紫贻贝和厚壳贻贝杂交及F1代杂交优势初探

2005年-2007年对紫贻贝和厚壳贻贝种间进行了杂交生产性试验，通过亲贝强化培育、确认雌雄亲贝、改革育苗水体交换方法、投喂混合单细胞藻类、流水培养附着稚贝等技术，获得F₁代。结果表明，各试验组生长特性，正交F₁代﹥紫贻贝﹥厚壳贻贝﹥反交F₁代。其中正交F₁代获附着稚贝3.77亿粒，平均壳长为1.13 mm、平均壳高为0.78 mm，育苗成活率达35.57%。正交F₁除在孵化率上低于紫贻贝、厚壳贻贝外，在壳长、壳高、成活率等方面指标具有一定杂交优势，而反交F1代的杂交优势不明显。     

厚壳贻贝人工繁殖技术的研究

厚壳贻贝(Mytilus coruscus)是我国贻贝的主要养殖品种,其苗种主要依靠天然苗种和半人工采苗获得。2006年浙江海洋学院于嵊泗县石柱育苗厂,首次突破了厚壳贻贝规模化全人工繁殖和稚贝海区中间培育技术。研究结果表明,厚壳贻贝亲贝通过室内强化培养,经人工催产可获得成熟受精卵,受精率可达95%。在水温16℃时,受精卵在受精后25min出现第一极体,受精后39h50min发育至直线绞合幼虫期,胚胎孵化率达92%,在水温15.8～21℃的条件下,经39d室内人工培育,获平均壳长0.694mm的附着稚贝1304.7×104ind;附着稚贝经102d海区中间培育,获平均壳长13.95mm的稚贝404.46×104ind,海区保苗成活率达31%。该研究结果为今后厚壳贻贝大规模苗种生产奠定了重要理论和技术基础。     

杂交贻贝同工酶表达的初步分析

运用聚丙烯酰胺凝胶电泳技术,对2种贻贝(厚壳贻贝、紫贻贝)稚贝及通过人工培育所得的二者的杂交后代稚贝同工酶表达状况进行分析.结果显示,所检测5种酶(SOD、MDH、ME、ATP、ADH)在上述3种贻贝中的表达极为相似,MDH在3种贻贝中除r迁移率相同的条带外,在紫贻贝中表达出迁移率较小、表达较弱的条带;SOD和ATP仅在厚壳贻贝中表达出比另外2种多1条迁移率最大的条带,ME则表现为紫贻贝少1条中间的条带,ADH在杂交贻贝和紫贻贝中表达完全相同,厚壳贻贝虽也表现为3条酶带,但迁移率明显不同.试验结果说明,厚壳贻贝和紫贻贝具有较近的亲缘关系,其杂交后代与二者均有一定的相似性,这与传统的分类方法所得的结论相一致.     

硅烷化表面海洋细菌对厚壳贻贝稚贝附着的影响

为探讨厚壳贻贝稚贝对自然微生物膜中海洋细菌的附着行为反应,本论文研究了厚壳贻贝（Mytilus coruscus）稚贝附着与硅烷基化附着基表面、微生物膜密度以及细菌种属系统发育之间的相互关系。研究表明所测海洋细菌均能显著促进厚壳贻贝稚贝的附着;其中Staphylococcus sp. 1和Cobetia sp. 1表现出较低诱导活性,且这两株细菌形成的微生物膜密度与稚贝附着率之间无显著相关性;其他7株海洋细菌均表现出中等程度诱导活性,且所形成的微生物膜密度与稚贝附着率之间呈显著相关性。系统发育分析表明所测海洋细菌对厚壳贻贝稚贝附着的诱导活性与细菌种属无关。因而,硅烷基化表面的海洋附着细菌对厚壳贻贝附着有着显著性促进作用,本研究将为后续开展厚壳贻贝稚贝附着机制奠定了基础。     

