Bacterial culture and drug susceptibility testing are used to identify pathogen infections. Nevertheless, the process requires several days from collection to the identification of bacterial species and drug-resistance patterns. The digital PCR system is a rapidly developing quantitative detection technology widely applied to molecular diagnosis, including copy number variations, single nucleotide variant analysis, cancer biomarker discovery, and pathogen identification. This study aimed to use a droplet digital PCR system to identify bacteria in blood samples and explore its ability to identify pathogen in bacteremia. Then, we designed primers and probes of SWG-9 and COA gene for E. coli and S. aureus to identify in blood samples with the ddPCR system.
FIG: The dynamic range of the ddPCR-based assay. Detecting the bacterial nucleic acid by ddPCR system, (A) The 1D droplet spots of FAM fluorescence amplitude for eight detection sites for E. coli ATCC25922 (from A06 to H06: the estimated copy numbers of nucleic acid template were 105, 104, 103, 102, 25, 6.25, 1.56, and NC) were performed for each detection site. Linear fitting lines are shown on the right (B) The 1D droplet spots of VIC fluorescence amplitude for four detection sites for Staphylococcus aureus ATCC25923 (from A01 to H01: the estimated copy numbers of nucleic acid template were 104, 103, 102, and NC) were performed for each detection site. Linear fitting lines are shown on the right.
The system had demonstrated extremely high detection accuracy in blood samples, and the detection rate of E. coli was 13.1–21.4%, and that of S. aureus was 50–88.3%. Finally, blood samples containing both E. coli and S. aureus were tested to evaluate further the accuracy and applicability of this method, indicating the detection rates range from 18.1% to 97%. The ddPCR system is highly promising as a qualitatively and quantitatively screening method for rapidly detecting pathogen.
Zhang, T., Niu, Z., Wu, F. et al. Qualitative and quantitative detection of surgical pathogenic microorganisms Escherichia coli andStaphylococcus aureus based on ddPCR system. Sci Rep 11, 8771 (2021). https://doi.org/10.1038/s41598-021-87824-5