Evaluation of the MicroSeq 500 16S rDNA-based gene sequencing for the diagnosis of culture-negative bacterial meningitis.
Gene amplification using 16S rDNA primers has been proposed as a strategy for the diagnosis of bacterial meningitis. The aim of this study was to evaluate the performance of the MicroSeq 500 16S ribosomal DNA test (Applied Biosystems) from patients with suspected bacterial meningitis and CSF negative-culture in comparison to traditional methods. Twelve purulent culture-negative CSF samples were collected between January 2005 and January 2007. For DNA extraction, 500 microl of CSF samples were treated using the QIAamp mini kit (QIAGEN). The extracted DNA was examined amplifying 500 bp at the 5' end of 16S rRNA gene using MicroSeq500 16S rDNA Bacterial Identification PCR kit and the sequencing reactions were performed with the MicroSeq500 16S rDNA Bacterial Identification Sequencing kit (Applied Biosystems). The sequences were compared with those available in GenBank. For the culture-negative CSF samples the MicroSeq 500 16S rDNA yielded a positive result in 9 cases (75.0%): three samples were identified as Streptococcus. pneumoniae, three as Neisseria meningitidis, and the remaining 3 as Haemophilus influenzae, Abiotrophia defectiva and Porphyromonas gingivalis. The MicroSeq 500 16S ribosomal DNA test may improve the microbiological diagnosis of bacterial meningitis, especially when spinal fluid samples are obtained after the administration of antimicrobial therapy.
Arosio
Marco
M
USC Microbiology and Virology, AO Ospedali Riuniti of Bergamo, Italy. marosio@ospedaliriuniti.bergamo.it
Nozza
Francesca
F
Rizzi
Marco
M
Ruggeri
Maurizio
M
Casella
Pietro
P
Beretta
Gianluca
G
Raglio
Annibale
A
Goglio
Antonio
A
D000328:Adult; D000368:Aged; D001431:Bacteriological Techniques; D002555:Cerebrospinal Fluid / Q000382:microbiology; D002648:Child; D002675:Child, Preschool; D004269:DNA, Bacterial / Q000737:chemistry; D004275:DNA, Ribosomal / Q000737:chemistry; D006801:Humans; D007223:Infant; D016920:Meningitis, Bacterial / Q000134:cerebrospinal fluid / Q000175:diagnosis* / Q000382:microbiology; D008875:Middle Aged; D025202:Molecular Diagnostic Techniques; D016133:Polymerase Chain Reaction; D012336:RNA, Ribosomal, 16S / Q000235:genetics*; D012189:Retrospective Studies; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA
D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S