Background: Gestational diabetes mellitus (GDM) is one of the commonly occurring high-risk obstetric complications that accounts for 4%–9% of total pregnancies. The present study was an attempt to assess the effect of GDM on composition of the neonatal oral microbiota. Materials and Methods: In this study, oral samples from 155 full-term vaginally delivered newborns were collected with sterile swabs. Seventy-five mothers diagnosed with GDM group and 80 were nondiabetic mothers (control). The oral microbiota was evaluated and analyzed by SPSS software. Results: The mean gestational age in Group I was 38.1 weeks and in Group II was 39.6 weeks. Firmicutes was present in 38.1% in Group I versus 77.6% in Group II patients, Actinobacteria was seen in 15.2% in Group I and 7.4% in Group II, Bacteroidetes in 27.6% in Group I and 7.9% in Group II, Proteobacteria in 9.5% in Group I and 3.8% in Group II, and Tenericutes in 9.6% in Group I and 3.3% in Group II. There was a significant difference in major genera Prevotella, Bacteroidetes, Bifidobacterium, Corynebacterium, Ureaplasma, and Weissella in both groups (P < 0.05). Conclusion: There was increased bacterial microbiota in neonates born to mothers with GDM as compared to neonates born to nondiabetic mothers. Assessment of initial oral microbiota of neonates could help in assessing the early effect of GDM on neonatal oral microbial flora.