三疣梭子蟹B型清道夫受体蛋白的原核表达及细胞、组织分布研究

B型清道夫受体是清道夫受体超家族成员,可识别多种配体,在机体脂类转运及免疫防御过程中发挥重要作用。为深入研究三疣梭子蟹B型清道夫受体(Pt-SRB)的功能,本实验在前期工作基础上构建了Pt-SRB胞外结构域原核表达载体,并成功获得了重组蛋白rPt-SRB。利用亲和层析方法获得纯化的rPt-SRB,并将纯化rPt-SRB蛋白免疫大鼠获得抗rPt-SRB重组蛋白免疫抗血清以用于后续研究。SDS-PAGE检测发现,体外诱导表达重组rPt-SRB以包涵体的形式出现在大肠杆菌BL21裂解液的沉淀中,分子量大小约为49.18 ku。Western-Blot分析表明,大鼠抗rPt-SRB血清能与rPt-SRB特异性结合。本实验还利用免疫荧光技术,对Pt-SRB在三疣梭子蟹血淋巴细胞的定位及不同组织中的分布进行研究。结果显示,Pt-SRB在三疣梭子蟹消化道、肝胰腺、鳃、心脏、肌肉组织中均有分布,但不同组织中Pt-SRB的分布不同。免疫荧光结果显示,绿色荧光信号在消化道及腺体结缔组织中较强,另外在肝小管上皮、鳃丝上皮细胞中亦有较强的阳性信号,表明PtSRB蛋白在上述组织结构中分布较广;在血淋巴细胞中,绿色荧光信号主要分布于细胞质和细胞膜,在细胞核中没有明显荧光信号,提示Pt-SRB在三疣梭子蟹血淋巴细胞的细胞质和细胞膜上表达。本结果将为三疣梭子蟹B型清道夫受体蛋白的生理及免疫学功能的研究奠定基础。     

Supplementation of heat‐inactivated Bacillus clausii DE5 in diets for grouper,Epinephelus coioides,improves feed utilization,intestinal and systemic immune responses and not growth performance

In recent years, more and more attentions have been paid to the development and application of probiotics in aquaculture, and viable probiotics have been extensively studied, while rare information was available about inactivated probiotics in aquaculture. Therefore, in this study, a feeding trial was designed to investigate the effect of dietary supplementation of heat‐inactivated probiotic Bacillus clausii DE5 on growth performance, immune response and key immune genes expression in head kidney and intestine in grouper Epinephelus coioides. Fish were fed for 60 days with control diet (C) and two experimental diets containing 1.0 × 10⁸ CFU/g live (T1) and heat‐inactivated (T2) B. clausii DE5, respectively. The probiotic treatments did not affect the final weight, weight gain (WG) and specific growth rate (SGR) of E. coioides at days 30 and 60 (p > .05), while both heat‐inactivated and live B. clausii DE5 significantly decreased the feed intake and feed conversion ratio (FCR) at day 60 (p < .05). Serum lysozyme activity and complement C3 level in the two probiotic treatments were significantly higher than those in the control (p < .05). The lysozyme activity and complement C3 level at day 60 were significantly higher than those at day 30 (p < .05), while no significant interaction effect between diet and administration date was observed. Moreover, the heat‐inactivated B. clausii DE5 significantly improved the expression of TLR5, pro‐inflammatory cytokines (IL‐8 and IL‐1β) and TGF‐β1 in head kidney and intestine (p < .05), while the live probiotic did not show any significant effect on the expression of these key immune‐related genes in head kidney and intestine. These results indicate that dietary supplementation of heat‐inactivated B. clausii DE5 effectively improved feed utilization and both the local and systemic immune responses of E. coioides.     

Improvement of flesh quality in rainbow trout (Oncorhynchus mykiss) fed supranutritional dietary selenium yeast is associated with the inhibited muscle protein degradation

