title: Reduced turnaround time and improved diagnosis of invasive serogroup B Neisseria meningitidis and Streptococcus pneumoniae infections using a lyophilized quadruplex quantitative PCR.
abstract: Since 1996 the Meningococcal Reference Unit (MRU) in Manchester has provided a national service for PCR confirmation of meningococcal and pneumococcal disease. Neisseria meningitidis serogroup B is predominant in the UK, accounting for >60% of cases. In response to this, the MRU has developed a quadruplex quantitative PCR that detects N. meningitidis capsule transporter (ctrA), serogroup B sialyltransferase (siaDB), Streptococcus pneumoniae pneumolysin (ply) and an internal control. The assay was prepared in a ready-to-use lyophilized format by Applied Biosystems. Laboratory validation showed excellent performance in a specificity panel of 52 isolates and improved detection in comparison with the routine assay. Testing of 244 patient samples showed sensitivity of 93% [95% confidence interval (CI): 88-98%] for the ctrA assay, 95% (95% CI: 91-100%) for the siaDB assay and 100% (95% CI: 95-100%) for the ply assay. Specificity was 100% (95% CI: 98-100%) for both meningococcal targets and 95% (95% CI: 92-98%) for ply. The quadruplex also retained high performance in mixed samples and had acceptable reproducibility. After introduction of the quadruplex into routine use the turnaround time for N. meningitidis group B PCR confirmation reduced from 37 to 29 h and the internal control has proved useful for detecting inhibitory samples. The quadruplex assay provides rapid group B confirmation of meningococcal positive samples, enabling timely public health interventions for the most common disease-causing meningococcal serogroup in the UK.
authors.affiliation: Meningococcal Reference Unit, Public Health England, Manchester Laboratory, Manchester Royal Infirmary, Oxford Road, Manchester M13 9WL, UK.
authors.forename: Martin P
authors.initials: MP
authors.lastname: McHugh
authors.affiliation: Meningococcal Reference Unit, Public Health England, Manchester Laboratory, Manchester Royal Infirmary, Oxford Road, Manchester M13 9WL, UK.
authors.forename: Steve J
authors.initials: SJ
authors.lastname: Gray
authors.affiliation: Meningococcal Reference Unit, Public Health England, Manchester Laboratory, Manchester Royal Infirmary, Oxford Road, Manchester M13 9WL, UK.
authors.forename: Edward B
authors.initials: EB
authors.lastname: Kaczmarski
authors.affiliation: Meningococcal Reference Unit, Public Health England, Manchester Laboratory, Manchester Royal Infirmary, Oxford Road, Manchester M13 9WL, UK.
authors.forename: Malcolm
authors.initials: M
authors.lastname: Guiver
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008585:Meningitis, Meningococcal / Q000175:diagnosis* / Q000382:microbiology; D038541:Neisseria meningitidis, Serogroup B / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D011008:Pneumococcal Infections / Q000175:diagnosis* / Q000382:microbiology; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D013296:Streptococcus pneumoniae / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*
chemical_list: D001426:Bacterial Proteins