Research on the gut microbiota in irritable bowel syndrome (IBS) shows discordant results due to inconsistent study designs or small sample sizes. This study aimed to characterize how gut microbiota in IBS patients differs from that in healthy controls by performing a case-control study and cross- and mega-cohort analysis. Multiple publicly shared data sets were examined by using a unified analytical approach. We performed 16S rRNA gene (V3-4) sequencing and taxonomic profiling of the gut bacterial communities. Fecal samples from children with IBS ( = 19) and age-matched healthy controls ( = 24) were used. Next, we analyzed 10 separate data sets using a unified data-processing and analytical approach. In total, 567 IBS patients and 487 healthy controls were examined. In our data sets, no significant differences existed in stool α-diversity between IBS patients and healthy controls. After combining all the data sets using a unified data-processing method, we found significantly lower α-diversity in IBS patients than in healthy controls. In addition, the relative abundance of 21 bacterial species differed between the IBS patients and healthy participants. Although the causal relationship is uncertain, gut bacterial dysbiosis is associated with IBS. Further functional studies are needed to assess whether the change in gut microorganisms contributes to the development of IBS. Research on the gut bacteria in irritable bowel syndrome (IBS) shows discordant results due to inconsistent study designs or small sample sizes. To overcome these issues, we analyzed microbiota of 567 IBS patients and 487 healthy people from 10 shared data sets using a unified method. We demonstrated that gut bacteria are less diverse in IBS patients than in healthy people. In addition, the abundance of 21 bacterial species is different between the two groups. Altered bacterial balance, called dysbiosis, has been reported in several disease states. Although the causal relationship is uncertain, gut bacterial dysbiosis also seems to be associated with IBS.