Periphyton is a complex community of detritus, bacteria, fungi, and algae that grows attached to submerged substrates. Assemblage structure data for algae and cyanobacteria from periphyton samples were collected on a monthly basis from 2017 to 2021. Compiled data were reported as absolute abundance (cells) and biovolume per volume as well as absolute abundance (cells) and biovolume per area at four locations increasing in distance East of the L-67 canal in Water Conservation Area 3 (WCA 3) of the Florida Everglades immediately north of Everglades National Park. The periphyton assemblages were analyzed for five sample types including periphyton plates (Plates), invertebrate inclusion and exclusion plates (Inverts), biological oxygen [...]