement factors (C3 and factor B) and acute phase proteins (hepcidin, precerebellin and transferrin) was significantly up‐regulated after 24 h bath stimulation with β‐glucan (100 μg mL^?1). These innate immune factors may play a vital role in clearance of pathogens. The expression of most of genes showed both a dose‐ and time‐dependent response. A medium dose (100 μg mL^?1) induced a significant increase in expression of complement factors and acute phase proteins mainly at 24 h exposure, whereas the highest dose of β‐glucan (1000 μg mL^?1) down‐regulated the expression of most of the studied genes. The result from the present study indicates that β‐glucan bath exposure could be applied for enhancing the innate immune factors even in fry.     

The effects of different proportions of the 17β‐estradiol and 17α‐methyltestosterone on growth,sex reversal and skin colouration of the electric blue hap (Sciaenochromis ahli Trewavas, 1935)

This study was performed to investigate the effects of 17β‐estradiol (ES) and 17α‐methyltestosterone (MT) on growth, development, survival, sex ratio and colour change in the electric blue hap (Sciaenochromis ahli Trewavas, 1935). The hormones were not supplemented to the control feed, while six other feeds were prepared by adding 20, 40 and 60 mg kg^?1 17β‐ES or 20, 40 and 60 mg kg^?1 17α‐MT to each, resulting in seven different feed treatments. Average live weight of the fish supplemented with these diets was 0.42 ± 0.04 g. At the end of the study, the highest weight gain was observed in fish fed 60 mg kg^?1 17α‐MT group (2.62 ± 0.11 g) and the difference with the groups fed with 17β‐ES was found to be significant. All fish fed 17α‐MT were male, while the rates of feminization in fish fed 17β‐ES at 20, 40, 60 mg kg^?1 were 91.11%, 88.88% and 93.33% respectively. Survival rates were respectively determined as 80%, 95.56%, 84.44%, 93.33%, 77.78%, 84.44% and 84.44% for the control, 20, 40, 60 mg kg^?1 17β‐ES and 20, 40, 60 mg kg^?1 17α‐MT treatments. The best colouration was achieved in the 17α‐MT groups (P < 0.05). The L* values varied between 32.98 ± 4.44 and 61.35 ± 2.19, a* values between ?7.06 ± 0.22 and ?3.42 ± 0.11, and b* values between ?7.74 ± 0.10 and 11.65 ± 0.03, while Chroma (C*) and Hue (H°_ab) angle values varied between 7.54 ± 0.22 and 13.60 ± 0.01 and between 119.76 ± 0.05 and 239.73 ± 4.86. In conclusion, the 17α‐MT feeding was found to have a greater effect on the growth, feed conversion ratio, masculunization and pigmentation of the electric blue haps than the 17β‐ES treatment.     

Effects of α‐ketoglutarate on the growth performance,amino acid metabolism and related gene expression of mirror carp (Cyprinus carpio)

A 70‐day feeding experiment was conducted to assess the effects of α‐ketoglutarate (AKG) supplementation in different protein sources of diets on the growth performance, nitrogen metabolism and related gene expression of mirror carp. Two kinds of protein diets (soybean meal protein source and fish meal and soybean meal compound protein source) and two levels of AKG (0 g kg^?1 and 15 g kg^?1, respectively) were utilized. Each experimental diet was fed three times a day with five replicates of 20 fish per tank with an average individual weight of 217.93 ± 0.78 g. The results indicated that AKG significantly improved the protein efficiency ratio and decreased the feed conversion rate (FCR) (p < .05). Soybean protein significantly decreased the weight gain rate and increased the FCR (p < .05). In addition, diets with AKG significantly improved the concentration of glutamine and glutamic acid (Glu) in the foregut and hindgut (p < .05), as well as the glutamine synthetase (GS) activity in the foregut (p < .05). Soybean protein significantly decreased the concentration of Glu and GS activity in the hindgut (p < .05). AKG significantly improved the contents of threonine, alanine, proline and essential amino acid in muscle (p < .05). Dietary supplementation with AKG significantly improved GS gene expression in the intestine and reduced Rhesus glycoprotein (Rhag and Rhbg) gene expression in the gill (p < .05). From the results, we know that a diet with AKG is beneficial for the growth and amino acid metabolism of carp.     

