Biases and complementarity in gut viromes obtained from bulk and virus-like particle-enriched metagenomic sequencing
ABSTRACT Due to varying sequencing strategies, current gut virome findings show significant variability. Specifically, bulk- and virus-like particle (VLP)-enriched metagenomic sequencing (termed bulk and VLP, respectively) present unique advantages and limitations, affecting viral genome discovery,...
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| Main Authors: | , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
American Society for Microbiology
2025-08-01
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| Series: | Microbiology Spectrum |
| Subjects: | |
| Online Access: | https://journals.asm.org/doi/10.1128/spectrum.00013-25 |
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| Summary: | ABSTRACT Due to varying sequencing strategies, current gut virome findings show significant variability. Specifically, bulk- and virus-like particle (VLP)-enriched metagenomic sequencing (termed bulk and VLP, respectively) present unique advantages and limitations, affecting viral genome discovery, taxonomic annotation, and community structure analysis. A comprehensive comparison of these strategies is crucial for thoroughly understanding the gut virome. This study comprehensively compared gut viromes identified from paired bulk and VLP data from 151 adult and 141 infant fecal samples. The VLP method showed superior performance to bulk in viral genome discovery in both data sets by recovering longer and more complete viral genomes, with higher sensitivity for low-abundant ones, resulting in a higher taxonomic annotation rate. However, we observed no correlations in the viral community structure (i.e., Shannon diversities) between bulk- and VLP-derived viromes, implying biases introduced during VLP enrichment. Such biases could be caused by the bacterial host features, such as the structural differences in cell walls and the prevalence and abundance of the viruses. Viruses that are of low prevalence, low abundance, or have Gram-positive bacteria as their hosts were enriched in VLP-derived viromes, in both the adult and infant data sets. Significant complementarity was observed between bulk and VLP viromes, with only about a quarter (26.7% in infants; 29.3% in adults) of VLP-viral genomes overlapping with bulk viruses. Together, our study identifies causal factors underlying the biases of bulk and VLP strategies in human gut virome studies and advocates the use of both strategies to enhance a comprehensive understanding of gut viromes.IMPORTANCEThe two mainstream gut phageome profiling strategies, namely bulk and virus-like particle (VLP), generated significantly overlapped results and have their own merits and drawbacks. Particularly, VLP exhibits higher efficiency in obtaining more, longer, and more complete viral genomes. However, VLP sequencing has the potential to alter the natural structure of viral communities, often resulting in the identification of viruses with lower prevalence and those specifically associated with Gram-positive bacterial hosts. While bulk metagenome features a more stable and diverse community, which can well reveal the interactions between viruses and bacteria. Nevertheless, bulk sequencing can suffer from lower coverage, leading to fragmented sequences and potentially missing some viral species. Therefore, it is essential to recognize that these methods are complementary rather than competitive in the comprehensive characterization of the gut phageome. |
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| ISSN: | 2165-0497 |