Traditional identification of molds is challenging, cumbersome, and time-consuming, relying on non-specific phenotypic characteristics from culture or expensive and specialized DNA sequencing methods. MALDI-TOF MS has revolutionized clinical microbiology laboratories worldwide, enabling rapid identification of bacteria and yeast. This technology, however, has not been readily applied to mold identification due to technical challenges in developing an efficient protein extraction method and the limited availability of clinically comprehensive databases. Recently, we developed and optimized a simple protein extraction procedure for molds and constructed a highly stringent database comprising 410 individual isolates representing 90 genera and 220 species for the rapid identification of molds by MALDI-TOF MS. The NIH Mold Database is the most comprehensive database developed to date for mold identification and has filled a huge diagnostic gap in the field of clinical microbiology. The NIH Mold Database has now been shared with >100 clinical microbiology laboratories worldwide in an effort to improve the diagnosis of invasive fungal infections by providing faster identification (within 30 minutes of growth detection as opposed to weeks) that in turn can guide optimal antifungal therapy. During this fiscal year, we completed a ten-center national controlled study that evaluated the clinical performance and accuracy of two fungal databases (the NIH Mold Database and the Bruker Fungi Database) against 80 clinical mold isolates has come to a close. This study has highlighted important (previously undetermined) factors such as spectral acquisition settings required to improve and achieve successful/uniform implementation of MALDI-TOF MS for routine mold identification in clinical microbiology laboratories. In fact, in August 2019, the manufacturer of the mass spectrometer (Bruker Daltonics) released updated software with specific changes to acquisition parameters to improve mold identification. Data regarding poor inter-instrument reproducibility is supported by two recent publications from different groups. This phenomenon has not been demonstrated previously for large-scale bacterial and yeast studies, and results are surprising and warrant further evaluation. The results from this study have been accepted for publication in Frontiers Microbiology (in press, Sep 2019).