Abstract: Guangxi Province is located in the southern China and the upper and middle reaches of the Pearl River. It spans north tropical, south subtropical and central subtropical regions. The diversity of habitats in these regions promotes high fish species diversity. However, considering the reduction of global fishery resources due to numerous anthropogenic stressors, especially the over-construction of dams, studies on the spatial distribution of fish communities are limited and urgently needed. This study investigated fish diversity and distribution patterns of the Pearl River in Guangxi based on an investigation of 15 sample sites. A total of 67985 individual fishes were collected during the study period, including 134 taxa representing 23 families. The Cypriniformes order, representing 92 taxa, accounted for 69% of the total fish collected. A total of 11 non-native species were collected, of which tilapia was the highest abundant, almost all over the Guangxi. This suggests that the invasion of non-native species is serious in the Pearl River in Guangxi, and fish miniaturization is also serious. Significantly higher species diversity and evenness were recorded in the first order streams, and low species diversity and low evenness were recorded in the second order streams. The spatial variation in fish assemblages were significantly different in different river sections (P<0.05), such as the Hongshuihe River section with the lowest fishery resources. A comparison with historical data revealed that fish species have decreased sharply and the spatial distribution of fish communities has changed significantly. The present study is part of a long-term investigation of the Pearl River’s wild fishery resources. Therefore, understanding processes outlined in this study will help protect the diversity of fish communities, which is critical to the sustainability of fish diversity of the Pearl River in Guangxi Province.