The main objectives of this study were to assess the diversity of diatom species and explain how physico-chemical factors and trophic levels interact with diatom assemblages. An assemblage of diatom communities in a riverine environment was assessed through forthright sampling for a period of 7 months from September 2019 to March 2020. Three sampling sites from the upstream, midstream and downstream parts of a river located in the northwest part of Bangladesh were selected for water quality analysis. A variety of multivariate data analyses were done to evaluate the environmental and biological data as well as to observe their interrelationships. A total of 7 genera of Diatom namely Nitzschia, Diatoma, Melosira, Synedra, Tabillaria, Navicula, and Fragillaria were recorded. Nitzschia was the most dominant genus compared to the other genus at downstream with an abundance of 1.92× 10 3 cells/L while Tabillaria sp. was noted as a rare one with the lowest abundance of 0.37× 10 3 cells/L at the midstream site. The interrelationship analysis clarified that water temperature positively altered the distribution of Melosira, Navicula, and Tabillaria while its abundance was less dependent on dissolved oxygen and pH. The results from a 2-dimensional Nonmetric Multidimensional Scaling (NMS) revealed the abundance of diatom at downstream was statistically higher than upstream and midstream. Hence, the study concluded that the diatom composition was very sensitive to the physico-chemical parameters according to water direction and could be used as a bio-indicator to assess the surface water quality.