Faculty Opinions recommendation of A human gut microbial gene catalogue established by metagenomic sequencing.

Abstract Background Metagenomic sequencing has led to the identification and assembly of many new bacterial genome sequences. These bacteria often contain plasmids: usually small, circular double-stranded DNA molecules that may transfer across bacterial species and confer antibiotic resistance. These plasmids are generally less studied and understood than their bacterial hosts. Part of the reason for this is insufficient computational tools enabling the analysis of plasmids in metagenomic samples. Results We developed SCAPP (Sequence Contents-Aware Plasmid Peeler)—an algorithm and tool to assemble plasmid sequences from metagenomic sequencing. SCAPP builds on some key ideas from the Recycler algorithm while improving plasmid assemblies by integrating biological knowledge about plasmids. We compared the performance of SCAPP to Recycler and metaplasmidSPAdes on simulated metagenomes, real human gut microbiome samples, and a human gut plasmidome dataset that we generated. We also created plasmidome and metagenome data from the same cow rumen sample and used the parallel sequencing data to create a novel assessment procedure. Overall, SCAPP outperformed Recycler and metaplasmidSPAdes across this wide range of datasets. Conclusions SCAPP is an easy to use Python package that enables the assembly of full plasmid sequences from metagenomic samples. It outperformed existing metagenomic plasmid assemblers in most cases and assembled novel and clinically relevant plasmids in samples we generated such as a human gut plasmidome. SCAPP is open-source software available from: https://github.com/Shamir-Lab/SCAPP.

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A Protocol for Extraction of Infective Viromes Suitable for Metagenomics Sequencing from Low Volume Fecal Samples

Viruses ◽

10.3390/v11070667 ◽

2019 ◽

Vol 11 (7) ◽

pp. 667 ◽

Cited By ~ 4

Author(s):

Ling Deng ◽

Ronalds Silins ◽

Josué L. Castro-Mejía ◽

Witold Kot ◽

Leon Jessen ◽

...

Keyword(s):

Genome Sequencing ◽

Large Scale ◽

Metagenomic Sequencing ◽

High Input ◽

Human Gut ◽

Specific Host ◽

Substantial Progress ◽

Low Volume ◽

Made In

The human gut microbiome (GM) plays an important role in human health and diseases. However, while substantial progress has been made in understanding the role of bacterial inhabitants of the gut, much less is known regarding the viral component of the GM. Bacteriophages (phages) are viruses attacking specific host bacteria and likely play important roles in shaping the GM. Although metagenomic approaches have led to the discoveries of many new viruses, they remain largely uncultured as their hosts have not been identified, which hampers our understanding of their biological roles. Existing protocols for isolation of viromes generally require relatively high input volumes and are generally more focused on extracting nucleic acids of good quality and purity for down-stream analysis, and less on purifying viruses with infective capacity. In this study, we report the development of an efficient protocol requiring low sample input yielding purified viromes containing phages that are still infective, which also are of sufficient purity for genome sequencing. We validated the method through spiking known phages followed by plaque assays, qPCR, and metagenomic sequencing. The protocol should facilitate the process of culturing novel viruses from the gut as well as large scale studies on gut viromes.

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Interplay between the human gut microbiome and host metabolism

10.1101/561787 ◽

2019 ◽

Cited By ~ 1

Author(s):

Alessia Visconti ◽

Caroline I. Le Roy ◽

Fabio Rosa ◽

Niccolo Rossi ◽

Tiphaine C. Martin ◽

...

Keyword(s):

Gut Microbiome ◽

Metabolic Pathways ◽

Taxonomic Composition ◽

Metagenomic Sequencing ◽

Human Gut ◽

Metabolic Potential ◽

Microbial Ecosystem ◽

Microbiome Composition ◽

Shotgun Metagenomic Sequencing ◽

Key Species

AbstractThe human gut is inhabited by a complex and metabolically active microbial ecosystem regulating host health. While many studies have focused on the effect of individual microbial taxa, the metabolic potential of the entire gut microbial ecosystem has been largely under-explored. We characterised the gut microbiome of 1,004 twins via whole shotgun metagenomic sequencing (average 39M reads per sample). We observed greater similarity, across unrelated individuals, for functional metabolic pathways (82%) than for taxonomic composition (43%). We conducted a microbiota-wide association study linking both taxonomic information and microbial metabolic pathways with 673 blood and 713 faecal metabolites (Metabolon, Inc.). Metabolic pathways associated with 34% of blood and 95% of faecal metabolites, with over 18,000 significant associations, while species-level results identified less than 3,000 associations, suggesting that coordinated action of multiple taxa is required to affect the metabolome. Finally, we estimated that the microbiome mediated a crosstalk between 71% of faecal and 15% of blood metabolites, highlighting six key species (unclassified Subdoligranulum spp., Faecalibacterium prausnitzii, Roseburia inulinivorans, Methanobrevibacter smithii, Eubacterium rectale, and Akkermansia muciniphila). Because of the large inter-person variability in microbiome composition, our results underline the importance of studying gut microbial metabolic pathways rather than focusing purely on taxonomy to find therapeutic and diagnostic targets.

