Skip to Main content Skip to Navigation
Preprints, Working Papers, ...

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

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 qPCR Initiative, a working group of the International Society for Human and Animal Mycology, a multicentre and monocentre evaluation of PCP qPCR assays was performed. For the multicentre study, 16 reference laboratories from eight different countries, performing 20 assays analysed 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 monocentre 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
Complete list of metadatas

Cited literature [5 references]  Display  Hide  Download

https://hal-pasteur.archives-ouvertes.fr/pasteur-02337518
Contributor : Alexandre Alanio <>
Submitted on : Tuesday, October 29, 2019 - 2:53:09 PM
Last modification on : Wednesday, October 14, 2020 - 4:14:46 AM
Long-term archiving on: : Thursday, January 30, 2020 - 7:09:36 PM

File

Gits muselli Med Mycol 2019_fi...
Files produced by the author(s)

Identifiers

  • HAL Id : pasteur-02337518, version 1

Collections

Citation

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: towards a standard for a diagnostics assay. 2019. ⟨pasteur-02337518⟩

Share

Metrics

Record views

74

Files downloads

170