Abstract <p>Rapid urbanization has resulted in numerous ecological challenges, including biodiversity loss and species homogenization. Within this highly human-disturbed and interconnected ecosystem, the diversity and community assembly of microfauna (less than 2 mm), such as bdelloid rotifers, are anticipated to be primarily structured by niche processes and less by dispersal limitation. This view has recently been questioned. This study investigated bdelloid rotifer diversity and community assembly process in subtropical megacity of Guangzhou based on a long-term investigation (2017–2024) of 150 sites. A total of 108 species including three suspected new species were identified. Dispersal-Niche Continuum Index (DNCI) analysis across different habitats and regions clearly highlights the dominant effect of dispersal processes in driving the community assembly of bdelloid rotifers, with the degree varying among the genera, habitats, and geographic locations. Our findings indicate that their community assembly appears to be independent of their active dispersal abilities and geographical distances, and might be influenced by complex biotic interactions and dispersal media. Furthermore, significant differences in diversity and community composition across different habitats were identified, with several differential and habitat-preferring species being identified through similarity percentage (SIMPER) and Indicator Value (IndVal) analyses, indicating contributions of niche effects. In conclusion, this study observed no loss of bdelloid rotifer biodiversity due to urbanization. It offers a new perspective on the dispersal process-dominant assembly of microfaunal communities in megacities, differing from the general notion of their ubiquity, and will aid in the evaluation and development of conservation policies for microfaunal biodiversity in urban ecosystems.</p>