MALDI-TOF MS Identification of Candida auris

Overnight cultured C. auris isolates grown on SDA were used for spectra acquisition. Full extraction was performed according to the MALDI Biotyper standard protocol as described elsewhere [54 (link),55 (link),56 (link)]. Briefly, from each isolate, biomass was taken with a 10 µL inoculation loop and suspended in 500 µL distilled water and centrifuged 3 min at 14,000× g, the supernatant was discarded, and 1 mL of 70% ethanol was added. The suspension was homogenized and then centrifuged at 3 min at 14,000× g. Following this, based on the pellet size, equal amounts of 70% formic acid and 100% acetonitrile were added. After centrifugation for 5 min at 14,000× g, 1 μL of supernatant was spotted eight times onto a polished steel target plate (Bruker Daltonik) and overlaid with 1 µL MALDI matrix (10 mg/mL of α-cyano-4-hydroxy-cinnamic acid (α-HCCA) in 50% acetonitrile–2.5% trifluoroacetic acid; Bruker Daltonik) [57 (link)]. MALDI-TOF MS spectra were acquired with a Microflex LT/SH mass spectrometer (Bruker Daltonik) calibrated with the Bruker Bacterial Test Standard in the mass range between 2 and 20 kDa [25 (link)]. Up to 24 raw spectra were analyzed by the MALDI Biotyper Compass Explorer 4.1 software (Bruker Daltonik) to generate the reference spectra (MSP—Main Spectra Projection) and a UPGMA dendrogram was created with BioloMICS v12 (BioAware).