Five aminoglycoside antibiotics including gentamicin, sisomicin, tobramycin, amikacin and netilmicin were comparatively tested in vitro against 283 clinical isolates of Pseudomonas, Enterobacteriaceae and Gram-positive cocci, using the agar dilution technique according to I.C.S. recommendations. In respect to potency by weight, netilmicin proved the most active aminoglycoside against E. coli, K. pneumoniae, Enterobacter, Serratia spp. and Staphylococcus aureus, followed by sisomicin or tobramycin in relation to various bacterial species. Against Pseudomonas aeruginosa, sisomicin and tobramycin had the lowest MICs (minimum inhibitory concentrations). Amikacin had overall less activity but the widest spectrum. Rate of killing curves showed few differences among the aminoglycosides tested. Against gentamicin-resistant strains (MIC greater than or equal to 16 micrograms/ml) the activity of netilmicin was comparable to or higher than that of amikacin, except for P. aeruginosa, P. cepacia and indole-positive Proteus, which were inhibited by amikacin only. Netilmicin also showed the highest "therapeutical index" calculated as the ratio between MIC and blood levels for each aminoglycoside against each most important bacterial specie.