Effect of dietary supplementation of phospholipids and highly unsaturated fatty acids on reproductive performance and offspring quality of Chinese mitten crab, Eriocheir sinensis (H. Milne-Edwards), female broodstock

This study was conducted to determine the optimum phospholipids: highly unsaturated fatty acids (PL/HUFA) ratio in maturation diets for female Chinese mitten crab, Eriocheir sinensis broodstock. Five isolipidtic and isonitrogenous diets were formulated to contain different amounts of pork lard, fish oil and soy lecithin with resulting PL/HUFA levels (%dry weight) of 1.4/0.3 (Diet 1), 1.6/2.5 (Diet 2), 2.3/2.0 (Diet 3), 3.8/1.5 (Diet 4) and 5.1/0.8 (Diet 5). Diet 1 lacked supplemental fish oil and soy lecithin and served as the control. Each of the five formulated diets was fed to a total of 150 female crabs for a period of 7 months. All diet treatments were triplicated with 50 crabs stocked in a pond as a replicate. After 6 months, male crabs were introduced into the ponds where females were kept, mating, spawning and egg hatching (in seawater) occurred in the following month. The nutritional value of various formulated diets was assessed based on survival, gonadosomatic index (GSI), egg production per female, fecundity (eggs/g female weight) of the females fed those diets and egg and larval (newly hatched zoea I larvae) quality.Females fed Diet 1 had the lowest GSI, egg production per female and fecundity while no significant difference were found for survival, hepatosomatic index (HSI) and the percentage of female spawned among the diet treatments (P > 0.05). Although there were no significant differences in egg diameter, egg wet or dry weights among all treatments (P > 0.05), the highest and the second highest proportions of essential fatty acids (EFA), i.e. 20:5n-3 (EPA), 22:6n-3 (DHA) and HUFA were found in the eggs produced by the females fed Diet 3 and Diet 2, respectively. Statistical analysis showed that EPA, DHA and HUFA in eggs produced by females fed Diet 3 were significantly higher than those from the other treatments (P < 0.05). Meanwhile, zoea I larvae from crabs fed Diet 3 had significantly larger carapace length than those from the other four treatments (P < 0.05). The zoea I larvae from crabs fed Diet 2 and Diet 3 also showed generally better tolerance to starvation and osmotic shock.In conclusion, our results indicated that diet included levels of PL/HUFA around 1.6/2.5 or 2.3/2.0 (% dry weight) fed female E. sinensis broodstock supported increased fecundity and elevated level of HUFA in egg, which in turn resulted in improved overall quality of newly hatched larvae.     

Biochemical composition of pond-reared and lake-stocked Chinese mitten crab Eriocheir sinensis (H. Milne-Edwards) broodstock

The morphological characteristics, tissue indices, proximate and fatty acid compositions of hepatopancreas, gonad and muscle of pond‐reared (PR) and lake‐stocked (LS) Eriocheir sinensis broodstock were compared. Lake‐stocked crabs are characterized by a blue carapace, white abdomen, golden legs and yellow setae while PR crabs have a brown carapace, grey abdomen, black legs and similar yellow setae. The carapace length and width of PR crabs were significantly less than those of LS crabs of the same wet weight (WW). The results showed that the hepatosomatic index (HSI) of PR males was significantly higher, but the gonadosomatic index (GSI) was significantly lower for PR females when compared with LS individuals of the same sex. Regardless of their origin, the muscular index (MI) of males was higher than that of females. The hepatopancreas of LS crabs had higher protein levels and lower lipid levels compared with PR crabs. The results of proximate analysis of male gonads from two different sources were similar, while the protein level in the ovary of LS females was higher than that of PR females. The tissues of males had higher water content but lower lipid and protein levels compared with females regardless of their source. The LS crabs had higher highly unsaturated fatty acid (HUFA) levels than PR crabs in their gonad and hepatopancreas. In conclusion, our results suggest that the nutritional quality of LS crabs is better than that of PR crabs, the nutritional quality of males is poorer than that of females. The reproductive performance and larval quality of crabs from these two rearing systems should be further verified.     

