Long- and short-read metabarcoding technologies reveal similar spatiotemporal structures in fungal communities
Long- and short-read metabarcoding technologies reveal similar spatiotemporal structures in fungal communities
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DOI：
10.1111/1755-0998.13387
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发表时间：
2021-05-06
影响因子：
7.7
通讯作者：
Ryberg, Martin
中科院分区：
文献类型：
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作者：
Furneaux, Brendan;Bahram, Mohammad;Ryberg, Martin
真菌形成多样的群落，在许多陆地生态系统中发挥着重要作用，但从环境序列中对真菌的分类和系统发育定位存在方法论上的挑战。为了解决这些挑战，我们利用土壤元条形码技术研究了真菌群落的时空结构，采用四种不同的测序策略：利用Illumina MiSeq、Ion Torrent Ion S5和PacBio RS II对ITS2区域进行短扩增子测序（300-400 bp），并利用PacBio RS II对ITS和部分LSU区域进行长扩增子测序（1200-1600 bp）。由此产生的群落结构和多样性更多地依赖于统计方法而不是测序技术。使用长扩增子测序可以从元条形码读取构建系统发育树，这有利于序列的分类鉴定。然而，在不同的社区中，长读取给去噪算法带来了问题。我们提出了一种解决方案，即在去噪之前将reads分割成更短的同源区域，然后重建完整的去噪reads。在选择短扩增子和长扩增子时，我们建议采用混合方法，使用短扩增子来采样宽度和深度，使用长扩增子来表征本地物种池，以改进鉴定和系统发育分析。
Fungi form diverse communities and play essential roles in many terrestrial ecosystems, yet there are methodological challenges in taxonomic and phylogenetic placement of fungi from environmental sequences. To address such challenges, we investigated spatiotemporal structure of a fungal community using soil metabarcoding with four different sequencing strategies: short-amplicon sequencing of the ITS2 region (300-400 bp) with Illumina MiSeq, Ion Torrent Ion S5 and PacBio RS II, all from the same PCR library, as well as long-amplicon sequencing of the full ITS and partial LSU regions (1200-1600 bp) with PacBio RS II. Resulting community structure and diversity depended more on statistical method than sequencing technology. The use of long-amplicon sequencing enables construction of a phylogenetic tree from metabarcoding reads, which facilitates taxonomic identification of sequences. However, long reads present issues for denoising algorithms in diverse communities. We present a solution that splits the reads into shorter homologous regions prior to denoising, and then reconstructs the full denoised reads. In the choice between short and long amplicons, we suggest a hybrid approach using short amplicons for sampling breadth and depth, and long amplicons to characterize the local species pool for improved identification and phylogenetic analyses.