Reverting Multidrug-resistant Phenotypes of Escherichia coli Isolated from Cattle Using 1-(1-Naphthylmethyl)-Piperazine

Introduction: The extensive use of antimicrobial agents in both the health and food sector has led to the emergence of multidrug resistant (MDR) bacteria, a development of importance to public health. Efflux pumps extrude antimicrobial compounds from cells contributing to the development of resistance. Chemosensitizers with the capacity to modulate efflux pump activity are being studied as adjuvants in efforts to reverse resistant phenotypes. However, little is known about their efficacy and mechanism of action.

Purpose: The purpose of this study was to systematically analyze the MDR reversal activity of the chemosensitizer 1-(1-naphthylmethyl)-piperazine (NMP), when applied as an adjuvant with antibiotics on both planktonic and sessile Escherichia coli cells.

Methods: Bovine E. coli isolates from the UCD Veterinary Hospital were screened for their MDR phenotype. A panel of 12 isolates, resistant to different classes of antibiotics including fluoroquinolones, tetracyclines, and chloramphenicol, were further studied. All were characterized by whole genome sequencing. The ability to form biofilm and fimbriae was, also, determined. Minimum inhibitory concentration (MIC) for each antibiotic, alone or in combination with NMP at sub-MIC levels, was determined by broth microdilution, using planktonic and sessile-grown cells. Transmission electron microscopy (TEM) using NMP was performed.

Results: Isolates had diverse AMR and virulence gene profiles. Based on these data the gsp operon was mainly associated with strong biofilm formers. In planktonic cells, using NMP as the adjuvant, the MIC of ciprofloxacin, chloramphenicol, and tetracycline exhibited a two-, six- and ten-fold reduction, respectively, in comparison to the antibiotic alone. In the case of sessile cells, half showed reductions in biofilm biomass when tetracycline was combined with NMP. TEM imaging demonstrated cell wall damage with NMP.

Significance: Our findings showed that NMP damages the cell wall, increasing drug permeabilization. Use of NMP and NMP-like structures has the potential to reverse MDR in bacteria.