A major goal of green infrastructure is the reduction of stormwater in the urban and suburban landscape, which is accomplished by the design—particularly the growing substrate and plants within it. Engineered growth substrate is often used because it is a known quantity that can hold specific volumes of water while still being lightweight. Natural growth substrate (soil) is a more unknown quantity but has the benefit of being able to support more plant species and introduce native soil organisms into the system. The interactions between growth substrate, plant, and soil biota have the potential to be able to bring additional benefits to the urban environment besides just stormwater reduction, such as air and water purification, and providing biological habitat. This study aims to examine how different types of substrate, plants, and the addition of soil organisms called mycorrhizal fungi impact the quality and quantity of stormwater. Located at the Cleveland Industrial Innovation Center, 39 different square meter plots were built on a low height roof. Three different substrate types: a bioretention grade soil, a worm casting compost, and a conventional engineered media (RoofliteTM) were used in combination with two separate plant communities, a native prairie and a designed community. Half of these were inoculated with symbiotic mycorrhizal fungi and the other half were left uninoculated. Water runoff was collected bi-monthly over the course of a year and total N and P content were measured. Preliminary analyses show that substrate type had a very significant effect on the volume of water runoff, while plant communities significantly impacted the amount of nitrogen runoff. All the treatments showed high amounts of phosphorus runoff, which could potentially impact downstream water quality if not addressed. However, further analyses on the plants, substrate, and soil biota are still in progress and the information may help improve green roof health and functioning.