Fungal ITS1 Deep-Sequencing Strategies to Reconstruct the Composition of a 26-Species Community and Evaluation of the Gut Mycobiota of Healthy Japanese Individuals.
Fungal ITS1 Deep-Sequencing Strategies to Reconstruct the Composition of a 26-Species Community and Evaluation of the Gut Mycobiota of Healthy Japanese Individuals.
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DOI：
10.3389/fmicb.2017.00238
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发表时间：
2017
影响因子：
5.2
通讯作者：
Nakamura S
中科院分区：
文献类型：
--
作者：
Motooka D;Fujimoto K;Tanaka R;Yaguchi T;Gotoh K;Maeda Y;Furuta Y;Kurakawa T;Goto N;Yasunaga T;Narazaki M;Kumanogoh A;Horii T;Iida T;Takeda K;Nakamura S
与细菌微生物组研究相比，由于缺乏足够的参考菌株和数据库，真菌生物群的研究仍然相对未被探索。内转录间隔区（ITS）区域的深度测序是真菌多样性分析的事实上的标准。然而，由于ITS区域的序列长度多种多样以及高通量测序（HTS）技术的复杂性，结果往往存在偏差。在这项研究中，一个策展的ITS数据库，ntF-ITS 1的构建。该数据库可用于真菌群落成员的分类分配。我们通过使用该数据库评估了真菌组分析策略的有效性，并使用三个HTS平台（Illumina MiSeq（MiSeq），Ion Torrent Personal Genome Machine（IonPGM）和Pacific Biosciences（PacBio））使用ITS 1测序表征了由代表15个属的26个物种组成的模拟真菌群落。我们的评估表明，PacBio的循环共识测序超过8个完整的通道最准确地重建了模拟社区的组成。使用这种策略对健康日本人的肠道菌群进行深度测序分析，发现了两种主要的菌群类型：由白色念珠菌或酿酒酵母组成的单种类型和多种类型。在这项研究中，我们提出了三个测序平台的最佳处理策略，其中PacBio平台可以最准确地估计真菌群落。本文描述的数据库和方法为新兴的真菌生物群研究领域提供了关键工具。
The study of mycobiota remains relatively unexplored due to the lack of sufficient available reference strains and databases compared to those of bacterial microbiome studies. Deep sequencing of Internal Transcribed Spacer (ITS) regions is the de facto standard for fungal diversity analysis. However, results are often biased because of the wide variety of sequence lengths in the ITS regions and the complexity of high-throughput sequencing (HTS) technologies. In this study, a curated ITS database, ntF-ITS1, was constructed. This database can be utilized for the taxonomic assignment of fungal community members. We evaluated the efficacy of strategies for mycobiome analysis by using this database and characterizing a mock fungal community consisting of 26 species representing 15 genera using ITS1 sequencing with three HTS platforms: Illumina MiSeq (MiSeq), Ion Torrent Personal Genome Machine (IonPGM), and Pacific Biosciences (PacBio). Our evaluation demonstrated that PacBio’s circular consensus sequencing with greater than 8 full-passes most accurately reconstructed the composition of the mock community. Using this strategy for deep-sequencing analysis of the gut mycobiota in healthy Japanese individuals revealed two major mycobiota types: a single-species type composed of Candida albicans or Saccharomyces cerevisiae and a multi-species type. In this study, we proposed the best possible processing strategies for the three sequencing platforms, of which, the PacBio platform allowed for the most accurate estimation of the fungal community. The database and methodology described here provide critical tools for the emerging field of mycobiome studies.