中华绒螯蟹的摄食节律

为了探究中华绒螯蟹(Eriocheir sinensis)最适宜的投喂时间,本实验将120只初重约68 g的幼蟹随机分养在24个塑料缸(70 cm×40 cm×50 cm)中。采用一次饱食投喂(将昼夜分为8个时间点:10:00、13:00、16:00、19:00、22:00、01:00、04:00、07:00,每个时间段作为1个处理组,每天每个处理组饱食投喂1次)的方法研究中华绒螯蟹的昼夜摄食节律。结果显示,幼蟹在不同投喂时间点表现出24 h周期性的摄食节律,且总摄食高峰期在22:00。从其中6d的平均摄食量可以看出,河蟹主要在19:00~22:00时间段摄食;在肝胰腺酶活性方面,淀粉酶活性在19:00~22:00内高于其他各组,且在22:00获得最高值;脂肪酶、碱性磷酸酶和钠钾ATP酶酶活性有相似的变化趋势,在13:00~22:00时间段这些酶的酶活呈现上升的趋势,且最高值均在22:00获得。在相关摄食基因表达方面,肝胰腺中Leptin基因的相对表达量在22:00点显著低于13:00和16:00,与其他各组相比差异不显著(P0.05);肠道中胆囊收缩素(CCK)基因的相对表达量在22:00点显示出最低值,与10:00、13:00、04:00和07:00点相比差异显著(P0.05)。综上所述,中华绒螯蟹存在明显的摄食节律,这可能与光照紧密相关,最适宜的投喂时间为22:00。     

不同饵料对二次抱卵期间中华绒螯蟹生殖性能和HUFA组成的影响

中华绒螯蟹(Eriocheir sinensis)经一次受精便可多次产卵.与其他的虾类一样,在卵巢发育过程中,中华绒螯蟹的肝胰脏需从外界汲取许多能量和营养物质,通过血淋巴转移到卵巢以供胚胎发育和幼体孵化.亲蟹的营养需求可以通过第一次和第二次卵巢发育过程加以研究.第一次抱卵之后亲蟹消耗了大量的营养和能量物质,需要在二次抱卵前的短时间内得到及时补充.通常来说,二次抱卵蟹的产卵力低下,幼体质量差.因此研究饵料对二次抱卵期间中华绒螯蟹生殖性能和幼体质量的影响是很有意义的.本实验分4组,分别以3种天然饵料缢蛏(TC)、沙蚕(TS)、杂鱼(TT)和一种人工饵料(TA)投喂中华绒螯蟹亲蟹,投喂周期为2个月(从初始一次抱卵至二次抱卵结束).天然饵料和人工饵料的日投喂量分别为亲蟹体重的10%和1%～3%,每日下午4点投喂.用PVC板将8m2的玻璃纤维槽隔成4个单元作为试验槽.每个单元底面积为2m2,水深40 Cm.同一槽中的3个单元用做一个试验组的3个重复,每个重复为10只蟹.海水盐度为20 g·L,pH 8.20～8.30,溶解氧为8.40～8.90 mg·L-1,依据水中氨氮和亚硝酸氮的浓度(分别小于1 mg·L-1和0.2 mg·L-1),每2至3天换一次水,水温为15～17℃.实验结束后,对二次抱卵亲蟹的生殖性能(相对饱卵量、卵径和亲蟹成活率)、孵化出的幼体质量(蚤Ⅰ幼体在饥饿状态下的成活率)以及卵、卵巢和肝胰脏的脂肪酸组成进行了测定.结果表明,卵、卵巢和肝胰脏的高级不饱和脂肪酸(HUFA)组成与试验饵料的组成具有很好的相关性,其中卵的DHA与EPA水平、肝胰脏DHA水平与饵料中相应的HUFA水平成正相关(R2＞0.90),说明外界营养物质的吸收和转移对卵巢的二次发育相当重要.统计学分析表明,投喂杂鱼的亲蟹(TT)孵出的蚤Ⅰ幼体对饥饿的耐受力以及投喂缢蛏的亲蟹(TC)的卵径显著高于其他组,各组之间其他指标(相对抱卵量和成活率等)均无显著差异.二次抱卵的中华绒螯蟹的卵、卵巢和肝胰脏中HUFA的水平与试验饵料中HUFA水平具有很好的相关性,说明无论在第一次卵巢发育过程中积聚了多少营养物质,第一次抱卵后消耗殆尽的亲蟹需要从外界食物中汲取大量的能量和营养物质,通过肝胰脏转运到卵巢供二次发育之用.本研究结论与其他作者对其他甲壳类的研究结果是一致的.此外组织器官的HUFA水平不只局限于饵料HUFA水平,还受器官本身对这些脂肪酸特定需求的影响.肝胰脏作为甲壳类动物的脂肪存储器,在饵料中的脂类物质被吸收和利用之前首先将其存储起来.甲壳类动物具有合成更长链的脂肪酸能力,虽然这种能力很有限,但仍能重新合成一定数量的脂肪酸.由于卵巢和卵中的n-3 HUFA,尤其是EPA和DHA含量高于肝胰脏,因此有理由相信它们在中华绒螯蟹繁殖过程中起着非常重要的作用.用人工饵料投喂对中国对虾的研究表明,n-3 HUFA的水平与亲本的生殖性能有很大关系,其中EPA与产卵量有很大的关系,而DHA在胚胎发育早期和幼体孵化起着很大的作用.本实验结果可以推论,EPA和DHA对中华绒螯蟹的卵巢的发育和生殖性能具有相当重要的作用,其中EPA的作用更加明显.遗憾的是本实验没有得出非常明显的相关性结论.应该指出的是本实验所用的不同饵料,不但HUFA含量不同,而且在蛋白质、总脂和微量营养素(维他命、矿物质、胆固醇、磷脂、钙盐等)的组成上差异很大,因此不宜下确定的结论.     

