Molecular diagnosis of bacterial meningitis by multiplex real time PCR in Tunisian children
Introduction: Bacterial meningitis is a medical emergency requiring a fast and reliable diagnosis. Molecular methods such as real-time PCR (rt-PCR) offer an attractive alternative. Thus, this study aims to establish multiplex rt-PCRs detecting N. meningitidis, S. pneumoniae and H. influenzae b from...
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| Format: | Article |
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The Journal of Infection in Developing Countries
2018-04-01
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| Series: | Journal of Infection in Developing Countries |
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| Online Access: | https://jidc.org/index.php/journal/article/view/9650 |
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| author | Sondes Haddad-Boubaker Marwa Lakhal Cyrine Fathallah Aida Bouafsoun Maher Kharrat Monia Khemiri Amel Kechrid Hanen Smaoui |
| author_facet | Sondes Haddad-Boubaker Marwa Lakhal Cyrine Fathallah Aida Bouafsoun Maher Kharrat Monia Khemiri Amel Kechrid Hanen Smaoui |
| author_sort | Sondes Haddad-Boubaker |
| collection | DOAJ |
| description |
Introduction: Bacterial meningitis is a medical emergency requiring a fast and reliable diagnosis. Molecular methods such as real-time PCR (rt-PCR) offer an attractive alternative. Thus, this study aims to establish multiplex rt-PCRs detecting N. meningitidis, S. pneumoniae and H. influenzae b from cerebrospinal fluid in Tunisian children beyond neonatal age.
Methodology: Using bioinformatic tools and experimentation, we validated the specificity and optimal criteria of PCRs for primers and probes of plyA (S. pneumoniae), ctrA and sodC (N. meningitidis) and bexA genes (H. influenzae b). We performed one multiplex RT-PCR for detection of S. pneumoniae and N. meningitidis targeting plyA and ctrA, sodC genes respectively, simultaneously with a singleplex RT-PCR for H. influenzae b. The sensitivity and specificity of our methods were assessed. Then, we tested our methods for 122 CSF samples collected from suspected meningitis cases between 2014 and 2016 in Bechir Hamza Children’s Hospital of Tunis.
Results: Our results have shown the sensitivity of the designed PCRs was up to 10-4 DNA dilution and the specificity was 100%. PCR evaluation has shown 51 positive samples: 38 of pneumococcal cases, 12 meningococcal cases, 1 case of H. influenzae b with 8.57% and 50% of supplementary positive cases rates respectively.
Conclusions: Our assay proved to be very sensitive, specific and rapid for bacterial meningitis diagnosis. In the recent context of Hib vaccination, the possibility of detecting S. pneumoniae and N. meningitidis separately constitute an attractive opportunity. Nevertheless, simultaneous detection of Hib remains relevant in specific clinical context and for epidemiologic study.
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| format | Article |
| id | doaj-art-e882f3ed70264cfd947dcbbbae9d2032 |
| institution | Kabale University |
| issn | 1972-2680 |
| language | English |
| publishDate | 2018-04-01 |
| publisher | The Journal of Infection in Developing Countries |
| record_format | Article |
| series | Journal of Infection in Developing Countries |
| spelling | doaj-art-e882f3ed70264cfd947dcbbbae9d20322025-08-20T03:52:42ZengThe Journal of Infection in Developing CountriesJournal of Infection in Developing Countries1972-26802018-04-01120410.3855/jidc.9650Molecular diagnosis of bacterial meningitis by multiplex real time PCR in Tunisian childrenSondes Haddad-Boubaker0Marwa Lakhal1Cyrine Fathallah2Aida Bouafsoun3Maher Kharrat4Monia Khemiri5Amel Kechrid6Hanen Smaoui7Béchir Hamza Children’s Hospital, University Tunis El Manar, Tunis, TunisiaBéchir Hamza Children’s Hospital, University Tunis El Manar, Tunis, TunisiaBéchir Hamza Children’s Hospital, University Tunis El Manar, Tunis, TunisiaBéchir Hamza Children’s Hospital, University Tunis El Manar, Tunis, TunisiaUniversity Tunis El Manar, Tunis, TunisiaUniversity Tunis El Manar, Tunis, TunisiaBéchir Hamza Children’s Hospital, University Tunis El Manar, Tunis, TunisiaBéchir Hamza Children’s Hospital, University Tunis El Manar, Tunis, Tunisia Introduction: Bacterial meningitis is a medical emergency requiring a fast and reliable diagnosis. Molecular methods such as real-time PCR (rt-PCR) offer an attractive alternative. Thus, this study aims to establish multiplex rt-PCRs detecting N. meningitidis, S. pneumoniae and H. influenzae b from cerebrospinal fluid in Tunisian children beyond neonatal age. Methodology: Using bioinformatic tools and experimentation, we validated the specificity and optimal criteria of PCRs for primers and probes of plyA (S. pneumoniae), ctrA and sodC (N. meningitidis) and bexA genes (H. influenzae b). We performed one multiplex RT-PCR for detection of S. pneumoniae and N. meningitidis targeting plyA and ctrA, sodC genes respectively, simultaneously with a singleplex RT-PCR for H. influenzae b. The sensitivity and specificity of our methods were assessed. Then, we tested our methods for 122 CSF samples collected from suspected meningitis cases between 2014 and 2016 in Bechir Hamza Children’s Hospital of Tunis. Results: Our results have shown the sensitivity of the designed PCRs was up to 10-4 DNA dilution and the specificity was 100%. PCR evaluation has shown 51 positive samples: 38 of pneumococcal cases, 12 meningococcal cases, 1 case of H. influenzae b with 8.57% and 50% of supplementary positive cases rates respectively. Conclusions: Our assay proved to be very sensitive, specific and rapid for bacterial meningitis diagnosis. In the recent context of Hib vaccination, the possibility of detecting S. pneumoniae and N. meningitidis separately constitute an attractive opportunity. Nevertheless, simultaneous detection of Hib remains relevant in specific clinical context and for epidemiologic study. https://jidc.org/index.php/journal/article/view/9650N. meningitidisS. pneumoniaeH. influenzae bmultiplex real-time PCRbacterial meningitischildren |
| spellingShingle | Sondes Haddad-Boubaker Marwa Lakhal Cyrine Fathallah Aida Bouafsoun Maher Kharrat Monia Khemiri Amel Kechrid Hanen Smaoui Molecular diagnosis of bacterial meningitis by multiplex real time PCR in Tunisian children Journal of Infection in Developing Countries N. meningitidis S. pneumoniae H. influenzae b multiplex real-time PCR bacterial meningitis children |
| title | Molecular diagnosis of bacterial meningitis by multiplex real time PCR in Tunisian children |
| title_full | Molecular diagnosis of bacterial meningitis by multiplex real time PCR in Tunisian children |
| title_fullStr | Molecular diagnosis of bacterial meningitis by multiplex real time PCR in Tunisian children |
| title_full_unstemmed | Molecular diagnosis of bacterial meningitis by multiplex real time PCR in Tunisian children |
| title_short | Molecular diagnosis of bacterial meningitis by multiplex real time PCR in Tunisian children |
| title_sort | molecular diagnosis of bacterial meningitis by multiplex real time pcr in tunisian children |
| topic | N. meningitidis S. pneumoniae H. influenzae b multiplex real-time PCR bacterial meningitis children |
| url | https://jidc.org/index.php/journal/article/view/9650 |
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