Recovery of Metagenome-Assembled Genomes from a Human Fecal Sample with Pacific Biosciences High-Fidelity Sequencing

Oñate, Florian Plaza; Roume, Hugo; Almeida, Mathieu

doi:10.1128/mra.00250-22

Cited by 6 publications

(4 citation statements)

References 16 publications

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“…Therefore, some of the results (e.g., performance of each assembler) and the cMAGfilter parameters may not be generalized. While this manuscript is being peer reviewed, a public release of nine cMAGs assembled by HiFi sequencing was announced by independent research group 27 . However, the study did not present their systematic evaluations on nucleotide accuracy, taxonomic diversity, and ability in large genome assembly, probably due to the insufficient number of retrieved cMAGs.…”

Section: Discussionmentioning

confidence: 99%

HiFi metagenomic sequencing enables assembly of accurate and complete genomes from human gut microbiota

Kim

Lee

2022

Nat Commun

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Advances in metagenomic assembly have led to the discovery of genomes belonging to uncultured microorganisms. Metagenome-assembled genomes (MAGs) often suffer from fragmentation and chimerism. Recently, 20 complete MAGs (cMAGs) have been assembled from Oxford Nanopore long-read sequencing of 13 human fecal samples, but with low nucleotide accuracy. Here, we report 102 cMAGs obtained by Pacific Biosciences (PacBio) high-accuracy long-read (HiFi) metagenomic sequencing of five human fecal samples, whose initial circular contigs were selected for complete prokaryotic genomes using our bioinformatics workflow. Nucleotide accuracy of the final cMAGs was as high as that of Illumina sequencing. The cMAGs could exceed 6 Mbp and included complete genomes of diverse taxa, including entirely uncultured RF39 and TANB77 orders. Moreover, cMAGs revealed that regions hard to assemble by short-read sequencing comprised mostly genomic islands and rRNAs. HiFi metagenomic sequencing will facilitate cataloging accurate and complete genomes from complex microbial communities, including uncultured species.

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Section: Discussionmentioning

confidence: 99%

HiFi metagenomic sequencing enables assembly of accurate and complete genomes from human gut microbiota

Kim

Lee

2022

Nat Commun

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“…OF03 18AA and Mediterraneibacter sp. 210,702 DFI 5 30, both belonging to the phylum Firmicutes, are relatively new and comes under the group of uncharacterized isolates of Clostridium ( 41 ) and Mediterraneibacter , respectively ( 42 ). Hence, their increase in abundance or its functional association post-supplementation of pea protein and enzymes-probiotics blend needs further evaluation.…”

Section: Discussionmentioning

confidence: 99%

Study of amino acids absorption and gut microbiome on consumption of pea protein blended with enzymes-probiotics supplement

Rathi,

Gaonkar,

Dhar

et al. 2024

Front. Nutr.

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The current randomized, double-blind, crossover clinical trial was conducted to evaluate changes in the amino acid absorption and gut microbiota on consumption of pea protein supplemented with an enzymes-probiotics blend (Pepzyme Pro). A total of 15 healthy subjects were instructed to take test (pea protein + Pepzyme Pro) or placebo (pea protein + maltodextrin) for 15 days with a 30-day washout period. Blood samples were analyzed for plasma-free amino acids, insulin, and C-reactive protein (CRP). Additionally, nitrogen levels in urine and feces, along with the composition of gut microbiota, were evaluated. On day 15, the test arm showed a tendency to increase the rate of absorption and total absorption (AUC) of amino acids compared with the placebo arm, though the increase was statistically insignificant. In addition, 15-day test supplementation showed a tendency to reduce Tmax of all the amino acids (statistically insignificant except alanine, p = 0.021 and glycine, p = 0.023) in comparison with the placebo supplementation. There were no changes in urine and fecal nitrogen levels as well as serum CRP levels in the test and placebo arm. The increase in serum insulin level after 4 h was statistically significant in both arms, whereas the insulin level of the placebo and test arm at 4 h was not statistically different. Supplementation showed changes with respect to Archaea and few uncharacterized species but did not show statistically significant variations in microbiome profile at the higher taxonomic levels. A study with large sample size and detailed gut microbiome analysis is warranted to confirm the results statistically as well as to characterize altered species. However, the current study could provide an inkling of a positive alteration in protein digestibility, amino acid absorption, and gut microbiome with regular consumption of protein and enzymes-probiotics blend.Clinical Trial Registration:clinicaltrials.gov/; identifier [CTRI/2021/10/037072].

