UNLABELLED: The colonization and development of the gut microbiota during early life, especially , may be influenced by maternal bacterial communities, including those of human milk. However, the interaction of bacteria in mother-infant dyads during breastfeeding remains unclear. This study focused primarily on the characteristics and dynamics of the infant gut and human milk microbiota within the first month of life on the basis of a birth cohort and explored the interaction of the microbiota derived from the two niches by sequencing and culture-based methods, especially as the representative dominator in the infant gut. Infant feces and human milk samples from 21 mother-infant dyads were collected on days 0, 7, and 30 postpartum. The bacterial composition was identified by sequencing the 16S rRNA gene, and the contributions of the bacterial communities were estimated via SourceTracker2. Bifidobacterial strains were isolated from infant feces and human milk via culture-based methods. The suspected strains were identified through Sanger sequencing and genotyped via multilocus sequence typing (MLST). The bacterial communities were distinct between infant feces and human milk. Human milk microbes contribute 63.89%-77.61% to the infant's gut within the first month of life, whereas in the infant's gut contributes more (80.18%-84.30%) to human milk. A total of 60 bifidobacterial isolates were obtained from 10 pairs of mother-infant samples, 48 isolates from 10 out of 27 infant feces samples, and 12 isolates from 4 out of 27 human milk samples. Among these, 30 isolates were identified as and 18 were identified as subsp. . Strains belonging to from a single mother-infant pair were found to be monophyletic (ST: BRE-1), whereas this strain was found much earlier in infant feces across the three time points (collected on days 0, 7, and 30) than in human milk (collected on day 30). Our data suggest that during very early breastfeeding, human milk contributes a significant proportion of the overall bacterial population to the infant's gut, whereas the infant's gut selectively contributes a greater proportion of to human milk. Certain bifidobacterial strains, such as , are retrogradely transmitted from the infant's gut to the mother's human milk during breastfeeding, implying a potential challenge regarding the reliability of the source when potential probiotics are isolated from human milk. IMPORTANCE: Understanding how microbes, especially beneficial bacteria such as are shared between mothers and infants during breastfeeding is crucial for promoting infant health. Although most research has focused on transmission from mother to child, our study reveals a novel and significant reverse route: from the infant gut to breast milk. By combining microbiome sequencing with culture-based techniques, we provide evidence that specific strains of , especially , may transmit back to the mother during breastfeeding. This insight reshapes our understanding of microbial exchange within the mother-infant dyad and highlights breastfeeding as a bidirectional process that influences both maternal and infant microbiota. These findings may have important implications for designing probiotics and supporting early-life microbial development through maternal health interventions.