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Candida glabrata is often the second most common causative agent for candidiasis following Candida albicans. Despite the importance of C. glabrata infections, few epidemiological studies have been conducted on this issue. The goal of this study was genotyping of clinical isolates of C. glabrata by multilocus sequence typing (MLST) technique for determination of the endemic prevalent genotypes and any association between isolation source and drug resistance. A total of 50 C. glabrata clinical isolates from Iran were analyzed by MLST and tested for in-vitro susceptibilities to amphotericin-B, caspofungin, fluconazole, and voriconazole according to the Clinical Laboratory Standards Institute (CLSI) M27-A4 document guidelines. Among these isolates, 16 distinct STs were identified, indicating a discriminatory power index of 0.9029. The three major sequence types (STs) were ST-59, ST-74, and ST-7 with 10, 8, and 7 isolates, respectively. Furthermore, a total of 11 new sequences were found, to which no allele numbers were assigned in the MLST database. All the isolates were susceptible to amphotericin B and caspofungin. Fluconazole resistance was shown in four isolates. Also, a sole isolate was voriconazole resistant. This study shows that the population structure of C. glabrata in Iran consists of groups closely related to the global database as well as to some new clonal clusters and STs. Regarding the high prevalence of 11 new sequences found in this study, it can be concluded that, these new alleles are among the endemic genotypes of Iran. The genotypes or STs were independent of drug susceptibility and anatomic sources.