Effects of PSP-Derived Antimicrobial Peptides on Enterococcus faecalis Conjugation

Objectives: Enterococcus faecalis is a Gram positive opportunistic pathogen associated with failed root canal treatments and nosocomial infections including endocarditis. A peptide-pheromone based high-efficiency plasmid transfer system contributes to widespread antibiotic resistance. The system relies on hydrophobic peptides for induction of cell-cell contact/agglutination.  Interference with this transfer could prevent the spread of antibiotic resistance in this species.

We recently designed cationic, hydrophobic, antimicrobial peptides based on the human salivary protein PSP. The peptide GL13NH₂ causes bacterial agglutination while GL13K is bactericidal to oral and non-oral bacteria. We hypothesize that these PSP-peptides could prevent the transfer of plasmids by competitively blocking agglutination or signaling in E. faecalis .

Methods: Donor bacteria, OG1RF-pCF10 (plasmid containing), were cultured under inducing or non-inducing conditions. Induced donor cells are expected to have a high transfer rate (10^-1-10^-2) due to the presence of pheromone. Non-induced donors have a lower rate (~10^-5) due to the presence of an inhibitor peptide. Induced and un-induced donor cultures were incubated without PSP-peptides or with a sublethal concentration (32µg/ml) of GL13K or GL13NH₂ and incubated 2h/35°C. Donor cultures were then combined 1:10 with recipients (OG1SSp-plasmid free) to form mating mixtures. After 10-min incubation to allow conjugation, bacterial mixtures were plated on selective media for enumeration of CFUs. The efficiency of plasmid transfer was determined by the transconjugant/donor ratio.

Results:   In induced cultures, the transconjugant/donor ratio  was 2.5×10^-2 (control) and  5.5×10^-3 in the presence of GL13K. With GL13NH₂ the ratio was 1.1×10^-2  vs. 1.8x10^-2 for the peptide-free control. Uninduced cultures yielded similar qualitative results. The transconjugant/donor ratio did not differ significantly for any mating combination (P>0.2).    

Conclusion: The PSP-peptides GL13K and GL13NH₂ had no effect on the conjugation system of E. faecalis and are not suited to prevent plasmid transfer.