Enrichment techniques for clinical metagenomics.

Area of Science:

• Clinical diagnostics
• Microbiology
• Genomics

Background:

• Metagenomic next-generation sequencing (mNGS) enables hypothesis-free pathogen identification in clinical samples.
• High host nucleic acid content in samples challenges mNGS sensitivity and increases costs.
• Pathogen-specific enrichment techniques improve mNGS performance but reduce its hypothesis-free nature.

Purpose of the Study:

• To comprehensively review and analyze key mNGS enrichment techniques for clinical applications.
• To evaluate methods based on methodology, cost, sensitivity, specificity, and clinical integration.
• To discuss current limitations and future directions for mNGS enrichment strategies.

Main Methods:

• Review of PCR-based enrichment, CRISPR-Cas9 enrichment, molecular inversion probes (MIP), nanopore adaptive sequencing (AS), and hybridization capture.
• Comparative analysis of enrichment methods across various performance metrics.
• Assessment of applications in pathogen detection, antimicrobial resistance profiling, and whole-genome sequencing.

Main Results:

• Enrichment techniques significantly improve sensitivity and specificity of mNGS for pathogen detection.
• Methods vary in cost, complexity, and suitability for different clinical workflows.
• CRISPR-Cas9 and nanopore adaptive sequencing show promise for targeted mNGS applications.

Conclusions:

• Enrichment strategies are crucial for optimizing mNGS in clinical diagnostics, balancing breadth with sensitivity.
• Further refinement and standardization of these techniques are needed for broader clinical implementation.
• Future research should focus on cost-effective, highly sensitive, and hypothesis-free enrichment methods.