Reproductive performance and offspring quality of Chinese mitten crab Eriocheir sinensis (H. Milne-Edwards) females fed an optimized formulated diet and the razor clam Sinonovacula constricta

After feeding female Eriocheir sinensis on an optimized formulated diet or fresh razor clam Sinonovacula constricta for 7 months, their reproductive performance and offspring quality were compared. To evaluate diet nutrient contents, the proximate, fatty acid and amino acid compositions of the formulated diet and the razor clam were analysed. The nutritional value of the diets was determined by assessing survival, gonadosomatic index (GSI) and hepatosomatic index (HSI) of female crabs from both diet treatments, together with the percentage of females that spawned, total egg production per female and fecundity (number of eggs g^?1 female wet weight). Furthermore, the quality of eggs and newly hatched larvae from the two dietary treatments were determined using the following parameters: egg diameter, wet weight and dry weight, hatchability, proximate and fatty acid profile of eggs, larval carapace length, resistant to starvation and osmotic shock, larval survival and development to the zoea II stage. Higher protein, phospholipids (PL) and amino acids (AA) contents were found in the razor clam while the formulated diet contains higher levels of ash, total lipid (TL) and 18:1n‐9, 18:2n‐6 and 22:6n‐3 fatty acids. Although female crabs fed the two different diets showed similar reproductive performances, newly hatched zoea I larvae produced by the crabs fed the formulated diet had significantly longer mean carapace length and shorter development time to the zoea II stage under identical culture condition (P<0.05). Moreover, dietary fatty acid appeared to have more significant effects on the fatty acid composition of the hepatopancreas than it did on mature ovaries or eggs. This suggests that the fatty acid profile of mature ovaries is indicative of the specific fatty acid required for ovarian development in E. sinensis. In conclusion, our results show that the optimized formulated diet developed in this laboratory can totally replace the razor clam, a broodstock food widely used in E. sinensis hatcheries in China. This encouraging result should facilitate more reliable hatchery production of this important aquaculture species.     

中华绒螯蟹微卫星多重PCR体系的建立及其亲子鉴定应用

利用自主开发的微卫星标记对中华绒螯蟹(Eriocheir sinensis)进行微卫星多重PCR体系构建,最终成功建立4组多重体系,每组体系包含4个微卫星位点,并成功应用于3个家系的亲子鉴定中。结果显示:(1)本研究筛选的16个微卫星标记,平均观测杂合度为0.8206,平均期望杂合度为0.8164,平均多态信息含量为0.7927,具有丰富的多态性;(2)运用Cervus 3.0软件对已知系谱信息的3个中华绒螯蟹家系共95个子代个体进行亲子鉴定分析,结果显示,选用任意两组微卫星多重PCR体系时,累积实际正确鉴定率均超过94.74%;使用任意三组微卫星多重PCR体系时,累积实际正确鉴定率均大于98.95%;使用四组微卫星多重PCR体系时,累积实际正确鉴定率达到100%。并且当选用组合1、2和组合3,或者选用组合1、3和组合4时,对3个家系的累积正确鉴定率达到了100%,因此选用这些组合不但可以获得准确的系谱信息,还能减少工作量,降低成本。本研究构建的微卫星多重PCR体系能为中华绒螯蟹的种群选育和家系管理提供便捷、高效的途径。     

