- Publication Date:
- NSF-PAR ID:
- 10256499
- Journal Name:
- PeerJ
- Volume:
- 9
- Page Range or eLocation-ID:
- e11447
- ISSN:
- 2167-8359
- Sponsoring Org:
- National Science Foundation
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Gralnick, Jeffrey A. (Ed.)ABSTRACT Reconstructing microbial genomes from metagenomic short-read data can be challenging due to the unknown and uneven complexity of microbial communities. This complexity encompasses highly diverse populations, which often includes strain variants. Reconstructing high-quality genomes is a crucial part of the metagenomic workflow, as subsequent ecological and metabolic inferences depend on their accuracy, quality, and completeness. In contrast to microbial communities in other ecosystems, there has been no systematic assessment of genome-centric metagenomic workflows for drinking water microbiomes. In this study, we assessed the performance of a combination of assembly and binning strategies for time series drinking water metagenomes that were collected over 6 months. The goal of this study was to identify the combination of assembly and binning approaches that result in high-quality and -quantity metagenome-assembled genomes (MAGs), representing most of the sequenced metagenome. Our findings suggest that the metaSPAdes coassembly strategies had the best performance, as they resulted in larger and less fragmented assemblies, with at least 85% of the sequence data mapping to contigs greater than 1 kbp. Furthermore, a combination of metaSPAdes coassembly strategies and MetaBAT2 produced the highest number of medium-quality MAGs while capturing at least 70% of the metagenomes based on read recruitment. Utilizing different assembly/binningmore »
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Background Metagenomics has transformed our understanding of microbial diversity across ecosystems, with recent advances enabling
de novo assembly of genomes from metagenomes. These metagenome-assembled genomes are critical to provide ecological, evolutionary, and metabolic context for all the microbes and viruses yet to be cultivated. Metagenomes can now be generated from nanogram to subnanogram amounts of DNA. However, these libraries require several rounds of PCR amplification before sequencing, and recent data suggest these typically yield smaller and more fragmented assemblies than regular metagenomes.Methods Here we evaluate
de novo assembly methods of 169 PCR-amplified metagenomes, including 25 for which an unamplified counterpart is available, to optimize specific assembly approaches for PCR-amplified libraries. We first evaluated coverage bias by mapping reads from PCR-amplified metagenomes onto reference contigs obtained from unamplified metagenomes of the same samples. Then, we compared different assembly pipelines in terms of assembly size (number of bp in contigs ≥ 10 kb) and error rates to evaluate which are the best suited for PCR-amplified metagenomes.Results Read mapping analyses revealed that the depth of coverage within individual genomes is significantly more uneven in PCR-amplified datasets versus unamplified metagenomes, with regions of high depth of coverage enriched in short inserts. This enrichment scales with the number of PCRmore »
Conclusions PCR-amplified metagenomes have enabled scientists to explore communities traditionally challenging to describe, including some with extremely low biomass or from which DNA is particularly difficult to extract. Here we show that a modified assembly pipeline can lead to an improved
de novo genome assembly from PCR-amplified datasets, and enables a better genome recovery from low input metagenomes. -
Abstract Background With the advent of metagenomics, the importance of microorganisms and how their interactions are relevant to ecosystem resilience, sustainability, and human health has become evident. Cataloging and preserving biodiversity is paramount not only for the Earth’s natural systems but also for discovering solutions to challenges that we face as a growing civilization. Metagenomics pertains to the in silico study of all microorganisms within an ecological community in situ
, however, many software suites recover only prokaryotes and have limited to no support for viruses and eukaryotes.Results In this study, we introduce the
Viral Eukaryotic Bacterial Archaeal (VEBA) open-source software suite developed to recover genomes from all domains. To our knowledge,VEBA is the first end-to-end metagenomics suite that can directly recover, quality assess, and classify prokaryotic, eukaryotic, and viral genomes from metagenomes.VEBA implements a novel iterative binning procedure and hybrid sample-specific/multi-sample framework that yields more genomes than any existing methodology alone.VEBA includes a consensus microeukaryotic database containing proteins from existing databases to optimize microeukaryotic gene modeling and taxonomic classification.VEBA also provides a unique clustering-based dereplication strategy allowing for sample-specific genomes and genes to be directly compared across non-overlapping biological samples. Finally,VEBA is the only pipeline that automates the detection of candidate phyla radiation bacteria and implements the appropriate genomemore »Conclusions The
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