Retrospective analysis of clinical data associated with patients enrolled in a molecular diagnostic feasibility study highlights the potential utility for rapid detection of bloodstream infection.

BACKGROUND: Measurement of pathogen DNA polymerase activity by enzymatic template generation and amplification (ETGA) has shown promise in detecting pathogens in bloodstream infection (BSI). We perform an in-depth analysis of patients with clinical BSI enrolled in ETGA feasibility experiments. METHODS: In addition to hospital blood cultures, 1 study aerobic culture bottle was drawn from patients with suspected BSI. The study bottle was split into 2 bottles and was additionally subjected to ETGA analysis. Enzymatic template generation and amplification sensitivity/specificity for BSI detection was determined against the Centers for Disease Control BSI definition. When split cultures were both positive, time course analysis was performed to determine time to detection. The records of patients with BSI were reviewed for presence of systemic inflammatory response syndrome, antibiotic timing and appropriateness, and organism identification. RESULTS: Of 307 enrollees, 38 met the Centers for Disease Control BSI definition. Seventy-four percent met systemic inflammatory response syndrome criteria on admission. Antibiotic coverage was adequate in 76% of patients. Antibiotics were more often delayed in afebrile patients (odds ratio, 5). Twenty-seven of the split study culture bottles were positive in at least 1 sample, and ETGA detected microbes within all samples (sensitivity/specificity, 70.3%/99.3%). Of these, 22 were culture positive in both split study bottles and underwent ETGA time course analysis. Enzymatic template generation and amplification detected microbes within these 3-fold faster than culture. CONCLUSIONS: Patients with BSI often have diagnostic and treatment delays. Enzymatic template generation and amplification provides clinically meaningful data more rapidly than cultures. Future development should focus on real-time application of assays that detect microbes at the molecular level.

Measurement of microbial DNA polymerase activity enables detection and growth monitoring of microbes from clinical blood cultures.

Surveillance of bloodstream infections (BSI) is a high priority within the hospital setting. Broth-based blood cultures are the current gold standard for detecting BSI, however they can require lengthy incubation periods prior to detection of positive samples. We set out to demonstrate the feasibility of using enzymatic template generation and amplification (ETGA)-mediated measurement of DNA polymerase activity to detect microbes from clinical blood cultures. In addition to routine-collected hospital blood cultures, one parallel aerobic blood culture was collected and immediately refrigerated until being transported for ETGA analysis. After refrigeration holding and transport, parallel-collected cultures were placed into a BACTEC incubator and ETGA time-course analysis was performed. Of the 308 clinical blood cultures received, 22 were BACTEC positive, and thus were initially selected for ETGA time course analysis. The ETGA assay detected microbial growth in all 22 parallel-positive blood cultures in less time than a BACTEC incubator and also yielded genomic DNA for qPCR-based organism identification. In summary, feasibility of detecting microbes from clinical blood culture samples using the ETGA blood culture assay was demonstrated. Additional studies are being considered towards development of clinically beneficial versions of this methodology.