Three ectomycorrhizal fungi, Cenococcwn graniforme, Suillus luteus, and Thelephora terrestris were grown in artificial nutrient media. Water potential of the media was varied by the use of the osmoticum polyethylene glycol (PEG) 4000 and measured by thermocouple psychrometry. Cenococcum graniforme was very tolerant of low water potentials and exhibited maximum growth at a potential of −15 bars. Maximum growth of S. luteus and T. terrestris occurred at −5 bars. The water potential of solutions containing PEG 4000 appears to consist of both an osmotic and matric component, making PEG 4000 ideally suited for simulation of soil moisture stress. It was neither metabolized nor readily absorbed by C. graniforme as inorganic salts or sugars might be.