The Guana River Marsh Aquatic Preserve is located within the Florida East Coast Drainage Basin which drains directly into the Atlantic Ocean or into coastal lagoons. This basin is divided into upper and lower sub-basins. The Upper East Coast sub-basin contains seven drainage areas, including the Tolomato and Guana Rivers. The Tolomato River has a drainage basin encompassing 84 sq.miles, and is part of the Atlantic Intracoastal Waterway (AIW). Smith, Deep, Sweetwater, Marshall, Stokes, and Casa Cola Creeks all drain in the Tolomato River from the west.