Despite the strong association between Cryptococcus neoformans infection and the Human immunodeficiency virus (HIV) status of patients globally, most cryptococcosis cases in Far East Asia occur in non-HIV individuals. Molecular epidemiological studies, using multilocus sequence typing (MLST), have shown that more than 95% of cryptococcal strains belong to a specific subtype of VNI. However, this association has never been specifically examined in other parts of Asia. Therefore, in this study, we investigated the VNIc/ST5 genotype distribution among cryptococcosis patients in Thailand. Fifty-one C. neoformans isolates were collected from clinical samples in Siriraj Hospital, Bangkok, Thailand. The strains were predominantly isolated from HIV-positive patients (88.57%) and all were molecular type VNI MATα. An MLST analysis identified five sequence types (ST) in Siriraj Hospital, of which ST4 (45.10%) and ST6 (35.29%) were most common, and ST5 (15.69%), ST32 (1.96%), and ST93 (1.96) were less common. Contrary to reports from Far East Asia, ST5 was predominantly (83.33%) found in HIV patients (P = 0.657), and there was no significant change in the prevalence of ST5 over the past 10 years (P = 0.548). A further analysis of comorbidities showed higher morbidity and delays in the cryptococcal diagnosis in patients with tuberculosis coinfection or without HIV. Our study suggests that although the Thai population is genetically closely related to the Far East Asian population, ST5 is not associated with non-HIV status in Thailand. Therefore, this association may not be related to the host’s genetic background. However, its mechanism remains unclear.