中湿度表面的海洋细菌对厚壳贻贝稚贝附着的影响

为研究自然微生物膜中海洋细菌对厚壳贻贝稚贝附着的影响,通过海洋贝类生物学、分子微生物生态学等手段调查附着基表面湿度、微生物膜的密度以及细菌种属系统发育与厚壳贻贝(Mytilus coruscus)稚贝附着关系。结果表明,所有测试海洋细菌形成微生物膜的最终密度随初始密度的增加而增加。所测海洋细菌均能不同程度地诱导厚壳贻贝稚贝的附着,其中Cobetia sp.3形成的微生物膜显示出最高诱导活性,其诱导的稚贝附着率为(70%±3%);Nautella sp.2、Pseudoalteromonas sp.9、Pseudoalteromonas sp.10、Bacillus sp.5和Pseudoalteromonas sp.11等5株细菌表现中等程度的诱导活性,其诱导的附着率范围为51%~60%。所有测试菌株所形成的微生物膜密度与稚贝附着均呈显著相关,尤其是Pseudoalteromonas sp.9和Cobetia sp.3诱导活性与附着率相关性极强,其相关性系数分别为0.774 1和0.723 3。系统发育分析结果表明,微生物膜密度和厚壳贻贝稚贝的附着率显著相关,然而海洋细菌诱导活性与细菌种属无关。因而,中湿度表面的海洋附着细菌对厚壳贻贝的附着有着明显的促进作用,本研究为后续开展厚壳贻贝稚贝的附着机制提供了理论依据。     

不同来源海洋弧菌微生物被膜对厚壳贻贝稚贝附着的影响

为探讨广泛存在于近海环境中海洋弧菌和贝类附着的相互作用关系,实验从自然微生物被膜和厚壳贻贝成体肠道内分离了海洋弧菌,测定其种属及亲缘性,调查了这些不同来源弧菌形成的微生物被膜与厚壳贻贝稚贝附着的调控作用。结果显示,测试弧菌形成的微生物被膜密度随着初始细菌密度的增加呈现不同程度的增加。源于自然微生物被膜和贻贝肠道内的10株弧菌均能显著促进厚壳贻贝稚贝的附着,不同菌株形成微生物被膜的诱导活性存在显著差异,附着率变化范围为17%~67%,其中稚贝在Vibrio sp.17微生物被膜上的附着率为67%±2%。微生物被膜对稚贝附着诱导活性与被膜密度呈显著相关性,其中弧菌V.crassostreae ECSMB14106的相关性系数最高,为0.8992。此外,微生物被膜的诱导活性与弧菌的来源无关。本实验初步探明了海洋弧菌对厚壳贻贝稚贝附着的影响,有助于进一步理解微生物被膜调控厚壳贻贝的附着分子机理。     

厚壳贻贝早期幼虫低温保存的影响研究

探讨低温(4 ℃)条件下的厚壳贻贝早期幼虫保存可能性,同时调查了不同培育密度对低温保存的影响。在正常条件下,继续培育低温保存后的幼虫,并调查其存活率和生长的变化。 结果表明,在低温保存后,早期幼虫存活率较高,超过95%;厚壳贻贝早期幼虫的壳长和壳高出现显著性的增长。不同培育组间,培育密度对厚壳贻贝早期幼虫的存活和生长影响不同,表明密度是幼虫低温保存的一个重要因素。在正常培育条件下,低温保存后的幼虫,3周后其存活率明显低于对照组,但仍超过50%,且其生长速度明显高于对照组。因此,低温培育是保存厚壳贻贝早期幼虫的有效方法,可用于今后贝类幼虫生物学实验和人工育苗技术的改善研究。     

翡翠贻贝人工养殖研究

翡翠贻贝(Pera virid is Linnaeus)是瓣鳃纲动物，隶属贻贝目(Mytiloida)贻贝科(Mytilidae)。贝壳较大、较长，壳质比厚壳贻贝(Mytilus coruscus Goyil)薄，比紫贻贝(M.galloprovincialis Lamarck)厚。壳表为翠绿色，尤以边缘部最鲜艳，壳里为瓷白色。本种为热带和亚热带种，垂直分布于低潮线至水深17米左右，但以水深5—6米处生长最密。     

