Investigating the adjuvant activity of the proline-rich antimicrobial lipopeptide C₁₂-PRP against...

Farm animals exposed to excessive amounts of antibiotics are reservoirs for antimicrobial-resistance genes and multidrug-resistant bacteria. The goal of this study was to investigate the adjuvant activity of a previously synthesized, proline-rich lipopeptide called C₁₂-PRP in combination with various antibiotics against multidrug-resistant E. coli isolated from chicken and cattle.

The microbroth dilution method was used to determine the minimum inhibitory concentration of 10 antibiotics against four E. coli isolates. The adjuvant activity of C₁₂-PRP was then tested in combination with the same antibiotics using a checkerboard assay. The best synergistic antibiotic-C₁₂-PRP combinations were tested with the most resistant E. coli isolate using the Galleria mellonella insect infection model. A fluorescence-based membrane permeabilization assay was used to investigate a possible mode of action for C₁₂-PRP.

The four E. coli isolates showed varying resistance to the antibiotics (0.008 µg/mL to ≥256 µg/mL), and overall, the addition of C₁₂-PRP lowered the minimum inhibitory concentration. The two most-potent antibiotic-C₁₂-PRP combinations were with novobiocin and erythromycin showing a susceptibility increase of 128-fold and up to 32-fold, respectively. G. mellonella survival was significantly improved when C₁₂-PRP was included with novobiocin or erythromycin during treatment of infected larvae. Membrane permeabilization of an E. coli isolate was markedly higher in the presence of C₁₂-PRP.

We demonstrated the synergistic ability of C₁₂-PRP to enhance antibiotic activity and reduce the virulence of the veterinary E. coli isolates, strengthening the potential promise of this adjuvant candidate as a therapeutic agent for multidrug-resistant bacteria.