Fig. 5: Resistance to nitroxoline is associated with efflux pump upregulation across species.

a Whole-genome sequencing of experimentally evolved nitroxoline-resistant strains (Supplementary Data 2). Nitroxoline MIC values are indicated below each strain. Mutation effects are colour-coded. Strains on which proteomics was performed (Fig. 5b) are indicated in bold. K. pneumoniae strains whose sensitive parental strain lacks oqxR are marked with an asterisk. The in-patient evolved K. pneumoniae clinical isolate 8_R1 is indicated in italics. b Protein abundance changes in nitroxoline-resistant strains. Selected proteins annotated as efflux pumps or porins are shown and clustered according to Pearson’s correlation. Hits are marked with an asterisk (adjusted p value ≤0.05 and at least twofold abundance change (Supplementary Fig. 6a, b and Supplementary Data 6). Species are colour-coded, as in Fig. 5a. c Wild-type emrR and oqxr complementation restores nitroxoline susceptibility. The experimentally evolved E. coli strain 1_R4 with emrR D109V mutation (Fig. 5a and Supplementary Fig. 6c) and K. pneumoniae strain 4_R1 harbouring the oqxR G60-L67 duplication (Fig. 5a, d and Supplementary Fig. 6d) are shown. Nitroxoline MIC was measured by broth microdilution. Mean and standard error across four biological replicates are shown. ns p > 0.05; *p ≤ 0.05 (Wilcoxon test. For E. coli: empty plasmid vs no-plasmid, p = 0.608; empty plasmid vs wild-type efflux pump, p = 0.042; no-plasmid vs wild-type efflux pump, p = 0.042. For K. pneumoniae: empty plasmid vs no-plasmid, p = 0.217; empty plasmid vs wild-type efflux pump, p = 0.042; no-plasmid vs wild-type efflux pump, p = 0.042). d Amino acid changes resulting from oqxR mutations. The domain annotation of OqxR was obtained from its closest annotated structural homologue NsrR (Methods). e Efflux pump inhibitors resensitize nitroxoline-resistant strains. Nitroxoline MIC was measured by broth microdilution. Mean and standard error across four biological replicates are shown. Resistant strains are shown as shaded plots next to their parental-sensitive strains. For results on all strains, see Supplementary Fig. 6g. p values are only shown when significant: *p ≤ 0.05; **p ≤ 0.01 (Wilcoxon test). Source data are provided as a Source Data file.