Predator-prey interactions in highly dynamic ecosystems such as non-perennial rivers and streams (NPRs) are relevant to understanding the effects of fragmentation and reshaping of aquatic habitat structure in interspecific relationships. In this context, our study offers a temporal snapshot of predator-prey interaction dynamics across different hydrological phases in an NPR. We sampled along 1.15 km of the NPR during flowing (23 sections) and dry (22 isolated pools) phases, identifying 18 fish species (predators) and 11 prey categories. Predator composition and abundance shifted between phases, with higher abundance during the flowing phase and increased environmental heterogeneity among isolated pools in the dry phase. The predator-prey interaction network exhibited a nested pattern, indicating hierarchical organisation where generalist and specialist predators coexist. In the dry phase, interaction patterns across isolated pools varied, including nested, modular, specialised, and/or random structures. Stochastic processes appear to shape interaction networks during the dry phase, as variation in species composition across isolated pools was not explained by predator richness. Our study shows that shifts in hydrological phases restructure predator-prey networks in NPRs. Given increasing climatic and human pressures on flow regimes, conserving natural hydrological variability is important to support biodiversity and ecosystem functioning in these ecosystems.