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“…Presumably due to the large amount of DNA required to perform long-read sequencing, we did not find any viral long-read metagenomes in public databases at the time of this writing. Instead, we used 19 PacBio HiFi cellular metagenomes from human and chicken guts as well as seawater samples described previously (42)(43)(44)(45)(46). Sequences were quality filtered using filtlong v0.2.1 (https://github.com/rrwick/Filtlong) with a minimum read length of 10 kbp and a minimum window quality of 99.…”

Section: Methodsmentioning

confidence: 99%

A natural ANI gap that can define intra-species units of bacteriophages and other viruses

Aldeguer-Riquelme,

Conrad,

Antón

et al. 2024

Preprint

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Despite the importance of intra-species variants of viruses for causing disease and/or disrupting ecosystem functioning, there is no universally applicable standard to define these. A 95% whole-genome average nucleotide identity (ANI) gap is commonly used to define species, especially for bacteriophages, but whether a similar gap exists within species that can be used to define intra-species units has not been evaluated yet. Whole-genome comparisons among members of 1,016 bacteriophage species revealed a region of low frequency of pairs around 99.2-99.8% ANI, showing 3-fold or fewer pairs than expected for an even or normal distribution. This second gap is prevalent in viruses infecting various cultured or uncultured hosts, and from a variety of environments, although a few exceptions to this pattern were also observed (~3.7% of the total species evaluated) and are likely attributed to cultivation biases. Similar results were observed for a limited set of eukaryotic viruses that are adequately sampled including SARS-CoV-2, whose ANI-based clusters matched well the WHO-defined Variants of Concern, indicating that they represent functionally and/or ecologically distinct units. The existence of sequence-discrete units appears to be predominantly driven by (high) ecological cohesiveness coupled to either recombination frequency for bacteriophages or selection and clonal evolution for other viruses such as SARS-CoV-2. These results indicate that fundamentally different underlying mechanisms could lead to similar diversity patterns. Based on these results, we propose the 99.5% ANI as a practical, standardized, and data-supported threshold for defining viral intra-species units of bacteriophages, for which we propose the term genomovars. Importance: Viral species are composed of an ensemble of intra-species variants whose dynamic may have major implications for human and animal health and/or ecosystem functioning. However, the lack of universally-accepted standards to define these intra-species variants has led researchers to use different approaches for this task, creating inconsistent intra-species units across different viral families and confusion in communication. By comparing hundreds of viral bacteriophage genomes, we show that there is a nearly universal natural gap in whole-genome average nucleotide identities (ANI) among genomes at around 99.5%, which can be used to define intra-species units. Therefore, these results advance the molecular toolbox for tracking viral intra-species units and should facilitate future epidemiological and environmental studies.

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Recovery of Metagenome-Assembled Genomes from a Human Fecal Sample with Pacific Biosciences High-Fidelity Sequencing

Cited by 6 publications

References 16 publications

HiFi metagenomic sequencing enables assembly of accurate and complete genomes from human gut microbiota

HiFi metagenomic sequencing enables assembly of accurate and complete genomes from human gut microbiota

Study of amino acids absorption and gut microbiome on consumption of pea protein blended with enzymes-probiotics supplement

A natural ANI gap that can define intra-species units of bacteriophages and other viruses

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