Abstract:[Background] Bifidobacterium bifidum, an obligate intestinal bacterium that specifically metabolizes human milk oligosaccharides (HMOs) and mucin glycans of host gut epithelium, is essential for early life health and development. However, little is known about the genetic differences of this bacterium among different populations. [Objective] To investigate whether the B. bifidum strains from genetically similar populations with similar diets in a limited geographic area have population-specific regularities and to reveal the genetic differences of B. bifidum among different populations. [Methods] A total of 115 strains of B. bifidum were isolated and identified from school-age children of Uyghur and Kazak ethnic minorities in Yining, Xinjiang. Further, we performed multilocus sequence typing (MLST) for 53 representative strains screened out by rep-PCR to understand the genetic differences of B. bifidum between different ethnic groups. [Results] The 53 strains belonged to 37 sequence types (STs), showing high genetic diversity. Among them, 17 STs were identified for 26 isolates from the children of Uyghur ethnic minority, while 20 STs for 27 isolates from the children of Kazak ethnic minority. Only a few homologous genetic recombination events were detected between strains from both ethnic groups. goeBURST showed that B. bifidum isolates from the same population were more likely to be assigned to a specific phylogenetic clade or clonal complex than the isolates from the other population. [Conclusion] B. bifidum isolates from different ethnic groups showed high genetic diversity. The host (ethnic group) specificity of their genetic structure needs to be confirmed by larger sampling. Our findings provide a theoretical basis for further screening of elite probiotic strains tailored to localized population by in vivo and in vitro experiments.