title: Evaluation of a PCR-immunoassay technique for detection of Neisseria meningitidis in cerebrospinal fluid and peripheral blood.
abstract: A multiplex PCR-immunoassay for the rapid diagnosis of bacterial meningitis from cerebrospinal fluid (CSF) or peripheral blood was compared with conventional microbiological techniques used in the routine diagnostic laboratory. The multiplex PCR was designed to detect simultaneously a universal conserved sequence coding for bacterial 16S rRNA and the Neisseria meningitidis porA gene. The PCR-immunoassay had a detection limit of (3-5) x 10(2) cfu/ml (equivalent to 1-3 cfu/PCR) with spiked CSF or blood samples, compared with (3-5) x 10(3) cfu/ml for PCR followed by conventional agarose gel electrophoresis for detection of PCR products. Of 294 CSF samples from patients suspected on clinical grounds of suffering from meningitis, the PCR-immunoassay detected bacterial DNA in 29 CSF samples, 15 of which were also positive for N. meningitidis DNA. The 29 positive CSF samples comprised 25 samples that were also reported positive and four that were reported negative by conventional methodology; the latter four were all positive for N. meningitidis by the PCR-immunoassay. Of 173 peripheral blood samples examined, the PCR-immunoassay detected bacterial DNA in 18 samples, 14 of which were also positive for N. meningitidis DNA. In comparison, only 10 samples were reported positive for N. meningitidis by conventional methodology. All negative PCR-immunoassay results correlated with those obtained by conventional methodology for both CSF and blood samples. The sensitivity and speed of the PCR-immunoassay system indicated that it could be used as a routine diagnostic test for meningococcal meningitis, enabling a diagnosis to be made within 4 h of receipt of the specimen.
authors.affiliation: Department of Microbiology and PHLS Laboratory, University Hospital, Queen's Medical Centre, Nottingham NG7 2UH.
authors.forename: R J
authors.initials: RJ
authors.lastname: Seward
authors.affiliation: Department of Microbiology and PHLS Laboratory, University Hospital, Queen's Medical Centre, Nottingham NG7 2UH.
authors.forename: K J
authors.initials: KJ
authors.lastname: Towner
mesh_terms: D016470:Bacteremia / Q000382:microbiology*; D002555:Cerebrospinal Fluid / Q000382:microbiology*; D004269:DNA, Bacterial / Q000097:blood / Q000134:cerebrospinal fluid; D004275:DNA, Ribosomal / Q000097:blood / Q000134:cerebrospinal fluid; D004587:Electrophoresis, Agar Gel; D005069:Evaluation Studies as Topic; D006801:Humans; D007118:Immunoassay; D008585:Meningitis, Meningococcal / Q000175:diagnosis*; D009345:Neisseria meningitidis / Q000235:genetics / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S