以單一顆粒基因體研究臺灣大型真核微生物

Abstract

隨著高通量定序技術的進步，真核生物基因體序列資料數據量顯著增加。真核生物演化樹中絕大多數是由微生物組成，但前人對於真核生物基因體的研究著重於動物、植物和真菌，導致真核微生物中有許多演化分支缺乏基因體紀錄。為了拓展對環境真核微生物的理解，本研究將形態與單一顆粒基因體分析整合，利用配有攝影設備的大顆粒流式細胞分選儀，或是手動使用微量移液管，來分選在臺灣水域樣本的單一大型真核微生物顆粒，進行全基因放大與定序，再分析其基因體、親緣關係與微生物交互作用。本研究成功從淡水樣本取得多個單一顆粒的環狀胞器基因體並分析其親緣關係。特別在矽藻方面找出獨特葉綠體序列的演化支系。另外，從綠島浮游性有孔蟲的單一顆粒中，分別組裝出有孔蟲與金色藻屬的環狀粒線體基因體，顯示了這兩個真核微生物間有潛在的交互作用。本研究也分析原核生物在海洋環境中的組成，發現綠島有孔蟲有特定的菌相。這些結果顯示單一大顆粒的基因體分析，對揭示真核微生物的基因體、演化多樣性及微生物交互作用具有很大的潛力。

The advancement of high-throughput sequencing has significantly increased our access to genomic datasets of eukaryotes. The vast majority of the eukaryotic Tree of Life (eToL) comprises microbial organisms with crucial roles in various ecosystems. However, previous studies on eukaryotic genomes have mainly focused on animals, plants, and fungi. Research on microeukaryote genomics and evolutionary diversity remains limited by their generally unculturable nature, with numerous lineages lacking genomic records, especially some that have large cell sizes. To enhance our understanding of their genomics, ecology and evolution, a workflow that integrates morphology with the analysis of single-particle genomes was utilized in this study, particles of interest from freshwater and marine environments in Taiwan isolated using either manual micro-pipetting or a large-particle flow cytometer/sorter equipped with a camera. This was followed by whole-genome amplification, sequencing, de novo assembly and genomic analyses. Circularized or nearly complete organellar genome sequences were successfully recovered, enabling the phylogenomic analyses of the isolated particles. By linking the morphological data of particles with their phylogenetic placements, large particles within Bacillariophyta (diatoms) were identified and found to represent lineages with distinct plastid genomes. Additionally, circularized mitochondrial genomes of planktonic foraminifera (Rhizaria) and Chrysochromulina sp. (Haptophyta) were assembled from a single particle, suggesting putative microbial interactions between these two microbes. Furthermore, multiple prokaryotic metagenome-assembled genomes were recovered from both foraminiferan single particles and the surrounding seawater, which differ in their bacterial community, suggesting foraminiferans may be associated with specific microbiomes. Overall, this study highlights the potential of large single-particle for unraveling the hidden genomic and evolutionary diversity of protists and their associated microorganisms.