超低温冷冻对中华绒螯蟹胚胎形态结构的影响

为了解超低温冷冻对胚胎形态结构的影响,以中华绒螯蟹为研究对象,运用石蜡切片、扫描电镜和透射电镜观察细胞分裂期、原肠期和原溞状幼体期胚胎,分别用玻璃化液处理和经超低温冷冻后外部形态和内部结构的变化。结果发现:(1)显微观察表明,细胞分裂期胚胎在玻璃化液中用二步平衡法处理后吸水膨胀明显,三步平衡后胚胎形态无明显变化,超低温冷冻后,卵黄物质从细胞中溢出,细胞破损严重;原肠期胚胎在玻璃化液中用二步平衡法处理后,外部形态与鲜胚无明显差异,经过冷冻后,所有胚胎内部变成粉红色,胚体由原来的透明变成不透明状,细胞膜边缘模糊似绒毛状;(2)扫描电镜观察,玻璃化液处理后的所有原肠期胚胎表面褶皱呈沟壑状,形成一层网状结构;透射电镜观察,处理后胚胎细胞内出现白色团块,细胞边缘变得粗糙有突起,细胞内冰腔清晰可见,空泡形成,80%以上线粒体解体,细胞破裂明显;(3)组织切片观察,原肠期胚胎细胞外面的膜脱落破损,胚层内有大小不一的冰腔,细胞内出现明显的空泡。原溞状幼体期胚胎经过玻璃化液处理后,外部形态与鲜胚间无明显区别,但经过超低温冷冻后,95%以上胚胎组织呈弥散状,部分卵黄物质碎裂成颗粒状,胚胎的细胞膜脱落,胚层内出现大量冰腔和空泡,90%胚胎表面皱缩凹陷,但仍有10%的胚胎表面保持光滑完整,表明原溞状幼体期胚胎是适合进行冷冻保存的时期。     

不同规格中华绒螯蟹亲本子一代在扣蟹阶段养殖性能的比较研究

为探究中华绒螯蟹(Eriocheir sinensis)亲本规格对其子代在扣蟹阶段养殖性能的影响,本试验在相同的养殖环境下将4种规格河蟹亲本(规格I:雌90~110 g、雄140~160 g;规格Ⅱ:雌115~135 g、雄160~180 g;规格Ⅲ:雌140~160 g、雄220~240 g;规格Ⅳ:雌190~210 g、雄290~310 g)所产的大眼幼体养殖至扣蟹,并比较各组扣蟹的体质量、增重率、特定生长率、成活率及最终产量差异。试验结果显示:(1)不同规格的河蟹亲本,其子一代在大眼幼体至豆蟹阶段的平均体质量、成活率以及增重倍数均无明显差异(P>0.05);(2)在豆蟹至扣蟹养殖阶段,各组幼蟹的生长速度差异大体上与其亲本规格的差异相一致,其中Ⅰ组幼蟹在8—10月期间的平均体质量均显著低于Ⅳ组(P<0.05);(3)养殖效果方面,各组扣蟹平均规格和最终产量均随着亲本规格的提高而提高,其中Ⅳ组扣蟹的最终平均体质量显著高于Ⅰ组扣蟹(P<0.05);(4)就规格分布而言,Ⅳ组扣蟹2.00~3.99 g规格的比例显著低于另外3组,而≥6.00 g规格的比例显著高于另外3组(P<0.05)。结果表明,中华绒螯蟹亲本规格对其后代的养殖性能存在影响,大规格河蟹亲本子一代在扣蟹生产阶段生长速度快、养殖效果好,因此建议在生产上选择大规格亲本的后代用于扣蟹养殖。     

长江水系野生和养殖中华绒螯蟹生殖性能、胚胎色泽和生化组成的比较

生殖性能是评价水生动物亲本质量的重要指标,本研究测定和比较了野生和养殖中华绒螯蟹亲本的生殖性能、胚胎质量、胚胎色泽、常规生化和脂肪酸组成。结果显示:①野生中华绒螯蟹亲本的生殖力、生殖指数和抱卵量略高于养殖亲本的,但2组间无显著性差异,野生和养殖中华绒螯蟹亲本的胚胎单卵湿重、单卵干重和卵径无显著性差异;②养殖组冻干胚胎的红度(a~*)、黄度(b~*)值和总类胡萝卜素含量显著高于野生组亲本,然而2组胚胎的亮度(L~*)和色差值(dE~*)无显著性差异;③2组亲本所产胚胎的水分、粗蛋白和粗脂肪含量均无显著性差异,但其脂肪酸组成差异显著,野生组胚胎的C18:1n9、C18:1n7、C20:4n6、C22:5n3和C22:6n3显著高于养殖组,但其C18:2n6和C18:3n3含量显著低于养殖组。研究表明,池塘养殖和野生亲本的生殖性能和胚胎中常规生化组成无显著差异,但2组胚胎的色泽、总类胡萝卜素含量和部分脂肪酸组成存在显著差异,该研究为中华绒螯蟹亲本选择、生殖性能评价和人工育苗提供了重要基础资料。     

