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Background <p>Bloodstream infections (BSI) are a significant cause of morbidity and mortality, but their diagnosis is limited by slow and often insensitive culture-based methods. While molecular diagnostics offer a faster and more accurate alternative, their performance requires efficient removal of human host DNA.</p> Methods <p>In this study, blood from healthy volunteers was spiked with either gram-negative or gram-positive bacteria. Three methods for bacterial separation—centrifugation, chemical lysis (Polaris), and enzymatic digestion (MolYsis) along with a standard DNA isolation method were evaluated. The performance of each separation method was assessed using RT-qPCR targeting the bacterial 16S rRNA gene and the human beta-2 microglobulin gene to assess bacterial recovery and host depletion, respectively.</p> Results <p>The centrifugation method achieved the lowest Ct values for bacterial detection, showing thus the highest bacterial recovery. It also achieved significantly more efficient depletion of host DNA compared to chemical and enzymatic methods. Additionally, centrifugation demonstrated the highest reproducibility, based on variability analysis. Overall, centrifugation method outperformed other tested techniques in terms of turnaround time, effectiveness, and reliability.</p> Conclusion <p>Centrifugation represents a fast, robust, and cost-effective method for bacterial separation and host DNA depletion from the whole blood. These characteristics make it particularly suitable for molecular diagnostic approaches for BSI.</p>

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Comparison of rapid bacterial separation techniques from blood

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年8月1日

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  • Róbert Lipták,
  • Michal Tamáš,
  • Tomáš Strečanský,
  • Barbora Tamášová,
  • Ľubomíra Tóthová,
  • Július Hodosy

摘要

Background

Bloodstream infections (BSI) are a significant cause of morbidity and mortality, but their diagnosis is limited by slow and often insensitive culture-based methods. While molecular diagnostics offer a faster and more accurate alternative, their performance requires efficient removal of human host DNA.

Methods

In this study, blood from healthy volunteers was spiked with either gram-negative or gram-positive bacteria. Three methods for bacterial separation—centrifugation, chemical lysis (Polaris), and enzymatic digestion (MolYsis) along with a standard DNA isolation method were evaluated. The performance of each separation method was assessed using RT-qPCR targeting the bacterial 16S rRNA gene and the human beta-2 microglobulin gene to assess bacterial recovery and host depletion, respectively.

Results

The centrifugation method achieved the lowest Ct values for bacterial detection, showing thus the highest bacterial recovery. It also achieved significantly more efficient depletion of host DNA compared to chemical and enzymatic methods. Additionally, centrifugation demonstrated the highest reproducibility, based on variability analysis. Overall, centrifugation method outperformed other tested techniques in terms of turnaround time, effectiveness, and reliability.

Conclusion

Centrifugation represents a fast, robust, and cost-effective method for bacterial separation and host DNA depletion from the whole blood. These characteristics make it particularly suitable for molecular diagnostic approaches for BSI.