Abstract: The spatio-temporal pattern of stream fish assemblages and the causal mechanism are the basis for scientific protection and sustainable utilization of fish species diversity. Both habitat structure and spatial position of streams are the mechanisms determining fish assemblages by influencing environmental capacity and fish immigration-extinction rate, respectively. In this study, spatial pattern in fish assemblages were studied by specimens electro-fishing collected from shallow streams (1–3 orders) of the Chang and Qingyi Rivers in the Huangshan Mountain, Anhui Province, in October, 2008 and 2009 respectively, and the effects of stream size (width and depth) and spatial position (order, link, C-link, and D-link) on fish assemblages were analyzed. Among the 35 fish species collected wholly, 26 and 29 species were involved in the Chang and Qingyi Rivers, respectively, which suggested 57.1% of Jaccard’s similarity index in species composition between the two rivers. Species richness and abundance each site were 4.69 ± 3.56 species and 26.46 ± 22.83 specimens in the Chang River, and 6.26 ± 3.09 species and 119.06 ± 90.90 specimens in the Qingyi River. Species richness and abundance of each site was significantly different between the two rivers. Along upstream-downstream gradient in stream orders, both species richness and abundance increased, while significant difference was only observed in species richness but not in abundance. Stream width and depth were both significantly correlated with stream orders, suggesting the significant difference in width and depth among 1–3 orders. Two-tailed Pearson’s correlation analysis detected that the factors significantly influencing species richness were stream order and D-link in the Chang River, while stream width, stream order, link and D-link in the Qingyi River. This analysis also detected that those influencing species abundance were both stream width in the Chang and Qingyi Rivers. Stepwise multiple regression analysis discovered the significant effects of D-link (Chang), D-link and stream width (Qingyi) on species richness, but stream width (both Chang and Qingyi) on abundance. Canonical correspondence analysis indicated that stream width, stream order, link, and D-link significantly determined fish assemblages in the Chang and Qingyi Rivers. In conclusion, downstream link (D-link) and stream width were the dominant factors affecting the spatial patterns in fish species richness and abundance, respectively, which suggested that spatial pattern in fish assemblages from shallow streams in the Chang and Qingyi Rivers were resulted from the combined roles of stream size and spatial position.