De novo Sequencing of Novel Mycoviruses From Fusarium sambucinum: An Attempt on Direct RNA Sequencing of Viral dsRNAs.
De novo Sequencing of Novel Mycoviruses From Fusarium sambucinum: An Attempt on Direct RNA Sequencing of Viral dsRNAs.
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DOI：
10.3389/fmicb.2021.641484
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发表时间：
2021
影响因子：
5.2
通讯作者：
Chiba S
中科院分区：
文献类型：
--
作者：
Mizutani Y;Uesaka K;Ota A;Calassanzio M;Ratti C;Suzuki T;Fujimori F;Chiba S
在包括真菌在内的多种生物体中不断发现越来越多的病毒。最近的研究揭示了微生物中广泛的病毒多样性以及这些病毒在自然环境中的潜在重要性。尽管短读长、高通量测序 (HTS) 加速了病毒探索，但由于 RNA 基因组的微观多样性和二级结构等多种生物/分子特征，病毒从头测序仍然具有挑战性。本研究使用传统 HTS 和长读长直接 RNA 测序 (DRS) 对从感染两种接骨木镰刀菌菌株 FA1837 和 FA2242 的真菌病毒中获得的多个双链 (ds) RNA (dsRNA) 元件进行了从头测序。对两种技术读取的数据进行从头组装，生成了病毒的近乎完整的基因组序列，序列同源性搜索和系统发育分析表明，这些病毒代表了低病毒科、整体病毒科和线粒体病毒科的新物种。然而，基于 DRS 的共有序列包含许多与 HTS 共有序列不同的插入缺失错误，这些错误妨碍了准确的开放阅读框 (ORF) 预测。尽管就其目前的性能而言，使用 DRS 来确定病毒基因组序列还为时过早，但 DRS 介导的测序由于其长读取能力和相对结构耐受性，显示出作为一种用户友好的平台对 RNA 病毒进行一次性全基因组测序的巨大潜力。
An increasing number of viruses are continuously being found in a wide range of organisms, including fungi. Recent studies have revealed a wide viral diversity in microbes and a potential importance of these viruses in the natural environment. Although virus exploration has been accelerated by short-read, high-throughput sequencing (HTS), and viral de novo sequencing is still challenging because of several biological/molecular features such as micro-diversity and secondary structure of RNA genomes. This study conducted de novo sequencing of multiple double-stranded (ds) RNA (dsRNA) elements that were obtained from fungal viruses infecting two Fusarium sambucinum strains, FA1837 and FA2242, using conventional HTS and long-read direct RNA sequencing (DRS). De novo assembly of the read data from both technologies generated near-entire genomic sequence of the viruses, and the sequence homology search and phylogenetic analysis suggested that these represented novel species of the Hypoviridae, Totiviridae, and Mitoviridae families. However, the DRS-based consensus sequences contained numerous indel errors that differed from the HTS consensus sequences, and these errors hampered accurate open reading frame (ORF) prediction. Although with its present performance, the use of DRS is premature to determine viral genome sequences, the DRS-mediated sequencing shows great potential as a user-friendly platform for a one-shot, whole-genome sequencing of RNA viruses due to its long-reading ability and relative structure-tolerant nature.
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