Muscovy duck reovirus (MDRV) is highly pathogenic to young Muscovy ducklings. Although MDRV infection results in ducklings' acute watery diarrhea, the effect of MDRV infection on the composition of host's intestinal microbiota remains poorly understood. This study was conducted to investigate the impacts of MDRV on the composition of Muscovy ducklings' intestinal bacterial community. Three-day-old Muscovy ducklings were inoculated with either the virulent MDRV strain MW9710 or sterile Hank's solution, respectively. The cecal microbiota was analyzed between control and mock MDRV-infected ducklings using Illumina MiSeq sequencing at 6 dpi and 17 dpi, respectively. The results indicated that MDRV infection damaged the intestinal mucosa. In addition, MDRV infection caused severe perturbations of gut microbiota by decreasing microbial richness, altering the abundance of certain genera of the gut microbiota at 6 dpi. Specifically, the relative abundance of short chain fatty acids-producing bacteria (including Shuttleworthia, Streptococcus, and Ruminococcus) was reduced in MDRV-infected ducklings than those of control group, whereas, with an enrichment of Enterobacteriaceae (including Plesiomonas, Escherichia_Shigella and Proteus). Furthermore, microbiota analysis showed that the gut microbiota dysbiosis caused by MDRV infection was basically recovered at 17 dpi. Collectively, this study demonstrated that the gut microbiota of Muscovy ducklings were altered due to MDRV infection, mainly featuring as a net loss of beneficial bacteria and a compensatory proliferation of pathogenic bacteria, which may lead to severe pathology to the intestinal mucosa, and ultimately acute diarrhea. These results will provide insights into the pathology of MDRV infection.