RT Journal Article
SR Electronic
T1 Quantification of bacterial DNA in blood using droplet digital PCR: a pilot study
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2022.12.02.518639
DO 10.1101/2022.12.02.518639
A1 Ana P. Tedim
A1 Irene Merino
A1 Alicia Ortega
A1 Marta Domínguez-Gil
A1 José Maria Eiros
A1 Jesús F. Bermejo-Martín
YR 2023
UL http://biorxiv.org/content/early/2023/01/10/2022.12.02.518639.abstract
AB 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 StatementThe authors have declared no competing interest.