Metagenomic analysis of microbial cell-free DNA from plasma of patients with suspected infections: performance and therapeutic impact in clinical routine

Abstract

Objectives: The sensitivity of blood cultures (BCs) is limited, especially when antimicrobial therapy already has been administered or when non-culturable pathogens are causing the disease. Metagenomic next-generation sequencing of cell-free DNA from plasma has the potential to compensate for the disadvantages of BC diagnostics.

Methods: We conducted a retrospective study in patients with suspected infections over a period of 3 months. Cell-free DNA from plasma was analysed by metagenomic next-generation sequencing (Illumina NextSeq, 25 million reads per sample, read length 75 base pairs) and sequences were analysed with DISQVER®, a CE-IVDD-labelled software algorithm and curated database. The data were compared with findings obtained with simultaneously taken BC and other microbiological results (±7 days).

Results: DISQVER® analysis was performed on 190 samples from 147 patients (124 adult and 23 paediatric). The median time-to-result including transport was 2 days (interquartile range, 2-3; range, 2-8). DISQVER® detected 158 pathogens (103 bacteria, 49 viruses, 4 fungi, and 1 parasite) in 80 plasma samples (positivity rate 42.1%). The median number of pathogens per positive sample was one (interquartile range, 1-2; range, 1-10). The most common bacteria were Enterobacterales (30.1%; 31/103), anaerobic bacteria (18.4%; 19/103), and Enterococcus spp. (15.5%; 16/103); the most frequent viruses were Epstein-Barr virus (28.6%; 14/49), human herpesvirus 6B (18.4%; 9/49), and human cytomegalovirus (18.4%; 9/49). Mycobacterium avium, Legionella pneumophila, Tropheryma whipplei, Rhizomucor pusillus, and Leishmania infantum were detected in one sample each. Simultaneous BC were positive in only 10.2% (18/176) of the samples, but were mostly (68.2%; 120/176) collected under antibiotic therapy. DISQVER® analysis resulted in 24 treatment changes in 20 patients (13.6%; 20/147; 9 start/escalation, 10 stop/de-escalation, 2 catheter replacements, and 3 other).

Discussion: DISQVER® significantly increased the detection rate of pathogens, led to the diagnosis of serious infections that otherwise would have been missed, and possibly improved the treatment of more than 10% of patients.

Keywords: Bacteria; Blood; Bloodstream infection; Culture; DISQVER; Karius; Molecular diagnostic; NGS; Noscendo; Sepsis; cfDNA.