1H NMR-based metabolomic study of a mother-infant dyad on the evolution of gut microbiota during the breastfeeding and weaning

Human milk is a buffet of fats, proteins and sugars, designed to be easily digestible and to provide an adequate supply of nutrients to support healthy growth and development of the infant. In addition, milk is rich of nonnutrient bioactive factors, such as cells, anti-infectious and anti-inflammatory agents, growth factors, and prebiotics, providing protection to the newborn. Among this plethora of components, human milk oligosaccharides (HMOs) were supposed to have anti-adhesive and antimicrobials properties, thus preventing pathogen attachment to infant mucosal surfaces and lowering the risk for viral, bacterial and protozoan parasite infections [1]. Furthermore, HMOs were recognized as a substrate to promote the growth of desired bacteria, such as bifidobacteria, in the infant’s intestine (“bifidogenic” or prebiotic effects) [2]. Early microbial colonization is essential for healthy intestinal and immunological development in neonates. The development of gut microflora during infancy consists in a complex succession of bacterial species, modulated by breastfeeding, perinatal antibiotic use and environmental factors. In a recent study, we have compared the changing fecal microbial composition with the fecal metabolome of newborns in the first thirty days of the breastfeeding [3]. The results showed a relation between microbiota and fecal biochemical composition, probably linked to the changes of breast milk composition during the maturation process.In this study, we have investigated, by 1H NMR spectroscopy, the changes of the neonatal fecal metabolome in relation to diet: breastfeeding and weaning. We have examined breast milk and newborn faeces of a single mother-infant dyad for 6 months, starting from the 4th month after the birth, in order to evaluate the changes in microbial metabolome during the diet evolution. The NMR spectroscopy of the breast milk allowed to recognize the mother’s phenotype as Secretor (FUT2 gene positivity) on the basis of the structure of fucosyl-HMO, and to confirm the stability of milk biochemical composition during the considered period of lactation. On the other hand, the NMR analysis of fecal samples showed that the microbiota HMO metabolism changed as a function of time, and furthermore the daily variability increased during the weaning period. Therefore, important experimental aspects are evidenced in order to obtain representative samples of faeces from newborns to compare metabolomics with metagenomics data. The results display the evolution of the infant gut microbiota metabolism in relation to changes of the diet composition.