To assess application of NGS to identify etiological agents in stored cerebrospinal fluid samples of presumed or definitive neurological infectious disease cases. 

Metagenomic next-generation sequencing (NGS) has led to development of high sensitivity methods such as virus capture based sequencing for all vertebrate viruses  (VirCapSeq-VERT) and bacterial capture sequencing (BacCapSeq). Both methods have shown promise in detecting a broad range of pathogens, though further understanding of utility in clinical settings is needed. 

Forty samples were selected for the study from following categories: (1) Patients with definitive CNS infection by gold standard testing; (2) Patients without definitive microbiological diagnosis who met clinal criteria by Brighton Criteria (BC) for meningoencephalitis (3) Patients who neither received a definitive microbiological diagnosis nor met BC but had infectious etiology high on differential by primary clinical team. CSF samples were de-identified and processed for further sequencing analysis. RNA extracts from CSF were used for VirCapSeq-VERT and DNA extracts were used for BacCapSeq analysis.

Among 9 samples from definitive CNS infections, NGS confirmed 3 diagnoses (33.33%), was negative in 5 (55.6%), and yielded alternative result in 1 (11.1%). The confirmed cases identified HHV-6, HSV-2, and VZV while the negative samples included Cryptococcus, PML, HSV-2, Powassan, and Lyme. In the 1 sample with alternate results, VZV was the definitive infection, however NGS detected Torque Teno Virus. In groups 2 and 3, 12/31 samples (38.7%) were positive for at least one pathogen, with 10 returning bacterial etiologies (32.26%) and 3 viral (9.68%).  The negative controls did not have any significant reads for any pathogens in both VirCapSeq-VERT and BacCapSeq analysis.

NGS  shows promise for pathogen identification in presumed CNS infectious diseases, however larger-scale prospective studies should be conducted to assess clinical value of this novel technique. Clinicians should further understand benefits and limitations of using this modality at bedside.