Integrative diagnosis of invasive pulmonary aspergillosis in non-neutropenic patients using BALF-tNGS–derived Aspergillus load and host risk factors: a multicenter study

Background Diagnosing invasive pulmonary aspergillosis (IPA) in non-neutropenic patients is challenging because of non-specific manifestations and limited diagnostic tools. Targeted next-generation sequencing (tNGS) of bronchoalveolar lavage fluid (BALF) enables rapid pathogen detection; however, its capacity for quantitative assessment of fungal load remains unclear. This study integrated BALF-tNGS fungal load with host risk factors to develop a diagnostic nomogram for IPA in non-neutropenic patients. Methods Non-neutropenic adults with suspected IPA were retrospectively enrolled at three tertiary hospitals (December 2020–December 2024). IPA was classified according to consensus definitions. Normalized Aspergillus reads from BALF-tNGS were stratified into low, medium, and high tiers. Clinical, radiological, and microbiological variables were analyzed, and a multivariable logistic regression model was built and internally validated via bootstrapping. Diagnostic performance was assessed using receiver operating characteristic analysis. Results Among 238 patients, 134 had IPA and 104 had non-IPA. Compared with non-IPA cases, patients with IPA had higher rates of diabetes (73.9%), corticosteroid exposure (87.3%), bacterial co-infection (94.8%), and intensive care unit (ICU) admission (72.4%) (all P < 0.01). IPA prevalence peaked in the medium-load group (75.4%) compared with the low-load (36.7%) and high-load (38.3%) groups ( P < 0.001). Seven independent predictors, Aspergillus load, diabetes, corticosteroid exposure, bacterial co-infection, ICU admission, nodular shadow, and positive BALF culture, were incorporated into the model, which showed excellent discrimination (AUC = 0.966; sensitivity, 91.8%; specificity, 95.2%) and good calibration. Conclusion Integrating BALF-tNGS–derived normalized Aspergillus reads with host factors substantially improves differentiation of IPA from colonization in non-neutropenic patients. This semi-quantitative framework supports early, individualized antifungal decision-making and merits external validation.