The treatment of choice for chronic atrophic candidiasis (CAC), also known as denture stomatitis, is topical antifungal therapy. This study aimed to isolate, identify, and assess the antifungal susceptibility of Candida species from mucosal sites in denture wearers with a diagnosis of CAC and determine the prevalence of associated variables. The sample consisted of 44 patients wearing complete or partial dentures who had a clinical diagnosis of CAC. Using sterile cotton swabs, specimens were collected from the oral mucosa of all patients and grown at 30ºC for 48 h in CHROMagar Candida, as a means of isolating and screening the species. The complementary identification of the species was performed using the VITEK 2 automated system (BioMérieux), as well as the determination of their susceptibility to antifungal agents. Data were analyzed using the chi-square test. STATA 13.1 was used for statistical analysis (α = 5%). Of 44 patients with CAC, 33 (75%) had lesions classified as Newton type II. Yeasts were isolated in 38 cases. The most prevalent species was Candida albicans. None of the isolates were resistant to the antifungals tested. Our findings suggest that current indications for antifungal agents are appropriate. Also, antifungal susceptibility testing and proper fungal identification can help dentists to determine the optimal course of treatment for CAC.