In the present study, we investigated the effect of different garlic concentrations on growth performance and intestinal microbiota of rainbow trout (Oncorhynchus mykiss). A total of 240 healthy rainbow trout were fed diets containing 0% (control), 1% (G1), 1.5% (G2) and 2% (G3) garlic extract for 120 days. At the end of the study, weight gain and specific growth rate of fish were significantly higher among garlic-supplemented groups compared to the control group. Moreover, genomic DNA was isolated from the intestinal mucus and the V4 region of the 16S rRNA gene was amplified and sequenced on the Illumina MiSeq platform. The results demonstrated that the most abundant operational taxonomic units (OTUs) were affiliated to the genera Deefgea (15.1%) and Aeromonas (17.4%) in the control group, whereas members belonging to the genera Deefgea (20.5-20.9%) and Mycoplasma (36.3-24.3%) were dominant in the G1 and G2 groups, respectively. Members belonging to the genus Aeromonas (44.6%), and to a lesser extent, the genera Deefgea (13.9%) and Exiguobacterium (14.8%) were dominant in the G3 group, which received the highest level of garlic in the diet. In conclusion, dietary garlic supplementation may promote growth performance and induce changes in the intestinal microbiota, conferring beneficial effects to the host.