Current research on defensins against Staphylococcus aureus

Defensins are small cysteine-rich cationic peptides, found in a broad range of species in animal and plant kingdoms. It has been identified three sub-classes of defensins (e.g. α, β, and θ) which have conserved six cysteine signature (1). They play an important role as host defense peptides with activity against bacteria, fungi and many enveloped and nonenveloped viruses. They are usually between 18 and 45 amino acids long, with three or four highly conserved disulfide bonds. In the last two years, several studies have been done on defensins against Staphylococcus aureus

Human defensins have been nicely reviewed by Fruitwala et al (2019)(2). In brief, human defensins are members of a large family of antimicrobial cationic peptides of approximately 30 amino acids. They can be classified according to their disulfide bond linkages into α-, and β- defensins. Most members of the same family share a similar structure. They play an important function in the immune response against pathogens due to their broad-spectrum antimicrobial activity. Human defensins are produced mainly by neutrophils and epithelial cells and to a lesser extent by monocytes, macrophages, dendritic cells, and lymphocytes.

Current works in this area focus on the human defensins HBD2-3, HNP-1, and Phd1-3. In a recent study, Bolatchiev (2020) investigated the effect of HNP-1, hBD-1, and hBD-3, and their combination with rifampicin or amikacin against clinical isolates of S. aureus (3). The most effective antimicrobial combination against S.aureus was hBD-3 with rifampicin, having a minimum inhibitory concentration (MIC) of 1 compared with microbial MIC values of 4 and 8 of HNP-1 and hBD-1, respectively. These antimicrobial peptides demonstrated a synergistic impact on the majority of isolates studied when they are used together with rifampicin and amikacin. This can be attributed to the ability of human defensins to enhance the permeability of bacterial membranes and by extent the access of antibiotics into the periplasm and cytoplasm of bacterial cells (4). Scudiero et al. (2020) reviewed the importance of antimicrobial peptides in the pathogenesis of S. aureus cutaneous diseases including psoriasis and atopic dermatitis. Expression of  HBD2 and 3 are downregulated in patients with atopic dermatitis while they are upregulated in patients with psoriasis and rosacea. The decreased expression could be explained by the activation of cytokines which repress the expression of antimicrobial peptides (5).

It has been demonstrated that some defensins (e.g. human enteric defensin 5) inhibit the pro-inflammatory cytokine TNF-α in LPS-stimulated macrophages by blocking the interaction between LPS and LPS-binding protein (6). However, the antibacterial activity of HBD 2-3, Phd1-3, and MPhd1-3 against Staphylococcus aureus is decreased in the presence of LPS. It seems that the native defensin amino acid sequence or structure is also not essential for this, however, the cationic charges are necessary for binding to LPS (7).

Rodriguez et al. (2019) identified two novel putative peptides (hBDconsensus and hBD10) that seemed to belong to the human β-defensin family showed antimicrobial activity against Staphylococcus aureus (8)

Plant defensins are an extensive family of small, highly stable, and cysteine-rich peptides with a conserved tertiary structure that consists of a triple-stranded antiparallel b-sheet and a-helix (9). Al Kashgry and colleagues demonstrated the recombinant peptide MzDef, a defensin from Maize, could be utilized as a potential antimicrobial peptide against different species of fungi and bacteria including S. aureus (10). In another study, it was investigated in vitro the broad-spectrum activities of recombinant defensin (SDmod) which sequence is based on the sequence of the gene sd2 from Sunflower. Their results indicated that Sdmod can absorb heavy metals like cadmium and zinc and have antibacterial activity against S. aureus (11)

Insect defensins compose approximately 40 amino acid (AA) residues, with an N-terminal loop, an α-helical fragment, followed by an anti-parallel β-sheet cross-linked by three disulfide bonds (CSαβ)(12). They have antimicrobial activity against several gram-positive bacteria, including methicillin-resistant S. aureus (MRSA) (13).

DLP4 is an insect defensin from the hemolymph of Hermetia illucens larvae was used as a starting template for the rational design of a novel class of “CSαβ” antimicrobial peptide. Of the designed peptides, ID13 demonstrated to have reduced cytotoxicity and antibacterial activity against S. aureus. This peptide could penetrate and damage the cell membrane of S. aureus, which would lead to increased potassium ion leakage. Measurement of the number of viable bacteria remaining at various times after exposure to the peptides in vitro showed that ID13 killed more than 99 percent of S. aureus in less than an hour. 10 mg/kg ID13 reduced the number of  S. aureus to 1.8 log₁₀ (CFU/g) (CFU: colony-forming units) in infected mouse thigh muscles and decreased the levels of TNF-α, IL-6, and IL-10 levels. These results suggest that ID13 has the potential to be a promising antimicrobial agent for therapeutic purposes (14).

One of the humoral effector molecules in ticks are defensins, which are a family of small peptides with a conserved γ-core motif, which is essential for their antimicrobial activity (15). A novel defensin-like peptide (Ds-defensin) from ticks showed potent antimicrobial activity against several gram-positive bacteria including Staphylococcus aureus (16). It seems that Ds-defensin formed pores after a 30-min exposure to the Ds-defensin peptide. This finding provides future perspectives for the development of antimicrobial therapeutic drugs. Ixodes holocyclus is a species of Australian tick that produces defensins with a preserved γ-core motif that is critical for their antimicrobial activity. Its transcriptome analysis revealed the presence of five defensins, of which two had antimicrobial activity against S. aureus. This study found a multigene defensin family in I. holocyclus with wide antimicrobial activity (15).

Big defensins are antimicrobial peptides related to β-defensins with a unique structure usually described in marine organisms, mainly mollusks. They composed of a highly hydrophobic N-terminal region and a cationic C-terminal region containing six cysteine residues involved in three internal disulfide bridges (17). Loth et al. (2019) tested the bactericidal activity of the defensins BigDef1 from oyster Crassostrea gigas against multidrug-resistant human clinical isolates of Staphylococcus aureus. Their finding indicates that the ancestral N-terminal domain confers salt-stable antimicrobial activity to the β defensin-like domain. It triggers the Cg-BigDef1 assembly, which captures and kills bacteria. It is important to note that BigDef1 does not appear to be cytotoxic to mammalian cells (18).

Keywords: defensin, antimicrobial peptide, staphylococcus aureus, human defensin, plant defensin.