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Specific detection of Treponema pallidum in clinical samples: validation of a qPCR assay combining two genomic targets
  1. Romain Salle1,2,
  2. Constance Mayslich1,
  3. Philippe Alain Grange1,2,
  4. Valentin Leducq3,
  5. Guillaume Ollagnier1,2,
  6. Ugo Heller4,
  7. Julie Saule5,
  8. Pervenche Martinet6,
  9. Jean-Luc Robert7,
  10. Nadjet Benhaddou8,
  11. Sebastien Fouere9,
  12. Nicolas Dupin1,2
  1. 1INSERM, Institut Cochin U1016-CNRS UMR8104, Équipe Biologie Cutané, Université de Paris, Paris, France
  2. 2Service de Dermatologie-Vénéréologie et CeGIDD, Hôpital Cochin, APHP, CNR IST Bactériennes - Expertise Syphilis, Paris, France
  3. 3Sorbonne Université, INSERM, Institut Pierre Louis d’Epidémiologie et de Santé Publique (iPLESP), AP-HP, Pitié Salpêtrière Hospital, Department of Virology, F-75013, Paris, France
  4. 4Service de Chirurgie Maxillo-Faciale, Hôpital Beaujon, APHP, Clichy, France
  5. 5CeGIDD-Conseil Départemental 13 Joliette, Marseille, France
  6. 6CeGIDD-Conseil Départemental 13 Joliette; CeGIDD-Conseil Départemental 13 Saint Adrien, Marseille, France
  7. 7CeGIDD-Conseil Départemental 13 d’Aix, Aix-en-Provence, France
  8. 8Service de Bactériologie, APHP, CNR Streptocoques, Paris, France
  9. 9Groupe Hospitalier Saint-Louis, Lariboisière, Fernand-Widal, CeGIDD, APHP, Paris, France
  1. Correspondence to Professor Nicolas Dupin, Institut Cochin, Paris, Île-de-France, France; nicolas.dupin{at}aphp.fr

Abstract

Objectives We evaluated a real-time quantitative PCR (qPCR) for detection of the Treponema pallidum (TP) genome in clinical samples through simultaneous detection of two genomic targets.

Methods We performed qPCR with TaqMan technology using two TP genes, polA and tpp47, as targets, with an internal positive control. The qPCR assay was compared with syphilis diagnosis based on a combination of clinical examination, serological results and inhouse nested PCR (nPCR). Samples were analysed at the National Reference Center for STIs at Cochin Hospital in Paris.

Results In total, from October 2010 to December 2016, 320 documented clinical samples (mucosal and cutaneous swabs) were collected from patients with or without syphilis attending STI centres in France. The qPCR had an overall sensitivity of 89% (95% CI 85.1% to 92.1%), a specificity of 100%, a positive predictive value of 100% and a negative predictive value of 88% (95% CI 84.3% to 91.5%). The agreement between qPCR and nPCR results was 94% (κ=0.88, 95% CI 0.83 to 0.93). Calibration of the qPCR assay, by cloning both the polA and tpp47 genes, defined the detection threshold as 1 copy/µL of DNA elution.

Conclusions We validated a new qPCR for detecting the TP genome in clinical samples with excellent sensitivity and specificity. The cloning of polA and tpp47 genes for calibration would be interesting in the evaluation of bacterial loads in samples.

  • syphilis
  • polymerase chain reaction
  • diagnosis

Data availability statement

All data relevant to the study are included in the article or uploaded as supplemental information.

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Data availability statement

All data relevant to the study are included in the article or uploaded as supplemental information.

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Footnotes

  • Handling editor Apostolos Beloukas

  • Contributors RS drafted the article and participated in data collection, analysis and interpretation. CM, VL, GO and NB participated in conception of the study. PG participated in conception of the study and data analysis and interpretation. UH participated in data analysis and interpretation. JS, PM, J-LR and SF participated in data collection. ND participated in conception of the study and data collection and gave final approval of the version to be published as guarantor.

  • Funding This work was supported by the French Society of Dermatology.

  • Competing interests None declared.

  • Provenance and peer review Not commissioned; externally peer reviewed.

  • Supplemental material This content has been supplied by the author(s). It has not been vetted by BMJ Publishing Group Limited (BMJ) and may not have been peer-reviewed. Any opinions or recommendations discussed are solely those of the author(s) and are not endorsed by BMJ. BMJ disclaims all liability and responsibility arising from any reliance placed on the content. Where the content includes any translated material, BMJ does not warrant the accuracy and reliability of the translations (including but not limited to local regulations, clinical guidelines, terminology, drug names and drug dosages), and is not responsible for any error and/or omissions arising from translation and adaptation or otherwise.