Polymerase chain reaction targeting 16S ribosomal RNA for the diagnosis of bacterial meningitis after neurosurgery

Abstract

OBJECTIVES: Bacterial and aseptic meningitis after neurosurgery can present similar clinical signs and symptoms. The aims of this study were to develop and test a molecular method to diagnose bacterial meningitis (BM) after neurosurgery. METHODS: A 16S ribosomal RNA gene PCR-based strategy was developed using artificially inoculated cerebrospinal fluid (CSF) followed by sequencing. The method was tested using CSF samples from 43 patients who had undergone neurosurgery and were suspected to suffer from meningitis, and from 8 patients without neurosurgery or meningitis. Patients were classified into five groups, confirmed BM, probable BM, possible BM, unlikely BM, and no meningitis. RESULTS: Among the samples from the 51 patients, 21 samples (41%) were culture-negative and PCR-positive. Of these, 3 (14%) were probable BM, 4 (19%) were possible BM, 13 (62%) were unlikely BM, and 1 (5%) was meningitis negative. Enterobacterales, non-fermenters ( Pseudomonas aeruginosa and Acinetobacter baumannii ), Staphylococcus haemolyticus , Granulicatella , Variovorax , and Enterococcus cecorum could be identified. In the group of patients with meningitis, a good agreement (3 of 4) was observed with the results of cultures, including the identification of species. CONCLUSION: Molecular methods may complement the diagnosis, guide treatment, and identify non-cultivable microorganisms. We suggest the association of methods for suspected cases of BM after neurosurgery, especially for instances in which the culture is negative.

Etiology and Outcomes of Healthcare-Associated Meningitis and Ventriculitis—A Single Center Cohort Study

“…Our study demonstrated the potential of 16S rRNA gene PCR/sequencing (30% of tested, culture negative CSF samples were positive) that can identify the presence of bacteria even in patients who have already received antibiotics [ 14 , 15 , 16 ]. This method can also provide more rapid identification, especially of slow-growing and fastidious bacteria [ 17 , 18 ]. Multiple studies on broad range PCR methods in HCAMV report high specificity and often limited sensitivity [ 19 , 20 , 21 ].…”

A Refined, Controlled 16S rRNA Gene Sequencing Approach Reveals Limited Detection of Cerebrospinal Fluid Microbiota in Children with Bacterial Meningitis

“…In addition, culture is frequently unable to identify pathogenic organisms in patients with meningitis, despite the clear purulence of the resulting samples ( 12 ). Therefore, there is great interest in using culture-independent, DNA-based methods for pathogen identification in CSF samples ( 12 , 13 ). In this study, we found that the nearly consistent low bacterial abundances relative to the contamination levels of reagents greatly diminish the utility of molecular approaches, even with the inclusion of rigorous controls and computational removal of contaminant sequencing reads, regardless of how the samples are processed or extracted.…”

Comparison of Suspected and Confirmed Internal External Ventricular Drain-Related Infections: A Prospective Multicenter United Kingdom Observational Study

“…Some possible candidates include nucleic acid amplification assays, CSF cytokines (interleukin-6), CSF lactate, or CSF procalcitonin, which have had variable results in the literature so far [ 26–30 ]. 16S rRNA has had a number of studies evaluating its diagnostic value, and it may be of use for culture-negative suspected iERIs [ 31 , 32 ]. Future research could also include work on developing and validating scoring systems, diagnostic trees [ 11 ], or predictive algorithms, which have previously been found to have high accuracy in differentiating between bacterial and viral meningitides [ 33 ].…”