Protected estuaries provide reference conditions for assessing coastal biodiversity, but information on swimming crab assemblages in such systems remains limited. This study evaluated the composition, dominance structure, diversity, body size, and spatio-seasonal variation in Callinectes swimming crab assemblages in the Una do Prelado River estuary, within the Barra do Una Sustainable Development Reserve, south-eastern Brazil. Seasonal sampling was conducted at four sites along the estuarine gradient using baited traps. Species composition, abundance, diversity indices and multivariate patterns were analyzed in relation to site and climatic season. A total of 146 individuals from four species were recorded: Callinectes exasperatus, C. danae, C. sapidus and C. bocourti. The assemblage was strongly dominated by C. exasperatus, which represented 81.5% of the total catch. Species richness was low, consistent with the restricted taxonomic scope and environmental filtering expected in estuaries. Abundance varied more clearly than species composition, with higher catches in autumn and at the intermediate estuarine site. Diversity indices indicated low evenness where C. exasperatus was dominant, whereas multivariate analyses detected no significant compositional differences among sites or seasons. As the first study conducted in this protected estuary, our findings provide a baseline for future monitoring of this system. Given the spatial and temporal scope of the study, these results should not be generalized to other protected estuarine systems.