Culture and Isolation
1. Principle
Grow viable microorganisms on/in nutrient media, then separate them into pure colonies so they can be identified and tested for drug susceptibility.
2. Step-by-Step Procedure
- Collect appropriate specimen before antibiotics when possible; note site and clinical question.
- Inoculate nonselective and selective/differential media (e.g., blood agar, MacConkey, CNA).
- Incubate with correct atmosphere/temperature/time (O₂, CO₂, anaerobic jars; prolonged for slow growers).
- Inspect colonies; Gram stain representative morphologies.
- Subculture to purity → identify (biochemicals, MALDI-TOF, molecular) → Antimicrobial Susceptibility Testing as indicated.
3. Interpretation
- Positive: Growth of a plausible pathogen in a significant quantity/context.
- Negative: No growth — may still miss anaerobes, fastidious, or pretreated organisms.
- Pitfalls: Contamination with Normal Microbiota; overgrowth; wrong atmosphere; interpreting colonizers as pathogens.
4. Clinical Use Cases
- Gold standard for: Many bacterial infections when organism is cultivable; enables AST.
- Alternative to: Empiric NAAT/PCR when speed matters or organism is unculturable — see PCR.
5. Comparison with Other Methods
| Method | Pros | Cons |
|---|---|---|
| Culture | Viable organism + AST + epidemiology | Slow; misses non-culturables |
| Gram Stain | Fast morphology | No species/AST |
| PCR | Fast, sensitive | Colonization vs infection; no AST |
6. Mnemonic / Visual Aid
SIT: Specimen quality → Isolate pure → Test (ID + AST)