Host DNA Depletion for Improved Pathogen Detection in Clinical Metagenomic Sequencing: Current Strategies and Future Directions

Clinical metagenomic sequencing has transformed infectious disease diagnostics by enabling comprehensive profiling of microbial communities directly from patient samples. However, a major challenge limiting its sensitivity and efficiency is the overwhelming abundance of host DNA, which can dominate sequencing reads and obscure low-abundance pathogens. This review synthesizes evidence from systematic reviews, meta-analyses, and primary studies to examine the strategies developed to deplete host DNA in clinical specimens. Pre-extraction methods, including physical separation, selective lysis, and enzymatic degradation, aim to remove host cells or extracellular DNA before sequencing. Post-extraction approaches, such as methylation-based enrichment and restriction enzyme treatment, further refine microbial DNA recovery. Emerging technologies, including CRISPR/Cas-mediated depletion and real-time selective nanopore sequencing, show promise in reducing host interference while preserving microbial diversity. Despite these advancements, limitations remain, including potential loss of sensitive microbial taxa, sample loss in low-biomass specimens, and overlapping methylation patterns between host and microbes. Integrating multiple complementary approaches tailored to sample type and clinical context appears most effective for maximizing pathogen detection. Optimizing host DNA depletion not only improves sequencing efficiency but also reduces costs, accelerates turnaround times, and enhances the accuracy of microbial diagnostics. Future research should focus on refining these strategies, addressing technical biases, and validating methods across diverse clinical samples. Overall, host DNA depletion represents a critical step in advancing the practical utility of metagenomic sequencing for patient care.