Yes — for CSF it is best to centrifuge the whole sample (5,000–10,000 × g, 5–10 min, 4 °C) and use the pellet for DNA extraction. This concentrates bacteria and significantly improves PCR sensitivity in low-load samples. The more CSF volume you start with, the better.

CSF (Neisseria, Strep, H. influenzae): Spin, keep 50–200 µL with pellet, add lysozyme if S. pneumoniae, then proceed with DNeasy/QIAamp lysis (Proteinase K + Buffer AL, 56 °C, 15–30 min), wash, and elute in 30–50 µL.

Whole blood (EDTA): Use 200 µL directly with kit protocol, or enrich buffy coat from 1–3 mL blood if bacterial load is low. Avoid heparin.

177 / 5.000

It is not necessary, if it is isolated in culture media the concentration will be higher and you will obtain a good pool for DNA for sequencing, but if it is only for diagnosis, PCR will be useful.

Centrifugation of CSF concentrates bacterial cells into a pellet, increasing the DNA yield and improving the sensitivity of PCR detection. This is especially beneficial when bacterial concentrations are low, as it reduces the inhibitory effects of host DNA and other substances present in the CSF. Studies have demonstrated that DNA extraction from CSF pellets can achieve detection limits as low as 1–2 × 10² CFU/mL for pathogens like Neisseria meningitidis and Streptococcus pneumoniae .

Effect of Sample Volume on Extraction Method

The volume of CSF collected can influence the DNA extraction process. Larger volumes may require longer centrifugation times or the use of larger tubes to ensure efficient pelleting of bacterial cells. It's essential to adjust centrifugation conditions accordingly to maximize bacterial recovery.

🧪 Optimized DNA Extraction Protocols

For CSF Samples

For Whole Blood Samples (EDTA Tubes)

These protocols are compatible with the QIAamp DNA Mini Kit and the DNeasy Blood & Tissue Kit. Ensure to follow the manufacturer's instructions for each kit to achieve optimal results.

Additional Considerations

• Enzymatic Pretreatment: Incorporating enzymatic treatments, such as lysozyme or mutanolysin, can enhance bacterial cell wall lysis, potentially increasing DNA yield .
• Inhibition Removal: If PCR inhibitors are suspected, consider using a DNA clean-up step or inhibitor-removal columns to purify the extracted DNA before amplification.
• Automation: For high-throughput processing, consider using automated systems like the QIAcube Connect to standardize and streamline the extraction process.

By following these optimized protocols and considerations, you can enhance the sensitivity and reliability of PCR-based diagnostics for bacterial meningitis pathogens.