Molecular Characterization of the Von Willebrand Factor Type D Domain of Vitellogenin from Takifugu flavidus

The von Willebrand factor type D (VWD) domain in vitellogenin has recently been found to bind tetrodotoxin. The way in which this protein domain associates with tetrodotoxin and participates in transporting tetrodotoxin in vivo remains unclear. A cDNA fragment of the vitellogenin gene containing the VWD domain from pufferfish (Takifugu flavidus) (TfVWD) was cloned. Using in silico structural and docking analyses of the predicted protein, we determined that key amino acids (namely, Val¹¹⁵, ASP¹¹⁶, Val¹¹⁷, and Lys¹²²) in TfVWD mediate its binding to tetrodotoxin, which was supported by in vitro surface plasmon resonance analysis. Moreover, incubating recombinant rTfVWD together with tetrodotoxin attenuated its toxicity in vivo, further supporting protein–toxin binding and indicating associated toxicity-neutralizing effects. Finally, the expression profiling of TfVWD across different tissues and developmental stages indicated that its distribution patterns mirrored those of tetrodotoxin, suggesting that TfVWD may be involved in tetrodotoxin transport in pufferfish. For the first time, this study reveals the amino acids that mediate the binding of TfVWD to tetrodotoxin and provides a basis for further exploration of the molecular mechanisms underlying the enrichment and transfer of tetrodotoxin in pufferfish.     

Dietary protein requirement of juvenile obscure puffer,Takifugu obscurus

A 10‐week feeding experiment was conducted to determine the optimum dietary protein requirement of juvenile obscure puffer (Takifugu obscurus). Six isoenergetic (20 MJ kg^?1 gross energy) diets were formulated to contain graded levels of 34%, 38%, 42%, 46%, 50% or 54% crude protein (as dry matter basis). The results showed final body weight, weight gain and specific growth rate (SGR) increased significantly with increasing protein levels up to 42% and then decreased thereafer. Second‐order polynomial regression analysis (y = ?0.0024x² + 0.1788x ? 1.3196, R² = 0.9032) indicated a maximum SGR at protein level of 37%. Feed conversion ratio (FCR) decreased with increasing levels of dietary protein up to 42% and increased thereafter. Second‐order polynomial regression analysis (y = 0.0054x² ? 0.4351x + 10.391, R² = 0.753) indicated a minimum FCR at protein level of 40%. Protein efficiency ratio (PER) of fish fed the 34%, 38% and 42% diets was significantly higher than that of fish fed the 46%, 50% and 54% diets, and broken‐line analysis indicated PER tended to decrease when dietary protein level was higher than 40%. Generally, whole body lipid content, total cholesterol, low‐density lipoprotein cholesterol and triacylglycerol decreased with increasing levels of dietary protein. Fish fed the 42% protein diet showed the highest essential amino acids (histidine, isoleucine, leucine, lysine and threonine) and non‐essential amino acids (aspartic acid and glutamic acid) in muscle. Based on the second‐degree polynomial regression analysis of SGR and FCR and broken‐line analysis of PER, the optimal dietary protein level of obscure puffer is estimated to be between 37% and 40% (% as dry matter basis).     

饲料中胆汁酸对红鳍东方鲀脂肪酸组成及抗氧化能力的影响

为研究饲料中添加胆汁酸对红鳍东方鲀脂肪酸组成及抗氧化能力的影响，以鱼油为主要脂肪源，配制五种等氮饲料，以初始体重14g的红鳍东方鲀进行为期56天的投喂养殖实验。设置适宜脂肪含量(8%)的对照组，并在对照组添加0.02%及0.10%的胆汁酸，分别获得低胆汁酸添加组(BA)和高胆汁酸组(HBA)。另设一高脂组(脂肪含量12.5%，HL组)及高脂+0.02%胆汁酸组(HLBA组)。结果表明，添加胆汁酸的处理组肌肉和肝脏MUFA均低于对照组，其中HBA组显著低于对照组(P<0.05)。肌肉中n-6PUFA含量在HLBA和HL组显著低于其他组(P<0.05)。在适宜脂肪含量的三个处理组间，肝脏中的C20:0、C16:1n-7、C18:1n-9、C18:2n-6、C20:4n-6、C18:3n-3、C18:4n-3、C20:5n-3和C22:6n-3都呈现随着胆汁酸添加量增加而降低的趋势。血清超氧化物歧化酶活性和谷胱甘肽过氧化物酶活性都有随着胆汁酸添加而升高的趋势，且在高脂组显著高于对照组(P<0.05)；血清谷胱甘肽还原酶则呈现相反的趋势。肝脏中谷胱甘肽过氧化物酶1的mRNA相对表达量在对照组显著高于其他组，而BA组氧化还原酶1的基因表达量显著高于对照组；在高脂组添加胆汁酸显著升高了过氧化氢酶的基因表达量。综上所述，饲料中胆汁酸的添加降低了饲料及鱼体组织中一系列长链脂肪酸的含量；胆汁酸对血清抗氧化蛋白活性的影响与其对肝脏抗氧化相关基因表达的影响结果不一致。     

