长牡蛎食物组成的高通量测序分析

为了更加细致地甄别滤食性贝类的食物组成,于2019年8月,以北方规模化典型养殖海湾——桑沟湾养殖的长牡蛎(Crassostrea gigas)为研究对象,运用Illumina高通量测序技术对长牡蛎的胃含物及所处养殖水体中的真核生物进行分析研究。结果显示,扩增18S rDNA V4区平均得到111,359个有效序列短片段,在97%相似性水平上划分OTUs (operational taxonomic units),聚类后得到239个类别。其中,长牡蛎胃含物中的真核生物分属于34个门,绿藻门(Chlorophyta)、甲藻门(Pyrrophyta)、链形植物(Streptophyta)、硅藻门(Bacillariophyta)和原生动物(Protozoa)等为主要类群。所处养殖水体中的真核生物分属于37个门,绿藻门、脊索动物门(Chordata)、节肢动物门(Arthropoda)、甲藻门和硅藻门等为主要类群。结果表明,浮游植物是长牡蛎的主要食物来源,链型植物和原生动物也有一定的贡献,分别占总食物贡献量的10.43%和4.11%。研究结果为深入认识滤食性贝类的摄食生态学及其在养殖生态系统物质循环和能量流动中发挥的作用提供了数据支撑。     

近江牡蛎和长牡蛎的配子兼容性及合子育性

为了评估长牡蛎(GG)和近江牡蛎(AA)的配子兼容性和合子育性,本实验探究了盐度(16、20、24、28和32)、温度(18、21、24、27和30 ℃)和精子浓度(10⁰、10¹、10²、10³和10⁴个/μL)对受精率、胚胎畸形率和孵化率的影响。结果显示,随盐度的增加,GG组的受精率和孵化率逐渐上升,而AA组和AG组的受精率和孵化率先上升后下降;GA组和AG组分别在盐度28和24时具有较优的配子兼容性和合子育性,受精率、畸形率、孵化率分别为7.45%±5.05%、2.00%±0.90%、63.60%±9.88%和14.26%±9.26%、1.74%±0.93%、66.16%±9.43%。随温度上升,AA、GA和GG组的受精率和孵化率先升高后下降,而畸形率先下降后上升;GA组和AG组均在27 ℃时表现出较优的配子兼容性和合子育性,此时GA组和AG组的受精率、畸形率、孵化率分别为7.95%±4.04%、1.79%±1.04%、57.11%±9.95%和14.70%±7.27%、1.66%±0.85%、67.25%±15.19%。随精子浓度的增加,GG组的受精率逐渐上升,孵化率逐渐下降,GA组和AG组的受精率逐渐上升;GA组和AG组均在10⁴个/μL时表现出较优的配子兼容性和合子育性,此时二者的受精率、畸形率、孵化率分别为9.09%±7.53%、1.59%±0.48%、67.97%±19.96%和14.30%±6.04%、1.06%±0.68%、67.33%±12.65%。研究表明,温度、盐度和精子浓度对长牡蛎和近江牡蛎种间配子兼容性影响较小,但对合子育性影响较大。     

香港牡蛎不同育肥方式的育肥效果

香港牡蛎营养丰富,味道鲜美,在上市前会通过集中育肥提高风味口感和经济价值。为探明不同育肥方式对香港牡蛎的育肥效果,对经过池塘育肥和外海育肥后香港牡蛎的壳高、壳长、壳宽、体质量、软体质量、出肉率、糖原含量、甜菜碱含量、蛋白含量等指标进行检测,并与内海对照组牡蛎进行比较分析。检测结果显示,经过池塘育肥,香港牡蛎的壳高、体质量、糖原含量和蛋白含量均显著升高( P <0.01),甜菜碱含量显著降低( P <0.01),壳长、壳宽、软体质量、出肉率无显著变化;经过外海育肥的香港牡蛎糖原含量显著升高( P <0.01),壳高、壳长、体质量、软体质量( P <0.01)和甜菜碱( P < 0.05 )含量显著降低,壳宽、出肉率和蛋白含量无显著变化。试验结果表明,池塘育肥比外海育肥效果要好;不论哪种育肥方式,育肥后牡蛎的糖原含量均显著升高,甜菜碱含量均显著降低,出肉率均无显著变化。     

