【作者】 黄瑛；

【导师】 唐学玺； 朱丽岩；

【作者基本信息】 中国海洋大学 ， 生态学， 2008， 博士

【摘要】 三丁基锡（Tributyltin TBT）是毒性最大、污染最严重的有机锡污染物之一,对多种海洋生物具有长期有效的杀生效果,同时也影响许多非靶生物,对经济水产和海洋生态环境构成了严重的威胁。桡足类作为海洋生态系统中承上启下的关键种类,研究其个体和种群对环境污染物的响应情况,对污染物的生态效应评价具有重要意义。本论文以火腿许水蚤为实验动物,建立了用于实验室内生态毒理学研究的多世代培养体系,在进行桡足类生物学研究的基础上,以毒性最大的三丁基锡种类-三丁基氧化锡（TBTO）为污染物,研究了火腿许水蚤个体和种群的响应情况。主要的研究结论如下:1.为了尽可能地节约成本和空间,获得高数量、营养和遗传具有一致性且发育同步的桡足类用于生态毒理学实验,需要筛选高效的饵料种类和饵料浓度,并且对火腿许水蚤的生殖节律进行控制。在本实验选取的4种单胞藻中,金藻是最高效的种类。在桡足类的多世代培养过程中,为了提高饵料多样性、防止饵料过度增殖,建议采用金藻和三角褐指藻混合投喂（投喂比例倾向于金藻）,建议饵料浓度为5×104 cells ml-1。另外,可以通过短期禁食（48 h）后恢复投饵和定时收集（第5天、第8天和第10天）的方法控制火腿许水蚤的生殖节律。2.本实验通过组织学研究方法和个体生活史实验研究了火腿许水蚤的繁殖生物学特征。火腿许水蚤雌体采取怀卵囊、周期性的生殖方式,生殖系统主要包括卵巢和输卵管两个部分,生殖周期可分为4个阶段,其间卵巢的形态基本没有改变,而输卵管的形态发生变化。环境中的盐度和温度对桡足类发育和生殖参数都有显著影响:随着盐度的升高,存活率升高,雌体的发育减缓,生殖量减少;而随着温度的升高,存活率降低,雌体发育加快。因此,在火腿许水蚤应用于生态毒理学暴露实验时,应确定环境条件,以提高实验的可重复性和数据的可比性。3. TBTO的急性暴露实验结果表明,火腿许水蚤对有毒污染物非常敏感,雌体96h-LC50值仅为0.42μg l^-1,而雌雄个体的敏感性并无显著差异。另外,环境因素对火腿许水蚤的敏感性有较大的影响,其中温度的影响最为明显,随着温度的升高,桡足类的敏感性显著提高;而盐度的影响相对较小,影响情况比较复杂;投喂饵料的实验组桡足类死亡率比不投饵的实验组显著较低,可见TBTO对火腿许水蚤主要的毒性作用途径更多的是水体中TBTO的影响。另外,桡足类密度对TBTO的毒性效应也有一定影响。因此,在设计相关的暴露实验时,应该综合考虑实验的可行性和实际的环境因素,选择合适的温度、盐度、饵料浓度和实验动物密度等条件。4.桡足类的世代暴露实验是评价环境污染物生态效应的必要方法。随着TBTO暴露浓度的升高和暴露时间的延长,火腿许水蚤的存活、发育和生殖都受到了显著影响。如火腿许水蚤幼体在5 ngTBTO l-1暴露环境中发育到成体的存活率即显著下降,其中无节幼虫期的敏感性比桡足幼体期更高。TBTO对火腿许水蚤性别比的影响主要体现在F1和F2世代,而且仅表现在20 ngTBTO l-1实验组中,这可能和TBTO的遗传毒性有关。在本实验的暴露浓度下,雌体的生殖量和世代时间并没有受到显著影响,但是在较高的暴露浓度组（40 ngTBTO l-1和60 ngTBTO l-1）出现了较大数量不能产生卵囊或流产的雌性个体。切片染色观察结果表明,暴露雌体不能产生卵囊可能是由于输卵管的末端堵塞导致卵母细胞不能排出引起的。TBTO对火腿许水蚤性别比率和卵囊产生的作用机制还需要进一步研究。5.本研究在传统Euler-Lotka方程的基础上,针对桡足类等有性生殖的种类进行改进,结合火腿许水蚤的世代暴露实验结果,预测了TBTO对火腿许水蚤种群的影响。结果表明,在20 ngTBTO l-1以上暴露浓度下,火腿许水蚤的种群内禀增长率显著降低。虽然与生活史参数相比,种群内禀增长率并不是最敏感的评价指标,但是具有更大的生态学意义。另外,本研究还通过群体培养实验对理论预测的结果进行了验证,得到了基本一致的结果。这说明,通过对桡足类个体水平存活和生殖参数的影响研究,结合理论模型分析,能够较好地预测污染物对桡足类种群的影响。通过以上实验结果,我们认为火腿许水蚤对TBTO等环境污染物有较高的敏感性,适合作为低浓度持久性污染物生态效应评价的模式生物;通过多世代暴露实验和编制生命表的研究方法能够较好地预测环境污染物对桡足类种群的影响。更多还原

