Topoisomerase IV mutations in quinolone-resistant salmonellae selected in vitro.

The development of high-level fluoroquinolone resistance has rarely been observed in salmonellae and, in contrast to other Gram-negative 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 epidemiologically 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 > or = 8 microg/ml). All first-step mutants examined had a single gyrA mutation (affecting either Ser83 or Asp87). Additional topoisomerase mutations affecting gyrA (Asp87), gyrB (Ser464), and parC (Gly78) were detected in second- and third-step mutants. Introducing into the respective mutants the corresponding plasmid-coded quinolone-susceptible allele of either gyrA, gyrB, or parC resulted in reduction 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 dilution steps, providing evidence that an efflux-mediated resistance mechanism contributes to the development of high-level fluoroquinolone resistance in salmonellae.