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The etiology of central nervous system (CNS) infections such as meningitis and encephalitis remains unknown in a large proportion of cases partly because the diversity of pathogens that may cause CNS infections greatly outnumber available test methods. We developed a metagenomic next generation sequencing (mNGS)-based approach for broad-range detection of pathogens associated with CNS infections suitable for application in the acute care hospital setting. The analytical sensitivity of mNGS performed on an Illumina MiSeq was assessed using simulated cerebrospinal fluid (CSF) specimens (n = 9). mNGS data were then used as a training dataset to optimize a bioinformatics workflow based on the IDseq pipeline. For clinical validation, residual CSF specimens (n = 74) from patients with suspected CNS infections previously tested by culture and/or PCR, were analyzed by mNGS. In simulated specimens, the NGS reads aligned to pathogen genomes in IDseq were correlated to qPCR C T values for the respective pathogens (R = 0.96; p < 0.0001), and the results were highly specific for the spiked pathogens. In clinical samples, the diagnostic accuracy, sensitivity and specificity of the mNGS with reference to conventional methods were 100%, 95% and 96%, respectively. The clinical application of mNGS holds promise to benefit patients with CNS infections of unknown etiology.

Background. Neisseria meningitidis continues to be an important cause of invasive bacterial disease among children and young adults worldwide. In Canada, N. meningitidis strains that bear serogroups B and C polysaccharide capsules predominate. We report the first documented case of invasive meningococcal disease in an immunocompetent host caused by an acapsular strain of N. meningitidis containing the capsule null locus (cnl). Methods. Analysis of the isolate was performed with use of serological and molecular methods, including multilocus sequence typing and cnl gene identification. Analysis of 16S ribosomal RNA (rRNA) and porA genes was also performed to confirm the identity of the bacterium. Results. The patient was a healthy, immunocompetent 13-year-old child, and N. meningitidis was recovered from a sample of her cerebrospinal fluid before death. The isolate was nontypeable by both conventional antisera and indirect whole-cell enzyme-linked immuosorbent assay methods using antibodies to serogroups B, C, Y, and W135. The isolate was further identified as a cnl strain, serotype 15 (ST-198). N. meningitidis-specific DNA was identified in the isolate and in the pre- and postmortem specimens by 16S rRNA and porA gene analysis. Conclusions. This is the first reported case of fatal meningococcal disease caused by an acapsular cnl strain of N. meningitidis that was isolated from an immunocompetent host. Routine molecular diagnostic methods targeted at the cnl locus failed to detect this organism, indicating a need to determine the incidence of infection with cnl strains among patients with culture-negative invasive disease. [ABSTRACT FROM AUTHOR]

An assessment was made of the utilization and impact of a diagnostic polymerase chain reaction (PCR) assay for the diagnosis of herpes simplex viruses (HSV) 1 and 2 in cerebrospinal fluid of children who attended a Canadian pediatric referral centre. One hundred and three assays were performed on specimens from 103 patients during the period August 1997 to September 1998. Patient ages ranged from newborn to 16 years. Indications for HSV PCR included seizures with or without fever (56.3%), aseptic meningitis (16.5%), and encephalopathy with or without fever (10.7%). Only 2 of 103 (1.9%) assays were positive, including one each for HSV1 and HSV2. Control specimens that were seeded with virus indicated inhibition for 24.3, 8.8, and 6.8% of assays for HSV1, HSV2, and both HSV1 and HSV2, respectively. The mean turn-around time for HSV PCR was 2.5 days, and 90.3% were completed in less than 5 days. Acyclovir was administered to 78.6% of the patients overall; the results of the HSV PCR impacted on the treatment courses for 36 individuals. Nevertheless, 16.5% of patients continued to receive extended courses of antiviral therapy despite negative HSV PCR assays. Although it is desirable to decrease the frequency of PCR inhibitions and to further decrease the interval to assay completion, HSV PCR does have a significant impact on antiviral use in this setting.Key words: herpes simplex, polymerase chain reaction, central nervous system.Nous avons évalué l'utilisation et l'impact d'un test de réaction de polymérase en chaîne (PCR) à des fins de diagnostic du virus herpes simplex (HSV) 1 et 2 dans le liquide céphalo-rachidien d'enfants qui ont fréquenté un centre canadien de pédiatrie. Cent trois tests ont été effectués sur des spécimens provenant de 103 patients pendant la période allant de août 1997 à septembre 1998. L'âge des patient allait du nouveau-né jusqu'à 16 ans. Les indications prescrivant le PCR HSV incluaient des crises avec ou sans fièvre (56,3%), une méningite aseptique (16,5%) et une encéphalopathie avec ou sans fièvre (10,7%). Seulement 2/103 (1,9%) des tests furent positifs, incluant un chacun pour HSV1 et HSV2. Des spécimens témoins ensemencés avec des virus ont démontré une inhibition de respectivement 24,3, 8,8, et 6,8% des tests pour le HSV1, HSV2, et les deux. Le délai moyen pour l'obtention des résultats de PCR HSV était de 2,5 jours, et 90,3% des tests ont été complétés avant 5 jours. De l'acyclovir fut administré à 78,6% du nombre total de patients; les résultats de PCR HSV a eu un impact sur le déroulement des traitements de 36 individus. Néanmoins, 16,5% des patients ont continué de recevoir des traitements prolongés aux antiviraux malgré des test de PCR HSV négatifs. Bien qu'il soit souhaitable de diminuer la fréquence d'inhibitions de PCR et de diminuer davantage le délai de l'accomplissement du test, le PCR HSV a bel et bien un impact important sur l'utilisation d'antiviraux dans ce contexte.Mots clés : herpes simplex, réaction de polymérase en chaîne, système nerveux central.[Traduit par la Rédaction] [ABSTRACT FROM AUTHOR]