Changes in the Ultrastructure of Staphylococcus aureus Treated with Cationic Peptides and Chlorhexidine

Antimicrobial Peptides, including synthetic ones, are becoming increasingly important as a promising tool to fight multidrug-resistant bacteria. We examined the effect of cationic Peptides H₂N-Arg₉-Phe₂-C(O)NH₂ and H₂N-(Lys-Phe-Phe)₃-Lys-C(O)NH₂ on Staphylococcus aureus, which remains one of the most harmful pathogens. Antiseptic chlorhexidine served as reference preparation. We studied viability of S. aureus and examined its ultrastructure under treatment with 100 µM of R9F2 or (KFF)3K Peptides or chlorhexidine using transmission electron microscopy of ultrathin sections. Bacterial cells were sampled as kinetic series starting from 1 min up to 4 h of treatment with preparations. Both Peptides caused clearly visible damage of bacteria cell membrane within 1 min. Incubation of S. aureus with R9F2 or (KFF)3K Peptides led to cell wall thinning, loss of cytoplasm structure, formation of mesosome-derived multimembrane structures and "decorated fibers" derived from DNA chains. The effect of R9F2 Peptides on S. aureus was more severe than the effect of (KFF)3K Peptides. Chlorhexidine heavily damaged the bacteria cell wall, in particular in areas of septa formation, while cytoplasm kept its structure within the observation time. Our study showed that cell membrane damage is critical for S. aureus viability; however, we believe that cell wall disorders should also be taken into account when analyzing the effects of the mechanisms of action of antimicrobial Peptides (AMPs).

S. aureus; cationic peptides; cell membrane; cell wall; chlorhexidine; cytoplasm damage; transmission electron microscopy.