Comparison of human DNA depletion and bacterial enrichment methods from large blood volumes to improve bloodstream infection diagnosis

Culture-independent methods have the potential to improve diagnosis of bloodstream infections by directly identifying microorganisms from whole blood without the need for culture. However, performance is limited by abundant human DNA, inhibitory components, and low concentrations of circulating microorganisms. Human DNA depletion and bacterial enrichment methods may overcome these challenges by selectively lysing human cells or by directly isolating bacteria, enabling the use of larger blood volumes. We compared three manual methods processing up to 10 mL of whole blood: Polaris and Ultra Deep Microbiome Prep10 deplete human DNA through chemical lysis, and SwiftX Sepsis enriches bacteria through paramagnetic-based extraction. Whole blood samples were spiked with Klebsiella pneumoniae and Staphylococcus aureus at concentrations similar to 1000, 100, 10 and 1 CFU/mL blood. Bacterial recovery and human DNA depletion were measured by quantitative real-time PCR. Resulting Cq values were converted to genome equivalents per mL blood using a defined formula. Polaris and Ultra Deep outperformed SwiftX for recovery of S. aureus, with Polaris generally yielding better results than Ultra Deep. For K. pneumoniae, Polaris recovered the highest bacterial amounts and significantly outperformed SwiftX at the three highest concentrations, while Ultra Deep showed no significant differences compared with the other methods. Ultra Deep achieved greatest depletion of human DNA. Polaris and SwiftX had similar processing times, whereas Ultra Deep took 1.5 & times; longer. In conclusion, Polaris demonstrated the highest overall bacterial recovery. These methods may improve culture-independent bloodstream infection diagnostics by enabling the processing of larger blood volumes, comparable to those used in routine blood cultures.