岩牡蛎正常和低盐胁迫下定量PCR内参基因的筛选与验证

实时荧光定量 PCR 已广泛用于基因表达的分析,合适的内参基因选择是获得准确分析结果的关键。本研究选用正常生理状态下岩牡蛎(Crassostrea nippona)的鳃、外套膜、闭壳肌和内脏团 4 种组织以及盐度 10、20 和 30暂养 1 周后的鳃组织为材料,对已报道的常见内参基因(EF1A、GAPDH、RO21、TUB 和 TUA)的稳定性通过 3 种方法(geNorm、NormFinder 和 BestKeeper)进行分析和筛选,发现针对鳃单一组织在不同盐度胁迫下, GAPDH 和RO21 可以作为合适内参基因,而在不同组织中,需要更多的内参基因联合分析才能获得准确的定量表达结果。本研究是首次利用 q-PCR 对岩牡蛎进行内参基因的筛选和验证,为今后该物种低盐胁迫相关的基因表达和功能基因的研究提供重要依据。     

太平洋牡蛎(Crassostrea gigas)类α-1,2-岩藻糖基转移酶的密码子优化与原核表达

牡蛎消化组织内存在的类A型血型组织抗原是其特异性富集诺如病毒的主要原因,FUT2(Fucosyltransferase 2,α-1,2-岩藻糖基转移酶)是A型血型组织抗原合成的关键酶.本研究在前期克隆了太平洋牡蛎(Crassostrea gigas)类FUT2基因cDNA全长的基础上,根据大肠杆菌密码子偏爱性优化并合成了类FUT2基因,插入原核表达载体pRSET A构建pRSET-mof,将其转化大肠杆菌BL21,用异丙基-β-D-硫代半乳糖苷(IPTG)诱导.经SDS-PAGE分析显示,在37℃、IPTG终浓度为0.8 mmol/L的条件下,诱导4h后出现大小约为46 kDa的特异性目的条带.利用His亲和层析柱纯化及超滤管浓缩目的蛋白,得到单一条带,说明纯化效果良好.Western blot分析显示,目的蛋白与抗6×His标签单克隆抗体、抗人FUT2单克隆抗体均能发生特异性反应,表明优化后的太平洋牡蛎类FUT2基因在大肠杆菌系统中成功表达.本研究结果为今后研究太平洋牡蛎类FUT2基因的功能,进一步探索牡蛎特异性富集诺如病毒的分子机理奠定了基础.     

升温与聚苯乙烯微塑料复合暴露对长牡蛎血细胞功能、免疫基因表达和能量代谢的影响

为阐明全球气候变暖和微塑料复合胁迫对长牡蛎(Crassostrea gigas)免疫应答、氧化应激和能量代谢的影响,本研究采用3个微塑料(microplastics, MPs)水平[无微塑料、小粒径聚苯乙烯微塑料(SPS-MPs,6μm)和大粒径聚苯乙烯微塑料(LPS-MPs,50～60μm)]和2个温度水平(20℃和25℃)对长牡蛎进行了为期21 d的单一和复合暴露,检测分析了各组长牡蛎血细胞功能[吞噬活性、活性氧(reactive oxygen species,ROS)含量]、糖原含量以及免疫相关基因表达的变化。研究结果表明,SPS-MPs暴露能增加长牡蛎血淋巴细胞中ROS含量,降低血细胞吞噬活性,揭示SPS-MPs毒性作用更强。升温与微塑料的协同作用增加了长牡蛎消化腺组织中的糖原含量。实时荧光定量PCR结果显示,升温与SPS-MPs复合暴露组长牡蛎消化腺组织通过上调热休克蛋白90 (heat shock protein90, HSP90)、核因子κB抑制蛋白(inhibitor of NF-κB, IκB)和p53基因表达量进行免疫应答;升温与微塑料的拮抗作用增加了鳃组织p53基...     

