Based on an analysis of the key scientific problems that need to be solved in river microbial ecology, including the mechanisms of community composition, ecological function, and environmental interaction, the frontier theoretical research on distribution patterns, ecological construction, environmental response, and food web construction of river microbial communities, as well as their interaction with matter transformation is discussed. The current situation of microbial community technology applied in the construction of river ecological health indication, assessment, and diagnosis system, pollutant source tracing, and river remediation is summarized. It is concluded that the future research on river microbial ecology should focus on innovative methods, large research scales, multidimensional study, and transformation of theoretical results into technical application.