Identification of nonpoint source (NPS) pollution is essential for effective water management. In this study we used a combined approach of hierarchal cluster analysis (HCA) and positive matrix factorization (PMF) to identify NPS pollution for Huaihe River basin in China. NH₃-N, COD, DO and pH were regularly monitored weekly over 2 years (2011–2012) from 27 monitoring stations subjected to high anthropogenic and natural changes. As identified by multiple correspondence analyses, the monitoring stations #3, #9 and #21 are located away from the rest of sites. HCA classified all the stations into 4 groups. PMF identified four factors on each group and each season. They were associated with the major causes of Huaihe River water quality deterioration resulted by discharges inputs from urban, agricultural and industrial land uses. Seasonal NPS pollution variation was found, and it is possibly linked with natural processes, for instance hydrological regime. This research work demonstrates the usefulness of PMF model for the identification of NPS pollution in surface waters. Furthermore, our study also shows that urban, agricultural and industrial land uses were the main factors impairing surface water quality, and limiting NPS pollution would be critical for enhancing surface water quality in the study area.