Coilia nasusis influenced by various external pressures during spawning migration and these anadromous transitions can lead to specific gut microbiome characteristics that affecting the host biological process. Therefore, the purpose of this study was to determine the variations of components and functions in the gut prokaryotic microbiome during spawning migration as well as the key factors that triggered the changes. The gut microbiome inC. nasuswas mainly consisted ofProteobacteria,Bacteroidetes,Firmicutes,Deinococcus-ThermusandFusobacteriavia 16S rRNA Gene Amplicon Sequencing. The relative abundance ofAcinetobacterandClostridiumincreased, whileCorynebacterium,Actinomyces,Bacillus,KlebsiellaandOchrobactrumdecreased after entering freshwater, indicated the preference ofC. nasusgut microbial members transferred from seawater to freshwater. Additionally, the proportion ofFirmicutessignificantly decreased and then increased, as well as the arise of some soil bacteria in gut,corresponding to the phenomenon thatC. nasusare fasting during the upstream process and refeeding after entering the spawning grounds. The function prediction of gut microbiome was also consistent with the above results. The present study generally demonstrated the gut microbiome dynamics and the significant correlation between the gut microbiome and salinity and feeding behavior in the spawning migration ofC. nasus.