培养基对微生物被膜形成和厚壳贻贝附着的影响

为探讨微生物被膜生物学特性、培养基以及大型生物附着三者之间的相互关系,本实验通过微生物分子生态学和海洋贝类生态学等方法调查了不同培养基对海洋细菌所形成微生物被膜的影响及其对厚壳贻贝幼虫附着的影响。结果显示,厚壳贻贝稚贝率与培养基类型和细菌初始密度显著相关,进一步分析表明,Zo Bell 2216E和Seawater Luriabertani(SWLB)条件下分别有6株和9株细菌形成的微生物被膜密度与稚贝率显著相关。扫描电镜结果显示,Staphylococcus sp.3在Zo Bell 2216E培养基条件下形成微生物被膜的细菌分布较为紧密,Pseudoalteromonas sp.8则在SWLB培养基条件下微生物被膜细菌分布较为紧密,且形态变为短杆状。SDS-PAGE结果显示,相比Zo Bell 2216E培养基,Staphylococcus sp.3在SWLB培养基条件下9条条带蛋白显著下降,其中2个条带完全消失;Pseudoalteromonas sp.8则在SWLB培养基条件下明显增加5条蛋白条带。研究表明,微生物被膜的形成受到培养基的影响,培养基的不同导致微生物被膜的形态结构、分布和蛋白有所差异,最终导致微生物被膜诱导厚壳贻贝幼虫附着变态的活性差异,本研究为后续开展厚壳贻贝附着的分子机制奠定良好基础。     

弧菌生物被膜的动态演替对厚壳贻贝附着的影响

为探讨生物被膜动态演替过程中如何影响海洋无脊椎动物附着,实验选取了对厚壳贻贝附着具有不同诱导活性的弧菌Vibrio cyclitrophicus、V. chagasii和Vibrio sp. 22形成单一生物被膜,观察弧菌动态演替中生物被膜细菌密度、膜厚度和胞外产物等生物学特性变化,探究其对厚壳贻贝稚贝附着的影响。结果显示,弧菌生物被膜动态演替过程中,被膜细菌随着时间推移出现聚集现象,细菌密度和膜厚度也随着时间变化呈先增多后减少。除了Vibrio sp. 22,V. cyclitrophicus和V. chagasii生物被膜细菌密度和膜厚度与稚贝附着均有不同程度的相关性。所测弧菌生物被膜胞外产物的显微激光共聚焦结果分析发现,胞外多糖随着时间先增多,然后开始下降。相对比而言,胞外蛋白和胞外脂质无显著性变化。因而,胞外多糖变化规律与稚贝在被膜上附着变化相一致,表明胞外多糖是生物被膜动态演替过程中调控厚壳贻贝附着的重要因素。本实验初步探讨了生物被膜动态演替特征及其对厚壳贻贝稚贝附着的影响,对于后续进一步在海区开展生物被膜的动态演替与海洋底栖动物附着相互关系研究具有重要的学术价值,同时对于人工鱼礁礁体生物附着机理的研究具有重要的实践价值。     

Performance of Mussels,Mytilus edulis,Mytilus trossulus,and Their Hybrids in Cultivation at Three Scottish Lochs

Mussel aquaculture is a growing industry in Scotland, dominated by the production of Mytilus edulis. However, Mytilus trossulus, Mytilus galloprovincialis, and their hybrids with M. edulis have recently been detected in Scotland. At some farms, M. trossulus have been associated with significant losses in production because of presenting low meat yields and fragile shells. In the present study, M. edulis, M. trossulus, and their hybrids were examined from three cultivation areas in order to investigate shell and meat characteristics and advise on future management of mixed‐species areas in Scotland. Results suggest that M. edulis is likely to consistently outperform M. trossulus and hybrids in mixed‐species areas, but that M. trossulus meat yields from different areas can vary considerably. The ability to classify mussels into two (M. edulis and non‐edulis) groups suggests that separation of M. trossulus genotypes could potentially be achieved by grading the harvested mussels. In the most affected areas, however, the fallowing of cultivation sites and importation of unispecific M. edulis spat appear as an alternative. Management of this problem in Scotland is likely to involve the implementation of area‐specific measures, and the establishment of clear guidance on good practice aiming at preventing further distribution of M. trossulus.     

