Development of a PCR algorithm to detect and characterize Neisseria meningitidis carriage isolates in the African meningitis belt

MenAfriCar Consortium

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7 Scopus citations

Abstract

Improved methods for the detection and characterization of carried Neisseria meningitidis isolates are needed. We evaluated a multiplex PCR algorithm for the detection of a variety of carriage strains in the meningitis belt. To further improve the sensitivity and specificity of the existing PCR assays, primers for gel-based PCR assays (sodC, H, Z) and primers/ probe for real-time quantitative PCR (qPCR) assays (porA, cnl, sodC, H, E, Z) were modified or created using Primer Express software. Optimized multiplex PCR assays were tested on 247 well-characterised carriage isolates from six countries of the African meningitis belt. The PCR algorithm developed enabled the detection of N. meningitidis species using gel-based and real-time multiplex PCR targeting porA, sodC, cnl and characterization of capsule genes through sequential multiplex PCR assays for genogroups (A, W, X, then B, C, Y and finally H, E and Z). Targeting both porA and sodC genes together allowed the detection of meningococci with a sensitivity of 96% and 89% and a specificity of 78% and 67%, for qPCR and gel-based PCR respectively. The sensitivity and specificity ranges for capsular genogrouping of N. meningitidis are 67% - 100% and 98%-100% respectively for gel-based PCR and 90%-100% and 99%-100% for qPCR. We developed a PCR algorithm that allows simple, rapid and systematic detection and characterisation of most major and minor N. meningitidis capsular groups, including uncommon capsular groups (H, E, Z).

Original languageEnglish (US)
Article numbere0206453
JournalPloS one
Volume13
Issue number12
DOIs
StatePublished - Dec 1 2018

Bibliographical note

Funding Information:
This study was funded by the Meningitis Research Foundation (www.meningitis.org) under the auspices of the MenAfriCar project. The MenAfriCar Consortium was supported by grants from the Wellcome Trust (086546) and the Bill & Melinda Gates Foundation (OPP51251). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. K.D. Is a Wellcome Trust Tropical Fellow. O.B.H. and M.C.J.M. were supported by the Wellcome Trust (grant 087622). This research made use of the Neisseria Sequence Typing database website (http://pubmlst.org/neisseria/) developed by Keith A. Jolley and Martin C. J. Maiden, hosted by the University of Oxford, and supported by the Wellcome Trust (grant 104992).

Publisher Copyright:
This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.

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