Diagnostic microbiology laboratories in Australia and the UK commonly incorporate p-nitrophenylglycerol (PNPG) into solid susceptibility testing media in order to prevent the swarming of Proteus spp. We have investigated the effects of PNPG and adjusting the cation concentrations of the media to physiological levels on the MICs of aminoglycosides and other antibiotics for 128 strains of Pseudomonas aeruginosa isolated from the sputa of children with cystic fibrosis. The addition of PNPG to the media led to higher MICs of gentamicin and tobramycin for up to 23% of the isolates. Depending on the base medium, supplementation with the cations, calcium and magnesium, also increased the MICs of these aminoglycosides for 12-27% of the strains tested. Both incorporation of PNPG and cation adjustment led to higher MICs for 25-53% of isolates, again depending on the base medium. The MICs of ticarcillin, ciprofloxacin and colistin (on Iso-Sensitest agar) for significant numbers of strains were lower in the presence of PNPG, while those of ceftazidime varied from higher to lower, according to the concentration of the drug and the base medium. In many instances these changes would have altered the way in which the susceptibilities of the organisms would have been reported. PNPG clearly exerts an important effect when the in-vitro activities of various antibiotics against P. aeruginosa are determined by the agar dilution method. Recommendations for the inclusion of PNPG in susceptibility testing media should therefore be reviewed.