【Abstract】 Tributyltin(TBT) is one of the most toxic organotin type and has been used extensively. Tributyltin is so effective as a biocide and its toxicity is non-specific. It has a high bioaccumulation factor and a persistence that poses a great problem to the marine environment. Crustacean zooplankton such as copepods form the keystone link between primary producers and fish stocks, making it critical to understand their responses to toxic pollutants in marine. In this paper, we established a multi-generation cultivation system for colanoid copepod Schmackeria poplesia, and investigated the responses of S. poplesia to bis(tributyltin) oxide (TBTO). The main conclusions as follows:1. To produce copepods in large quantity and nutritional, developmental and genetic uniformity, experiments were carried out to manipulate and control egg production in females. Results showed that Isochrysis galbana was a more efficacious diet, and a 2:1 mixed diet of I. galbana and Phaeodactylum tricornutum in a density of 5×104 cells ml-1 was recommended in multi-generation culturing system and life-cycle toxicity tests. In addition, by starvation and reintroduction of food as well as timed collection, the production of copepods can be well controlled.2. The female reproductive system of S. poplesia consists of a single ovary with two diverticula, and we established a classification system of four ganad maturation stages. During a spawning cycle, distinct modifications of the gonad morphology were observed, while the ovary did not change, neither in size nor in cytological composition. Environmental factors such as temperature and salinity can significantly affect the development and reproduction of S. poplesia. At higher salinities, the survival increased, and the development was delayed. While at higher temperatures, the survival decreased and the development was advanced. So we should make invariable conditions in the exposed tests for reproducible work.3. The acute exposure test revealed that S. poplesia was very sensitive for TBTO exposure. The 96h-LC50 value for females was 0.42μgTBTO l-1, and no sex-specific differences were observed. The effects of temperature, salinity, food concentration and copepod density were detected for TBTO exposures. The results shows that increase in temperature increased the toxicity of TBTO, but the effect of salinity was more complex. Low salinity and high salinity increased mortality of S. poplesia. In addition, food concentration and copepod density have significant effects on the mortality of copepods. Therefore, the eco-toxicity tests should be conducted at a range of environmentally realistic regimes, as this will enhance the sensitivity of the test and improve the safety margin in line with the precautionary principle.4. Life-cycle exposure tests are essential method for risk assessment. The survival, development and reproduction of S. poplesia are affected when exposed to TBTO for three generations. The results indicate that nauplii are more sensitive than copepodites, and the survival of copepods exposed to 5 ngTBTO l-1 was decrease. The female-to-male ratio in F1 and F2 generations exposed to 20 ngTBTO l-1 concentration were significantly reduced. The first egg sac for females and the fecundity was not affected, while in higher TBTO concentration treatments, there were some females couldn’t produce egg sacs or aborted. Histological examinations suggest that the exposure to TBTO block the posterior end of the diverticula and inhibit the production of egg sacs. A full explanation of the effect of TBTO on copepods requires further investigation.5. A modified Euler-Lotka equation was used to calculate a population-level endpoint, the intrinsic rate of natural increase, from individual life-table endpoints, i.e. mortality rate, time of release of first brood, sex ratio, the fraction of ovigerous females among all females as well as the number of nauplii per ovigerous female. The intrinsic rate of natural increase of S. poplesia was significantly decrease when expose to TBTO concentrations higher than 20 ngTBTO l-1. Compared with individual life-table endpoints, the population effects the relatively weak. While a more sensitive end-point does not necessary have to be more ecologically relevant than a less sensitive endpoint. The results of colony exposure tests reveal a similar phenomenon that the population exposed to higher than 20 ngTBTO l-1 concentrations decline. The results from the present study demonstrate that ecologically relevant toxicity data on population dynamics can be gained from a relatively short and cost-effective full life-cycle toxicity test, which can improve the assessment of ecosystem risk of chemicals.These results show that the colanoid copepod Schmackeria poplesia is sensible for toxic pollutants such as TBTO, and is a promising species for use in ecotoxicology test. The full life-cycle toxicity test combined with the study of population growth rate structure can indicate the toxicity of pollutants more effectively.更多还原