Candida species are the utmost prevalent opportunistic fungal pathogens in the immune-compromised patients and bone marrow transplants patients, as well as in Children suffering from orofacial birth defects called Orofacial Clefts that affect the lip, palate, or both, and are more prone to oral candidiasis. Consequently, this study aimed to identify Candida dubliniensis and its virulence gene, Candida dubliniensis mitochondrial gene using various culture media, PCR, and ELISA. This study was extended between January 2023 to August 2024 at Al-Diwaniya Children’s Hospital /Iraq. A 75 oral swab and blood samples were collected from cleft lip/palate patients (with and /without candidiasis). Candida dubliniensis was identified via culture om CHROM and Sabouraud agar and PCR targeting the Cdub gene. Serum IL-17/IL-23 levels were measured by ELISA. Statistical analysis used SPSS (chi-square). Among the tested specimens (57.33%) were positive for C. dubliniensis, while (42.67%) were negative. All C. dubliniensis isolates from patients with orofacial clefts carried the Candida dubliniensis mitochondrial gene, whereas no mitochondrial gene was detected in patients without candidiasis. The cytokine levels revealed that oral candidiasis significantly increased the concentration of cytokine Interleukin-17 to 115.91 ± 15.42 pg/ml, in comparison to 68.85 ± 10.59 pg/ml and 21.58 ± 2.33 pg/ml in orofacial cleft patients without candidiasis and the control group respectively. Similarly, Interleukin -23 levels were significantly elevated to 77.28 ± 8.94 pg/ml in the candidiasis group, compared to 53.84 ± 8.93 pg/ml and 8.45 ± 2.46 pg/ml in orofacial cleft patients without candidiasis and the control group respectively. In Conclusion: This study demonstrates a relatively high prevalence of Candida dubliniensis in children with orofacial clefts, with a significant association between the presence of the Candida dubliniensis mitochondrial gene and candidiasis. The study also approved the immunological impact of C. dubliniensis infections in orofacial clefts patients with Candida dubliniensis, as evidenced by significantly elevated levels of Interleukin-17 and Interleukin -23 compared to non-candidiasis orofacial cleft patients and control group.