Table_1_Metagenomic 18S rDNA reads revealed zonation of eukaryotic communities in the Yongle blue hole.DOCX

The Yongle blue hole (YBH), situated in the South China Sea, represents a compelling subject of study in marine microbiology due to its unique redox-layered microbial ecosystems. However, the diversity and ecology of microbial eukaryotes within the YBH remains underexplored. This study endeavors to bridge this gap through the application of the in situ microbial filtration and fixation (ISMIFF) device to collect 0.22–30 μm microbial samples from 21 water layers of YBH. Subsequent extraction of 18S rRNA metagenomic reads of 21 metagenomes and 10 metatranscriptomes facilitated a comprehensive analysis of community structures. Findings revealed a pronounced superiority in the diversity and richness of eukaryotic microorganisms in the oxic zone compared to its suboxic and anoxic counterparts. Notably, Dinophyceae and Maxillopoda emerged as the predominant taxa based on the analysis of the 18S rRNA reads for the V4 and V9 regions, which showed stratification In their relative abundance and suggested their potential role in the thermo-halocline boundaries and oxic-anoxic interface. Specifically, In these eukaryotic microbial communities, Dinophyceae exhibited significant abundance at 20 m (20.01%) and 105 m (26.13%) water depths, while Maxillopoda was prevalent at 40 m (22.84%), 80 m (23.19%), and 100 m (15.42%) depths. A part of these organisms, identified as larvae and protists, were likely attracted by swarming chemosynthetic bacterial prey prevailing at the thermo-halocline boundaries and oxic-anoxic interface. Furthermore, the phylogenetic relationships of the major 18S operational taxonomic units (OTUs) showed a close adjacency to known species, except for three Dinophyceae OTUs. In conclusion, this study provides critical insights into the vertical distribution and transcriptional activity of <30-μm eukaryotic microbes, shedding light on the taxonomic novelty of eukaryotic microorganisms within the semi-enclosed blue holes.

位于南海的永乐蓝洞（Yongle Blue Hole, YBH）因其独特的氧化还原分层微生物生态系统，成为海洋微生物学研究中极具吸引力的课题。然而，目前针对永乐蓝洞内部真核微生物的多样性与生态特征的研究仍较为匮乏。本研究通过应用原位微生物过滤固定装置（in situ microbial filtration and fixation, ISMIFF），从永乐蓝洞的21个水层中采集了粒径0.22~30μm的微生物样品，以填补这一研究空白。随后对21个宏基因组和10个宏转录组的18S核糖体RNA（18S rRNA）宏基因组读段进行提取，从而实现对群落结构的全面解析。研究结果显示，与缺氧亚区和厌氧区相比，有氧区的真核微生物多样性与丰富度显著更高。值得注意的是，通过对18S rRNA的V4与V9可变区读段的分析，甲藻纲（Dinophyceae）和颚足纲（Maxillopoda）成为优势类群；二者的相对丰度呈现明显的分层特征，且其分布可能与温盐跃层边界及有氧-厌氧界面存在关联。具体而言，在该真核微生物群落中，甲藻纲在20m（20.01%）和105m（26.13%）水深处丰度显著，而颚足纲则在40m（22.84%）、80m（23.19%）及100m（15.42%）水深处占据优势。部分被鉴定为幼虫与原生生物的类群，可能是被温盐跃层边界及有氧-厌氧界面处大量聚集的化能合成细菌猎物所吸引。此外，对主要18S操作分类单元（operational taxonomic units, OTUs）的系统发育分析显示，除3个甲藻纲OTUs外，其余类群均与已知物种具有较近的亲缘关系。综上，本研究深入解析了粒径小于30μm的真核微生物的垂直分布与转录活性，为半封闭蓝洞内部真核微生物的分类学新发现提供了重要依据。