Grepafloxacin (OPC-17116) and three other fluoroquinolones were tested against 461 bacterial isolates representing 44 species. Grepafloxacin was superior to ciprofloxacin, ofloxacin and fleroxacin in its activity against the Gram-positive cocci, including staphylococci, pneumococci and hemolytic streptococci. Attempts to select grepafloxacin-resistant mutants from a methicillin-resistant Staphylococcus aureus strain were unsuccessful, although ciprofloxacin readily selected quinolone-resistant mutants of that strain. Among Gram-negative bacilli other than Escherichia coli, mutants were readily selected by in vitro exposure to either grepafloxacin or ciprofloxacin. All such mutants showed complete cross resistance to six other quinolones. The four study drugs were active against nalidixic acid-susceptible strains belonging to 13 species of the Enterobacteriaceae, but ciprofloxacin was the most potent drug studied. Against Acinetobacter spp. and Stenotrophomonas maltophilia, grepafloxacin was two- to four-times more active than ciprofloxacin. However, against Pseudomonas aeruginosa, ciprofloxacin was two- to four-times more potent than grepafloxacin. Grepafloxacin and ciprofloxacin were both found to be bactericidal agents.