栉孔扇贝Chlamys farreri生态免疫机制的基础研究

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	栉孔扇贝Chlamys farreri生态免疫机制的基础研究
其他题名	Mechanism of Ecological Immunity in Chlamys farreri
	陈慕雁
学位类型	博士
	2007-06-06
学位授予单位	中国科学院海洋研究所
学位授予地点	海洋研究所
关键词	栉孔扇贝chlamys Farreri 生态免疫环境胁迫应激激素血细胞免疫功能
摘要	针对目前栉孔扇贝Chlamys farreri养殖亟待解决的种质、病害、环境和产品质量等日趋严重的问题，从栉孔扇贝本身的防御机制和神经内分泌机制入手，较为系统地研究了环境胁迫，应激激素与血细胞免疫功能之间的相互作用机制；从生态免疫学角度，探讨了栉孔扇贝大规模死亡的原因，为栉孔扇贝病害防治和种质优化提供了一些理论依据。主要研究结果如下： 1.较为系统地综述了贝类生态免疫机制的研究进展。分析了应激激素对贝类细胞免疫活性的抑制作用，以及生态免疫过程中免疫成本的投入与其他生态因子之间的内在联系，分析了病原体与宿主之间的相互作用机制，提出了贝类生态免疫机制研究新见解和新思路。 2.筛选出一种较适用的抗凝剂配方：Glucose 20.8 gL-1，EDTA 20mM, Sodium chloride 20 gL-1，Tris-HCl 0.05M，pH=7.4。 3.模拟研究了栉孔扇贝养殖过程中的主要环境胁迫因子，包括露空胁迫（5°C，17°C和25°C露空最长持续24小时），急性温度胁迫（从17°C分别直接升至23°C和28°C或降至11°C），急性盐度胁迫（从盐度31直接升至盐度35或降至盐度25和20），饥饿胁迫（持续40天）和密度胁迫（分为低、中和高密度），对栉孔扇贝血细胞免疫功能的影响，养殖过程中的露空胁迫对栉孔扇贝的血细胞免疫功能具有抑制作用，从而削弱了扇贝胁迫后恢复的最初24小时中抗击病原体的能力。高温下（25°C）的露空胁迫能够显著地降低扇贝的成活率。急性升温胁迫（从17°C突变至28°C）会严重的破坏栉孔扇贝的内稳态，损伤其血细胞免疫功能，从而增加了扇贝对病原体的易感性。而扇贝对快速的降温胁迫（从17°C突变至11°C）则具有较高的耐受性。盐度20的低盐胁迫能够显著抑制栉孔扇贝的血细胞防御功能，同时低盐有利于许多病原体的繁殖，两方面的协同作用，将大大增加扇贝大规模死亡的几率。饥饿胁迫（40天）能够显著地抑制血细胞的免疫活性，然而在实验室饵料充足的条件下，养殖密度除了对血细胞的吞噬活性有一定的抑制作用外，对血细胞其他的免疫活性影响不明显。 4.揭示了环境胁迫因子，包括露空胁迫（17°C 露空24小时），温度胁迫（从17°C分别直接升至28°C或降至11°C持续7天），低盐胁迫(从盐度31直接降至盐度20持续7天)和饥饿40天胁迫，对栉孔扇贝血细胞超微结构的影响，露空胁迫（17°C 露空24小时），低盐胁迫(盐度20持续7天)和饥饿40天胁迫严重损伤了血细胞的膜系统及各种细胞器的结构。 5.利用酶联免疫法测定了栉孔扇贝血淋巴中应激激素（肾上腺素，去甲肾上腺素和多巴胺）的基础浓度，分别为0.088±0.11, 18.63±1.96 和 2.59±0.46ng/ml。研究了血淋巴中应激激素对环境胁迫（包括露空，急性升温和急性降盐）的响应水平，急性露空，升温和降盐能够显著提高血淋巴中肾上腺素和去甲肾上腺素的浓度，而多巴胺浓度变化却呈现出完全相反的趋势。 6.肾上腺素和去甲肾上腺素体外诱导栉孔扇贝血细胞研究结果表明：浓度为30ng/ml或50ng/ml的去甲肾上腺素能够显著抑制血细胞的吞噬活性，浓度为50ng/ml的去甲肾上腺素能够显著抑制血细胞的活性氧产物，而肾上腺素对血细胞免疫功能的影响则不显著。
其他摘要	Deteriorations of germplasm, disease, environment and product quality are becoming more serious problems, not yet to be resolved in Chlamys farreri culture. In view of this, studies on defence and neuroendocrine mechanism of C. farreri were carried out. The interaction among environmental stress, stress hormone and haemocyte immune functions was investigated systematically. To explore the cause of massive mortality of C. farreri from the ecological immunology point of view, it will be benefit for disease prevention and germplasm optimization in the long term. The main results are as follows: 1.The recent progress derived largely from the mechanism analysis of molluscan ecological immunity was reviewed in this paper. The immune suppression effect of stress hormones on the molluscan immunity has been analyzed, and the relationship between the cost investment in defence system and other ecological factors was expounded. The interaction mechanism between pathogen and host was clarified. Furthermore, the future studies in molluscan ecological immunology for inspiring the new thoughts in controlling mollusks disease were suggested. 2.According to the special characteristic of C. farreri haemocyte, a desirable anticoagulant solution was selected (Glucose 20.8 gL-1，EDTA 20mM，Sodium chloride 20 gL-1，Tris-HCl 0.05M，pH=7.4). 