ObjectiveTo examine gut microbiotal diversity in the Han Chinese and Yugur populations of Sunan County, Gansu Province, living in the same environmental conditions, and to analyze possible causes of differences in diversity.MethodsWe selected 28 people, ages 18–45 years old, all of whom were third-generation pure Yugur or Han Chinese from Sunan County. Fresh fecal samples were collected, and total bacterial deoxyribonucleic acid (DNA) was extracted. We performed 16S ribosomal ribonucleic acid (16S rRNA) high-throughput sequencing (HTS) and bioinformatics to study the relationships among between gut microbiota structure, genetics, and dietary habits in Yugur and Han Chinese subjects.ResultsWe found 350 differential operational taxonomic units (OTUs) in Han Chinese and Yugur gut microbiota, proving that gut microbiota differed between the two populations. That were less abundant among Yugurs than Han Chinese werePrevotella_9andAlloprevotella. That were more abundant among Yugurs than Han Chinese wereAnaerostipesandChristensenellaceae_R-7_group. And they were significantly associated with a high-calorie diet In addition. we found differences in predicted gut microbiota structural functions (The main functions were metabolic and genetic information) between the two populations.ConclusionYugur subjects demonstrated differences in gut microbiotal structure from Han Chinese subjects, and this difference influenced by dietary and may be influenced by genetic influences. This finding will provide a fundamental basis for further study of the relationships among gut microbiota, dietary factors, and disease in Sunan County.