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Rapid inference of direct interactions in large-scale ecological networks from heterogeneous microbial sequencing data

10.1101/390195 ◽

2018 ◽

Cited By ~ 4

Author(s):

Janko Tackmann ◽

João Frederico Matias Rodrigues ◽

Christian von Mering

Keyword(s):

Graphical Models ◽

Large Scale ◽

Study Data ◽

Microbial Interactions ◽

Data Sets ◽

Metagenomic Sequencing ◽

Sequencing Data ◽

Human Gut ◽

Data Set ◽

Seamless Integration

AbstractThe recent explosion of metagenomic sequencing data opens the door towards the modeling of microbial ecosystems in unprecedented detail. In particular, co-occurrence based prediction of ecological interactions could strongly benefit from this development. However, current methods fall short on several fronts: univariate tools do not distinguish between direct and indirect interactions, resulting in excessive false positives, while approaches with better resolution are so far computationally highly limited. Furthermore, confounding variables typical for cross-study data sets are rarely addressed. We present FlashWeave, a new approach based on a flexible Probabilistic Graphical Models framework to infer highly resolved direct microbial interactions from massive heterogeneous microbial abundance data sets with seamless integration of metadata. On a variety of benchmarks, FlashWeave outperforms state-of-the-art methods by several orders of magnitude in terms of speed while generally providing increased accuracy. We apply FlashWeave to a cross-study data set of 69 818 publicly available human gut samples, resulting in one of the largest and most diverse models of microbial interactions in the human gut to date.

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Assessment of fecal DNA extraction protocols for metagenomic studies

GigaScience ◽

10.1093/gigascience/giaa071 ◽

2020 ◽

Vol 9 (7) ◽

Author(s):

Fangming Yang ◽

Jihua Sun ◽

Huainian Luo ◽

Huahui Ren ◽

Hongcheng Zhou ◽

...

Keyword(s):

Dna Extraction ◽

Cost Effective ◽

Metagenomic Sequencing ◽

Human Gut ◽

Shotgun Metagenomics ◽

Fecal Samples ◽

Bead Size ◽

Standardized Protocol ◽

Fungal Dna ◽

Fungal Dna Extraction

Abstract Background Shotgun metagenomic sequencing has improved our understanding of the human gut microbiota. Various DNA extraction methods have been compared to find protocols that robustly and most accurately reflect the original microbial community structures. However, these recommendations can be further refined by considering the time and cost demands in dealing with samples from very large human cohorts. Additionally, fungal DNA extraction performance has so far been little investigated. Results We compared 6 DNA extraction protocols, MagPure Fast Stool DNA KF Kit B, Macherey Nagel™ NucleoSpin™®Soil kit, Zymo Research Quick-DNA™ Fecal/Soil Microbe kit, MOBIO DNeasy PowerSoil kit, the manual non-commercial protocol MetaHIT, and the recently published protocol Q using 1 microbial mock community (MMC) (containing 8 bacterial and 2 fungal strains) and fecal samples. All samples were manually extracted and subjected to shotgun metagenomics sequencing. Extracting DNA revealed high reproducibility within all 6 protocols, but microbial extraction efficiencies varied. The MMC results demonstrated that bead size was a determining factor for fungal and bacterial DNA yields. In human fecal samples, the MagPure bacterial extraction performed as well as the standardized protocol Q but was faster and more cost-effective. Extraction using the PowerSoil protocol resulted in a significantly higher ratio of gram-negative to gram-positive bacteria than other protocols, which might contribute to reported gut microbial differences between healthy adults. Conclusions We emphasize the importance of bead size selection for bacterial and fungal DNA extraction. More importantly, the performance of the novel protocol MP matched that of the recommended standardized protocol Q but consumed less time, was more cost-effective, and is recommended for further large-scale human gut metagenomic studies.

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Effect of the Degree of Polymerization of Fructans on Ex Vivo Fermented Human Gut Microbiome

Nutrients ◽

10.3390/nu11061293 ◽

2019 ◽

Vol 11 (6) ◽

pp. 1293 ◽

Cited By ~ 5

Author(s):

Erola Astó ◽

Iago Méndez ◽

Maria Rodríguez-Prado ◽

Jordi Cuñé ◽

Jordi Espadaler ◽

...

Keyword(s):

Ex Vivo ◽

Alpha Diversity ◽

Degree Of Polymerization ◽

Metagenomic Sequencing ◽

Beneficial Bacteria ◽

Human Gut ◽

Gastrointestinal Health ◽

Healthy Donors ◽

Acidification Activity ◽

Other Regarding

Prebiotic supplements are used to promote gastrointestinal health by stimulating beneficial bacteria. The aim of this study was to compare the potential prebiotic effects of fructans with increasing degrees of polymerization, namely fructooligosaccharides (FOS) and inulins with a low and high polymerization degree (LPDI and HPDI, respectively), using an ex vivo fermentation system to simulate the colonic environment. The system was inoculated with pooled feces from three healthy donors with the same baseline enterotype. Changes in microbiota composition were measured by 16S metagenomic sequencing after 2, 7, and 14 days of fermentation, and acid production was measured throughout the experiment. Alpha-diversity decreased upon inoculation of the ex vivo fermentation under all treatments. Composition changed significantly across both treatments and time (ANOSIM p < 0.005 for both factors). HPDI and LPDI seemed to be similar to each other regarding composition and acidification activity, but different from the control and FOS. FOS differed from the control in terms of composition but not acidification. HDPI restored alpha-diversity on day 14 as compared to the control (Bonferroni p < 0.05). In conclusion, the prebiotic activity of fructans appears to depend on the degree of polymerization, with LPDI and especially HPDI having a greater effect than FOS.

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