Whole-Genome Sequencing
1. Principle
Determine nearly the entire DNA sequence of an isolate (or metagenome) to identify species, resistance/virulence genes, plasmids, and relatedness for outbreak investigation.
2. Step-by-Step Procedure (clinical overview)
- Start from quality specimen/isolate (Sample Types and Specimen Quality).
- Extract DNA (DNA Extraction; see also Plasmid DNA Extraction for special plasmid work).
- Quantify (Nucleic Acid Quantification) → NGS Library Preparation (± Targeted Enrichment for low-load targets).
- Sequence (short and/or long read — Sequencing Technologies).
- Bioinformatic assembly/mapping → species ID, MLST/cgMLST, resistome, virulome, plasmid typing (WGS Bioinformatics Pipeline · Clinical WGS Pipelines).
- Interpret in clinical/epidemiologic context; correlate AMR genes with Antimicrobial Susceptibility Testing.
Culture-independent path: Metagenomic NGS from primary specimens.
3. Interpretation
- Positive utility: High-resolution relatedness; comprehensive gene catalog.
- Limits: Gene presence ≠ phenotypic resistance always; assembly gaps; contamination; turnaround and expertise needs.
- Pitfalls: Over-calling resistance without AST correlation; misassigning plasmid vs chromosome without long reads.
4. Clinical Use Cases
- Gold standard for: Outbreak genomic epidemiology; reference characterization of unusual pathogens.
- Alternative to: PFGE/older typing; complements Antimicrobial Susceptibility Testing and PCR.
5. Comparison with Other Methods
| Method | Pros | Cons |
|---|---|---|
| WGS | Maximal genomic info | Cost, time, bioinformatics |
| PCR | Fast targeted answers | Narrow scope |
| AST | Phenotype for therapy | No transmission genomics |
6. Mnemonic / Visual Aid
Sequence → Species → Resistome → Relatedness — four questions WGS can answer.
Related Concepts
- Horizontal Gene Transfer · Plasmid · Carl Woese · Kary Mullis
- Pipelines: WGS Bioinformatics Pipeline · Genome Assembly · AMR Gene Databases · Phylogenomics and Outbreak Typing
- AI: Machine Learning for AMR Prediction · MOC - AI in Microbiology
- Papers: Paper - AMR Database M.Centner 2026
- MOCs: MOC - Diagnostic & Lab Methods · MOC - Bioinformatics in Microbiology · MOC - Antimicrobial Resistance (AMR)
- Figure: Figure - WGS Bioinformatics Pipeline