Abstract: Spawn quality is the most important factor in the production of edible mushroom (which is fast gaining prominence in Nigeria and Africa at large). In order to determine the effects of substrates spawn preparation on mycelial growth of oyster mushroom species, the experiment was conducted in a factorial experiment design at randomized completely with three replications. In this experiment, first and second factors, respectively were substrates (Wheat, yellow maize, guinea corn, millet, red sorghum and white maize, Bende local and oyster mushroom species (Pleurotus tuber-regium and pelurotus pulmonarius). The results clearly demonstrated that between various substrates used, maximum and minimum growth rate were recorded for white maize (Bende Local) and least mycelial extension and fresh weight on wheat. The second best grain for both species used was Red Sorghum.