An investigation of the effects of an antimicrobial peptide on the survival of Acanthamoeba and intracellular bacteria associated with Cystic Fibrosis

Pseudomonas aeruginosa, Staphylococcus aureus and Haemophilus influenzae are the most notorious pathogens associated with pulmonary infections in Cystic Fibrosis (CF) lung. Treatment is problematic and re-infection is relatively common. It is vital to develop alternative therapies. Antimicrobial peptides are predominant candidates for combatting pathogenic bacteria. Free Living Amoebae is widely distributed in the environment and may also contaminate CF lung. It is well known that Acanthamoeba feed on and protect bacteria. The role of Acanthamoeba in CF lung infections has not yet been investigated. It is possible that the presence of Acanthamoeba in CF lung is one of the key factors for such re-infections. In this study, antimicrobial peptide, magainin II, was tested against clinical bacterial strains isolated from CF patients and Acanthamoeba. Magainin II has been shown to have antimicrobial efficacy on microorganisms employed in this study. P. aeruginosa, Escerichia coli, methicillin-sensitive Staphylococcus aureus, Staphylococcus epidermidis from CF lung showed ability to bind with, invade, survive and multiply inside Acanthamoeba. However, other two clinical isolates H. influenzae and Streptococcus pneumoniae were able to bind. They were found to have capsules that enables bacteria remain un-phagocytized by predators. In nature, bacteria living together with Acanthamoeba compete for nutrition and space. They develop strategies to combat each other. Bacterial conditioned media were tested for amoebicidal and amoebistatic abilities. P. aeruginosa, Str. pneumoniae and H. influenzae conditioned media demonstrated amoebistatic and amoebicidal effects. Acanthamoeba conditioned media exhibited bactericidal efficacy only against Str. pneumoniae. Magainin II in combination with silver nitrate or vancomycin/ciprofloxacin was successful in eradicating bacteria. It was essential to perform experiments in environment similar to CF lung. For this purpose artificial sputum medium was prepared. All bacteria and Acanthamoeba were able to grow in it. Results revealed sub-lethal concentrations of magainin II inhibited bacterial biofilm formation in artificial medium.