The development of high-level fluoroquinolone resistance has rarely been obsd. in salmonellae and, in contrast to other Gram-neg. bacteria mutations affecting topoisomerase IV, a known secondary target of quinolones in Escherichia coli has not been described except for one recent report. The present study used quinolone-susceptible field isolates representing epidemiol. relevant serovars and phage types Salmonella hadar and Salmonella typhimurium DT104 and DT204c to select fluoroquinolone-resistant mutants in vitro. Three selection steps were necessary to obtain high-level fluoroquinolone-resistant mutants (MICCip >=8 mg/mL). All first-step mutants examd. had a single gyrA mutation (affecting either Ser83 or Asp87). Addnl. topoisomerase mutations affecting gyrA (Asp87), gyrB (Ser464), and parC (Gly78) were detected in second- and third-step mutants. Introducing into the resp. mutants the corresponding plasmid-coded quinolone-susceptible allele of either gyrA, gyrB, or parC resulted in redn. of quinolone resistance, indicating a role for these mutations in quinolone resistance. In the presence of an inhibitor of RND-type efflux pumps, the susceptibilities to ciprofloxacin and chloramphenicol of second- and third-step mutants increased by two to four serial diln. steps, providing evidence that an efflux-mediated resistance mechanism contributes to the development of high-level fluoroquinolone resistance in salmonellae.