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Introduction: Infant formulas with reduced iron levels and lactoferrin (Lf) supplementation might mimic the beneficial effects of breast milk on the oral microbiome. This study aimed to investigate the impact of a bovine Lf-supplemented and iron-reduced formula on the oral microbiota in infants at 4, 6 and 12 months.

Methods: In a double-blind controlled trial, 6-week-old formula-fed infants were randomized to receive either a formula with reduced iron levels (2 mg/L) and Lf supplementation (1 g/L) (n = 72), the same formula without Lf (n = 72), or a standard formula (8 mg iron/L) (n = 36). A breast-fed reference group (n = 72) was also included. The oral microbiota was analyzed at 4 (n = 244), 6 (n = 216) and 12 (n = 229) months of age using the Oxford Nanopore Technology of the 16S rRNA gene annotation (eHOMD database).

Results: Neither the within- or between-group diversities nor overall microbiota pattern assessment revealed any statistically significant differences in microbiota composition between the formula groups. However, single species were significantly associated with specific formula-fed groups. At 6 months, breast-fed infants exhibited significantly lower species richness and distinct microbiota composition compared to the formula-fed groups.

Conclusions: The effects of reduced iron levels and lactoferrin supplementation of infant formula on the oral microbiome were inconclusive.