title: Diagnostic accuracy of two multiplex real-time polymerase chain reaction assays for the diagnosis of meningitis in children in a resource-limited setting.
abstract: BACKGROUND
  Accurate etiological diagnosis of meningitis is important, but difficult in resource-limited settings due to prior administration of antibiotics and lack of viral diagnostics. We aimed to develop and validate 2 real-time multiplex PCR (RT-PCR) assays for the detection of common causes of community-acquired bacterial and viral meningitis in South African children.
  
  
  METHODS
  We developed 2 multiplex RT- PCRs for detection of S. pneumoniae, N. meningitidis, H. influenzae, enteroviruses, mumps virus and herpes simplex virus. We tested residual CSF samples from children presenting to a local paediatric hospital over a one-year period, whose CSF showed an abnormal cell count. Results were compared with routine diagnostic tests and the final discharge diagnosis. We calculated accuracy of the bacterial RT-PCR assay compared to CSF culture and using World Health Organisation definitions of laboratory-confirmed bacterial meningitis.
  
  
  RESULTS
  From 292 samples, bacterial DNA was detected in 12 (4.1%) and viral nucleic acids in 94 (32%). Compared to CSF culture, the sensitivity and specificity of the bacterial RT-PCR was 100% and 97.2% with complete agreement in organism identification. None of the cases positive by viral RT-PCR had a bacterial cause confirmed on CSF culture. Only 9/90 (10%) of patients diagnosed clinically as bacterial meningitis or partially treated bacterial meningitis tested positive with the bacterial RT-PCR.
  
  
  CONCLUSIONS
  In this population the use of 2 multiplex RT-PCRs targeting 6 common pathogens gave promising results. If introduced into routine diagnostic testing, these multiplex RT-PCR assays would supplement other diagnostic tests, and have the potential to limit unnecessary antibiotic therapy and hospitalisation.
authors.affiliation: Division of Medical Microbiology, Department of Pathology, University of Cape Town, Cape Town, South Africa.
authors.forename: Jermaine
authors.initials: J
authors.lastname: Khumalo
authors.affiliation: Division of Medical Microbiology, Department of Pathology, University of Cape Town, Cape Town, South Africa.
authors.forename: Mark
authors.initials: M
authors.lastname: Nicol
authors.affiliation: National Health Laboratory Service, Johannesburg, South Africa.|Division of Virology, Department of Pathology, University of Cape Town, Cape Town, South Africa.
authors.forename: Diana
authors.initials: D
authors.lastname: Hardie
authors.affiliation: Department of Paediatrics, University of Cape Town, Cape Town, South Africa.
authors.forename: Rudzani
authors.initials: R
authors.lastname: Muloiwa
authors.affiliation: Department of Paediatrics, University of Cape Town, Cape Town, South Africa.
authors.forename: Phindile
authors.initials: P
authors.lastname: Mteshana
authors.affiliation: Division of Medical Microbiology, Department of Pathology, University of Cape Town, Cape Town, South Africa.|National Health Laboratory Service, Johannesburg, South Africa.
authors.forename: Colleen
authors.identifier: http://orcid.org/0000-0001-6236-879X
authors.initials: C
authors.lastname: Bamford
mesh_terms: D001419:Bacteria / Q000235:genetics / Q000302:isolation & purification*; D002648:Child; D002675:Child, Preschool; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification; D004770:Enterovirus / Q000235:genetics / Q000302:isolation & purification; D005260:Female; D006193:Haemophilus influenzae / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D008297:Male; D016920:Meningitis, Bacterial / Q000134:cerebrospinal fluid / Q000175:diagnosis* / Q000453:epidemiology / Q000382:microbiology*; D008587:Meningitis, Viral / Q000134:cerebrospinal fluid / Q000175:diagnosis* / Q000453:epidemiology / Q000821:virology*; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D009109:Mumps virus / Q000235:genetics / Q000302:isolation & purification; D009345:Neisseria meningitidis / Q000235:genetics / Q000302:isolation & purification; D009696:Nucleic Acids / Q000235:genetics / Q000302:isolation & purification; D012680:Sensitivity and Specificity; D018139:Simplexvirus / Q000235:genetics / Q000302:isolation & purification; D013019:South Africa; D013296:Streptococcus pneumoniae / Q000235:genetics / Q000302:isolation & purification; D014780:Viruses / Q000235:genetics / Q000302:isolation & purification*
chemical_list: D004269:DNA, Bacterial; D009696:Nucleic Acids