Abstract South America is home to more miniature fishes (<26 mm in standard length) than any other continent. Despite this diversity, the ecology of miniature fishes is poorly studied. To promote the study of miniature fish ecology, we investigated patterns in total richness, assemblage structure and environmental correlates for miniature fishes in the rio Jacund drainage of the Lower Amazon River basin, Par State. Based on multi-pass dip-netting of leaf litter at 20 locations distributed across two sites, we collected miniature species and used rarefaction to estimate 9 to 14 species might be present. The miniature fish assemblage at the upstream site was a nested subset of the downstream site, and water pH and canopy cover, two features known to be altered by deforestation, correlated most strongly with assemblage variation. Our work represents one of the first quantitative assessments of environmental correlates with miniature fish assemblages and highlights research topics that should be investigated further to promote conservation and preservation of the overlooked and understudied Amazonian diminutive freshwater fish fauna.