期刊
MICROBIOME
卷 10, 期 1, 页码 -出版社
BMC
DOI: 10.1186/s40168-022-01415-8
关键词
Nanopore sequencing; Long-read metagenomics; Activated sludge microbiome; Reference-quality genome reconstruction; NanoPhase; Prophage
类别
资金
- Theme-based Research Scheme - University Grants Committee of Hong Kong, China
- [T21-705/20-N]
NanoPhase is an open-source tool that enables the reconstruction of reference-quality genomes from complex metagenomes using only Nanopore long reads. It improves the quality and contiguity of MAGs, allowing for a more comprehensive investigation of target microbiomes.
Background: The accurate and comprehensive analyses of genome-resolved metagenomics largely depend on the reconstruction of reference-quality (complete and high-quality) genomes from diverse microbiomes. Closing gaps in draft genomes have been approaching with the inclusion of Nanopore long reads; however, genome quality improvement requires extensive and time-consuming high-accuracy short-read polishing. Results: Here, we introduce NanoPhase, an open-source tool to reconstruct reference-quality genomes from complex metagenomes using only Nanopore long reads. Using Kit 9 and Q20+ chemistries, we first evaluated the feasibility of NanoPhase using a ZymoBIOMICS gut microbiome standard (including 21 strains), then sequenced the complex activated sludge microbiome and reconstructed 275 MAGs with median completeness of 90%. As a result, NanoPhase improved the MAG contiguity (median MAG N50: 735 Kb, 44-86X compared to conventional short-readbased methods) while maintaining high accuracy, allowing for a full and accurate investigation of target microbiomes. Additionally, leveraging these high-contiguity reference-quality genomes, we identified 165 prophages within 111 MAGs, with 5 as active prophages, indicating the prophage was a neglected source of genetic diversity within microbial populations and influencer in shaping microbial composition in the activated sludge microbiome. Conclusions: Our results demonstrated that NanoPhase enables reference-quality genome reconstruction from complex metagenomes directly using only Nanopore long reads. Furthermore, besides the 16S rRNA genes and biosynthetic gene clusters, the generated high-accuracy and high-contiguity MAGs improved the host identification of critical mobile genetic elements, e.g., prophage, serving as a genomic blueprint to investigate the microbial potential and ecology in the activated sludge ecosystem.
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