Supranutritional dietary selenium (Se) has been demonstrated beneficial for fish health, however, its effects on fish flesh quality remain unclear. This study investigated the effects of supranutritional dietary Se on the flesh quality of rainbow trout (Oncorhynchus mykiss) and their potential mechanism. Fish were fed a basal diet supplemented with or without graded Se yeast for 10 weeks. Results showed that Se supplementation significantly increased fillet crude protein and enhanced fillet water‐holding capacity as well as fillet firmness. Supplementing with both 2 and 4 mg/kg Se significantly downregulated the expressions of two autophagy–lysosome‐related genes (autophagy‐related 12‐like and gamma‐aminobutyric acid type A receptor‐associated protein‐like 1) in fish muscle, while supplementing with 4 mg/kg Se also significantly downregulated the expression of two ubiquitin–proteasome‐related genes (muscle RING finger 2 and F‐box protein 25). Correlation analysis indicated that the improved fillet quality parameters were closely correlated with the expressions of these differentially expressed genes. This study revealed that dietary Se was effective for the improvement of rainbow trout flesh quality, and the improved fish flesh quality was associated with the inhibited protein degradation in fish muscle.     

Effect of dietary fish oil on enhanced inflammation and disturbed lipophagy in white adipose tissue caused by a high fat diet

Fisheries Science - Fish oil containing omega-3 polyunsaturated fatty acids (PUFA) attenuates chronic inflammation found in obesity, leading to a reduction in insulin resistance. The effect of fish...     

方斑东风螺“急性死亡症”的病原病理研究

对海南发生的方斑东风螺（Babylonia areolata）“急性死亡症”进行了病原分离纯化、人工感染试验,以及病原16SrDNA分析和生理生化鉴定。结果显示病原菌为哈维弧菌（Vibrioharveyi）;药敏试验结果显示,该病原菌对常见的抗菌药物如恩诺沙星、复合磺胺等敏感,而对青霉素表现为耐药;组织病理结果显示鳃叶、腹足、肝胰腺、消化道、肾等器官的细胞出现变性、坏死、血细胞浸润等现象。     

Immunostimulatory effects of dietary fructooligosaccharides on red swamp crayfish,Procambarus clarkii (Girard)

Fructooligosaccharides (FOS) are widely researched as prebiotics and immunostimulants in humans and terrestrial animals. However, limited researches on immune functions of FOS were reported in crustacean. In the present study, red swamp crayfish Procambarus clarkii were fed with different concentrations of FOS (2.0, 5.0, 8.0, 10.0 g kg^?1 diet), and then the immunostimulatory effects of FOS were evaluated by measuring immune parameters and examining immune‐related gene expression. PO and SOD activities were significantly increased in groups fed with 8.0 or 10.0 g kg^?1 of FOS compared with those in control group. In addition, significant elevations of SOD and phagocytic activities were also observed in group fed with 5.0 g kg^?1 of FOS. Among examined immune‐related genes, expressions of crustin1, lysozyme, SOD and pro‐PO were significantly up‐regulated by feeding with 8.0 or 10.0 g kg^?1 of dietary FOS. Survival rates of crayfish challenged with Aeromonas hydrophila were significantly improved by feeding with 5.0, 8.0 or 10.0 g kg^?1 of dietary FOS for 30 days. Phagocytic activities and bacterial clearance efficiencies were also significantly increased in groups fed with 8.0 or 10.0 g kg^?1 of FOS. The results indicated that dietary FOS effectively induced innate immune responses of red swamp crayfish in a dose‐dependent way and could be used as potential immunostimulant in crustacean.     

Apparent digestibility coefficients of selected feed ingredients for juvenile snakehead,Ophiocephalus argus

Apparent digestibility coefficients (ADCs) of dry matter, crude protein, crude lipid, gross energy, phosphorus and amino acids in Peruvian fish meal, poultry by‐product meal, meat and bone meal, spray‐dried blood meal, hydrolysed feather meal, corn gluten meal, soybean meal, peanut meal, cottonseed meal and rapeseed meal were determined for juvenile snakehead (Ophiocephalus argus) with initial mean body weight of 78.1 g. A reference diet and test diets that consisted of a 70 : 30 mixture of the reference diet to test ingredient were used with 5 g kg^?1 Cr₂O₃ as an external indicator. Fish meal, poultry by‐product meal and corn gluten meal had higher ADCs of dry matter, crude protein, and gross energy among ingredients tested. Dry matter ADCs ranged 61.9–81.5% for animal ingredients and corn gluten meal and ranged 52.2–68.0% for soybean meal, peanut meal, cottonseed meal and rapeseed meal. Energy ADCs of ingredients followed similar trends to differences in dry matter digestibility. Protein ADCs of animal and plant ingredients ranged 73.6–92.8% and 75.3–85.6%, respectively. Amino acid ADCs generally reflected protein digestibility. Lipid ADCs were relatively high for the ingredients tested. Phosphorus ADCs of animal and plant ingredients ranged 39.5–65.2% and 38.7–57.1%, respectively.     

