The genetic profiles of 50 Candida tropicalis isolates serially collected from 14 patients during a prospective surveillance study in adult intensive care units (ICUs) were characterized by multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) of NaeI restriction fragments. A total of 21 diploid sequence types (DSTs) and 43 genotypes were differentiated by MLST and PFGE, respectively. Significant correlations were found between PFGE genotypes and DST types (P<0.05). Dendrograms generated by either MLST or PFGE-NaeI showed that most isolates from the same patient co-clustered with high similarity regardless of the anatomical source of isolation. Maintenance, microvariation or replacement of C. tropicalis isolates could be observed within the individual patients by further analysis of variations in MLST sequence data. Antifungal susceptibility testing revealed that 17 (34%) of 50 isolates presented high MICs to flucytosine (MIC> or =8 microg/mL). Sixteen (94%) of these isolates belonged to DST 164, and these were collected from four patients with different PFGE genotypes. Isolates sharing the same DST may represent a common clone that underwent extensive mutation over time to cope with drug selection pressure, different hosts or different geographic environments.