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About: Abstract The incidence of invasive fungal infections is on the rise worldwide due to the growth of the immunocompromised population. We report here the use of a diagnostic assay that utilizes a universal extraction method, broad spectrum PCR amplification and analysis via electrospray ionization mass spectrometry (PCR/ESI-MS) to detect and identify more than 200 pathogenic fungi directly from bronchoalveolar lavage (BAL) specimens in less than 8 hours. In this study, we describe both analytical and clinical performance of the assay, when run with prospectively collected clinical BAL specimens. In 146 patients with probable and possible fungal infections defined by EORTC/MSG (European Organization for Research and Treatment of Cancer/Mycoses Study Group) criteria, the PCR/ESI-MS assay demonstrated a sensitivity of 90.9% (95% CI: 76.4–96.9%) and a specificity of 82.3% (95% CI: 74.2–88.2%). This data demonstrates the utility of a non-culture based broad fungal targets molecular diagnostic tool for rapid and accurate diagnosis of invasive fungal infections in patients at risk of developing fungal diseases.

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