互补宏基因组学方法改进森林土壤中微生物多样性的重建

Complementary Metagenomic Approaches Improve Reconstruction of Microbial Diversity in a Forest Soil.

作者信息

Alteio L V, Schulz F, Seshadri R, Varghese N, Rodriguez-Reillo W, Ryan E, Goudeau D, Eichorst S A, Malmstrom R R, Bowers R M, Katz L A, Blanchard J L, Woyke T

机构信息

Graduate Program in Organismic and Evolutionary Biology, University of Massachusetts Amherst, Amherst, Massachusetts, USA.

Department of Energy, Joint Genome Institute, Berkeley, California, USA.

出版信息

mSystems. 2020 Mar 10;5(2):e00768-19. doi: 10.1128/mSystems.00768-19.

Abstract

Soil ecosystems harbor diverse microorganisms and yet remain only partially characterized as neither single-cell sequencing nor whole-community sequencing offers a complete picture of these complex communities. Thus, the genetic and metabolic potential of this "uncultivated majority" remains underexplored. To address these challenges, we applied a pooled-cell-sorting-based mini-metagenomics approach and compared the results to bulk metagenomics. Informatic binning of these data produced 200 mini-metagenome assembled genomes (sorted-MAGs) and 29 bulk metagenome assembled genomes (MAGs). The sorted and bulk MAGs increased the known phylogenetic diversity of soil taxa by 7.2% with respect to the Joint Genome Institute IMG/M database and showed clade-specific sequence recruitment patterns across diverse terrestrial soil metagenomes. Additionally, sorted-MAGs expanded the rare biosphere not captured through MAGs from bulk sequences, exemplified through phylogenetic and functional analyses of members of the phylum Analysis of 67 sorted-MAGs showed conserved patterns of carbon metabolism across four clades. These results indicate that mini-metagenomics enables genome-resolved investigation of predicted metabolism and demonstrates the utility of combining metagenomics methods to tap into the diversity of heterogeneous microbial assemblages. Microbial ecologists have historically used cultivation-based approaches as well as amplicon sequencing and shotgun metagenomics to characterize microbial diversity in soil. However, challenges persist in the study of microbial diversity, including the recalcitrance of the majority of microorganisms to laboratory cultivation and limited sequence assembly from highly complex samples. The uncultivated majority thus remains a reservoir of untapped genetic diversity. To address some of the challenges associated with bulk metagenomics as well as low throughput of single-cell genomics, we applied flow cytometry-enabled mini-metagenomics to capture expanded microbial diversity from forest soil and compare it to soil bulk metagenomics. Our resulting data from this pooled-cell sorting approach combined with bulk metagenomics revealed increased phylogenetic diversity through novel soil taxa and rare biosphere members. In-depth analysis of genomes within the highly represented phylum provided insights into conserved and clade-specific patterns of carbon metabolism.

摘要

土壤生态系统中存在着多种多样的微生物,但由于单细胞测序和全群落测序都无法完整呈现这些复杂群落的全貌,所以对其特征的了解仍不全面。因此,这一“未培养的大多数”的遗传和代谢潜力仍未得到充分探索。为应对这些挑战,我们应用了基于池化细胞分选的宏基因组学方法,并将结果与整体宏基因组学进行比较。对这些数据进行信息分类后,得到了200个宏基因组组装基因组(分选宏基因组组装基因组,sorted-MAGs)和29个整体宏基因组组装基因组(MAGs)。相对于联合基因组研究所的IMG/M数据库,分选的和整体的MAGs使土壤分类单元已知的系统发育多样性增加了7.2%,并在不同的陆地土壤宏基因组中显示出特定分支的序列招募模式。此外,分选宏基因组组装基因组扩展了通过整体序列的MAGs未捕获到的稀有生物圈,这通过对某一门成员的系统发育和功能分析得到了例证。对67个分选宏基因组组装基因组的分析显示,四个分支中存在保守的碳代谢模式。这些结果表明,宏基因组学能够对预测的代谢进行基因组解析,并证明了结合宏基因组学方法来挖掘异质微生物群落多样性的实用性。微生物生态学家历来使用基于培养的方法以及扩增子测序和鸟枪法宏基因组学来表征土壤中的微生物多样性。然而,微生物多样性研究中仍然存在挑战,包括大多数微生物难以在实验室培养以及来自高度复杂样本的序列组装有限。因此,未培养的大多数仍然是未开发遗传多样性的宝库。为应对与整体宏基因组学相关的一些挑战以及单细胞基因组学的低通量问题,我们应用了流式细胞术辅助的宏基因组学来从森林土壤中捕获扩展的微生物多样性,并将其与土壤整体宏基因组学进行比较。我们从这种池化细胞分选方法结合整体宏基因组学得到的数据显示,通过新的土壤分类单元和稀有生物圈成员,系统发育多样性增加。对高度代表性的某一门内基因组的深入分析提供了对保守和特定分支碳代谢模式的见解。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/46fe/7065516/98af8eb2d785/mSystems.00768-19-f0001.jpg

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