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Jamuna River is one of the vital rivers in Bangladesh for its rich aquatic biodiversity. A total of 52 fish species were recorded during the study period, belonging to 9 orders, 20 families, from the three selected locations of the river. Fish populations are the highest during the dry winter season and the lowest during the monsoon season. The mean values of Shannon-Weaver diversity, Margalef's richness, and Pielou's evenness indices recorded as 3.29± 0.02, 0.58±0.01, and 7.00±0.10, respectively, varied from season to season. Nonmetric MDS and Canonical Correspondence analysis were used to evaluate the effect of environmental factors on the structure of the fish assemblage. The Nonmetric Multidimensional Scaling (nMDS) had attained in two groups in the cluster analysis, one of which showed the association of the fish samples between the summer and winter seasons, and the other included the species from the monsoon season among the sampling sites. The fish species responsible for the fluctuation in seasonal abundance up to a cumulative dissimilarity of 21.85%, and four species were responsible for this dissimilarity. Canonical correspondence analysis on the fish community was used to demonstrate the fascinating functions of alkalinity, total dissolved solids, electrical conductivity, and dissolved oxygen in Jamuna River.