Abstract
Aim To use genus/species-specific genes droplet digital PCR (ddPCR) assays to detect/quantify bacterial DNA from Escherichia coli, Klebsiella pneumoniae, Staphylococcus aureus and Enterococcus spp in blood samples.
Methods and Results Bacterial DNA from clinical strains (4<n<12) was extracted, quantified and diluted (10-0.0001ng/μL) and ddPCR assays were performed in triplicate. These ddPCR assays showed low replication variability, low detection limit (1–0.1pg/μL) and high genus/species specificity. ddPCR assays were also used to quantify bacterial DNA obtained from spiked blood (1×104-1CFU/mL) of each bacterial genus/species. Comparison between ddPCR assays and bacterial culture was performed by Pearson correlation. There was an almost perfect correlation (r≥0.997, p≤0.001) between the number of CFU/mL from bacterial culture and the number of gene copies/mL detected by ddPCR. The time from sample preparation to results was determined to be 3.5-4h.
Conclusions The results demonstrated the quantification capacity and specificity of the ddPCR assays to detect/quantify four of the most important bloodstream infection (BSI) bacterial pathogens directly from blood.
Significance and Impact This pilot study results reinforce the potential of ddPCR for the diagnosis and/or severity stratification of BSI. Applied to patients’ blood samples it can improve diagnosis and diminish sample-to-results time, improving patient care.
Competing Interest Statement
The authors have declared no competing interest.
Footnotes
The article was updated to accommodate the format of the journal it has been submitted to.