Plasma and Serum Metagenomic Sequencing of Microbial Cell-Free Nucleic Acids for Suspected Central Nervous System Infection: A Systematic Review

Abstract

Background/Objectives: Untargeted metagenomic sequencing of microbial cell-free DNA or RNA in plasma or serum offers pathogen detection without lumbar puncture or biopsy, but its value in suspected central nervous system (CNS) infection is uncertain. We reviewed diagnostic yield, agreement with CNS-site testing, clinical impact, and failure modes. Methods: PubMed/MEDLINE was searched with seven documented strings (inception to 2 September 2026), supplemented by citation and web searching. Eligible reports were primary human studies of untargeted plasma or serum metagenomic sequencing with a separable result in patients with suspected or confirmed infectious CNS syndromes. Selection, extraction, and design-specific risk-of-bias appraisal followed PRISMA 2020 and were performed independently by two authors with disagreements resolved by discussion; the search was restricted to PubMed/MEDLINE and no protocol was registered. Synthesis was narrative because designs and reference standards were incompatible. Results: Of 190 reports assessed, 69 studies were included (18 denominator-based cohorts, 8 small series, 43 case reports; 51 from the United States). Positive CNS-specific clinical impact in commercial assay cohorts ranged from 0 of 24 adult to 5 of 24 pediatric meningoencephalitis tests. Among 51 case-level reports, the plasma or serum result was concordant with CNS-site testing in 28, positive when CNS-site testing was negative in 5, unconfirmed in 12, and negative or missed the pathogen in 6. All five plasma-only detections came from single case reports, which contributed to no yield or agreement denominator; excluding the four abstract-level case reports moved no proportion by more than 1.3 percentage points. RNA viruses, low-burden focal mould disease, and chromosomally integrated HHV-6 were recurrent sources of false-negative or misleading results. Conclusions: Plasma or serum metagenomic sequencing is a useful adjunct in selected difficult CNS infections, particularly disseminated, fungal, amoebic, or fastidious bacterial disease, but it does not replace CNS-site diagnostics.