Oral microbiota assessment at different time points during the first year of life: delivery mode and breastfeeding shape the bacterial community

Abstract

Background: The assembly of the oral microbiota during the first year of life is a critical process that may influence long-term health and immunological development. While the gut microbiome has been extensively studied, early oral colonization patterns remain less understood. This study aimed to characterize the oral microbial succession in infants and to evaluate the impact of two primary clinical factors—delivery mode and breastfeeding practices—on microbial composition from birth through the first 12 months of life. Methods: Forty-eight newborns were prospectively recruited between January and June 2022 through the Basque Health Service (Osakidetza, Spain). Oral samples were collected at five time points (4–10 days, 1 month, 4 months, 6 months, and 12 months). Feeding status was categorized as exclusive breastfeeding (EBF) or mixed feeding (MF) for the first 4 months, after which the EBF group transitioned to a “Breast milk -No Formula” (BM-NF) group to account for the complementary feeding. Microbial communities were assessed via 16S rRNA amplicon sequencing. To compare alpha diversity (Shannon, Chao1, ACE, Simpson, Observed, and Fisher indices) between groups, Mann–Whitney or Kruskal–Wallis tests were employed. Beta diversity was evaluated via non-metric multidimensional scaling (NMDS) based on Bray-Curtis distances. PERMANOVA was employed to assess community shifts over time, and relative abundance differences across delivery and lactation type´s groups were analyzed using Mann–Whitney tests. Results: Oral bacterial diversity increased significantly over time as the oral cavity transitioned towards a more complex community. Streptococcus, Ralstonia, Gemella, and Rothia were the primary early colonizers. Significant taxonomic shifts were associated with birth and feeding modes. C-section infants exhibited a higher abundance of Veillonella and Neisseria during the first days of life. Regarding nutrition, mixed feeding (MF) infants showed higher proportions of Rothia, Prevotella, and Veillonella during the initial months. Conversely, Streptococcus remained significantly predominant in infants receiving breast milk (EBF/BM-NF) at the 6-month mark. Conclusions: These findings demonstrate that delivery mode and lactation significantly shape the neonatal oral ecosystem. Specific microbial signatures up to 12 months suggest that early-life factors and feeding choices are key in establishing the mature oral microbiota.