Background/Objectives: Candida species are among the leading causes of fungal infections, especially in immunosuppressed patients. In recent years, the increase in non-albicans species has altered the epidemiological landscape, requiring updated local surveillance data. Therefore, this study aims to analyze the epidemiological pattern of Candida spp. involved in various clinical manifestations. Methods: This was a retrospective observational descriptive study of clinical Candida isolates recovered from patients at a single tertiary care hospital in Puebla, Mexico, between 2021 and 2022. Sociodemographic and clinical data were collected from medical records. Candida spp. isolates were recovered from the hospital’s clinical specimen collection and identified using conventional methods and phylogenetic analysis. Results: Ninety isolates were recovered from a total of 88 patients during the study period. The majority of patients were women (60.2%), and among the age groups, the largest proportion corresponded to older adults (≥60 years, 25%). Candida was most frequently isolated from the urinary tract (47.7%), followed by the respiratory tract (19.3%) and the vulvovaginal tract (14.7%). Candida albicans was the predominant species (51.1%), followed by Nakaseomyces glabratus (32.2%) and other species such as Pichia kudriavzevii (5.5%), Candida tropicalis (3.3%), Candida dubliniensis (3.3%), Candida parapsilosis (2.2%), and Candida orthopsilosis (1.1%). The study showed discrepancies between conventional identification methods and molecular analysis. Of the 33 isolates evaluated using culture methods, only 21 matched the identification by PCR and phylogenetic analysis. In the case of the BD Phoenix automated system, 20 matches with and 13 discrepancies from the molecular tests were recorded. Conclusions: Our findings revealed the significant presence of species belonging to the non-albicans family in specific clinical contexts, highlighting the urgent need to monitor the epidemiological profile. The observed discrepancies between phenotypic and molecular identification methods suggest that species identification based solely on phenotypic approaches should be interpreted with caution.