Abstract: Cascade development transforms natural rivers into continuous river-reservoir ecosystems, altering the communities of aquatic organisms. To understand these changes, we conducted field surveys in the mid-lower reaches of the Hanjiang River during the dry and wet seasons of 2023, combined with historical data from the Wuhan reach of the Yangtze River mainstem. This study compared the community composition of macrozoobenthos across different habitats and seasons, investigated their response to longitudinal connectivity, and analyzed influencing factors. A total of 132 taxa of macrozoobenthos were recorded. From the mid-lower Hanjiang River to the Wuhan reach of mainstem, taxa number and density generally declined but slightly recovered in the confluence zone. The lake zone and the transition zone exhibited higher taxa number and standing crops, while the mainstem zone had the lowest. The community structure showed a parabolic response to longitudinal connectivity, with total taxa number and Shannon-Wiener diversity index peaking at intermediate connectivity levels (0.95—0.97) during the wet season. The highest community similarity was observed between the lake zone and the confluence zone, whereas the mainstem significantly differed from other habitats (P<0.05). The community composition differences among different habitats during the wet season (P=0.025) were greater than those during the dry season (P=0.038), and there was a highly significant difference between the dry and wet seasons (P=0.001). Seasonal variations (P=0.001) were greater than habitat variations (P=0.015). During the dry season, water temperature and dissolved oxygen dominated the distribution of communities, whereas it was regulated by transparency and longitudinal connectivity during the wet season. This study demonstrates that cascade development reshapes communities of the macrozoobenthos by altering longitudinal connectivity and hydrological regimes, with the confluence zone serving as a critical ecotone in maintaining regional biodiversity.