Objective  Mucormycosis is a life-threatening invasive fungal infection with high mortality, yet traditional diagnostic methods are limited by low positivity rates. This study aims to evaluate the diagnostic performance and clinical utility of metagenomic next-generation sequencing (mNGS) in mucormycosis.

Methods  A retrospective analysis was conducted on 135 patients with mNGS results positive for Mucorales fungi at West China Hospital of Sichuan University from November 1, 2022, to October 31, 2024. Based on comprehensive clinical diagnostic criteria (including proven and probable cases), patients were classified into a confirmed mucormycosis group and a non-mucormycosis group. Receiver operating characteristic (ROC) curve analysis was used to evaluate the diagnostic performance of normalized read counts (lgRPM) from different specimen types. Fungal species distribution and laboratory parameters were compared between the two groups.

Results  Among the 135 patients with positive mNGS results for Mucorales, 100 (74.1%) were ultimately diagnosed with mucormycosis. ROC curve analysis revealed that the diagnostic performance of mNGS varied by specimen type. For blood specimens, the area under the curve (AUC) was 0.772, with a specificity of 87.5% at the optimal cutoff value of 0.11 RPM. For bronchoalveolar lavage fluid specimens, the AUC was 0.717, with a sensitivity of 76.5% at the optimal cutoff value of 0.02 RPM. Combined analysis of all specimens showed that at the optimal cutoff value of 0.08 RPM (approximately 8 reads/100M), the sensitivity and specificity were 62.0% and 71.4%, respectively. Species distribution analysis showed that the proportions of Cunninghamella elegans (11.0% vs. 2.9%) and Rhizomucor pusillus (9.0% vs. 2.9%) were significantly higher in the confirmed group than in the non-mucormycosis group (P < 0.05). Levels of C-reactive protein and interleukin-6 were also significantly higher in the confirmed group (P < 0.05). Notably, all seven renal perfusion fluid samples yielded false-positive mNGS results.

Conclusion  mNGS technology can effectively improve the diagnostic yield for mucormycosis. However, results should be interpreted in conjunction with specimen type, read count, and clinical characteristics. BALF specimens offer high sensitivity, making them suitable for screening, while blood specimens demonstrate high specificity, making them valuable for confirmation. Positive results from low-biomass samples such as renal perfusion fluid warrant caution against false positivity. Fungal species identification and inflammatory markers may serve as adjunctive evidence for clinical diagnosis.