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Article Dans Une Revue Medical Mycology Année : 2020

The Fungal PCR Initiative's evaluation of in-house and commercial Pneumocystis jirovecii qPCR assays: Toward a standard for a diagnostics assay

P Lewis White
  • Fonction : Auteur
Carlo Mengoli
  • Fonction : Auteur
Sharon Chen
  • Fonction : Auteur
Brendan Crowley
  • Fonction : Auteur
Gijs Dingemans
  • Fonction : Auteur
Emilie Fréalle
Rebecca L Gorton
  • Fonction : Auteur
Malcom Guiver
  • Fonction : Auteur
Ferry Hagen
  • Fonction : Auteur
Catriona Halliday
  • Fonction : Auteur
Gemma Johnson
  • Fonction : Auteur
Katrien Lagrou
  • Fonction : Auteur
Martina Lengerova
  • Fonction : Auteur
Willem Melchers
  • Fonction : Auteur
Lily Novak-Frazer
  • Fonction : Auteur
Riina Rautemaa-Richardson
  • Fonction : Auteur
Joerg Steinmann
  • Fonction : Auteur
Mario Cruciani
  • Fonction : Auteur
Rosemary Barnes
  • Fonction : Auteur
J Peter Donnelly
  • Fonction : Auteur
Juergen Loeffler
  • Fonction : Auteur
Stéphane Bretagne
Alexandre Alanio

Résumé

Abstract Quantitative real-time PCR (qPCR) is increasingly used to detect Pneumocystis jirovecii for the diagnosis of Pneumocystis pneumonia (PCP), but there are differences in the nucleic acids targeted, DNA only versus whole nucleic acid (WNA), and also the target genes for amplification. Through the Fungal PCR Initiative, a working group of the International Society for Human and Animal Mycology, a multicenter and monocenter evaluation of PCP qPCR assays was performed. For the multicenter study, 16 reference laboratories from eight different countries, performing 20 assays analyzed a panel consisting of two negative and three PCP positive samples. Aliquots were prepared by pooling residual material from 20 negative or positive- P. jirovecii bronchoalveolar lavage fluids (BALFs). The positive pool was diluted to obtain three concentrations (pure 1:1; 1:100; and 1:1000 to mimic high, medium, and low fungal loads, respectively). The monocenter study compared five in-house and five commercial qPCR assays testing 19 individual BALFs on the same amplification platform. Across both evaluations and for all fungal loads, targeting WNA and the mitochondrial small sub-unit (mtSSU) provided the earliest Cq values, compared to only targeting DNA and the mitochondrial large subunit, the major surface glycoprotein or the beta-tubulin genes. Thus, reverse transcriptase-qPCR targeting the mtSSU gene could serve as a basis for standardizing the P. jirovecii load, which is essential if qPCR is to be incorporated into clinical care pathways as the reference method, accepting that additional parameters such as amplification platforms still need evaluation.

Dates et versions

hal-03529754 , version 1 (17-01-2022)

Identifiants

Citer

Maud Gits-Muselli, P Lewis White, Carlo Mengoli, Sharon Chen, Brendan Crowley, et al.. The Fungal PCR Initiative's evaluation of in-house and commercial Pneumocystis jirovecii qPCR assays: Toward a standard for a diagnostics assay. Medical Mycology, 2020, 58 (6), pp.779-788. ⟨10.1093/mmy/myz115⟩. ⟨hal-03529754⟩
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