Antimicrobial activity and mechanistic insights into the tick peptide persulcatusin against vancomycin-susceptible and vancomycin-resistant Enterococcus faecium strains

Introduction The emergence of multidrug-resistant bacteria, particularly vancomycin-resistant enterococci (VRE), represents a critical threat to global public health. Therefore, the development of novel therapeutic strategies is urgently required. Antimicrobial peptides (AMPs) are promising candidates for combating these infections. Persulcatusin (IP), an AMP from the hard tick Ixodes persulcatus , exhibits potent activity against Staphylococcus aureus via membrane disruption. However, IP’s efficacy and mechanism against enterococcal species remain unclear. This study aimed to evaluate the antibacterial potency of IP against clinical isolates of E. faecalis and E. faecium , including vancomycin-resistant strains, and determine whether its mode of action involves membrane disruption, as observed in S. aureus. Methods To assess the antibacterial efficacy of IP against enterococcal strains, including VRE, we determined the minimum inhibitory concentrations (MICs) and conducted short-term killing assays using clinical isolates of E. faecalis and E. faecium . Furthermore, we conducted electron microscopic observation of bacterial morphology, intracellular localization analysis using 5-carboxyfluorescein-labeled IP, and DNA-binding assays to elucidate its mechanism. Results IP exhibited the lowest MIC value (1.25 μg/mL) against vancomycin-susceptible E. faecium among the five AMPs tested (range: 10 to >40 μg/mL), showing growth-inhibitory activity comparable to that of vancomycin (1.25 μg/mL). IP also effectively inhibited the growth of vancomycin-resistant E. faecium (VREfm) strains with an MIC of 0.625 μg/mL. Conversely, IP showed no growth inhibition against any tested E. faecalis strains, revealing species-specific selectivity. Unlike the effects on S. aureus , IP did not induce membrane damage in Enterococcus strains. Instead, the growth-inhibitory activity of IP correlated with its cellular penetration capability (in the order: S. aureus > E. faecium > E. faecalis ). Additionally, IP demonstrated high binding affinity to bacterial DNA. Discussion This study is the first to demonstrate that IP effectively inhibits the growth of E. faecium , including VREfm, through a mechanism distinct from its known membrane-disrupting action. Given its high cellular penetration and DNA-binding capacity, the antimicrobial action of IP against E. faecium is likely to involve an intracellular target, such as genomic DNA. Our results highlight IP as a promising therapeutic agent for treating challenging VREfm infections.