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This study aimed to explore the effect of exercise or tai chi on Internet addiction disorder (IAD) among college students and clarified the abundance and population changes of gut microbiota in different groups. Thus explore the potential role of gut microbiota between exercise and IAD. A total of 93 subjects diagnosed with mild IAD were randomly assigned to the exercise group, the tai chi group, and the control group. The intervention groups received exercise or tai chi for 8 weeks and the control group was evaluated without any intervention. Fecal samples were collected after the intervention. 1) Analysis found a significant intervention effect with the exercise group showing an average decrease of 8.84 points on the Internet addiction test (IAT) compared with the control group (95%CI −15.41 to-2.27, P = 0.004). But there was no significant difference between the control group and the tai chi group. 2) Both exercise (P = 0.018) and tai chi (P = 0.026) could significantly relieve fatigue symptoms. 3) The relative abundance of the Betaproteobacteria, Porphyromonadaceae, Sutterellaceae, and Alistipes were significantly decreased in the exercise group compared with the control group, and the relative abundance of Escherichia was significantly increased in the exercise group. 4) The relative abundance of Betaproteobacteria, Sutterellaceae, and Escherichia had significant differences between the improved group and the no-improved group. Exercise intervention has a considerable effect on treating IAD. Exercise and tai chi might have effectiveness in relieving the symptoms of fatigue. Exercise intervention regulates the gut microflora and changes the abundance of microflora to improve IAD. ClinicalTrials.gov , identifier NCT05529368. • This study was a randomized, 3-arm, parallel clinical trial. • Exercise could significantly relieve the symptoms of Internet addiction (IA). • Dizziness and fatigue symptoms improved significantly after intervention. • Exercise regulates the gut microflora to improve IA. • The improvement of IA was associated with the change of the gut microbiota. [ABSTRACT FROM AUTHOR]