Pathogen identification in culture-negative cerebrospinal fluid specimens of patients with purulent meningitis using...

Antimicrobial therapy necessitates fast and accurate detection of pathogens in purulent meningitis (PM) diagnosis. Although considered a gold standard, the traditional bacterial detection and identification methods based on routine bacterial culture are time-consuming and show low sensitivity and specificity. In order to reduce PM-related deaths and improve the treatment efficiency, we employed trace DNA extraction technology and combined it with Next-Generation Sequencing technology for bacteria identification. We extracted DNA from 31 cerebrospinal fluid (CSF) samples from people with PM. The samples were collected by lumbar puncture at Xi'an Children Hospital under a stringent protocol. Ion PGM^TM System Next-Generation Sequencing was used to screen for pathogens in the CSF samples based on their 16S ribosomal RNA (rRNA) genes. Of the 31 CSF samples, 29 were culture negative. Streptococcus was detected in 2 samples by culture-based methods. The species identified by Next-Generation Sequencing showed a diverse bacterial population that included Streptococcus (22.6%), Escherichia (15.9%), Peptostreptococcus (10.7%), Pseudomonas (10.5%), Rothia (8.8%), Acinetobacter (4.9%), Prevotella (4.1%), Bacillus (3.3%), Neisseria (2.5%), Catonella (2.4%), Acitinomyces (2.0%), and Citrobacter (2.0%). Clustering analysis revealed three major bacterial groups, namely Streptococcus, Escherichia and Acinetobacter, each group with a different proportion of bacterial species. The results indicated that PM may result from infections by multiple bacterial species as opposed to a single species, with infectivity determined by abundance of the dominant species. Furthermore, Next-Generation Sequencing allowed a sensitive and specific etiological basis for identification of pathogens not only in the cerebrospinal fluid specimens, but also in other clinical samples.