Bifidobacterium breve is one of the dominant Bifidobacterial species in children. In the current work, 46 strains of B. breve isolated from fecal samples of Chinese children were analyzed using whole-genome sequencing and comparative genomics to explore their genetic diversity, as well as genotype and phenotype analysis for carbohydrate utilization and antibiotic tolerance. The phylogenetic tree was independent of region, age and feeding mode, and without any regularity in the clustering of carbohydrates and antibiotics at the genetic level. Based on genotypic-phenotypic correlation analysis, the diversity of glycosyl hydrolases and the ability of strains to metabolize carbohydrates corroborated the predominance of B. breve in the children's intestines. Simultaneously, the sensitivity of strains to antibiotics increased the understanding of its genetic features and provided a potential basis for safety evaluation.