Microbial community assembly processes are closely related to the composition, structure, and distribution of microbes. The changes in environmental conditions and species dispersal capacity induced by hydrological connectivity may significantly impact the microbial community assembly process in surface water, but the mechanisms remain unclear. To reveal how hydrological connectivity affects microbial community assembly processes, surface water samples were collected from the study watershed during periods of low, intermediate, and high hydrological connectivity. An integrated 16S amplicon sequencing technology and phylogenetic null model approach were used to identify the assembly processes of the bacterial communities. The results showed an inverse relationship between hydrological connectivity and environmental heterogeneity, with the highest environmental heterogeneity observed at low connectivity level. Bacterial alpha diversity under high hydrological connectivity gradient significantly exceeded those under low and intermediate hydrological connectivity. Beta diversity exhibited a trend toward biotic homogenization as hydrological connectivity increased. The co-occurrence network of bacterial communities under low hydrological connectivity were characterized by robust clustering and intricate interactions, whereas those under intermediate hydrological connectivity tended to form more straightforward network. Furthermore, stochastic processes play a crucial role in bacterial community assembly, accounting for approximately 80% of the observed patterns. This was substantiated by piecewise structural equation modelling, which showed that environmental factors and biotic interactions exerted minimal influence on the bacterial community assembly. As hydrologic connectivity increases, the assembly process shaping bacterial community appears more stochasticity. Moreover, the contributions of drift and heterogeneous selection in assembly processes was found to increase with hydrological connectivity, while the impact of dispersal limitation and homogenous selection diminished. These insights provide a deeper understanding of the ecological mechanisms that govern microbial distribution pattern and succession in watershed surface water.