Comparative analysis of 16S rRNA gene and metagenome sequencing in pediatric gut microbiomes

Peterson, Danielle; Bonham, Kevin S.; Rowland, Sophie; Pattanayak, Cassandra Wolos; Klepac‐Ceraj, Vanja

doi:10.1101/2021.02.20.432118

Cited by 9 publications

(12 citation statements)

References 53 publications

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“…Shotgun sequencing provides higher taxonomic resolution, is less prone to bias, can pro le eukaryotic and viral constituents, and provides information on the functional potential of the native community (36,37). However, it is signi cantly more costly due to requiring deeper sequencing depth to nd rarer taxa and genes and to compensate for hostassociated reads (38). It can also be limited by the available genomes present in the database (e.g., NCBI).…”

Section: Discussionmentioning

confidence: 99%

“…It can also be limited by the available genomes present in the database (e.g., NCBI). 16S rRNA gene sequencing is cost-effective and has less complex bioinformatic requirements (38). However, it is prone to primer biases which can skew the results or miss certain taxonomic groups and only provides taxonomic information on bacterial and some archaeal members (36,37,39).…”

Section: Discussionmentioning

confidence: 99%

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Insights from shotgun metagenomics into bacterial species and metabolic pathways associated with NAFLD in obese youth

Testerman¹,

Galuppo³

et al. 2021

Preprint

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Background and Aims NAFLD is the most common form of liver disease and is often the precursor for more serious liver conditions such as NASH and cirrhosis. The gut microbiome has been implicated in the development of NAFLD, but studies have often compared healthy individuals to those with the disease. In this study, we aimed to examine the taxonomic and functional differences between the microbiomes of obese youth with and without NAFLD. Approach and Results Shotgun metagenome sequencing was used to profile the microbial communities of 36 subjects, half of which were diagnosed with NAFLD. Beta diversity analysis showed community-wide differences between the cohorts (p = 0.002). Specific taxonomic differences included the species Fusicatenibacter saccharivorans (p = 0.042), Romboutsia ilealis (p = 0.046), and Actinomyces sp. ICM47 (p = 0.0009) elevated in the NAFLD-positive group and Bacteroides thetaiotamicron (p = 0.0002) elevated in the NAFLD-negative group. At the phylum level, Bacteroidetes (p < 0.0001) was elevated in the NAFLD-negative group. Functionally, branched chain amino acid (p = 0.01343) and aromatic amino acid (p = 0.01343) synthesis pathways were elevated in the NAFLD-positive group along with numerous energy utilization pathways including pyruvate fermentation to acetate (p = 0.01318). Conclusions The link between obesity and NAFLD development in younger individuals has rarely been probed, with most studies focusing on adults and comparing healthy individuals with obese, NAFLD-positive individuals. This study supports the idea that the NAFLD phenotype displays a differentiated microbial and functional signature from the obesity phenotype.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Insights from shotgun metagenomics into bacterial species and metabolic pathways associated with NAFLD in obese youth

Testerman¹,

Galuppo³

et al. 2021

Preprint

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“…Raw and processed data is available through SRA (NCBI BioProject PRJNA695570 ) and at OSF.io ( Peterson et al, 2021 ).…”

Section: Data Availability Statementmentioning

confidence: 99%

Comparative Analysis of 16S rRNA Gene and Metagenome Sequencing in Pediatric Gut Microbiomes

et al. 2021

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The colonization of the human gut microbiome begins at birth, and over time, these microbial communities become increasingly complex. Most of what we currently know about the human microbiome, especially in early stages of development, was described using culture-independent sequencing methods that allow us to identify the taxonomic composition of microbial communities using genomic techniques, such as amplicon or shotgun metagenomic sequencing. Each method has distinct tradeoffs, but there has not been a direct comparison of the utility of these methods in stool samples from very young children, which have different features than those of adults. We compared the effects of profiling the human infant gut microbiome with 16S rRNA amplicon vs. shotgun metagenomic sequencing techniques in 338 fecal samples; younger than 15, 15–30, and older than 30 months of age. We demonstrate that observed changes in alpha-diversity and beta-diversity with age occur to similar extents using both profiling methods. We also show that 16S rRNA profiling identified a larger number of genera and we find several genera that are missed or underrepresented by each profiling method. We present the link between alpha diversity and shotgun metagenomic sequencing depth for children of different ages. These findings provide a guide for selecting an appropriate method and sequencing depth for the three studied age groups.

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“…Limitations remain for short read analyses in metagenomics, despite the undoubted progress over the last decade or more. We have already alluded to the difficulties of reliably detecting bacterial taxa, especially at species or strain level, which are even more exacerbated in 16S rRNA amplicon sequencing (Peterson et al 2021), but detecting DNA reads from a pathogen is not necessarily associated with infection (Bender and Dien Bard 2018). An attempt has been made to alleviate this issue by integrating growth rates into the pipeline, with the idea that quickly growing rather than non-growing pathogenic bacteria are more likely to play a role in disease (Vich Vila et al 2018).…”

Section: Discussionmentioning

confidence: 99%

Wochenende - modular and flexible alignment-based shotgun metagenome analysis

Rosenboom

Friedrich

Poertner

et al. 2022

Preprint

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Shotgun metagenome analysis provides a robust and verifiable method for comprehensive microbiome analysis of fungal, viral, archaeal and bacterial taxonomy, particularly with regard to visualization of read mapping location, normalization options, growth dynamics and functional gene repertoires. Current read classification tools use non-standard output formats, or do not fully show information on mapping location. As reference datasets are not perfect, portrayal of mapping information is critical for judging results effectively. Our alignment-based pipeline, Wochenende, incorporates flexible quality control, trimming, mapping, various filters and normalization. We observe stringent filtering of mismatches and use of mapping quality sharply reduces the number of false positives. Further modules allow genomic visualization, as well as integration and subsequent plotting of pipeline results. Our novel normalization approach additionally allows calculation of absolute abundance profiles by comparison with reads assigned to the human host genome. Wochenende has the ability to find and filter alignments to all kingdoms of life using both short and long reads, and requires only good quality reference genomes. Wochenende is available at https://github.com/MHH-RCUG/Wochenende.

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Comparative analysis of 16S rRNA gene and metagenome sequencing in pediatric gut microbiomes

Cited by 9 publications

References 53 publications

Insights from shotgun metagenomics into bacterial species and metabolic pathways associated with NAFLD in obese youth

Insights from shotgun metagenomics into bacterial species and metabolic pathways associated with NAFLD in obese youth

Comparative Analysis of 16S rRNA Gene and Metagenome Sequencing in Pediatric Gut Microbiomes

Wochenende - modular and flexible alignment-based shotgun metagenome analysis

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