豆科植物與根瘤菌的互動關係受到廣泛研究，主因為兩者共生固氮的能力在生態系統中氮循環、永續農業發展以及植物與微生物間的共演化中具有其重要性。透過雙方基因體所決定的訊息交換，宿主與其微生物建立共生的專一性，以此發展多樣且複雜的共生關係。過往的研究多利用分離培養、擴增特定序列或短序列霰彈槍定序法來了解根瘤菌，但這些方法可能會導致潛在的結果偏差或無法取得完整的基因體資訊。長讀取定序被認為是定序技術的第三次革命，使我們不僅獲得高品質的共生細菌基因體，且能同時分析來自宿主的基因體，達到共生總基因體 (hologenome) 分析的層級。含羞草屬 (Mimosa) 隸屬於豆科蘇木亞科的含羞草分支 (mimosoid)，具有和beta-rhizobia共生的能力，被認為是beta-rhizobia的研究熱點，其中刺軸含羞木 (Mimosa pigra) 為屬內其中一種廣布全球的入侵植物。然而，現今研究對於入侵植物的在地適應了解甚少。臺灣作為研究與含羞草屬共生的beta-rhizobia先驅，我們使用長讀取定序法分析了十二個來自刺軸含羞木根瘤的共生總基因體，用以對比前人利用分離培養進行鑑定的結果。我們的調查中發現原先只被記錄在原生地的其中兩個變種Mimosa pigra var. asperata 和 Mimosa pigra var. dehiscens可能早已擴張到臺灣。此外，我們還驗證了Cupriavidus 和 Paraburkholderia能透過保守的共生固氮基因簇來和這兩個變種結瘤，其中除了C. taiwanensis以外，我們也在刺軸含羞木的根瘤中發現兩種全新的Cupriavidus物種。值得注意的是，這些Cupriavidus物種通常攜帶非典型的質體型態，這可能暗示臺灣分布的刺軸含羞木開始選擇新質體型態的Cupriavidus物種共生，不同於過去發現刺軸含羞木根瘤菌是以Paraburkholderia優勢共生的結果。總的來說，共生總基因體資訊幫助我們了解豆科植物與根瘤菌的共生與功能性基因庫，長讀取定序技術是一個有潛力且持續進展的方法來拓展我們對於非模式豆科植物與其共生根瘤菌互動的認知。

Legume-rhizobia relationship is extensively studied due to the importance of biological nitrogen fixation in sustainable agriculture, nitrogen cycle in ecosystem, and the co-evolution between plants and microorganisms. Through signal exchange determined by both genomes, the host and its microorganisms establish the specificity, forming diverse symbiotic relationships. Previous studies have shown rhizobia within nodules through isolation, amplification, and short-read shotgun metagenomic sequencing, yielding inherent biases and incomplete genomic information. Long-read sequencing, the third sequencing revolution, enables the acquisition of high-quality bacterial genomes and partial host genomes, achieving the hologenome analysis. Mimosa, known for symbiotic capabilities with beta-rhizobia, is considered a hotspot in beta-rhizobia research. Among its members, Mimosa pigra is a globally invasive plant. However, little research has been conducted to show the local adaptations of invasive plants. Here, we utilized long-read sequencing to dissect 12 hologenomes of M. pigra in Taiwan, where it has played a pioneering role in the study of symbiosis between Mimosa and beta-rhizobia, and then compared results with prior investigation. Our findings suggested Mimosa pigra var. asperata and Mimosa pigra var. dehiscens had been introduced to Taiwan based on morphological features and chloroplast sequences. Moreover, we evidenced Cupriavidus and Paraburkholderia can equally nodulate both M. pigra varieties with conserved nif/nod gene clusters, inconsistent with prior Paraburkholderia predominance. Two novel Cupriavidus species and C. taiwanensis were identified as symbionts of M. pigra. Notably, most of these symbionts carried novel plasmid types, indicating an effective way to form symbiotic relationship with Cupriavidus. Overall, Hologenome information provided insights into the symbiotic and functional gene repertoire. Long-read sequencing technologies are promising and continually advancing approaches to expand our understanding of the legume-rhizobia relationship, especially in non-model legumes with their symbiotic rhizobia.

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