The clinical study reported here was conducted in order to analyze the anti-obesity effect of the steamed root of Rehmannia glutinosa Libosch (Shu Dihuang), a Chinese herbal medicine widely used for the treatment of metabolic diseases. To understand the mechanism(s) underlying the beneficial effects of R. glutinosa, we investigated the changes in gut microbiota after herbal intervention through 16S rRNA gene-based pyrosequencing analyses. Twelve female middle-aged subjects (40-65 years old) with body mass index (BMI) over 25 kg m(-2) were included in this study which involved intake of steamed roots of R. glutinosa for eight weeks. The subjects were visited for anthropometric measurements every two weeks and fecal samples were collected before and after the intervention. All subjects showed a significantly decreased waist circumference following the herbal treatment. The relative abundances of fecal microbiota suggested an association of intake of R. glutinosa with increases in phylum Actinobacteria and genus Bifidobacterium, while phylum Firmicutes and genus Blautia were decreased in response to the herbal treatment. Based on these findings, it is conceivable that a decrease in the waist circumferences after R. glutinosa intake may be related to alteration of intestinal microbiota and the herbal intervention has the potential as a prebiotic.