半咸水人工湿地净化越冬养殖循环水的效果

为拓展人工湿地在水产养殖上的应用,该文研究了半咸水人工湿地的循环水越冬系统中暗纹东方鲀越冬养殖效果和净化效能。结果表明,经过166d越冬养殖,暗纹东方鲀越冬成活率(76.94±13.87)%,饵料系数5.49±1.77。在无任何外源能源加温的情况下,系统水温保持在暗纹东方鲀幼鱼的生存最低温度以上。在半咸水环境下(7.6‰～9.9‰),人工湿地对养殖水有良好的净化效果:总氮去除率为30.5%、总氨氮为69.6%、亚硝酸盐为96.1%、硝酸盐为9.7%、总磷为18.4%、COD为38.8%、总悬浮物为79.7%,循环水经过人工湿地后水质符合国家渔业用水标准;越冬期间,越冬养殖池水质状况良好;除TN和NO3-N在越冬期间的去除率比在夏季的明显低外,其他主要污染物的去除率与夏季比较没有明显变化。在试验负载范围内,系统去除量随进水污染负载量的增加而增加;通过建立预测模型预测出湿地养殖面积比值0.36～0.97,与实际值1.26相比,养殖密度尚有提高的空间。因此,人工湿地能持续有效地去除暗纹东方鲀越冬循环水养殖系统中的主要污染物,说明该系统能在越冬养殖生产实际应用。     

光唇鱼网箱养殖试验

网箱单养、混养光唇鱼的试验结果表明,在25m2网箱中,单养光唇鱼以200～220尾/m2的放养密度比较合适,单位面积产量可达12.7～12.9 kg/m2;瓯江彩鲤网箱中混养光唇鱼,在瓯江彩鲤保持180尾/m2的放养密度不变的情况下,混养20尾/m2的光唇鱼,其效果比单养瓯江彩鲤更好,单位面积产量可达103.2kg/m2.     

外源人绒毛膜促性腺激素诱导黑边石斑鱼排卵的研究

【目的】了解外源人绒毛膜促性腺激素(human chorionic gonadotropin,HCG)诱导黑边石斑鱼(Epinephelus fasciatus)排卵的影响。【方法】以成熟的黑边石斑鱼为研究对象,测量其体长、体质量和肥满度后,在2018年7—8月,使用HCG激素(剂量为200 IU/kg或500 IU/kg)注射分别对6尾雌鱼进行成熟诱导。【结果】在200 IU/kg HCG处理中,注射24 h前有5尾雌鱼的卵母细胞处于第三次卵黄球前期阶段;注射48 h后2尾雌鱼出现核移动卵母细胞,1尾雌鱼的卵母细胞为第三次卵黄球后期阶段;注射60 h后1尾雌鱼出现排卵现象,其余5尾雌鱼均为第三次卵黄球后期阶段。在200 IU/kg HCG处理的排卵个体中,注射时其卵母细胞直径为477.0 μm,注射后60 h卵母细胞的直径增加至624.4 μm。此外,经催产、排卵、受精和孵化后共获得总卵数16 906粒,受精率为68.7%,孵化率为43.0%。而500 IU/kg HCG处理的黑边石斑鱼排卵失败。【结论】使用剂量为200 IU/kg的外源HCG激素可诱导黑边石斑鱼成熟且排卵,但为了提高排卵率、受精率和孵化率,其注射剂量和效应时间还需要进一步调节。     

池养无毒暗纹东方鲀的人工繁殖

研究了温室(18～25℃)池养暗纹东方鲀性腺发育、人工繁殖和河鲀毒素.试验结果表明池养暗纹东方鲀雄鱼成熟系数二、三月份达到最大值分别为0.1480±0.0100和0.1317±0.0l40;三、四月份雌鱼成熟系数达到最大值分别为0.2440±0.0400和0.3058±0.0460.与野生的暗纹东方鲀相比较,经统计检验,P＞0.05,成熟系数无显著性差异.池养暗纹东方鲀不经降海洄游,性腺完全能发育成熟,能进行人工催产并孵化得苗,并且证明活体各部分均无毒或弱毒,这对于大规模开展暗纹东方鲀的人工养殖具有重要意义.     

红鳍东方鲀仔鱼期摄食与生长的研究

在水温 ( 1 6± 1 )℃条件下 ,红鳍东方 (Takifugurubripes)仔鱼孵出后 4d开始摄食 ,此时其卵黄囊体积由初孵时的 0 636mm3减少到 0 375mm3;1 2d时 ,卵黄完全耗尽。最高初次摄食率出现在卵黄耗尽时 ,可达 1 0 0 % ;仔鱼能维持 9d高达 90 %以上的初次摄食率。初次摄食强度最大值也出现在 1 2d。饥饿不可逆点 (PNR)出现在 1 5～ 1 6d。在内营养期日平均增长率为 0 2 1 7mm/d ;在PNR前的摄食期 ,饥饿仔鱼的日平均生长率为 0 0 1 7mm/d ;而摄食仔鱼的日平均生长率为 0 0 95mm/d ;PNR后至死亡前 ,饥饿仔鱼呈负增长 ,而摄食仔鱼的日平均生长率达 0 41 0mm/d。     

条石鲷胚胎及仔稚鱼的发育

通过对条石鲷胚胎发育和仔鱼发育形态的观察，确定了条石鲷早期发育的进程。条石鲷的受精卵为圆球形、透明、浮性，卵径为0．90～0．95mill，油球1个，卵裂为盘状卵裂。在水温23～24℃，盐度26～28的条件下，受精后7h 10mirh胚胎发育至原肠期；受精后12h，眼囊形成；受精后20h 30min，心跳开始搏动；受精后26h 30min仔鱼孵出。初孵仔鱼全长1．78～2．03min，至5日龄全长3．25～3．30min时，卵黄囊完全消失；至8～15日龄，全长3．50～4．97mm时，鳔形成；至18．20日龄，全长7．10～7．31min时，尾鳍鳍条发育完整。