Effects of Neutral Phytase Supplementation on Biochemical Parameters in Grass Carp,Ctenopharyngodon idellus,and Gibel Carp,Carassius auratus gibelio,Fed Different Levels of Monocalcium Phosphate

Two 8‐wk studies were conducted to evaluate the effects of neutral phytase supplementation on hemato‐biochemical status, liver biochemical parameter, and intestinal digestive enzyme activity of grass carp, Ctenopharyngodon idellus, and gibel carp, Carassius auratus gibelio, fed with different levels of monocalcium phosphate (MCP). The control diet was prepared with 2% MCP but without phytase (P2.0). The other three experimental diets were prepared with the addition of 1.5, 1.0, and 0.5% MCP, respectively, when supplemented with 500 U/kg neutral phytase in each diet and designated as PP1.5, PP1.0, and PP0.5, respectively. The results indicated that the serum alkaline phosphatase (ALP), alanine transaminase (ALT), and aspartate transaminase (AST) activities, as well as the albumin (ALB) content were increased in grass carp (P < 0.05) and gibel carp (P > 0.05) fed with phytase‐supplemented diets. Meanwhile, the serum cholesterol, high‐density lipoprotein, and total protein contents of the two species of fish were increased in comparison to the control. In addition, dietary phytase inclusion did not significantly affect hepatic ALP, ALT, and AST activities in the two species of carp fed with different levels of MCP. Amylase activity increased in foregut and hindgut of both species when fed with the phytase‐supplemented diets while lipase activity was reduced in the foregut and hindgut in both fish. This study suggests that neutral phytase supplementation increases serum ALP, ALT, and AST activities but does not notably affect these enzyme activities in the liver of the two species of carp when fed different levels of MCP. On the other hand, amylase activity increased while lipase activity was reduced in the intestine of the species of carp fed with phytase‐supplemented diets.     

Effects of Haematococcus pluvialis on the water quality and performance of Litopenaeus vannamei using artificial substrates and water exchange systems

Two kinds of culture systems were constructed, namely, an artificial substrate system (novel sponge biocarrier, SB) and a water exchange system (WE), to evaluate the effects of the microalga Haematococcus pluvialis on the water quality and performance of whiteleg shrimp (Litopenaeus vannamei). The results showed that the average NH₄⁺-N concentration with H. pluvialis was lower than those without it. The average concentration of NO₂^?-N and NO₃^?-N in SB system were significantly decreased than those present in WE system, but there was no significant difference in SB system. The growth performance of shrimp with H. pluvialis was superior to that without it. The final average weight and growth rate of shrimp in SB1 and WE1 tanks were differed significantly among that present in SB2 and WE2 tanks (p?<?0.05). SB1 tended to have the highest average weight gain and lowest feed conversion ratio. The shrimp survival was significantly higher in SB1 and WE1 tanks (89.06% and 82.29%, respectively) than that present in SB2 and WE2 tanks (78.13% and 48.44%, p?<?0.05). The high-throughput sequencing results showed that the diversity of bacterial communities and the abundance of bacteria related to the nitrogen cycle and the degradation of organic matter were increased. The present study showed that H. pluvialis had positive effects on shrimp culture in using artificial substrate system and water exchange system.

     

点带石斑鱼对盐度适应性的初步研究

通过对体长10.4～8.0cm、平均体长8.6cm;体重19.5～12.2g平均体重16.63g的点带石斑鱼的淡化试验,分别采用盐度突变及渐变淡化两种方式,在水温27.5～29.0℃,经过30d的试验,结果是：将点带石斑鱼鱼苗从盐度（‰）30的自然海水中直接投放至盐度2.6的水体中时,其成活率100%;直接投放至盐度1.3的水体中的成活率80%。而渐变淡化,在盐度降至0.3时的成活率100%,进入纯淡水时的3d后才有逐渐死亡现象出现,10d后的成活率55%。点带石斑鱼的生长速度从纯海水至盐度5.2,随着盐度的降低其生长速度加快。研究结果证明,点带石斑鱼对盐度变化具有很强的适应能力,不但能在海水中养殖而且也可以在河口及低盐地区进行养殖。