O-Demethylfortimicin A (compound A49759) was tested against 445 bacteria, and the results were compared with thooe obtained with fortimicin A, amikacin, gentamicin, netilmicin, sisomicin, and tobramycin. A49759 was found to be active and bactericidal against the Enterobacteriaceae, nonfermentative gram-negative bacilli, and Staphylococcus aureus. A49759 was two-to fourfold more active than fortimicin A against most species tested, but generally fourfold less active than amikacin against this population of Pseudomonas aeruginosa (85% inhibited at -16 ug of amikacin per ml and 85% inhibited at c64 ,ug of A49759 per ml). Only amikacin and A49759 were resistant to most aminoglycoside-inactivating enzymes and also had significant antipseudomonal activity. Amikacin was inactivated by aminoglycoside 6'-acetyltransferase, and A49759 was inactivated by aminoglycoside 3-acetyltransferase. The minimal inhibitory concentrations of all tested aminoglycosides were increased by augmenting the inoculum size. Several recent reports have described the in vitro antimicrobial activity of the pseudodisaccharide aminoglycoside antibiotics (4-8, 10, 11, 13, 14). Among these new compounds, the fortimicin group has been most extensively tested against recent clinical isolates (7). Numerous chemical and natural modifications of fortimicin have been presented (6,8,10,14,15). Fortimicin aminoglycosides are generally resistant to inactivating enzymes (6, 10, 13), but lack significant inhibitory activity against pseudomonads (6, 7). In recently presented abstracts no ototoxicity and very low renal toxicity for fortimicin A in the animal models were described (C. L