APX001A is the active moiety of the first-in-class drug candidate APX001. So far, most susceptibility testing studies have examined ≤30 isolates/species, and only one used the EUCAST method. Here, we investigated the activity of APX001A and five comparators against 540 candidemia and 122 isolates. Isolates (17 and 3 yeast species) were identified using CHROMagar, matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF) and, when needed, internal transcribed space (ITS) sequencing. EUCAST E.Def 7.3.1 susceptibility testing included APX001A, amphotericin B, anidulafungin, micafungin, fluconazole, and voriconazole. Wild-type upper limits (WT-UL) were established following the EUCAST principles for epidemiological cutoff value setting for APX001A, allowing classification as wild type (WT) or non-WT. APX001A MIC values (mg/liter) were as follows: ,, and , 0.004 to 0.008; and , 0.016;, 0.06; and ,>0.5. APX001A MICs against the rare species varied from ≤0.0005 () to >0.5 (). APX001A was equally or more active than the comparators against all species except and Four isolates were APX001A non-WT; all were fluconazole resistant. A correlation was observed between APX001A and fluconazole MICs across all species except and , and when comparing high and low fluconazole MIC isolates of, ,, , and APX001A showed promising activity against most and other yeast species, including , compared to five comparators. WT-UL were suggested for the common species, and a new and unexplained correlation to fluconazole susceptibility was observed.