Study of the reversal of metallo-β-lactamase-mediated resistance in Gram-negative bacteria by EDTMP

Metallo-β-lactamases (MBLs) are major causes of carbapenem resistance in Gram-negative bacteria. In this study, a simple and practical method was developed to synthesize the fluorescent probe CE-HF, which was proven effective for detecting the activity of the MBL NDM-1. Using this probe, ethylenediamine tetra(methylene phosphonic acid) (EDTMP) was identified as a potent MBL inhibitor with strong activity (IC₅₀ = 0.68 μM against NDM-1). Antibacterial assays, including inhibition zone, MIC, and MBC tests, revealed that EDTMP significantly enhanced the efficacy of meropenem and other β-lactam antibiotics by lowering their MIC values and exhibiting synergistic effects. Checkerboard assays further demonstrated that EDTMP reduced the MIC of meropenem by 256-fold at 8 μg/mL, whereas zinc ion rescue experiments confirmed that EDTMP exerts its inhibitory effect by chelating zinc ions at the active site of MBLs. Moreover, EDTMP restored the antibacterial activity of meropenem against recombinant E. coli strains expressing L1, ImiS, IMP-1, and VIM-2, as well as clinical isolates of Stenotrophomonas maltophilia producing VIM-2. These findings highlight EDTMP as a broad-spectrum MBL inhibitor and offer a new strategy to combat β-lactamase-mediated antibiotic resistance.