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This work systematically investigated the microbial community structure in the river sediments from upstream of Guanting Reservoir, Beijing, China. A total of 6 wastewater treatment plants (WWTPs) locate along the main rivers connected to the reservoir. Water and sediment samples were collected at sites near the effluents of WWTPs (regarded as W groups) or at the upstream/downstream rivers (R groups) to reveal the roles of the reclaimed water recharge. Multivariate techniques including typical statistical analysis, redundancy analysis (RDA), nonmetric multidimensional scaling analysis, and molecular ecological network analysis were used to evaluate the results and their relationships. The representative C/N/P water parameters and concentrations of target organic contaminants kept stable for W and R sites, while the microbial community parameters varied greatly for two groups. The microbial population at W sites were higher but with a lower biological diversity (with a lower Shannon index) than that at R sites, indicating WWTPs greatly altered the microbial community structure at the local reach. RDA results revealed that total organic carbon (TOC) and organophosphorus pesticides (OPPs) were two dominant factors affecting the function and composition of microbial communities at the phylum level. The network analysis revealed that the microbes with the most interactions mainly from R sites and they had closer relationships with each other.