Unraveling the Antimicrobial and Antibiofilm Activity of Ag-phendione and Cu-phendione Against Multidrug-resistant Klebsiella pneumoniae Using Shotgun Proteomics.

INTRODUCTION

Multidrug-resistant (MDR) Klebsiella pneumoniae infections are associated with high mortality, prolonged hospitalization, and limited treatment options, representing a global healthcare challenge. The coordination compounds [Ag(1,10-phenanthroline-5,6- dione)2]ClO4 (Ag-phendione) and [Cu(1,10-phenanthroline-5,6-dione)3(ClO4)2] (Cu-phendione) have previously demonstrated antimicrobial and antibiofilm activity against MDR Gram-negative bacilli.

METHODS

This study investigated their mechanisms of action by combining classical microbiology with shotgun proteomics. Minimum inhibitory and bactericidal concentrations (MIC/MBC), timekill assay, and antibiofilm effects were determined, while nanoESI-LC-MS/MS identified proteins modulated in MDR K. pneumoniae.

RESULTS

MIC values ranged from 3.125 mg/L to 12.5 mg/L for Ag-phendione and from 12.5 mg/L to 50 mg/L for Cu-phendione. Both compounds exhibited early antimicrobial effects ( ≤ 3 h), inhibited biofilm formation, and disrupted mature biofilms in a dose-dependent manner. Proteomic analysis revealed modulation of proteins related to NAD⁺ synthesis, carbohydrate metabolism (notably trehalose), DNA replication and repair, oxidoreductase activity, and plasmid partitioning.

DISCUSSION: The modulation of these proteins correlates with the observed antimicrobial and antibiofilm activity of the coordination compounds, suggesting potential targets for further validation. Interestingly, several proteins modulated by these compounds are also affected by classical antibiotics such as meropenem and colistin, suggesting partially convergent mechanisms involving central metabolism and redox homeostasis.

CONCLUSION

Ag-phendione and Cu-phendione are promising candidates for the development of novel therapeutics against MDR K. pneumoniae. Future studies should validate key targets through genetic knockouts or enzyme activity assays to clarify the mechanisms underlying these effects.