Persistent candidemia refers to the continued isolation of the same Candida species in the blood of a candidemic patient despite appropriate therapy. Despite the clinical importance of persistent candidemia, studies have superficially addressed the biological conditions behind this phenomenon. The aim of this study was to evaluate the correlation between the biofilm-forming ability by Candida bloodstream isolates and the persistence of infection. A total of 55 isolates of Candida were tested and characterized in two groups: (i) group I, which included seven patients with persistent candidemia, and (ii) group II, which included 18 patients with nonpersistent candidemia. Microorganisms were identified at the species level by sequencing the internal transcribed spacer (ITS) region of ribosomal DNA (rDNA). Biofilm quantification was evaluated by the crystal violet staining method and confocal scanning laser microscopy (CSLM). Molecular tests confirmed the identification of Candida albicans (92% group I and 94% group II) and Candida dubliniensis isolates (8% group I and 6% group II). All 55 isolates were able to form biofilms, but a higher biofilm mass was produced by C. albicans/C. dubliniensis strains cultured from the persistent group (P < .05). Our data suggest that Candida sp. biofilm production should be considered a relevant biologic variable in explaining patients who fail to clear a bloodstream infection despite adequate antifungal treatment with triazoles.