IntroductionOwing to urbanization, living habits have changed widely, leading to alterations in the intestinal microbiota of urban residents. However, there are few studies on the characteristics of intestinal microbiota of adolescents living in different urbanized areas in China.MethodsA total of 302 fecal samples collected from adolescent students in eastern China were examined. 16S rRNA high-throughput sequencing was used to identify the fecal microbiota. These data were combined with questionnaire survey results to investigate the effect of urbanization on the intestinal microbiota of adolescents in eastern China. Moreover, the role of lifestyle habits in this relationship was also evaluated.ResultsThe results revealed significant differences in the structure of the intestinal microbiota among adolescents living in regions with different levels of urbanization. Adolescents living in urban regions had a significantly higher proportion ofBacteroides(p<  0.001, FDR = 0.004), whereas those living in towns and rural regions had higher proportions ofBifidobacterium(p< 0.001, FDR < 0.001) andPrevotella(p< 0.05, FDR = 0.019). The diversity of the intestinal microbiota was higher in urban residents than in adolescents living in towns and rural regions (p<  0.05). In addition, the differences in intestinal microbiota across individuals living in cities, towns, and rural regions were related to dietary preferences, flavor preferences, and sleep and exercise durations. Adolescents who ate more meat had moreDorea(LDA = 3.622,p= 0.04), while the abundance ofEscherichia–Shigellais higher among adolescents who ate more condiments (LDA = 4.285,p= 0.02). The abundance ofDialisterwas significantly increased in adolescents with longer sleep durations (LDA = 4.066,p =0.03). Adolescents who exercised for a long duration had moreFaecalibacteriumthan those who exercised for a shorter duration (LDA = 4.303,p= 0.04).DiscussionOur research has preliminarily demonstrated that there were differences in the composition of Gut microbiome in stool samples of adolescents living in different urbanized areas, and provide a scientific basis for the maintenance of a healthy intentional microbota in adolescences.