Short, Synthetic Cationic Peptides Have Antibacterial Activity against Mycobacterium smegmatis by Forming Pores in Membrane and Synergizing with Antibiotics

Abstract

Multicellular organisms are constantly exposed to a multitude of pathogenic microbes. Infection is inhibited in vivo by the innate and adaptive immune system. Mycobacterium species have emerged that are resistant to most antibiotics. We identified several naturally occurring cationic antimicrobial peptides that were active at low micromolar concentrations against Mycobacterium smegmatis. Human-derived cathelicidin LL-37 is well characterized and studied against M. smegmatis; we compared LL-37 with Chinese cobra-derived cathelicidin NA-CATH and mouse cathelicidin (mCRAMP). Two synthetic 11-residue peptides (ATRA-1A and ATRA-2) containing variations of a repeated motif within NA-CATH were tested for their activity against M. smegmatis along with a short synthetic peptide derivative from the human beta-defensin hBD3 (hBD3-Pep4). We hypothesized that these smaller synthetic peptides may demonstrate antimicrobial effectiveness with shorter length (and at less cost), making them strong potential candidates for development into broad-spectrum antimicrobial compounds or use in combination with antibiotics. These peptides have antimicrobial activity with EC50 ranging from 0.05 to 1.88 μg/mL against Mycobacterium smegmatis. The ATRA-1A short peptide was found to be the most effective antimicrobial peptide (AMP) (EC50 = 0.05 μg/mL). High bactericidal activity correlated with bacterial membrane depolarization and permeabilization activities. The efficacy of the peptides was further analyzed through Minimal Inhibitory Concentration (MIC) assays. The MICs were determined by the microdilution method. The peptide mCRAMP showed the best MIC activity at 15.6 μg/mL. Neither of the effective short synthetic peptides demonstrated synergy with the antibiotic rifampicin, although both demonstrated synergy with the cyclic peptide antibiotic polymyxin B. The peptides LL-37 and mCRAMP displayed synergism with rifampicin in MIC assays, whereas antibiotic polymyxin B displayed synergism with LL-37, ATRA-1A, and hBD3-Pep4. In further studies, polymyxin B synergized with LL-37, ATRA-1A, and hBD3-Pep4 while Rifampicin synergized with LL-37 and mCRAMP for intracellular killing of mycobacteria residing inside macrophages. These studies provide the foundation for the potential development of synthetic cationic antimicrobial peptides with activity against mycobacteria.

Antimicrobial Activity of Protein Fraction from Naja ashei Venom against Staphylococcus epidermidis

“…In other cases, the second component somehow strengthens the action of the first component by facilitating access to the molecular target. This is how antibacterial peptides work, which cause permeabilization of the membrane, which in turn causes the release of antibiotics into the periplasm and cytoplasm of bacterial cells [42,[51][52][53][54][55][56]. It is also known that Staphylococci have an efflux pump, which determines drug resistance to a large extent because it is responsible for removing from cells various types of substances, which are unnecessary for metabolism or harmful to cells [57].…”

Bactenecin and Its Three Improved Derivatives for Enhancement of Antibacterial Activity Against Escherichia coli

“…Previous studies on synthetic forms of bactenecin reported the increased efficiency of this peptide with changes in its sequence and structure (25,26). The MIC test with E. coli (ATCC 25922) was conducted here for each peptide (14)(15)(16)(17)(18)(19).…”

Hitchhiking with Nature: Snake Venom Peptides to Fight Cancer and Superbugs

“…• Na-CATH: this CATH from the venom gland of the Chinese cobra N. atra [76] also demonstrated powerful, salt-resistant, antimicrobial activity against Gram-positive and Gram-negative bacteria, including Francisella novicida (the non-virulent strain in humans related to Francisella tularensis, the causative agent of tularemia) [98], E.coli, Aggregatibacter actinomycetemcomitans, Bacillus cereus [99], P. aeruginosa [100], and S. aureus [101] at low concentrations (EC 50 < 3 µg/mL). Na-CATH is also active against Mycobacterium smegmatis [102], Burkholderia thailandensis (closely related to B. pseudomallei, the causative agent of melioidosis) [103] and Bacillus anthracis (anthrax) [104]. The last two strains are of particular relevance due to their potential use as biological weapons.…”

“…In addition, Na-CATH is not only active against planktonic bacteria, but also inhibits S. aureus and B. thailandensis biofilm formation [101,102], while inducing minimal hemolysis (<2% at 100 µg/mL) [99]. However, Na-CATH did not inhibit Pseudomonas biofilm formation [100].…”