Pathology and mass mortality of Pacific oysters, Crassostrea gigas (Thunberg), in 2005 at the East Frisian coast, Germany

In 2005, Pacific oysters, Crassostrea gigas , were collected from May to September along the East Frisian coast and processed for histology. Because of mass mortalities in September, additional samples of moribund oysters and apparently healthy blue mussels, Mytilus edulis , were subjected to virological and ultrastructural investigation. The oysters displayed a variety of pathological conditions including viral gametocytic hypertrophy which is reported here for the first time from the German coast. Haemocyte aggregations in the digestive tract, in the intestinal mucosa and submucosa, in the mid-gut gland and in the ventricle of the heart were commonly observed at some stations. In association with mass mortalities, severe gill necrosis occurred which may have contributed to the high mortality rates. Total mortality rates of up to approximately 60% were seen. All size classes and thus age classes of oysters were affected, with highest mortality rates within the youngest age classes which had just reached sexual maturity (shell lengths <40 mm). The smallest dead oysters had shell lengths of 10 mm. The phenomenon was mainly restricted to C. gigas stocks in harbours, probably because of favourable conditions for infection, i.e. limited water exchange, less food availability, reduced oxygen content and higher pollution levels.     

Seasonal variation in the condition index of Pacific oyster postlarvae (Crassostrea gigas) in a land‐based nursery in Sonora,Mexico

This study examined the seasonal variation in the condition index (CI) of Crassostrea gigas postlarvae (<5 mm) that were cultivated at a commercial hatchery. Oysters were sampled weekly at the nursery using seawater from a lagoon for the grow‐out that precedes commercialization. Temperature, salinity, seston, chlorophyll a, oxygen and pH were recorded at each sampling and water samples were taken to identify phytoplankton groups and their abundance. High levels of primary productivity, chlorophyll a and seston were detected during summer, but the highest CI occurred in winter. During winter, elevated phytoplankton biomass was composed by diatoms and phytoflagellates, which served as the main food source and promoted weight gain in this season. Variations in salinity, oxygen and pH were not related to differences in the CI. However, it appears that the wide temperature variation affected functions, such as feeding activity, apparently enhancing ingestion during winter (mean 16.5±1.4 °C) and reducing ingestion during summer (mean 31±1.5 °C). Winter production resulted in postlarvae with a homogeneous size range and a high CI, indicating that winter is more favourable to start cultivation. The CI represents a practical means to determine the physiological state of postlarvae before transfer to cultivation sites.     

Comparative studies of the growth,survival, and reproduction of diploid and triploid Kumamoto oyster,Crassostrea sikamea

Triploid oysters have been used for farming to improve growth but have not been created in the Kumamoto oyster, Crassostrea sikamea, which is one of the crucial aquaculture species on the southern coast of China. In the present study, triploids were created using cytochalasin B to inhibit polar body II release in C. sikamea, with the untreated oysters as controls. Triploidy rates of 87 and 57.67%, on average, were obtained in larvae and adults, respectively. Larval growth and survival of the triploid were significantly lower than that of the controls (p < 0.05). In contrast, the triploid postlarvae and adults had a significant growth advantage over the controls (p < 0.05) during the period of 180 (December) to 450 days (September of the next year). Moreover, the triploids clearly exhibited significant sterility in the reproductive season. The glycogen and triglyceride contents in the gonad, adductor muscle, mantle, and gill were higher in triploids than in controls from 180 to 450 days. As a result, high physiological energy supply was strongly correlated with superior growth and reduced reproduction in triploid C. sikamea. The triploid C. sikamea is an excellent oyster species and can be used to improve growth for C. sikamea farming.