3组常用鱼类eDNA宏条形码通用引物对三亚水环境样品的物种检出效果比较  

Comparison of fish species detection effect of three sets of commonly used eDNA metabarcoding primers on Sanya water samples

作　　者：郭瑶杰 万武波 王海山 叶乐 陈治 GUO Yaojie;WAN Wubo;WANG Haishan;YE Le;CHEN Zhi(Hainan Tropical Ocean University/Yazhou Bay Innovation Institute/Key Laboratory of Utilization and Conservation for Tropical Marine Bioresources,Ministry of Education,Sanya 572022,China)

机构地区：[1]海南热带海洋学院/崖州湾创新研究院/热带海洋生物资源利用与保护教育部重点实验室,海南三亚572022

出　　处：《南方水产科学》2025年第1期66-76,共11页South China Fisheries Science

基　　金：海南省自然科学基金项目(422RC717,424QN298);国家自然科学基金青年科学基金项目(32002389);2023年海南热带海洋学院水产南繁联合开放课题(2023SCNFKF05);三亚市重大科技计划项目(ZDKJ-SY-2020-001)。

摘　　要：环境DNA (Environmental DNA, eDNA)宏条形码技术是一种高效率、高灵敏度且无侵入性的物种调查工具。目前基于eDNA宏条形码技术调查鱼类多样性的研究众多,但是该技术发展尚不完善,对不同引物的实际使用效果缺乏共识。为降低测序成本、筛选出实际使用效果最优的通用引物,选取三亚市鱼市场和亚特兰蒂斯水族馆共8个站位的水样,比较了3组常用鱼类eDNA宏条形码通用引物(MiFish-U、AcMDB07、Ac12S)在检测鱼类多样性方面的差异。结果表明:1) 3组引物在质控后总序列数、鱼类序列数、总可操作分类单元(Operational taxonomic units, OTUs)数、鱼类OTUs数和鱼类序列占比方面均存在极显著性差异(p<0.01),MiFish-U的扩增成功率和对鱼类物种的靶向性最高;2) MiFish-U检出的物种数最多(140种),AcMDB07和Ac12S则分别检出128和97种;3) Ac12S和AcMDB07的参考数据库尚不完善,两者分别有72.76%和42.11%的OTUs无法注释到种水平;4) Ac12S检出的特有鱼类很少(仅4种),暗示在实际使用过程中更容易被另外2组引物替代,而MiFish-U的可替代性最低;5) 3组引物反映鱼类丰度的总趋势相似,但对具体物种却存在一定差异。结果表明,综合OTUs注释等多种因素,特别是现有参考数据条件下,MiFish-U的物种检出效果优于AcMDB07和Ac12S。Environmental DNA(eDNA)metabarcoding is a high-efficiency,high-sensitivity and non-invasive species investiga-tion tool.At present,there are many studies on the investigation of fish diversity based on eDNA metabarcoding which however is not well-developed,with a lack of consensus on the actual use of different primers.In order to reduce the cost of sequencing and screen out the universal primers with the best practical effect,we selected the water samples from eight sites in Sanya fish markets and Atlantis Aquarium,then compared the differences of three sets of universal primers(MiFish-U,AcMDB07 and Ac12S)for fish eDNA.The results show that:1)There were significant differences in the reads number after quality control,the fish reads number,the total Operational taxonomic units(OTUs)number,the fish OTUs number and the ratio of fish reads among three sets of primers.MiFish-U had the highest amplification efficiency and targeting to fish species;2)MiFish-U had the highest number of species(140 species),while AcMDB07 and Ac12S had 128 and 97 species,respectively;3)The reference databases of Ac12S and AcMDB07 were not perfect,and 72.76%and 42.11%of the OTUs belonging to Ac12S and AcMDB07 could not be annotated to the species level,respectively;4)There are very few endemic fish detected by Ac12S(Only four species),suggesting that it was easier to be replaced by the other two sets of primers in the actual use process,and MiFish-U had the lowest substitutability;5)The three sets of primers reflected similar general trends in the fish abundance,but there were some differences in the specific species.The results show that MiFish-U is superior to AcMDB07 and Ac12S in species detection,considering various factors such as OTUs annotation,especially the conditions of existing reference data.

关 键 词：环境DNA宏条形码 鱼类多样性 通用引物 MiFish-U 海南 

分 类 号：S931.2[农业科学—渔业资源]