Biochemical parameters in the blood of gilthead sea bream (Sparus aurata Linnaeus, 1758) supplemented with commercially available β‐glucan‐based product (IMUNO‐2865®)

The aim of this study was to evaluate the effects of a promising new immune stimulant in aquaculture (IMUNO‐2865^®) on biochemical parameters in sea bream during the winter stress period. A total of 640 sea bream were fed throughout 90 days with diets containing 0 (Group 1), 1 (Group 2), 10 (Group 3) and 25 g (Group 4) of IMUNO‐2865^® kg^?1 of feed. Samples were taken each month and 90 days after supplementation. No statistical differences among treatment groups were noticed for the following biochemical parameters: glucose (GLU), aspartate aminotransferase (AST), plasma cholesterol (CHOL), lactate dehydrogenase (LDH), urea (URE) and creatinine (CREA). At the final sampling, total ammonia (NH₃) was higher in Groups 3 and 4 compared to the control and the low supplementation group (p < .05), while total proteins (TP) was significantly higher in Group 4 compared to all other groups, and in Groups 2 and 3 compared to the control group (p < .05). Blood Ca⁺⁺ levels were significantly higher after 60 days of feeding in all treatment groups compared to the control, and remained elevated in Group 4 even after 90 days following cessation of supplementation (p < .05). The results of this study described the increase of biochemical parameters in the blood of sea bream after use of IMUNO‐2865^® but future research is needed to evaluate its potentially immunostimulative effect on fish in commercial aquaculture.     

Effect of dietary β‐1,3‐glucan on the immune response of Litopenaeus vannamei exposed to nitrite‐N

β‐1,3‐Glucan with 1.0 g kg^?1 was supplemented to a basal diet [control (C)] with different feeding schedules: permanently β‐1,3‐glucan diet (BG) and 14 days BG–14 days control diet (C + BG). Growth and immunological responses [respiratory burst (RB), superoxide dismutase (SOD), catalase (CAT), lysozyme and total protein] in haemolymph, hepatopancreas and muscle of Litopenaeus vannamei were recorded after 84‐day feeding and exposure to nitrite‐N (20 mg L^?1) for 120 h. β‐1,3‐Glucan administration did not affect growth performance. However, as compared with control, elevated CAT and lysozyme activities were seen in haemolymph of both BG groups, while significantly higher activities of SOD, lysozyme and RB in hepatopancreas, and higher activities of CAT and lysozyme in muscle were only seen in C + BG group. After nitrite‐N stress, significantly higher haemolymph protein, and hepatopancreas activities of lysozyme and RB were observed in both BG groups, but significantly higher activities of haemolymph SOD was only seen in C + BG group. The mortality in groups BG and C + BG was significantly lower than that in group C, but C + BG group showed a trend of higher nitrite‐N resistance compared with BG group. Considering dietary cost and immunostimulatory effects, the feeding schedule with 14 days BG–14 days control diet is more recommended.     

Selection and characterization of an improved strain (ST‐2) of Pyropia haitanensis by 60Co‐γ ray irradiation

