In a previous report (1), a novel bacterial type II topoisomerase inhibitor NBTI 5463 with activity against Gram negative pathogens was described. First-step resistance mutants in Pseudomonas aeruginosa arose exclusively in the nfxB gene, a regulator of the MexCD-OprJ efflux pump system. The present report describes further resistance studies with NBTI 5463 in both Pseudomonas aeruginosa and Escherichia coli. Second-step mutations in P. aeruginosa arose at aspartate 82 of the gyrase A subunit, and led to 4-8 fold increases in the MIC over those seen in the parental strain with a first step nfxB efflux mutation. A third step mutant resulted in additional GyrA changes, with no changes in topoisomerase IV. Despite... More
In a previous report (1), a novel bacterial type II topoisomerase inhibitor NBTI 5463 with activity against Gram negative pathogens was described. First-step resistance mutants in Pseudomonas aeruginosa arose exclusively in the nfxB gene, a regulator of the MexCD-OprJ efflux pump system. The present report describes further resistance studies with NBTI 5463 in both Pseudomonas aeruginosa and Escherichia coli. Second-step mutations in P. aeruginosa arose at aspartate 82 of the gyrase A subunit, and led to 4-8 fold increases in the MIC over those seen in the parental strain with a first step nfxB efflux mutation. A third step mutant resulted in additional GyrA changes, with no changes in topoisomerase IV. Despite repeated efforts, resistance mutations could not be selected in E. coli. Genetic introduction of the Asp82 mutations observed in P. aeruginosa did not significantly increase the MIC in E. coli. However, with the aspartate 82 mutation present, it was possible to select second-step mutations in topoisomerase IV that did lead to MIC increases of 16 and 128 fold. As with the gyrase aspartate 82 mutation, the mutations in topoisomerase IV did not by themselves raise the NBTI MIC in E. coli. Only the presence of mutations in both targets of E. coli led to an increase in NBTI MIC values. This represents a demonstration of the value of balanced dual-target activity in mitigating resistance development.
