Interpreting combined BALF-galactomannan and next-generation sequencing in pulmonary aspergillosis and mucormycosis: a two-center clinical decision framework.

OBJECTIVES: To identify which patients with pulmonary aspergillosis or mucormycosis (PAM) derive the greatest incremental diagnostic benefit from next-generation sequencing (NGS) beyond conventional microbiology, and to derive a GM × NGS interpretive framework that identifies the patient subgroup in which NGS adds the most information, based on routinely available bedside variables. Secondary objectives were to characterize the pathogen spectrum detected by NGS, quantify pairwise concordance with BALF galactomannan and culture, and assess the relationship between quantitative Aspergillus reads and BALF-GM intensity. METHODS: Two-center retrospective case-cohort study of 595 adults with NGS-tested pulmonary aspergillosis or mucormycosis (PAM; proven, probable, or possible per EORTC/MSGERC and ECMM-MSGERC criteria). Primary method-comparison analysis was restricted to the strict subset with NGS, BALF-galactomannan (GM), and culture all available (n = 434). Total Aspergillus reads were defined as the per-case sum of species-level Aspergillus reads parsed from free-text NGS reports, with Mucorales reads excluded. Multivariable logistic regression identified predictors of GM-negative status among NGS-positive cases (n = 353; complete-case n = 342). The prospective component was registered at the Chinese Clinical Trial Registry (ChiCTR2500099900). RESULTS: In the strict 434-case subset, NGS fungal-signal positivity was 81.3%, BALF-GM (≥0.8) 58.1%, and culture 43.1%; pairwise overlap was substantial but incomplete. The GM-negative/NGS-positive subgroup (n = 150) showed a milder phenotype than GM-positive/NGS-positive cases (severe pneumonia 28.7% (43/150) vs. 52.7% (107/203); mortality 6.7% (10/150) vs. 19.7% (40/203); both p < 0.001). Severe pneumonia was independently inversely associated with GM-negative status (adjusted OR 0.44, 95% CI 0.26-0.75, p = 0.002), while immunocompromise was independently positively associated (adjusted OR 2.01, 95% CI 1.17-3.47, p = 0.012). NGS Aspergillus positivity was essentially flat across BALF-GM intensity bands (79-82%); read counts correlated only weakly with BALF-GM titer [Spearman ρ = 0.185 (mNGS-only, n = 246)]. CONCLUSION: When BALF-GM and NGS are co-ordered on the same lavage in routine PAM workup, the four GM × NGS quadrants define distinct clinical phenotypes with concrete management implications: NGS positivity in GM-negative cases (34.6%) carries the highest incremental yield and a milder phenotype; conversely, a negative NGS in GM-positive cases must not be used to rule out severe PAM. Across quadrants, NGS adds species- and co-pathogen-level information that changes therapy.