Clinical impact of microbial cell-free DNA next-generation sequencing for invasive mold infection—a single-center retrospective observational study

Background: Diagnosing invasive mold infections (IMI) is challenging because they typically occur as opportunistic infections in immunocompromised patients who often present with non-specific symptoms. Furthermore, no single test can definitively diagnose IMI, and a proven diagnosis often requires invasive sampling. This sampling can be unsafe and difficult to perform, especially in the immunocompromised population most at risk for these infections. Objectives: The objective of this study is to assess the clinical utility of plasma microbial cell-free DNA next-generation sequencing (mcfDNA-NGS) for diagnosing invasive mold infections in the context of conventional diagnostic methods. Design: Retrospective observational study at a quaternary care center (2017-2025). Methods: The charts of 30 patients with mold-positive mcfDNA-NGS (Karius™ Spectrum; Redwood City, CA, USA) were reviewed, with IMI adjudicated per 2020 EORTC/MSGERC criteria. Provider documentation, medication orders, and patient outcomes were used to assess clinician interpretation of mcfDNA-NGS result accuracy (true positive vs false positive) and its impact on diagnostic and therapeutic decision-making. Turnaround time (TAT) and molecules per microliter (MPM) were summarized. Results: = 0.86). mcfDNA-NGS changed diagnostic classification in 7/30 (23%) and antimicrobial management in 16/30 (53%). Conclusion: mcfDNA-NGS provided noninvasive, actionable information, informing diagnosis and therapy. Future studies should define optimal stewardship and cost-effectiveness.