Background The diagnosis of infective endocarditis (IE) can be difficult, particularly if blood cultures fail to yield a pathogen. This study evaluates the potential utility of microbial cell-free DNA (mcfDNA) as a tool to identify the microbial etiology of IE. Methods Blood samples from patients with suspected IE were serially collected. mcfDNA was extracted from plasma and underwent next-generation sequencing (NGS). Reads were aligned against a library containing DNA sequences belonging to >1400 different pathogens. mcfDNA from organisms present above a statistical threshold were reported and quantified in molecules/mL (MPM). Additional mcfDNA was collected on each subject every 2-3 days for a total of 7 collections or until discharge. Results Of 30 enrolled patients with suspected IE, 23 had Definite IE, 2 had Possible IE, and IE was Rejected in 5 patients by modified Duke Criteria. Only the 23 patients with Definite IE were included for analysis. Both mcfDNA and blood cultures achieved a sensitivity of 87%. The median duration of positivity from antibiotic treatment initiation was estimated to be approximately 38.1 days for mcfDNA vs 3.7 days for blood culture (proportional Odds 2.952, p= 0.02771), using a semi-parametric survival analysis. mcfDNA (log10) levels significantly declined (-0.3 MPM log10 units, 95% credible interval -0.45, -0.14) after surgical source control was performed (pre- vs post-procedure, posterior probability >0.99. Conclusion mcfDNA accurately identifies the microbial etiology of IE. Sequential mcfDNA levels may ultimately help to individualize therapy by estimating a patient’s burden of infection and response to treatment.

Microbial cell-free DNA (mcfDNA) sequencing is an emerging infectious disease diagnostic tool which enables unbiased pathogen detection and quantification from plasma. The Karius Test, a commercial mcfDNA sequencing assay developed by and available since 2017 from Karius, Inc. (Redwood City, CA), detects and quantifies mcfDNA as molecules/μL in plasma.

Microbial cell-free DNA (mcfDNA) sequencing is an emerging infectious disease diagnostic tool which enables unbiased pathogen detection from plasma. The Karius Test®, a commercial mcfDNA sequencing assay developed by and available since 2017 from Karius, Inc. (Redwood City, CA), detects and quantifies mcfDNA as molecules/μl in plasma. The commercial sample data and results for all tests conducted from April 2018 through mid-September 2021 were evaluated for laboratory performance metrics, reported pathogens, and data from test requisition forms. A total of 18,690 reports were generated from 15,165 patients in a hospital setting among 39 states and the District of Columbia. The median time from sample receipt to reported result was 26 hours (IQR 25–28), and 96% of samples had valid test results. Almost two-thirds (65%) of patients were adults, and 29% at the time of diagnostic testing had ICD10 codes representing a diverse array of clinical scenarios. There were 10,752 (58%) reports that yielded at least one taxon for a total of 22,792 detections spanning 701 unique microbial taxa. The 50 most common taxa detected included 36 bacteria, 9 viruses, and 5 fungi. Opportunistic fungi (374Aspergillusspp., 258Pneumocystis jirovecii, 196Mucorales, and 33 dematiaceous fungi) comprised 861 (4%) of all detections. Additional diagnostically challenging pathogens (247 zoonotic and vector borne pathogens, 144Mycobacteria, 80Legionellaspp., 78 systemic dimorphic fungi, 69Nocardiaspp., and 57 protozoan parasites) comprised 675 (3%) of all detections. We report the largest cohort of patients tested using plasma mcfDNA sequencing. The wide variety of pathogens detected by plasma mcfDNA sequencing reaffirm our understanding of the ubiquity of some infections while also identifying taxa less commonly detected by conventional methods.