The presence of Escherichia coli (E. coli) in river and seawater may cause different levels of infections, and constitutes a risk to public health. In this study, water samples were collected from 15 sites along the Kelantan River, estuaries and its adjacent coastal waters to investigate the prevalent and diversity of E. coli. Membrane filtration technique was used to enumerate. E. coli and phylogenetic grouping was performed using triplex PCR. E. coli abundance ranged from 3.1 × 10 to 1.6 × 105 CFU 100 mL−1, and total suspended solids correlated significantly with E. coli abundance (r2 = 0.165, p < 0.001) and rainfall (r2 = 0.342, p < 0.001). Phylogenetic group B1 and A (59.4%) were the most prevalent whereas groups B2 and D were least abundant. The higher abundance of phylogenetic group D at upstream sites of Kelantan River suggested fecal contamination mainly of animal origin. Canonical-correlation analysis showed phylogenetic group B2 Phylogenetic groups A and D were greater in waters with higher inorganic nutrients e.g. NH4, NO2 and NO3 whereas phylogenetic group B1 appeared to have better salinity tolerance amongst phylogenetic groups.