Temporal variations over short and medium time scales play an important role in fish assemblage dynamics, but have been poorly investigated in tropical estuaries. This study evaluates the hypothesis that fishes co-occurring on a tidal mudflat have different patterns of temporal segregation at short- and medium-term scales that optimize resource use and habitat partitioning. A total of 6222 individuals and 66 fish species were caught during different hours covering the entire 24 h cycle, tidal regimes and the wet and dry seasons. Biomass and species richness, and to a lesser extent CPUE and evenness, showed statistically significant interactions across short- and medium-term scales. Biomass was higher during the dry season and its oscillation along tidal cycles revealed distinct patterns over the photoperiod in each season. A similar complex pattern was also observed for species richness, which showed distinct temporal patterns between high and low tides over the photoperiod in each season. Overall, shorter-term variations on fish assemblage attributes were correlated mainly with photoperiod and, to a lesser extent, tidal regime. Medium-term variations in fish abundance and species richness, in contrast, could result from seasonality in recruitment patterns and higher availability of allochthonous food resources during the wet period.