Comparison of the physiological energetics between Mytilus chilensis,Mytilus galloprovincialis and their hybrids,under laboratory conditions

Laboratory‐produced juvenile individuals of the species Mytilus chilensis, M. galloprovincialis and their hybrids were subjected to physiological measurements under an experimental diet of Isochrysis galbana (30 × 10⁶ cells L^?1), 13°C temperature and a salinity of 30 psu. Pure species individuals showed a higher clearance rate (CR). Mytilus chilensis had a CR of 1.13 L h^?1, while M. galloprovincialis registered only 0.78 L h^?1. Also, pure taxa registered higher values (above 70%) of absorption efficiency when compared with hybrid individuals. Ammonia excretion in M. chilensis and M. galloprovincialis was 1.5% and 0.4%, respectively, while hybrids registered significantly lower values. Under these experimental conditions, M. chilensis registered the highest scope for growth (P < 0.05), compared with M. galloprovincialis and their hybrids. However, the net growth efficiency index (K ₂) in hybridization type I (♀Mg × ♂Mc) was higher (P < 0.05) than other experimental groups. The invasive mytilid M. galloprovincialis showed values that are very similar to those obtained with the hybridization I group (♀Mg × ♂Mc). Finally, we discuss that water temperature is an important factor in the biogeographic separation of both species and the potential effects that the settlement of the invasive species may have for Chilean mussel production.     

Effect of temperature on survival,growth and development of Mytilus galloprovincialis larvae

Mussel aquaculture is widely prevalent worldwide, but generally relies on natural seed collection, which does not always meet the needs of the producers. Thus, development of mussel hatcheries is of economic interest in some parts of the world, such as Europe; it provides opportunities not only on annual reliability of seed but also on genetic improvements. To broaden knowledge on mussel larval physiology, we carried out temperature treatments (17, 20 and 24 °C) on Mytilus galloprovincialis larvae under laboratory conditions. The trials ended when 30% of the larval population was in the post‐larval stage. The temperature coefficient Q₁₀ indicated a strong relationship between temperature and increase in growth from 17 to 20 °C, but not between 20 and 24 °C. Exposure of M. galloprovincialis larvae to 17 °C resulted in poor growth, low survival and a delayed development and was considered to be inadequate for M. galloprovincialis larval culture. Rearing the larvae at 20 or 24 °C produced better growth, higher survival rates and faster metamorphosis as compared with 17 °C. The temperature region within 20 and 24 °C was suggested as adequate for the mussel M. galloprovincialis larval culture, and implications of these results on the development of commercial hatcheries were discussed.     

翡翠贻贝的耐冷性

采用持续性降温(在海水中从)和突发性降温(在-1℃下的冷海水中冷处理1h)对翡翠贻贝进行冷处理,然后在室温下复苏,观察其复苏情况:采用持续性降温(在海水中,从21℃持续性降沮10h 9℃,从21℃持续性降沮18h 3℃)和突发性降沮(在-1.5℃、0℃、5℃、9℃的冷海水中冷处理10min)后进行冰藏,然后,每隔24h随机取出部分样品检定其存活情况,并观察其复苏情况,对翡翠贻贝的耐冷性进行了研究,结果证明翡翠贻贝具有较强的耐冷性,在≤0℃时,仍能存活,通过冰藏可达到保活保鲜之目的。     

