Point mutations in the ygiV promoter region lead to cystobactamid resistance and reduced virulence in E. coli

Abstract

Antimicrobial resistance is one of the major health threats of the modern world. Thus, new structural classes of antimicrobial compounds are needed in order to overcome existing resistance. Cystobactamids as such a new compound class inhibit the well-established target bacterial type II topoisomerases while exhibiting superior antibacterial and resistance-breaking properties. Understanding potential mechanisms of emerging resistances is crucial in the development of novel antibiotics as they directly impact the future therapeutic application and market success. Therefore, the frequency and molecular basis of cystobactamid resistance in Escherichia coli was analyzed. High-level resistant E. coli mutants were selected and found to harbor single nucleotide polymorphisms in the promotor region of the ygiV gene, causing an upregulation of the respective protein. These stable mutations are contrary to what was observed as resistance genotype in the structurally related albicidins, where ygiV gene amplifications were identified causing resistance. Overexpression of YgiV in the mutants was additionally amplified upon cystobactamid exposition, showing further adaptation to this compound class under treatment. YgiV binds cystobactamids with high binding affinity, thereby preventing their interaction with the antimicrobial targets topoisomerase IV and DNA gyrase. Additionally, we observed stabilization of the gyrase cleavage complex by YgiV itself, suggesting a further protective effect against topoisomerase poisoning. Moreover, tested cystobactamid-resistant mutants show reduced motility and fimbrial protein expression, indicating an overall decreased virulence and potentially pathogenicity, associated with ygiV promotor mutations.