Cyprinidae (Teleostei: Cypriniformes) is the predominate family among Bhutan's freshwater fishes, yet significant gaps exist in the knowledge of their diversity and assemblage. Cyprinids were sampled from 54 plots across four river basins (Aiechhu, Jaldakachhu, Mangdechhu, and Punatsangchhu) using systematic sampling at an interval of 5 km. A total of 850 individuals belonging to 22 species and 13 genera were recorded. Aiechhu had the highest diversity and evenness (H’= 1.66±0.28, J’= 0.95±0.03), followed by Punatsangchhu (H’= 1.58±0.34, J’= 0.94±0.04). Meanwhile, Jaldakachhu and Mangdechhu exhibited the lowest diversity and evenness (H’= 0.90±0.66, J’= 0.84±0.54 and H’= 0.54±0.70, J’= 0.38±0.49, respectively). There was a significant difference in diversity among river basins (χ2 (3)= 20.98, p < 0.001) with specific variation between Mangdechhu and Punatsangchhu (Z= 3.80, p= 0.00), Mangdechhu and Aiechhu (Z= 3.35, p= 0.01), and Jaldakachhu and Punatsangchhu (Z= 2.83, p= 0.03). Canonical correspondence analysis indicated that cyprinid assemblage was significantly correlated with environmental variables (r= 0.94, p= 0.001 in axis 1 and r= 0.82, p= 0.001 in axis 2) explaining 77% of variance. Axis 1 was positively correlated with temperature (r= 0.92), total dissolved solids (r= 0.53), conductivity (r= 0.51), and salinity (r= 0.39) and negatively correlated with elevation (r= −0.69). Axis 2 was negatively correlated with total hardness (r= −0.82) and dissolved oxygen (r= −0.65). Cluster analysis identified three clusters based on dominant species. Further studies exploring other river basins of Bhutan are needed to better understand the ecological dynamics of cyprinids in Bhutan.