The Yellow River flows through Lanzhou city and is the only drinking water source for 3.6 million residents. Yet, little is known regarding the safety and quality of the Yellow River for resident consumption. To address this knowledge-gap, water samples were collected from different sites within this section during the dry and wet seasons. Physico-chemical parameters and microbial community metrics were analyzed to assess the health risk associated with this Chinese mother river. Water quality of the river was better during the dry season (March–April) than in the wet season (September–October). Fifteen conventional physico-chemical and biological indices, such as NH3-N, NO2−-N, total nitrate (TN), five day biochemical oxygen demand (BOD5), chemical oxygen demand (COD), volatile phenol (VP), and coliform abundances, generally exceeded acceptable standards. The average abundance of coliforms was 2.8 times that of acceptable standards in the dry season and 4.6 times the standards in the wet season. The concentration of the toxic metal As was more than two times than that of the national standard in waters from the wet season. Microbial community analysis also indicated that community diversity and species richness were positively correlated with the concentrations of several physico-chemical parameters. The results indicate that As and Cr6+ pose potential risk for human health through consumption by residents. Further, the results indicated that human activities are the main causes of water pollution, and that long-term strict monitoring should be conducted to ensure the safety of drinking water consumption and the health of the environment.