An interdisciplinary and multiagency study was undertaken to study harmful algal bloom (HAB) dynamics on the Caloosahatchee River, which drains to the west from Lake Okeechobee into the Gulf of Mexico. Algae and cyanobacteria play crucial roles in aquatic ecosystems, but under favorable environmental conditions certain taxa may experience population growth booms resulting in HABs. Four independent mesocosm experiments were conducted in September 2019, June 2020, September 2020, and February 2021 to capture natural temporal variability in water quality and phytoplankton assemblages. The experimental installation consisted of three floating metal cradles each containing four fiberglass chambers filled with native river water. The twelve chambers were assigned to one of four treatments: phosphate (P), nitrate (N), ammonium (A), or control (C) – each with 3 replicates. The P, N, and A treatment chambers were enriched by applying a concentrated dosing solution of either dibasic dodecahydrate sodium phosphate, anhydrous sodium nitrate, or liquid ammonium hydroxide respectively to elevate concentrations above ambient levels. The mesocosms were treated and sampled over a 72-hour period (Time 0, Time 24, Time 48, Time 72). This dataset contains phytoplankton absolute and relative abundance measures at the genus level as natural units (NU/mL), cell densities (cells/mL), and biovolumes (um3/mL).