A disc containing 50 microgram fosfomycin and 25 microgram glucose-6-phosphate (G-6-P) has been developed for fosfomycin susceptibility testing. The present study was designed to examine the influence of G-6-P on the diffusion gradient of fosfomycin in Mueller-Hinton agar. Discs containing 50 microgram 3H-fosfomycin and/or 25 microgram 14C-G-6-P were placed in the center of agar plates. Radioactivity of agar cylinders removed along the radius of the Petri plate at different times from 30 sec to 24 hours was measured. Results showed that the fosfomycin/G-6-P ratio throughout diffusion remained close to the initial ratio in the disc, suggesting that loads of both substances borne by the discs are adequate for antibiotic sensitivity testing.