不同品系中华绒螯蟹土池生态育苗效果比较

为评估不同品系中华绒螯蟹（Eriocheir sinensis）的繁殖性能和育苗效果,分别采用中华绒螯蟹二龄早熟品系（雌体≥150 g,雄体≥210 g）、“长荡湖1号”A系（雌体≥200 g,雄体≥310 g）、“长荡湖1号”B系（雌体≥250 g,雄体≥360 g）及超大规格品系（雌体≥300 g,雄体≥400 g）作为亲本进行土池生态育苗试验,比较了各品系的抱卵蟹获得率、亲本孵化率、育苗期间的投饵量、幼体变态速度以及大眼幼体产量等指标。试验结果显示：（1）各品系越冬后的抱卵蟹获得率随着亲本规格的增大而明显降低,抱卵蟹成功孵化率随着亲本规格的增大同样呈降低趋势,但组间差异不明显（P>0.05）。（2）各品系投饵量在幼体培育期间呈现先增后减的变化趋势,其中,溞Ⅴ期最高,日投饵量约为75 g/m²;与之相反,幼体蜕皮周期在幼体培育过程中呈现出先降后增的变化趋势,溞Ⅲ-Ⅳ期蜕皮周期最短,仅为5～6 d。（3）除了个别产量异常的池塘外,各品系单蟹产苗量及饵料系数均随着亲本规格的增大而提高,且各品系大眼幼体淡化率差异较小,均在90%左右。结果表明,虽然大规格品系在...     

L-色氨酸和褪黑激素对中华绒螯蟹(Eriocheir sinensis)血清血糖水平及肝胰腺抗氧化能力的影响

为探究L-色氨酸和褪黑激素（MT）对中华绒螯蟹血糖水平及肝胰腺抗氧化能力的影响。实验选取初始体质量为（16.58±2.20）g的雌性中华绒螯蟹,随机分成8组（L-色氨酸和MT处理各4组）,实验周期均为30 d。（1）L-色氨酸饲料投喂实验4组：L-色氨酸所占饲料比重分别为0.36%（基础饲料即对照组）、0.47%、0.73%和1.05%（分别记为1、2、3和4组）,30 d后分别测定各组蟹的血糖水平及肝胰腺的T-AOC和SOD活性;（2）MT注射实验4组：每组分别在第1天和15天时注射生理盐水（即对照组,C组）和不同剂量的MT 10^-6 mol/只（μM组）、10^-9 mol/只（nM组）和10^-12 mol/只（pM组）。分别于第1天和15天时,收集血淋巴和肝胰腺,分别测定血糖水平、T-AOC和SOD活性。于第30天时,再次收集血淋巴以测定血糖水平。结果显示：不同含量L-色氨酸对中华绒螯蟹血糖水平无明显影响,其血糖含量约为（4.62±0.20）mmol/L。此外,0.73% L-色氨酸的饲料能显著提高肝胰腺SOD活性（82.86±1.07）U/mL;注射MT仅在第1天时对中华绒螯蟹的血糖水平有促进作用,并呈剂量依赖性的方式升高,其中μM组蟹血糖水平高达（7.56±0.36）mmol/L,其值显著高于C、pM和nM组。此外注射MT在第1天时,对肝胰腺T-AOC和SOD活性也有明显的促进作用。其中,μM组蟹肝胰腺T-AOC和SOD活性均显著高于C、pM和nM组。研究表明,饲喂0.73% L-色氨酸的饲料能够提高中华绒螯蟹的抗氧化能力,但对血糖水平的影响不明显;注射10^-6 mol/只MT可以在短时间内（1 d）提高中华绒螯蟹的血糖水平和抗氧化能力。     

大规格中华绒螯蟹亲本子代大眼幼体在微山湖地区的池塘扣蟹培育试验

为探究大规格中华绒螯蟹(Eriocheir sinensis)亲本子代大眼幼体在山东微山湖地区的池塘养殖效果,2019年在微山湖扣蟹基地将2种大规格河蟹亲本(规格I:雌190～210 g、雄280～300 g;规格Ⅱ:雌240～260 g、雄330～350 g)所产的大眼幼体饲养至扣蟹,并比较各组扣蟹的生长情况、养殖性...     

