Background <p><i>Candida albicans</i> is a major cause of invasive fungal infections in pediatric patients, including subgroups of neonates and young infants, particularly in intensive care settings where premature and low birth weight infants are at high risk. Management is increasingly complicated by antifungal resistance, notably to echinocandins. Understanding basal <i>FKS1</i> expression, antifungal susceptibility, and virulence traits in neonatal isolates, as a subgroup within pediatric patients, provides preliminary insights for clinical management.</p> Methods <p>A cross-sectional study was conducted on pediatric patients with suspected fungal infections admitted to NICU, PICU, IICU, and other specialized wards of the Children’s Medical Center, Tehran, Iran. Clinical specimens were collected from multiple sources including urine catheters, blood, abscesses, bronchoalveolar lavage, and other body fluids. Of 62 <i>Candida</i> isolates recovered, 50 (80.6%) were identified as <i>C. albicans</i>. For detailed molecular, antifungal susceptibility, enzymatic virulence, and <i>FKS1</i> expression analyses, 20 isolates from children under two years of age (7 newborns and 13 infants) were selected. Species identification was confirmed by ITS sequencing. Antifungal susceptibility testing followed CLSI M27-A4 documents with MICs determined based on 50% growth inhibition. Virulence was assessed via phospholipase and proteinase enzymatic activity, and <i>FKS1</i> gene expression was measured using real-time PCR.</p> Results <p>All 50 analyzed isolates were confirmed as <i>C. albicans</i> with &gt; 99% ITS sequence identity. Phylogenetic analysis revealed strong genetic relatedness among isolates. Caspofungin and amphotericin B demonstrated the highest in vitro efficacy, with 44 isolates (88%) susceptible to caspofungin and 27 isolates (54%) susceptible to amphotericin B. Resistance to fluconazole and ketoconazole was observed in 28 (56%) and 33 (66%) isolates, respectively. Virulence profiling showed that a subset of pediatric isolates, including neonatal cases, exhibited strong phospholipase and proteinase activity, suggesting elevated pathogenic potential within this subgroup. <i>FKS1</i> was detected in all isolates, with expression levels varying significantly; notably, two neonatal isolates showed higher basal <i>FKS1</i> expression, potentially indicating early adaptive responses to echinocandin exposure despite phenotypic susceptibility. All subsequent analyses, including molecular and functional assays, explicitly indicate the number of isolates evaluated for each experiment, ensuring clarity and reproducibility.</p> Conclusion <p>This study highlights the multifactorial pathogenicity of <i>C. albicans</i> in pediatric patients, with exploratory observations in neonates and young infants characterized by variable antifungal resistance and virulence factor expression. Variations in basal <i>FKS1</i> expression are described as preliminary findings. These data underscore the value of integrating molecular diagnostics, antifungal susceptibility testing, and virulence profiling to guide therapy, without making unsupported claims about echinocandin tolerance.</p>