Data from: MiFish, a set of universal PCR primers for metabarcoding environmental DNA from fishes: detection of >230 subtropical marine species

We developed a set of universal PCR primers (MiFish-U/E) for metabarcoding environmental DNA (eDNA) from fishes. Primers were designed using aligned whole mitochondrial genome (mitogenome) sequences from 880 species, supplemented by partial mitogenome sequences from 160 elasmobranchs (sharks and rays). The primers target a hypervariable region of the 12S rRNA gene (163–185 bp), which contains sufficient information to identify fishes to taxonomic family, genus and species except for some closely related congeners. To test versatility of the primers across a diverse range of fishes, we sampled eDNA from four tanks in the Okinawa Churaumi Aquarium with known species compositions, prepared dual-indexed libraries and performed paired-end sequencing of the region using high-throughput next-generation sequencing (NGS) technologies. Out of the 180 marine fish species contained in the four tanks with reference sequences in a custom database, we detected 168 species (93.3%) distributed across 59 families and 123 genera. These fishes are not only taxonomically diverse, ranging from sharks and rays to higher teleosts, but are also greatly varied in their ecology, including both pelagic and benthic species living in shallow coastal to deep waters. We also sampled natural seawaters around coral reefs near the aquarium and detected 93 fish species using this approach. Of the 93 species, 64 were not detected in the four aquarium tanks, rendering the total number of species detected to 232 (from 70 families and 152 genera). The metabarcoding approach presented here is non-invasive, more efficient, more cost-effective, and more sensitive than the traditional survey methods. It has the potential to serve as an alternative (or complementary) tool for biodiversity monitoring that revolutionises natural resource management and ecological studies of fish communities on larger spatial and temporal scales.

本研究开发了一套用于鱼类环境DNA（environmental DNA, eDNA）元条形码（metabarcoding）分析的通用PCR引物（MiFish-U/E）。该引物基于880个物种的完整线粒体基因组（mitogenome）比对序列设计，并补充了160种板鳃类（鲨及鳐类）的部分线粒体基因组序列。引物靶向12S核糖体RNA（12S rRNA）基因的一段高变区域（长度163–185 bp），该区域携带充足的分类鉴定信息，可将鱼类鉴定至科、属、种阶元，仅部分近缘同属物种除外。为验证该引物在多样鱼类类群中的通用性，我们从冲绳美丽海水族馆（Okinawa Churaumi Aquarium）的4个已知物种组成的水族箱中采集环境DNA，构建双索引文库，并采用高通量下一代测序（next-generation sequencing, NGS）技术对该目标区域进行双端测序。在自定义参考数据库收录的该4个水族箱所含的180种海洋鱼类中，我们成功检出168种（占比93.3%），这些物种隶属于59个科、123个属。这些鱼类不仅分类学多样性丰富，涵盖从鲨鳐类到高等硬骨鱼的多个类群，同时生态类型多样，包含栖息于浅海沿岸至深海水域的浮游性与底栖性物种。我们同时采集了水族馆附近珊瑚礁周边的天然海水样本，并通过该方法检出93种鱼类，其中64种未在4个水族箱的检测结果中出现，使得本次研究累计检出的物种总数达232种，隶属于70个科、152个属。本研究采用的元条形码技术相较于传统调查方法具有无创、高效、经济且灵敏度更高的优势，有望作为生物多样性监测的替代（或补充）工具，革新大空间与时间尺度下的鱼类群落生态学研究与自然资源管理工作。