Effect of lactic acid on enrofloxacin pharmacokinetics in Eriocheir sinensis (Chinese mitten crab)

As a representative acidifier, lactic acid (LA) is widely used in the diets of aquatic animals. LA is the main supporter of energy metabolism and may be associated with drug metabolism. This study was designed to explore the pharmacokinetics of enrofloxacin (ENR) in Eriocheir sinensis (Chinese mitten crab) fed diets containing 0.1%, 0.2% and 0.3% LA (designated groups LA1, LA2 and LA3 respectively). The concentrations of ENR in the haemolymph, hepatopancreas and muscle were determined by HPLC after a single oral dose of 10 mg/kg. Our results showed that LA had a significant effect on the peak ENR concentrations in all the tissues (p < 0.05) by one‐way ANOVA analysis. There was a trend that C_max (peak concentration) of ENR was elevated with LA levels increasing up to 0.3% in haemolymph, hepatopancreas, muscle and T_max (time‐point of the peak concentration of the drug), t_1/2β (elimination half‐life) and AUC_(0‐∞) of ENR were shortened. Taken together, 0.3% LA might be effective in improving ENR pharmacokinetics in E. sinensis. Furthermore, it can be speculated that the enhanced biotransformation of ENR in the hepatopancreas mediated by LA is responsible for the differences in the pharmacokinetics of ENR in E. sinensis.     

3种品系中华绒螯蟹扣蟹阶段池塘养殖性能比较

为探究不同品系中华绒螯蟹(Eriocheir sinensis)在扣蟹阶段的池塘养殖效果,于2020年在江苏东台进行了3个品系(2龄早熟品系、“长荡湖1号”A系和“长荡湖1号”B系)大眼幼体的池塘养殖试验,比较了各品系扣蟹的平均体质量变化情况、最终养殖性能、规格分布和经济效益。结果表明:(1)虽然3个品系大眼幼体在扣蟹阶段的平均体质量并无显著差异(P＞0.05),但各品系扣蟹平均体质量与亲本规格呈一定的正相关关系,总体上为“长荡湖1号”B系＞“长荡湖1号”A系＞2龄早熟品系;(2)正常扣蟹单位面积产量和扣蟹总产量与亲本规格呈正相关关系,即随着亲本规格的增大而显著增加(P<0.05);此外,“长荡湖1号”B系饲料系数(FCR)显著高于“长荡湖1号”A系(P<0.05);(3)3个品系扣蟹的体质量主要分布于6.0~7.9 g和8.0~9.9 g的规格区间内,其中2龄早熟品系在小规格区间(＜4.0 g和4.0~5.9 g)所占的比例较高,而“长荡湖1号”B系扣蟹在大规格区间(10.0~11.9 g和＞12.0 g)所占比例较高,但组间并无显著差异(P＞0.05);(4)各品系扣蟹的最终收益及投资回报率均随着亲本规格的增大而明显提高。综合来看,相对于2龄早熟品系,“长荡湖1号”A系和“长荡湖1号”B系在扣蟹养殖阶段具有产量高、投资回报率高等特点,市场应用和产业化前景较好。     

亲本强化培育对中华绒螯蟹雌体生殖性能和Z1幼体质量的影响

系统比较了亲本强化培育和常规培育对池塘养殖中华绒螯蟹的生殖性能和Z_1幼体质量的影响,同时分析2组Z_1幼体的脂肪酸组成。结果表明,①两种培育方式对亲本的成活率和产卵率均无显著影响,强化组亲本的GSI、HSI、产卵量和生殖力显著高于常规组;②两种培育方式对卵的卵径、水分含量、单卵干重和湿重影响不显著,但强化组的受精率和孵化率略高于常规组;③强化组Z_1幼体的头胸甲长度、变态速度和抗逆境能力均高于常规组,这可能和幼体中高含量的HUFA有关;④生产性实验结果表明强化组的出苗率高于常规组,且幼体培育时间减少4～5 d。因此,亲本强化培育可以显著提高绒螯蟹中华的生殖性能和Z_1幼体质量。     

