Related Experiment Video
Updated: Apr 1, 2026
Author Spotlight: Advancements in Multiplex Detection of Respiratory Viruses
Published on: November 10, 2023
Clinical validation of a new multi-Aspergillus species reverse transcriptase quantitative PCR: a diagnostic
Marion Benazra1, Wicem Mallek1, Théo Ghelfenstein-Ferreira2
1Institut Pasteur, Université Paris Cité, National Reference Center for Invasive Mycoses and Antifungals, Translational Mycology Research Group, Mycology Department, Paris, France.
Objectives:
Invasive aspergillosis is a difficult-to-diagnose fungal disease, the diagnosis of which relies on many criteria including mycological criteria. Recently, the European Organisation for Research and Treatment of Cancer/Mycoses Study Group Education and Research Consortium promoted Aspergillus PCR as a specific marker given the significant progress made in optimizing and standardizing Aspergillus real-time quantitative PCR (qPCR) assays. However, there is no unique and optimal assay in the literature, particularly for optimally detecting aspergillosis due to the main Aspergillus species responsible for invasive aspergillosis.
Methods:
We therefore designed, optimized, and validated a new type of real-time PCR assay using a diagnostic case-control study approach. This assay is based on reverse transcriptase qPCR methodology and multi-Aspergillus section-specific primers (multi Aspergillus species reverse transcriptase qPCR [MAS-RTqPCR]). This assay detects whole nucleic acids from species belonging to six Aspergillus sections (Fumigati, Flavi, Nigri, Terrei, Nidulantes, and Usti).
Results:
The limit of detection of this assay was five copies per reaction, and the analytical specificity was 100% based on the screening of >90 fungal species. We used three groups of patients to validate the assay, testing 120 patients and 223 plasma samples. The clinical sensitivity was 0.91 (95% CI, 0.71-0.98) and clinical specificity was 1.00 (95% CI, 0.93-1.00). Follow-up of positive patients revealed superiority of the MAS-RTqPCR assay compared with the reference assay. In addition, our MAS-RTqPCR assay detected three patients with non-Aspergillus fumigatus infections.
Conclusions:
Overall, this study demonstrates that our MAS-RTqPCR assay is a promising diagnostic for invasive aspergillosis and monitoring PCR-positive patients.
