Metagenomic Information Recovery from Human Stool Samples Is Influenced by Sequencing Depth and Profiling Method

Santiago-Rodriguez, Tasha M.; Garoutte, Aaron; Adams, Emmase; Nasser, Waleed; Ross, Matthew C.; La Reau, Alex; Henseler, Zachariah; Ward, Tonya; Knights, Dan; Petrosino, Joseph F.; Hollister, Emily B.

doi:10.3390/genes11111380

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Metagenomic Information Recovery from Human Stool Samples Is Influenced by Sequencing Depth and Profiling Method

Tasha M. Santiago-Rodriguez

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Diversigen Inc., Houston, TX 77021, USA

Alkek Center for Metagenomics and Microbiome Research, Baylor College of Medicine, Houston, TX 77030, USA

Department of Molecular Virology and Microbiology, Baylor College of Medicine, Houston, TX 77030, USA

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Diversigen Inc., Saint Paul, MN 55112, USA

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Department of Computer Science and Engineering, University of Minnesota, Minneapolis, MN 55455, USA

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Biotechnology Institute, College of Biological Sciences, University of Minnesota, Minneapolis, MN 55455, USA

Author to whom correspondence should be addressed.

Genes 2020, 11(11), 1380; https://doi.org/10.3390/genes11111380

Received: 3 November 2020 / Revised: 17 November 2020 / Accepted: 18 November 2020 / Published: 21 November 2020

(This article belongs to the Collection Microbiome Analysis Techniques and Discovery)

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Abstract

Sequencing of the 16S rRNA gene (16S) has long been a go-to method for microbiome characterization due to its accessibility and lower cost compared to shotgun metagenomic sequencing (SMS). However, 16S sequencing rarely provides species-level resolution and cannot provide direct assessment of other taxa (e.g., viruses and fungi) or functional gene content. Shallow shotgun metagenomic sequencing (SSMS) has emerged as an approach to bridge the gap between 16S sequencing and deep metagenomic sequencing. SSMS is cost-competitive with 16S sequencing, while also providing species-level resolution and functional gene content insights. In the present study, we evaluated the effects of sequencing depth on marker gene-mapping- and alignment-based annotation of bacteria in healthy human stool samples. The number of identified taxa decreased with lower sequencing depths, particularly with the marker gene-mapping-based approach. Other annotations, including viruses and pathways, also showed a depth-dependent effect on feature recovery. These results refine the understanding of the suitability and shortcomings of SSMS, as well as annotation tools for metagenomic analyses in human stool samples. Results may also translate to other sample types and may open the opportunity to explore the effect of sequencing depth and annotation method. View Full-Text

Keywords: alignment; marker gene; microbiome; shallow sequencing; shotgun metagenomic sequencing; virome

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MDPI and ACS Style

Santiago-Rodriguez, T.M.; Garoutte, A.; Adams, E.; Nasser, W.; Ross, M.C.; La Reau, A.; Henseler, Z.; Ward, T.; Knights, D.; Petrosino, J.F.; Hollister, E.B. Metagenomic Information Recovery from Human Stool Samples Is Influenced by Sequencing Depth and Profiling Method. Genes 2020, 11, 1380. https://doi.org/10.3390/genes11111380

AMA Style

Santiago-Rodriguez TM, Garoutte A, Adams E, Nasser W, Ross MC, La Reau A, Henseler Z, Ward T, Knights D, Petrosino JF, Hollister EB. Metagenomic Information Recovery from Human Stool Samples Is Influenced by Sequencing Depth and Profiling Method. Genes. 2020; 11(11):1380. https://doi.org/10.3390/genes11111380

Chicago/Turabian Style

Santiago-Rodriguez, Tasha M., Aaron Garoutte, Emmase Adams, Waleed Nasser, Matthew C. Ross, Alex La Reau, Zachariah Henseler, Tonya Ward, Dan Knights, Joseph F. Petrosino, and Emily B. Hollister. 2020. "Metagenomic Information Recovery from Human Stool Samples Is Influenced by Sequencing Depth and Profiling Method" Genes 11, no. 11: 1380. https://doi.org/10.3390/genes11111380

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Metagenomic Information Recovery from Human Stool Samples Is Influenced by Sequencing Depth and Profiling Method

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