紫贻贝磷脂的贮藏性能

对不同贮藏温度下紫贻贝磷脂含量及其组分的变化情况进行了研究。在30℃、4℃、-10℃、-20℃分别贮藏贻贝,随着温度的上升,磷脂含量的下降速度加快。对贮藏3d后的贻贝磷脂进行组分分析,磷脂各组分的相对百分比都发生了变化,并且随温度的升高PC的降解速度大大加快,所占的百分比快速下降,而LPC的比例有不同程度的增加。     

贻贝雌性生殖系统的组织学研究

从组织学方面研究了贻贝雌性生殖系统的构造。卵巢壁薄、由外膜和内生殖上皮构成。外膜主要由结缔组绢,意志支上皮和薄层肌肉构成。由其产生的许多延伸物伸入卵巢中把卵巢分成一个个不规则小区。内生死上皮不断增殖平生生殖带,其中含卵原细胞阁阶段雌性生殖细胞和滤泡细胞。卵原细胞较小,核圆形,核仁不明显。初级卵母细胞体积增加,核呈生发泡状,核仁１个,明显。次级卵母细胞卵圆形、细胞体积更大,核仁１个,十分明显。当卵母     

Toxicity of bacteria towards haemocytes of Mytilus edulis

Haemocytes of Mytilus edulis, allowed to attach to the plastic surface of a tissue culture plate in the presence of haemolymph, were observed by time-lapse video recording. When bacteria were added at concentrations of 10 or 50 bacteria per haemocyte, certain bacterial strains caused rounding of the cells within 2–3 h. Haemolymph was necessary for the rounding to occur; if bacteria were added in sterile seawater there was no significant difference in the number of rounded cells between control and bacteria-treated cultures for up to 4 h. The haemolymph factor required for this activity was active at 1/64 dilution in seawater, was sensitive to trypsin treatment, and activity was halved on heating at 56 or 100 °C for 30 min. For the most toxic bacteria tested, Vibrio alginolyticus NCMB 1339 and Vibrio anguillarum A7, haemocyte cell rounding appeared to be induced by a very small number of bacterial cells. Bacteria-free culture supernatant of V. anguillarum 2981 induced rounding of haemocytes in a dose-dependent manner, with 50 % of cells being rounded at a dilution of approximately 1/500 of the culture supernatant. In a survey of 226 bacterial isolates, those isolated from incidents of disease in a bivalve hatchery were significantly more toxic towards haemocytes than bacteria isolated from hatcheries without disease or from turbot hatcheries.     

Effects of larval cryopreservation on subsequent development of the blue mussels,Mytilus galloprovincialis Lamarck

In this study, the effects of three commonly used chemicals, dimethyl sulphoxide (DMSO), ethylene glycol (EG), propylene glycol (PG) and their combinations with trehalose, were evaluated on the cryopreservation of D‐larvae of the blue mussel Mytilus galloprovincialis. The larvae were harvested 30 h post‐fertilization at 21 °C and cryopreserved using a standard protocol in 5%, 10% or 15% of DSMO, EG and PG either as single chemical solutions or in combination with 0.2 M trehalose. Among these cryoprotectants, 5% DMSO resulted in the highest post‐thaw survival rate of 55.3±7.8%, although it did not significantly differ from those with 10% and 15% EG. The addition of 0.2 M trehalose did not improve the post‐thaw larval survival rates in all the combinations assessed. The cryo‐effects on subsequent development were evaluated using the D‐larvae frozen with 5% DMSO. The results showed that cryopreservation affected both larval survival and growth in this species. The relative daily mortality rate was significantly higher in treated than control groups over the period from 3 h post‐thaw to day 11 post‐fertilization. On day 6 post‐fertilization, the average larval length in the treated group was significantly smaller than that in the control. From day 11 post‐fertilization, and onwards, differences in these two traits were not significant between treated and control groups. On day 21 post‐fertilization, about 80% of the larvae in both treated and control groups developed eyes and the normalized survival rate in the treated group was 12.5%.