利用环境DNA探究中华鲟繁殖群体的分布与生物量动态

Along with the worsening plight of the natural population, natural breeding of Chinese sturgeon has been seriously affected. From 2013 to 2015 for three consecutive years in the middle and lower reaches of the Yangtze River, monitoring on natural breeding behaviors of Chinese sturgeon ended unsuccessfully. Distribution and dynamics of breeding groups are key issues in conservation of Chinese sturgeon, while low population density has made it difficult to collect enough samples and data. In recent years, environmental DNA test offered an uninvasive way to monitor wild species with high sensitivity. In this study, a highly sensitive environmental DNA quantitative analysis system of Chinese sturgeon will be developed using species specific fluorescent quantitative molecular markers, to study dynamic distribution and biomass of the breeding population below the Gezhouba dam on the basis of concentration variation of environmental DNA. Combined with findings of traditional investigation, we will further research the occurrence regularity of the natural breeding behaviors of Chinese sturgeon, and evaluate the effectiveness of environmental DNA detection system. This study will offer injury-free technological means in natural population monitoring of Chinese sturgeon, and provide scientific support for conservation of this species. In addition, the environmental DNA quantitative test system can also be used in monitoring and conservation of other aquatic organism.

随着中华鲟天然种群濒危处境日益加剧，其自然繁殖行为受到了严重影响，2013年至2015年连续三年未在长江中下游监测到中华鲟自然繁殖行为，当前繁殖群体的分布与动态成为中华鲟物种保护重点关注的问题，但种群数量稀少使样本和数据搜集面临难题，而近年来兴起的环境DNA检测技术则提供了一种非侵入式的高敏感度野外监测方法。本研究拟开发中华鲟物种特异性荧光实时定量分子标记，建立高敏感度的中华鲟环境DNA定量分析体系，在葛洲坝坝下江段进行中华鲟环境DNA实时监测，通过中华鲟环境DNA浓度的时空变化了解其繁殖群体的分布和生物量变动趋势，结合传统方法调查结果，探讨当前中华鲟自然繁殖行为的发生规律，评估环境DNA检测体系的有效性。本研究将为中华鲟天然群体监测提供无伤害的技术手段，为中华鲟物种保护提供科技支撑，并对其它水生生物的监测与保护具有借鉴意义。

近年来中华鲟自然繁殖行为出现间歇中断现象，表明其天然群体面临严重威胁，基于中华鲟繁殖群体监测需求，无损伤高效率的水样环境DNA检测技术为此提供了新的技术手段。本项目对长江水样环境DNA采样方法进行优化，基于中华鲟线粒体基因序列分析开发了中华鲟环境DNA荧光实时定量分子标记，筛选了Taqman探针和引物，建立了qPCR检测标准曲线，在中华鲟和近源鲟鱼物种组织DNA样本中进行体系有效性检测，在长江不同江段水样中进行对比检测。FAM探针和VIC探针在中华鲟组织DNA（包括子代和卵）中的检测阳性率分别为27%和73%，FAM特异性较高但扩增成功率较低，而VIC探针特异性较低但检测成功率较高，该位点的扩增产物DNA序列可明确区分中华鲟及其相近物种。连续三年在宜昌葛洲坝下中华鲟产卵场进行自然繁殖监测，设置不同的采样策略采集水样环境DNA样本，利用双探针荧光实时定量PCR进行样本检测，结果显示检测阳性率和DNA拷贝数在繁殖期前后具有一定的季节波动。本研究探索了长江水样环境DNA物种特异性定量检测方法，为中华鲟天然群体无损伤监测提供技术支撑，对其它濒危水生物种的监测具有借鉴意义。