In vitro activity of novel in silico‐developed antimicrobial peptides against a panel of bacterial pathogens

Abstract

Antimicrobial-peptide-based therapies could represent a reliable alternative to overcome antibiotic resistance, as they offer potential advantages such as rapid microbicidal activity and multiple activities against a broad spectrum of bacterial pathogens. Three synthetic antimicrobial peptides (AMPs), AMP72, AMP126, and also AMP2041, designed by using ad hoc screening software developed in house, were synthesized and tested against nine reference strains. The peptides showed a partial β-sheet structure in 10-mM phosphate buffer. Low cytolytic activity towards both human cell lines (epithelial, endothelial, and fibroblast) and sheep erythrocytes was observed for all peptides. The antimicrobial activity was dose dependent with a minimum bactericidal concentration (MBC) ranging from 0.17 to 10.12 μM (0.4-18.5 µg/ml) for Gram-negative and 0.94 to 20.65 μM (1.72-46.5 µg/ml) for Gram-positive bacteria. Interestingly, in high-salt environment, the antibacterial activity was generally maintained for Gram-negative bacteria. All peptides achieved complete bacterial killing in 20 min or less against Gram-negative bacteria. A linear time-dependent membrane permeabilization was observed for the tested peptides at 12.5 µg/ml. In a medium containing Mg²⁺ and Ca²⁺, the peptide combination with EDTA restores the antimicrobial activity particularly for AMP2041. Moreover, in combination with anti-infective agents (quinolones or aminoglycosides) known to bind divalent cation, AMP126 and AMP2041 showed additive activity in comparison with colistin. Our results suggest the following: (i) there is excellent activity against Gram-negative bacteria, (ii) there is low cytolytic activity, (iii) the presence of a chelating agent restores the antimicrobial activity in a medium containing Mg²⁺ and Ca²⁺, and (iv) the MBC value of the combination AMPs-conventional antibiotics was lower than the MBC of single agents alone.

In vitro antimicrobial activity of a gel containing antimicrobial peptide AMP2041, chlorhexidine digluconate and Tris‐EDTA on clinical isolates of Pseudomonas aeruginosa from canine otitis

“…The antimicrobial activity of AMP2041 has been reported previously 16,17 on different bacterial and fungal species and in particular, on different PA clinical isolates and ATCC strain 27853. In the present study, the antimicrobial activity of the undiluted and diluted product was evaluated.…”

“…Maintenance of activity at diluted concentrations is an important feature for antimicrobial agents used on the skin and in the ear canals. 19,20 A prior study of AMP2041 showed bactericidal effect against PA ATCC strain 27853 at an MBC value of 2.14 lg/mL after 2 h of contact time, 17 whereas reported MICs for chlorhexidine digluconate against PA range from 8 to >70 lg/mL. 21 Although Tris-EDTA alone is known to be bacteriostatic, its addition to topical products significantly lowers the MIC and MBC values for several antimicrobials.…”

“…Both compounds are strongly cationic in nature and bind strongly to bacteria, causing surface membrane damage. 17 Cases of chlorhexidine resistance have been reported over the past 30 years. 23 According to the Committee for Veterinary Medicinal Products of the European Agency for the Evaluation of Medicinal Products, 24 chlorhexidine is bactericidal above 100 lg/mL.…”

Strategies employed in the design and optimization of synthetic antimicrobial peptide amphiphiles with enhanced therapeutic potentials

“…A further 44 variants of the best candidate D28 were created through systematic charge and hydrophobicity mutations, of which 18 variants demonstrated improved activities against E. coli, B. cereus or S. aureus. In silico approaches that do not rely on the use of an original template sequence have also been reported [68][69][70][71][72]. Hancock's group reported a powerful strategy involving the combination of quantitative structural-activity relationships (QSAR) with machine learning techniques to predict the antimicrobial activities of 100, 000 virtual peptides [68][69][70].…”

Activity of AMP2041 against human and animal multidrug resistant Pseudomonas aeruginosa clinical isolates

“…The activity of AMP2041 against P. aeruginosa ATCC 27853 was also demonstrated, as well as additivity in combination with levofloxacin [9]. We hypothesized that the reported antimicrobial activity derived from a bacterial membrane damage, but a direct membrane damage was not previously investigated for P. aeruginosa .…”