Evaluation of metagenomic next-generation sequencing (mNGS) combined with quantitative PCR: cutting-edge methods for rapid diagnosis of non-invasive fungal rhinosinusitis

Abstract

Purpose: Fungal rhinosinusitis is a significant and growing health concern in arid regions, with an increasing incidence over recent decades. Without timely and appropriate management, it can lead to severe complications, including potential intracranial spread. This study aims to establish efficient and rapid diagnostics for non-invasive fungal rhinosinusitis (FRS), addressing the challenge of its difficult-to-culture diagnosis.

Methods: Twenty-eight patients suspected of FRS were studied using endoscopic sinus surgery to obtain tissue samples for histopathology, direct microscopy, fungal culture, quantitative PCR (qPCR) and metagenomic next-generation sequencing (mNGS) detection. A patented qPCR targeting prevalent Aspergillus species was evaluated.

Results: The patient cohort had a male-to-female ratio of 9:14, with disease duration up to 50 years. Histopathologically, 23 out of 28 cases were positive. Fungal culture exhibited a sensitivity of 21.74%, with one false positive. qPCR and mNGS showed 100% sensitivity and specificity, with a 100% consistency rate for identification at the species level (23/23), and potential detection of cases with co-infections. The most common pathogen was A. flavus, followed by A. fumigatus and A. niger. Two cases involved mixed infections of A. fumigatus and A. flavus.

Conclusion: qPCR and mNGS proved effective in rapidly identifying fungi from fresh sinus tissue that are challenging to culture, surpassing conventional methods. However, further evaluation and optimization with a larger cohort of patients are necessary. Histopathology is still recommended to confirm the clinical significance of the detected fungal species.

Keywords: Aspergillus; Allergy; Diagnostics; Fungal rhinosinusitis; Invasive infection; Metagenomics; qPCR.