Flow regimes of rivers globally have been affected by human activities. In particular, river regulation in many regions has led to a reduction and stabilization of flows, with the loss of over-bank flow events. Flow variability plays a critical role in shaping the hydraulic properties of habitat patches within river channels and across floodplains. Littoral areas, snags, and shallow floodplains are heterogeneous habitats that provide refuges from fast-flowing waters for biotic communities during high-flow events, including zooplankton. Zooplankton are pivotal in aquatic food webs, facilitating the transfer of energy from primary producers to higher trophic levels. Although floods are known to regularly affect the ecology of riverine biota, their influence on shaping riverine zooplankton communities among different habitat patches remains poorly understood. We evaluated the effect of river discharge and patch-specific current velocity on zooplankton communities of the lower Lachlan River (central New South Wales, Australia) before, during, and after a large over-bank flood. Zooplankton samples and physico-chemical measures were collected from five habitat patches (benthic, littoral, midstream, pelagic, and snags) at four sites over eight occasions during 2022 and 2023, encompassing the largest flood on record. A total of 15 microcrustacean and 43 rotifer genera were identified across all sites. We found that daily discharge had a significant positive impact on the mean abundance of zooplankton, with significant differences also observed among habitat types. In contrast, patch-specific water velocity did not have a detectable impact on zooplankton abundance. Zooplankton abundance was highest in littoral and pelagic habitats, with rotifers dominating the community across most sampling occasions and habitats. Microcrustaceans were consistently less abundant. Temporal variability in zooplankton distribution suggested that habitat patch and discharge are key drivers of community structure. Therefore, maintaining flow variability and habitat heterogeneity may enhance the resilience and productivity of zooplankton communities in riverine ecosystems.