ArticleNature microbiology2026
Benchmarking of shotgun sequencing depth reveals the potential and limitations of shallow metagenomics and strain-level analysis.
Article in Nature microbiology, 2026. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 5 papers.
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Who cites it
5 citing papers in PubMed.
- Observational study of the microbiome of perforated abomasal ulcers in unweaned beef calves in Canada.Veterinary research communications · 2026Article
- metaWEPP: leveraging biobank-scale intra-species phylogenies for near-haplotype resolution in metagenomic analysis.NAR genomics and bioinformatics · 2026Article
- Nanopore metagenomic sequencing links clinically relevant resistance determinants to pathogens.Microbial genomics · 2026Article
- ZipStrain Enables Rapid and Precise Strain-Resolved Metagenomics.bioRxiv : the preprint server for biology · 2026Article
- Decoding the power of the microbiome in human health.Frontiers in cellular and infection microbiology · 2026Review
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Authors and funding
8 authors.
Funding
Abstract
Shotgun metagenomics can provide both taxonomic and functional insights, but benchmarking is necessary to determine the sequencing depth appropriate for specific analyses. Here we used complex mixtures of DNA from cultured bacteria and analysed taxonomic composition, strain-level resolution and functional profiles at up to 11 sequencing depths (0.1-50.0 Gb). Reference-based analysis provided accurate strain-level taxonomy at 0.5-1.0 Gb. By contrast, de novo metagenome-assembled genome (MAG) reconstruction required deep sequencing (>10 Gb), and even MAGs deemed high quality by standard metrics were chimeric, with 54.5-81.8% accurately representing original strains, depending on the bioinformatic approach. Functionally, 2 Gb provided reliable insights at the pathway level for each of the mock communities tested, but sufficient proteome coverage was achieved only at or above 10 Gb. Library preparation and host DNA contamination were identified as confounders in shallow metagenomic analysis. This analysis highlights the potential and limitations of shallow metagenomics and provides guidance to accurately capture strain-level diversity using MAGs.
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