中华绒螯蟹幼体的栖息生境选择与行为特征

为探究中华绒螯蟹在早期发育阶段对不同栖息生境的选择偏好及其行为特征，实验以长江口近岸微生境类型为基础，在室内建立了藨草、泥底、铁板沙、砾石等4种模拟生境，选取中华绒螯蟹大眼幼体、Ⅰ、Ⅲ、Ⅴ期仔蟹分别开展了对不同生境选择的行为观察和定量分析。结果显示，中华绒螯蟹大眼幼体、Ⅰ、Ⅲ、Ⅴ期仔蟹在藨草生境中分布比例最高，显著高于其他三种生境。随着仔蟹的生长发育，Ⅲ、Ⅴ期仔蟹在泥底和铁板沙生境中占比增大。在藨草生境中，中华绒螯蟹幼体具有游泳、附着藨草、表栖、埋栖等4种行为，大眼幼体和Ⅰ期仔蟹主要表现为附着藨草行为，Ⅲ期仔蟹的表栖、埋栖行为显著增加。幼体在藨草生境中的蜕壳率显著高于泥生境，而死亡率显著低于泥生境。研究表明，中华绒螯蟹幼体偏好栖息于有利于其生存、发育的藨草等植被生境中，这可能与藨草等植被生境为中华绒螯蟹幼体提供了适宜的庇护场所有关。本研究结果指出了植被生境对中华绒螯蟹幼体阶段的重要性，丰富了早期生活史资料，并为其资源保护和栖息地生态修复提供了基础参考。     

Effect of dietary HUFA on tissue fatty acid composition and reproductive performance of Chinese mitten crab <Emphasis Type="Italic">Eriocheir sinensis</Emphasis> (H. Milne-Edwards) broodstock

This study was conducted to determine the optimum highly unsaturated fatty acid (HUFA) level in semi-purified diets for female Chinese mitten crab Eriocheir sinensis broodstock. Five isolipidic and isonitrogenous diets were formulated to contain different HUFA levels (0.14, 0.65, 1.15, 1.64, and 2.07%, respectively) by the supplementation of HUFA-rich fish oil. Diets were fed to triplicate groups of 25 female crabs (75 animals per treatment) stocked in concrete tanks for 8 months. The hepatopancreas, ovary, and muscle of the crabs were sampled after 3-month culturing, and the eggs were sampled after 8-month culturing. The reproductive performance of E. sinensis broodstock was evaluated in relation to the different dietary HUFA levels. Moreover, the fatty acid composition of the hepatopancreas, ovary, muscle, and eggs was determined in crabs fed various levels of HUFA. The results showed that a significant positive correlation was observed between the dietary HUFA level and the HUFA content in hepatopancreas, ovary, and egg, respectively. However, no significant difference in gonadosomatic index, hepatosomatic index, and total ovarian and hepatopancreatic lipid content among the treatments was observed after 3-month culturing. Also, the reproductive performance was similar for all treatments after 8-month culturing. The fact of lacking significant correlation between reproductive parameters and dietary HUFA level was discussed in respect of the catadromous life cycle of E. sinensis and warranted further research with more comprehensive approaches.     

Akt在中华绒螯蟹免疫中的功能

Akt作为PI3K-AKT 信号通路的核心分子,在细胞增殖、代谢、发育和存活等过程中发挥重要作用。本研究采用特异性引物及PCR技术,克隆获得了中华绒螯蟹Akt基因 (EsAkt)并进行生物信息学分析,实时定量PCR技术检测了EsAkt在中华绒螯蟹不同组织和不同免疫刺激后的表达水平,免疫荧光技术检测了EsAkt蛋白的细胞定位,双链RNA干扰技术分析了EsAkt干扰后凋亡基因的变化。结果显示,EsAkt分子包含PH结构域、中心催化结构域 (S_TKc)和羧基端疏水结构域 (S_TK_X) 3个保守的结构域。EsAkt在所有被检测的组织中呈现组成型表达,且在免疫相关组织 (如肝胰腺、鳃和血淋巴细胞)中的表达量显着高于胃和肠道。EsAkt蛋白主要以弥散状的形式分布在细胞质中。中华绒螯蟹注射脂多糖 (LPS)和嗜水气单胞菌后,显著诱导EsAkt转录本的表达,LPS免疫刺激12 h后,EsAkt响应达到峰值,为对照组的4.35倍,刺激48 h后,EsAkt的mRNA表达水平有所降低,但仍是对照组2.62倍。嗜水气单胞菌免疫刺激6 h后,EsAkt响应达到峰值,为对照组的9.05倍,刺激48 h后,EsAkt的mRNA表达量回落到正常水平。双链RNA干扰EsAkt后,EsAkt的mRNA表达量为对照组的0.38倍,而凋亡相关基因EsCaspase-3-like表达水平显著上调,为对照组的2.69倍。研究表明,EsAkt基因在中华绒螯蟹免疫中发挥重要作用,参与了机体对细菌引起的免疫应激过程。本研究丰富了中华绒螯蟹PI3K-AKT信号通路研究的基本资料,为进一步研究甲壳动物免疫防御机制奠定了基础。     

