High-resolution Microbiome Analysis of Host-Rich Samples Using 2bRAD-M Without Host Depletion

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Abstract

Background Characterizing human microbiota in samples is crucial for understanding host-microbe interactions but remains challenged by high host DNA contamination. Current depletion strategies are limited by DNA loss and require immediate processing. In this paper, we introduce 2bRAD-M, a reduced metagenomic sequencing method that offers an efficient and integrated analysis of the host-microbe ecosystem, overcoming issues of high host contamination. Results Validated on mock samples with > 90% human DNA, 2bRAD-M achieved over 93% in AUPR and L2 similarity metrics. In real saliva samples, it captured diurnal and host-specific microbiome patterns which matched whole metagenome sequencing (WMS) with only 5–10% sequencing effort. In an early childhood caries (ECC) study, 2bRAD-M identified key bacterial indicators and distinguished ECC from healthy subjects with an AUC of 0.92, demonstrating its utility in high host DNA contexts. Conclusions 2bRAD-M represents a significant technical advancement in microbiome research, particularly in tackling high host DNA contamination. Its ability to provide high-resolution microbial profiles without the need for immediate host depletion processing offers a practical and efficient alternative to existing methods.

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