Although the improved strain HR‐5 of Pyropia haitanensis has improvement in yield and quality, it shows earlier maturation and shortened growth period. In this study, blades of HR‐5 strain were treated with ⁶⁰Co‐γ ray irradiation, and a mutant strain ST‐2 was selected and characterized. The blade length of ST‐2 strain was 401.4 cm on day 70 post‐culture, which was 1.9‐fold longer than that of the parental strain HR‐5. The blades of HR‐5 strain matured earlier and most were matured on day 55, while the ST‐2 strain was not matured until day 90. When blades of HR‐5 and ST‐2 strains were firstly cultured at 23°C for 40 days and their blade discs cultured at 29°C for another 30 days, the mean length of ST‐2 strain was 2.0‐fold longer than that of HR‐5 strain. Besides, the contents of Chl. a and phycobiliprotein and the conchospore numbers in ST‐2 were comparable to that in HR‐5 strain. In summary, compared with HR‐5, we observed delayed maturation, increased length, weight and high‐temperature resistance of the blades, high content of photosynthetic pigments, and high production of conchospores in ST‐2, suggesting it might be a better strain for large‐scale production.     

Epitheliocystis in lake trout Salvelinus namaycush (Walbaum) is associated with a β‐proteobacteria

Lake trout Salvelinus namaycush (Walbaum) raised for stocking experienced yearly (2011–13) winter epizootics of epitheliocystis. Affected fish were dispersed on the bottom of the tank, had decreased feed and fright response, and mortality often reached 40%. Peak mortality occurred within 3 weeks of the appearance of clinical signs, and outbreaks typically lasted 6 weeks. Affected fish had no gross lesions but histologically had branchial epithelial necrosis and lamellar hyperplasia, with small to large numbers of scattered epithelial cells containing 10‐ to 20‐μm inclusions. A longitudinal study was undertaken of one annual outbreak, and lamellar hyperplasia was most closely associated with mortality. The number of inclusions was statistically greater (P < 0.05) before and during peak mortality, but inclusions were present in low numbers before clinical signs occurred. Results of histochemical staining, immunohistochemistry and transmission electron microscopy supported the presence of a β‐proteobacteria rather than a Chlamydiales bacterium within inclusions. PCR primers to identify Chlamydiales did not give consistent results. However, the use of universal 16S rDNA bacterial primers in conjunction with laser capture microdissection of inclusions demonstrated that a β‐proteobacteria was consistently associated with affected gills and is more likely the cause of the disease in lake trout.     

Effects of dietary β‐carotene levels on growth and liver vitamin A concentrations of the soft‐shelled turtle,Pelodiscus sinensis (Wiegmann)

A feeding trial was conducted to evaluate the effect of dietary β‐carotene level on the growth and liver vitamin A concentrations in soft‐shelled turtles, Pelodiscus sinensis, fed a vitamin A‐free diet. Soft‐shelled turtles were fed diets containing 0, 14.5, 26.5, 47.5, 87.3, 112.8 and 163.8 mg β‐carotene kg^?1 for 10 weeks. Although it was not statistically significant due to high deviation within each group, mean weight gain of soft‐shelled turtles fed the diet without β‐carotene supplementation was the lowest among all test groups. Vitamin A concentrations in liver of turtles significantly (P<0.05) increased when dietary β‐carotene level reached 47.5 mg kg^?1 indicating that soft‐shelled turtles were capable of converting β‐carotene to vitamin A. Analysed by regression modelling, dietary β‐carotene levels for optimal growth and maximal liver vitamin A contents of juvenile soft‐shelled turtles fed the vitamin A‐free diets were 49.1 and 88.7 mg kg^?1 respectively.     

Effect of β‐glucooligosaccharides as a new prebiotic for dietary supplementation in olive flounder (Paralichthys olivaceus) aquaculture

