Invasive fungal infections (IFIs) are a significant contributor to hospital acquired infections with immunocompromised patients being at the highest risk of such infections. Among the many fungal pathogens, species of Candida are the most common cause of IFIs and most commonly associated with nosocomial infections. Since 2009, C. auris has emerged as a major fungal pathogen responsible for HAIs, causing severe invasive infection with mortality rates ranging from 30 – 72%. C. auris infections can occur at multiple body sites, such as skin, urogenital tract, and respiratory tract and if not treated in time, infections can spread to the blood stream, which increases the mortality rate. It is unique among pathogenic fungi because of its ability to persist within clinical environments for long period of times. The best way to prevent spread of infection is with rapid and accurate identification of C. auris. The goal of this application is to demonstrate feasibility of a comprehensive molecular detection system for specific detection of C. auris in less than 30 minutes at point-of-care (POC). The proposed assay will be based on a novel “Duplex loop mediated isothermal amplification (Duplex RT-LAMP)” technology and performed on a commercially available benchtop instrument (AmpliFire®). While methods currently used for detection of LAMP amplification products are unable to differentiate between different targets, Duplex LAMP allows detection of two targets in the same reaction. This is feasible because of use of fluorescent labels (FAM, HEX etc.) on target specific primers and a specially formulated 10X Isothermal Master Mix. Another innovation that is the basis of this application is a new thermostable Bst polymerase which has been engineered for activity at higher temperatures (68°C - 74°C) and resistance to inhibitors. Total assay time will be 30 minutes including sample preparation with minimal hands-on time and without need of any additional equipment, such as pipettes, centrifuge etc. Results will be displayed on-screen as positive or negative for C. auris, minimizing errors caused by user interpretation. Successful completion of this project will lead to: • Development of a diagnostic kit for the detection of all 4 major clades of C. auris at POC. • Minimal hands-on and total assay time of 30 minute including sample preparation. • A simple and easy to use sample preparation method for processing of samples at POC. • Diagnostic sensitivity and specificity comparable to reference method. • Platform technology for rapid detection of other fungal pathogens at POC.