Multicenter assessment of microbial community profiling using 16S rRNA gene sequencing and shotgun metagenomic sequencing

Han, Dongsheng; Gao, Peng; Li, Rui; Tan, Ping; Xie, Jiehong; Zhang, Rui; Li, Jinming

doi:10.1016/j.jare.2020.07.010

Cited by 42 publications

(36 citation statements)

References 75 publications

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“…Workflows for metagenomics are however complex and prone to bias and errors at all steps, from sample collection and storage [ 6 , 7 ] to DNA extraction [ 8 , 9 ], sequencing and bioinformatics analysis [ 10 , 11 ]. Methodological bias can lead to substantial differences in observed microbiota profiles, resulting in considerable variability in results across studies and laboratories using different protocols [ 12 , 13 ]. To improve consistency and enhance confidence in the accuracy of measurement results, standardization of metagenomic analysis methods has thus been recognized as a pressing need by industrial and regulatory sectors [ 14 ].…”

Section: Introductionmentioning

confidence: 99%

Validation and standardization of DNA extraction and library construction methods for metagenomics-based human fecal microbiome measurements

et al. 2021

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Background Validation and standardization of methodologies for microbial community measurements by high-throughput sequencing are needed to support human microbiome research and its industrialization. This study set out to establish standards-based solutions to improve the accuracy and reproducibility of metagenomics-based microbiome profiling of human fecal samples. Results In the first phase, we performed a head-to-head comparison of a wide range of protocols for DNA extraction and sequencing library construction using defined mock communities, to identify performant protocols and pinpoint sources of inaccuracy in quantification. In the second phase, we validated performant protocols with respect to their variability of measurement results within a single laboratory (that is, intermediate precision) as well as interlaboratory transferability and reproducibility through an industry-based collaborative study. We further ascertained the performance of our recommended protocols in the context of a community-wide interlaboratory study (that is, the MOSAIC Standards Challenge). Finally, we defined performance metrics to provide best practice guidance for improving measurement consistency across methods and laboratories. Conclusions The validated protocols and methodological guidance for DNA extraction and library construction provided in this study expand current best practices for metagenomic analyses of human fecal microbiota. Uptake of our protocols and guidelines will improve the accuracy and comparability of metagenomics-based studies of the human microbiome, thereby facilitating development and commercialization of human microbiome-based products.

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Section: Introductionmentioning

confidence: 99%

Validation and standardization of DNA extraction and library construction methods for metagenomics-based human fecal microbiome measurements

et al. 2021

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“…In addition, the reliance on “universal” primers for DNA amplification may introduce biases whereby some species are amplified more than others, with taxonomic coverage reported from 11% to 93% depending on primer choice ( Thomas et al, 2012 ). Other sequencing artifacts, such as polymerase errors ( Cline et al, 1996 ), chimeras ( Haas et al, 2011 ; Eloe-Fadrosh et al, 2016 ), 16S rRNA copy number variation ( Louca et al, 2018 ), and laboratory contamination ( de Goffau et al, 2019 ; Han et al, 2020 ) are all exacerbated during PCR amplification.…”

Section: Compositional and Functional Characterization Of The Microbiome (Stage I)mentioning

confidence: 99%

Key Technologies for Progressing Discovery of Microbiome-Based Medicines

et al. 2021

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A growing number of experimental and computational approaches are illuminating the “microbial dark matter” and uncovering the integral role of commensal microbes in human health. Through this work, it is now clear that the human microbiome presents great potential as a therapeutic target for a plethora of diseases, including inflammatory bowel disease, diabetes and obesity. The development of more efficacious and targeted treatments relies on identification of causal links between the microbiome and disease; with future progress dependent on effective links between state-of-the-art sequencing approaches, computational analyses and experimental assays. We argue determining causation is essential, which can be attained by generating hypotheses using multi-omic functional analyses and validating these hypotheses in complex, biologically relevant experimental models. In this review we discuss existing analysis and validation methods, and propose best-practice approaches required to enable the next phase of microbiome research.

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“…16S rRNA gene sequence analysis can be used for a complete assessment of microbial diversity by selectively amplifying and sequencing the hypervariable regions of the 16S rRNA gene. It is a highly efficient and cost-effective technology easily accessible by various bioinformatics tools and has become a frequently used technique for profiling intricate microbial communities (Han et al, 2020). It can be used to identify novel, unculturable, and phenotypically unidentifiable microbes (Clarridge, 2004).…”

Section: Genomics and 16s Rrna For Bioremediationmentioning

confidence: 99%

“…It is the only way to study the microbial community with no markers like viruses (Quince et al, 2017;Vermote et al, 2018). It allows strain-level remodeling in the taxonomic analysis and pathway predictions for the functional annotation of the microbiome under study (Han et al, 2020).…”

Section: Shotgun Sequencingmentioning

confidence: 99%

Microbial Biodiversity and Bioremediation Assessment Through Omics Approaches

Chandran

Meena

Sharma

2020

Front. Environ. Chem.

110

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Industrialization, population burst, and changing lifestyles have resulted in the genesis of non-degradable pollutants languishing the environment and human health. Biological approaches using microorganisms are gaining importance as an eco-friendly and cost-effective substitute to mitigate the pollution load. Microorganisms can survive in a divergent environment and produce metabolites that can degrade and transform pollutants making it possible to revive contaminated sites naturally. Modern omics technologies like metagenomics, transcriptomics, proteomics, etc. have been used nowadays design strategies to study ecology and diversity of microorganisms and their application in environmental monitoring and bioremediation. The present article will focus on the omics techniques reportedly used in environmental monitoring to tackle the pollution load.

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Multicenter assessment of microbial community profiling using 16S rRNA gene sequencing and shotgun metagenomic sequencing

Abstract: Graphical abstract

Cited by 42 publications

References 75 publications

Validation and standardization of DNA extraction and library construction methods for metagenomics-based human fecal microbiome measurements

Validation and standardization of DNA extraction and library construction methods for metagenomics-based human fecal microbiome measurements

Key Technologies for Progressing Discovery of Microbiome-Based Medicines

Microbial Biodiversity and Bioremediation Assessment Through Omics Approaches

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