<i>In vitro</i> selection of cefiderocol-resistant mutants in <i>Acinetobacter baumannii</i> harbouring the most common carbapenemase genes

BACKGROUND: Carbapenem-resistant Acinetobacter baumannii (CRAB) represents a critical global health threat due to its high mortality rates in infections, limited treatment options and resistance to last-resort antibiotics. Cefiderocol, a novel siderophore-conjugated cephalosporin, has emerged as a promising therapeutic agent against such infections. However, resistance to this antibiotic has already been reported. OBJECTIVES: To investigate whether the production of common carbapenemases affects cefiderocol susceptibility in A. baumannii and to identify the genetic mechanisms underlying resistance development. METHODS: Using isogenic A. baumannii CIP 7010 strains carrying carbapenemase genes commonly identified in that species, namely blaNDM-1, blaOXA-23, blaOXA-40 or blaOXA-58, mutants were selected under increasing cefiderocol pressure. Mutants obtained were subjected to MIC determinations, WGS, complementation assays, real-time quantitative PCR (RT-qPCR) and fitness assays. RESULTS: WGS of cefiderocol-resistant isolates revealed recurrent mutations in genes encoding the global regulators BfmS and OxyR, rather than in genes directly related to iron uptake, PBPs or β-lactamases. Complementation assays with WT bfmRS or oxyR resulted in a reversion to the parental strain cefiderocol MICs. RT-qPCR indicated that these global regulators reduced expression of piuA and pirA genes involved in iron uptake, and hence were associated with a previously unknown mechanism that results in resistance to cefiderocol. CONCLUSIONS: Global regulators BfmS and OxyR were responsible for decreasing the expression of piuA and pirA genes, thereby contributing to cefiderocol resistance. This evolutionary analysis enhances our understanding of the mechanisms underlying cefiderocol resistance and identifies potential molecular targets for the development of new therapeutics against CRAB.