Objective To investigate antimicrobial susceptibility of Cryptococcus neoformans (C. neoformans) isolated from patients with acquired immunodeficiency syndrome(AIDS) in Yunnan Province. Methods 304 strains of C. neoformans isolated from AIDS patients in Yunnan Infectious Diseases Hospital between August 2007 and July 2018 were performed antimicrobial susceptibility testing by colorimetric microdilution method, susceptibility to 6 commonly used anti-cryptococcal agents was detected. Results Antimicrobial susceptibility testing results showed that MIC₅₀ of amphotericin B, 5-fluorocytosine, fluconazole, voriconazole, itraconazole, and posaconazole were 1, 2, 4, 0.03, 0.06, and 0.12 μg/mL respectively, and the MIC₉₀ were 1, 4, 8, 0.06, 0.12, and 0.25 μg/mL respectively, with geometric mean values of 0.75, 2.32, 3.28, 0.039, 0.053, and 0.09 μg/mL respectively. Among 6 antifungal agents, resistance rate of C. neoformans to voriconazole and amphotericin B was 1.32%, and to 5-fluorocytosine was 0.33%, no strain was found to be resistant to itraconazole and fluconazole. Conclusion C. neoformans isolated from AIDS patients in Yunnan Province have different susceptibility to different antimicrobial agents, which can provide reference for clinical antimicrobial choice.