Human milk and infant formula influence lactobacilli metabolism in a species-specific manner

The neonatal gut is colonized by commensal bacteria, including lactobacilli, that play a critical role in promoting host health. While lactobacilli are known for producing bioactive metabolites, species-specific responses to neonatal diets remain unclear. We hypothesized that human milk and infant formula would differentially influence lactobacilli metabolite production. Six species (Lactobacillus acidophilus, Levilactobacillus brevis, Lactobacillus johnsonii, Lacticaseibacillus paracasei, Limosilactobacillus reuteri, Lacticaseibacillus rhamnosus) were cultured in defined media with human milk, formula, or water, and supernatants were analyzed via untargeted LC-MS/MS metabolomics. Principal coordinates analyses showed distinct metabolomic profiles among lactobacilli species and across dietary treatments. Human milk enhanced the production of several metabolites, including hydroxyphenyllactic acid, tyrosine, indoles, vaccenic acid, and di-peptides. Interestingly, several unique metabolites were upregulated in the presence of infant formula, including isonicotinic acid. These results highlight the differential effects of human milk and infant formula on the metabolomic profiles of lactobacilli species, emphasizing significant species-specific variation and pronounced production of potentially beneficial compounds in response to human milk. This work underscores the importance of understanding diet-microbe interactions to optimize neonatal gut health.