育肥饲料中虾青素含量对雄性中华绒螯蟹肠道和鳃部可培养优势细菌数量和组成的影响

在育肥饲料中添加虾青素会影响中华绒螫蟹肠道及鳃内的可培养优势菌群以及机体的免疫力,本实验旨在探讨饲料中虾青素含量及养殖水体对其肠道和鳃部菌群的影响。采用不同虾青素含量的饲料(0.00、26.60、41.62、81.37和75.35 mg/kg,分别对应饲料1#~5#)对雄性中华绒螫蟹进行育肥70 d后,利用基础培养基和选择性培养基对其肠道、鳃及养殖水体中的细菌进行传统分离和纯化,所得菌株进行16S rRNA测序,再进行同源性分析后做系统进化树。结果发现,在分离获得的106株细菌(登录号:KU570293~KU570368,KU570370,KU570372~KU570383,KU601302~KU6013 16,KU720553)中,92株为优势菌株,其中,肠道、鳃部和水体各29、32和31株。肠道中的可培养优势细菌属于柠檬酸杆菌属、假单胞菌属和气单胞菌属;而鳃中可培养优势细菌属于柠檬酸杆菌属、芽孢杆菌属、假单胞菌属和气单胞菌属;养殖水体中可培养优势细菌属于芽孢杆菌属、气单胞菌属、柠檬酸杆菌属、假单胞菌属和黄杆菌属。饲料2#组肠道中可培养细菌总数最高(1.06×10~8cfu/g);饲料5#肠道中的潜在致病菌数量最低,但其可培养细菌总数显著高于其他饲料组;饲料3#组肠道和鳃中潜在致病菌数量相对较高。研究表明,饲料中添加不同含量的虾青素能够显著影响雄性中华绒螫蟹肠道和鳃中的菌群构成,但对水体中可培养细菌数量无显著影响。本研究首次对投喂不同含量虾青素饲料的育肥期雄性中华绒螫蟹肠道、鳃及养殖水体可培养的优势细菌数量和组成进行分析,探究了饲料中虾青素含量以及养殖水体与蟹肠道内可培养细菌之间的联系,为中华绒螫蟹营养免疫和菌群调控提供了理论基础。     

中华绒螯蟹对赖氨酸、蛋氨酸和精氨酸的需要量

试验以酪蛋白、白鱼粉、豆粕和晶体氨基酸为蛋白源,配制了15组试验饲料,分别测定了中华绒螯蟹幼蟹对赖氨酸(Lys)、蛋氨酸(Met)和精氨酸(Arg)的需要量。养殖试验在室内玻璃纤维水族缸中进行,试验水温为(28±3.0)℃,pH为7.4左右,试验周期为56 d。试验组1～试验组5为Lys试验,试验组6～试验组10为Met试验,试验组11～试验组15为Arg试验,试验用蟹平均初重分别为:Lys试验(2.050±0.027)g,Met试验(2.027±0.033)g,Arg试验(2.05±0.032)g。每个试验设5个氨基酸梯度,其中Lys梯度为:1.58%、1.98%、2.38%、2.78%和3.18%;Met梯度为:0.50%、0.75%、1.00%、1.25%和1.50%;Arg梯度为:1.39、2.09、2.79、3.49和4.19%。每个梯度设3重复。试验结果表明,Lys试验中,试验组3(含2.38% Lys)获得最大的增重率和特定生长率,但与试验组2(1.98%Lys)和试验组4(2.78%Lys)之间没有显著性差异;且试验组3的全蟹水分含量显著较低,而蛋白质含量显著较高(P<0.05)。Met试验中,试验组8(含1.00% Met)的增重率和特定生长率显著较高(P<0.05),而其它试验组的增重率和特定生长率差异不显著(P>0.05);试验组10 全蟹水分含量显著较高,而粗蛋白含量显著较低(P<0.05)。Arg试验中,试验组14(日粮含3.49% Arg)的增重率和特定生长率显著高于其它试验组(P<0.05);而各试验组间全蟹营养组成均没有显著性差异。根据试验的生长反应和二次曲线回归分析,确定了中华绒螯蟹幼蟹饲料中最适赖氨酸、蛋氨酸和精氨酸的需要量分别为2.34%,1.12% 和 3.62%。