摘要:底栖动物是淡水生态系统中物种多样性最高的类群，也是应用最广泛的水质监测指示生物之一。传统的底栖动物监测以形态学为基础，耗时费力，无法满足流域尺度大规模监测的需求。环境DNA-宏条形码技术是一种新兴的生物监测方法，其与传统方法相比优势在于采样方法简单、低成本、高灵敏度，不受生物样本和环境状况的影响，不依赖分类专家和鉴定资料，能够快速准确地对多个类群进行大规模、高通量的物种鉴定。然而，在实际应用中该方法的效果受诸多因素的影响，不同的方法、流程往往会产生差异较大的结果。鉴于此，着重分析总结了应用环境DNA-宏条形码技术监测底栖动物的关键影响因素，包括样品采集与处理流程、分子标记选择、引物设计、PCR偏好性、参考数据库的完整性及相应的优化。并基于此探讨了提高环境DNA-宏条形码技术在底栖动物监测效率和准确率的途径，以期为底栖动物环境DNA-宏条形码监测方案的制定提供可靠的参考。最后对该技术在底栖动物监测和水质评价中的最新发展方向进行了展望。



Abstract:Macrozoobenthos are the most diversified organisms in freshwater ecosystem, playing a vital role in maintaining the structural and functional integrity of freshwater ecosystem. They are one of the most significant bioindicators that have been widely used in water quality monitoring and assessment. Nowadays, anthropogenic disturbance, pollution and global warming lead to a remarkable decline of the macrozoobenthos biodiversity. Therefore, how to provide substantial data for macrozoobenthos biodiversity monitoring rapidly, accurately, and reliably has become a major issue in water environment management. However, the traditional morphology-based biomonitoring survey method can hardly fulfill current biomonitoring requirements due to being costly, time consuming, and labor intensive. Moreover, such method is highly dependent on experienced taxonomists and often results in low precision and unverifiable taxonomic data. Environmental DNA metabarcoding (eDNA metabarcoding), a newly emerged approach, is considered as a promising survey tool for aquatic biodiversity monitoring. Compared with traditional biomonitoring survey methods, eDNA metabarcoding remarkably outperforms morphology-based method by its low cost, straightforward workflow, and high sensitivity in species detection of macrozoobenthos. Particularly, it circumvents the taxonomy literature and expertise, providing rapid, accurate, and high-throughput identification simultaneously to a large variety of taxa regardless of the organism conditions and environmental disturbances, thus permitting the easy access of large scale biomonitoring programs for freshwater ecosystem. Currently, the application of eDNA metabarcoding in macrozoobenthos biodiversity monitoring is largely affected by several technical factors, such as the sampling protocols, primer selection, and the completeness of reference database. Particularly, different methods and procedures often lead to contrasting results. Hence much attention is drawn to the field workflow and laboratory methodologies to be standardized to allow results to be compared across studies. In this paper, we reviewed the recent published studies referring to macrozoobenthos biodiversity monitoring using eDNA metabarcoding to discuss its definition, advantages, applications and its methodologies used in macrozoobenthos biodiversity biomonitoring and assessment. Especially we discussed the vital technical factors that influences eDNA metabarcoding, including sample collection protocol, gene marker selection, primer design, PCR bias, the completeness of reference database, and their corresponding recommendations to be optimized. Based on the summary above, we give several helpful suggestions for improving the efficiency and accuracy of macrozoobenthos monitoring using eDNA metabarcoding. We expect that this review will provide reliable guidelines for developing a sound macrozoobenthos biomonitoring design using eDNA metabarcoding. At last, we gave perspectives regarding several emerging applications and innovative methodologies of eDNA metabarcoding can be used in ecology.





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