A meta-analysis was conducted on the correlation between intestinal flora and gestational diabetes mellitus (GDM) to provide a theoretical basis for the treatment of GDM. The PubMed, Embase, MEDLINE, Ovid, Springer, and Web of Science databases were searched from the establishment of the databases to December 31, 2020, to retrieve randomized control trials (RCTs) involving GDM patients. The Cochrane Handbook for Systematic Reviews of Intervention 5.0.2 was used to assess the bias risk of the included articles, and Rev Man 5.3 was used for the meta-analysis. A total of 7 studies were included in the meta-analysis, comprising 665 study participants. The meta-analysis results showed that the GDM patients in the experimental group had a lower level of Bifidobacterium [MD (mean difference) =-2.49; 95% confidence interval (CI): -3.54 to -1.45; Z=4.66; P<0.00001], Lactobacillus [MD =-1.69; 95% CI: -1.84 to -1.53; Z=20.66; P<0.00001], Bacteroides [MD =-1.17; 95% CI: -1.45 to -0.89; Z=8.15; P<0.00001], Bacteroidetes [MD =-1.22; 95% CI: -1.71 to -0.72; Z=4.81; P<0.00001], and a higher level of Enterobacter [MD =1.79; 95% CI: 1.13 to 2.45; Z=5.3; P<0.00001], Enterococcus [MD =1, 29; 95% CI: 0.98 to 1.6; Z=8.06; P<0.00001], Fusobacterium [MD =0.03; 95% CI: -0.13 to 0.19; Z=0.37; P=0.71], tumor necrosis factor alpha (TNF-α) [MD =113.66; 95% CI: 52.01 to 175.31; Z=3.61; P=0.0003], interleukin (IL)-17 [MD =37.92; 95% CI: 29.74 to 46.1; Z=9.09; P<0.00001], and IL-6 [MD =66.38; 95% CI: 33.6 to 99.15; Z=3.97; P<0.0001] than those in the control group; however, no statistically significant difference was found in relation to Fusobacterium between the experimental group and the control group. Intestinal microecological changes are closely related to the occurrence of GDM in our study, which manifested as a decrease in the level of probiotics, an increase in the level of intestinal bacteria and other strains, and an increase in the level of inflammatory factors. Thus, special attention should be paid to changes in patients' intestinal flora to prevent GDM.