3.Investigating the effects of major environmental stressors, including air exposure (5°C，17°C and 25°C for 24h period), acute temperature variation (increased directly from 17°C to 23°C and 28°C, or decreased directly from 17°C to 11°C), acute salinity variation (increased directly from 31 to 35, or decreased directly from 31 to 25 and 20), starvation (sustained for 40 days) and stocking density (low, medium and high), on haemocyte immune conditions of C. farreri. The results showed that air exposure has a negative effect on haemocyte immune functions of scallops, this may lead to scallop vulnerability to fight against any pathogen during the first 24h after re-immersion. Air exposure at 25°C was demonstrated to markedly reduced survival rate in scallops. The homeostatic capabilities appear to be compromised for C. farreri transferred directly from 17°C to 28°C, it is supposed that the overall health of the scallop would be compromised and the likelihood of being affected by pathogens might increased. By contrast, scallops showed possessing high tolerance to a rapid decrease of seawater temperature (from 17°C to 11°C). Acute salinity decrease (from 31 to 20) could significantly inhibit the haemocyte immune conditions of scallop and accelerate the propagation of pathogen, which will greatly increase the probability of mass mortality of scallops. Starvation (sustained for 40 days) could also significantly inhibit the haemocyte immune functions. Considering the superfluous feeding factor in experimental condition, no significant effect of stocking density was observed on haemocyte immune functions except the percentage of phagocytic haemocytes. 4.The ultrastructural alterations taking place in C. farreri haemocytes resulting from air exposure (exposed to air for 24hours at 17°C), temperature variation (transferred directly from 17°C to 11°C and 28°C and sustained for 7 days), salinity variation (transferred directly from salinity 31 to 20 and sustained for 7 days) and starvation for 40 days, were examined by transmission electron microscopy (TEM). The results showed that serious damages appeared in the haemocytes ultrastructure after air exposure (17°C for 24hours), hypotonic (salinity 20 for 7days) and starvation stress (40 days). 5.Enzyme immunoassay revealed that stable presence of catecholamines (including adrenaline, noradrenaline and dopamine) in the haemolymph of the scallop, the basic values were 0.088±0.11, 18.63±1.96 and 2.59±0.46ng/ml respectively. The method was also used to investigate these catecholaminergic responses to environmental stress. The results showed that both adrenaline and noradrenaline concentrations increased significantly in response to all the stress, including air exposure, acute temperature increase (from 17°C to 28°C) and acute salinity decrease (from 31 to 20). However, the dopamine concentration showed reverse variation trend in response to stress compared with adrenaline and noradrenaline. 6.Based on the basic concentration and fluctuation amplitude of catecholamines in response to stress in circulating haemolymph, different concentrations of adrenaline and noradrenaline were designed for investigating the influence of these two catecholamines on haemocyte immune activities. The results showed that noradrenaline had a dose-dependent inhibitory effect on phagocytosis at physiological concentrations of 30ng/ml and 50ng/ml, and on reactive oxygen species at concentration of 50ng/ml. But no significant effect of adrenaline on haemocyte immune activities was observed.
页数	115
语种	中文
文献类型	学位论文
条目标识符	http://ir.qdio.ac.cn/handle/337002/1297
专题	海洋环流与波动重点实验室
推荐引用方式 GB/T 7714	陈慕雁. 栉孔扇贝Chlamys farreri生态免疫机制的基础研究[D]. 海洋研究所. 中国科学院海洋研究所,2007.

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