β‐Glucooligosaccharides (BGO), produced from barley β‐glucan, were used as a feed supplement (0.1%) for juvenile olive flounder (Paralichthys olivaceus) to identify and quantify its oral administration effects on innate immunomodulation and infectious disease protection. Juvenile flounders (14 ± 0.5 g) were divided into two groups fed either 0.1% BGO (treatment) or a standard diet (control) for 8 weeks. At the end of the experiment, investigation of the effects was carried out through systemic studies on growth performance, serum and mucus biochemical parameters, innate immunity, microvillus length, and relative pro‐inflammatory cytokine gene expression. The results demonstrated that the BGO diet produced slightly higher levels of growth performance, serum protein, microvillus length and pro‐inflammatory cytokine gene (tumour necrosis factor‐α, interleukin [IL]‐1β, and IL‐6) expression without any significant differences (p > .05). All innate immunity parameters were up‐regulated by BGO administration and, among these, respiratory burst, lysozyme and superoxide dismutase (SOD) were significantly different (p < .05). Fish in the both groups were challenged with Streptococcus iniae (1.35 × 10⁸ CFU/ml), and BGO group was focused to confirm the promotion of innate immunity parameters. The results showed a significantly (p < .05) lower death rate compared with that of the control. Therefore, BGO could be used as a new prebiotic in future olive flounder aquaculture as well as to control streptococcosis.     

Kinetics and isotherms of oxytetracycline adsorption on β‐cyclodextrin/carboxymethylcellulose hydrogel films

Oxytetracycline (OTC), widely used as an antibiotic substance in aquaculture, has an adverse effect on environmental and public health. The aim of this paper was to demonstrate that OTC can be removed by hydrogel films composed of β‐cyclodextrin/carboxymethylcellulose (β‐CD/CMC) with esterification‐crosslink method. Oxytetracycline adsorption experiments were performed with an initial pH of 1–12 and an initial concentration of 10–500 mg/L. The experimental data were fitted with Langmuir and Freundlich isotherms and compared using linear and nonlinear models. The experimental results indicate that OTC's adsorption capacity depends on the active β‐cyclodextrin content. The optimal pH was around 8.     

Effect of poly‐β‐hydroxybutyrate on growth,enzyme activity and intestinal microbial community of Chinese mitten crab,Eriocheir sinensis (Milne‐Edwards) juveniles

Poly‐β‐hydroxybutyrate (PHB) is microbial carbon and energy storage polymer, which can be degraded into water‐soluble β‐hydroxybutyric acid in the gastrointestinal tract of aquatic animals. A 60‐day culture experiment was performed using Chinese mitten crab, Eriocheir sinensis (Milne‐Edwards) juveniles with an average initial body weight of 0.74 ± 0.06 g which were fed a diet supplemented with 0%, 0.5%, 1%, 3% or 5% PHB. A PHB dietary supplementation of 1% and 3% significantly improved the body weight gain, moulting frequency and concomitantly reduced 2nd–3rd moulting intervals of the crabs (P < 0.05). The dietary PHB level positively related to hepatopancreatic pepsin, trypsin and lipase activity (P < 0.05). Increasing the dietary PHB also improved total superoxide dismutase activity, but reduced alkaline phosphatase and acid phosphatase activity in the serum of hemolymph (P < 0.05). A 16S rRNA gene analysis by denaturing gradient gel electrophoresis indicated that PHB supplementation led to a significantly higher range‐weighted richness, diversity and evenness of the gut bacterial community when dosed at 3% in the feed. The beneficial effects of PHB are discussed in terms of immune defense, metabolism and gut microbiota of the crabs.     

The effect of poly‐β‐hydroxybutyrate on the performance of Chinese mitten crab (Eriocheir sinensis Milne‐Edwards) zoea larvae

Two experiments were conducted regarding the effect of poly‐β‐hydroxybutyrate (PHB) on the survival and development of Chinese mitten crab (Eriocheir sinensis Milne‐Edwards) larvae. Different PHB delivery approaches (particulate, enriched in filter feeding prey, or a combination of both) and feeding levels were applied to crab larvae from the zoea 2 (Z2) up to the megalopa stage. Bio‐encapsulation into rotifers and Artemia at a PHB dose of 100 mg L^?1 seemed an efficient approach to deliver PHB to the larvae and resulted in a significantly enhanced survival, development rate and osmotic tolerance. The results indicate that PHB should only be applied as a feed additive rather than as a food source, and that the availability of an adequate amount of nutrients seems to be of major importance for the PHB to induce these beneficial effects.